U.S. patent number 10,385,313 [Application Number 15/026,313] was granted by the patent office on 2019-08-20 for human ipsc-derived vascular-related and hematopoetic cells for therapies and toxicology/drug screenings.
This patent grant is currently assigned to The USA, as represented by the Secretary, Department of Health and Human Services. The grantee listed for this patent is THE UNITED STATES OF AMERICA, AS REPRESENTED BY THE SECRETARY, DEPARTMENT OF HEALTH AND HUMAN SERVICES, THE UNITED STATES OF AMERICA, AS REPRESENTED BY THE SECRETARY, DEPARTMENT OF HEALTH AND HUMAN SERVICES. Invention is credited to Manfred Boehm, Guibin Chen, Andre Larochelle, Mahendra Rao.
United States Patent |
10,385,313 |
Boehm , et al. |
August 20, 2019 |
Human IPSC-derived vascular-related and hematopoetic cells for
therapies and toxicology/drug screenings
Abstract
Described herein are cells, cell culture methods, and cell
culture media compositions useful for producing and maintaining
iPSC-derived cell lines that are of higher purity and maintain cell
type integrity better than current iPSC-derived cell lines. Also
disclosed are methods of using the described cells and media, such
as therapeutic methods of use for the described cells. The
described cells include iPSC-derived mesodermal precursor cells
(MPC), which itself may differentiate into at least four different
cell types. When cultured under appropriate conditions, the
mesodermal precursor cells can be used to produce hematopoietic
stem cells (HSC), mesenchymal stem cells (MSC), smooth muscle cells
(SMC), or unlimited functional endothelial cells (UFEC). One
characteristic that makes the described cells desirable is that
they can be maintained in culture for a number of days, or
passages, without changing phenotype through differentiation.
Inventors: |
Boehm; Manfred (Bethesda,
MD), Chen; Guibin (Ellicott City, MD), Rao; Mahendra
(Timonium, MD), Larochelle; Andre (Bethesda, MD) |
Applicant: |
Name |
City |
State |
Country |
Type |
THE UNITED STATES OF AMERICA, AS REPRESENTED BY THE SECRETARY,
DEPARTMENT OF HEALTH AND HUMAN SERVICES |
Washington |
DC |
US |
|
|
Assignee: |
The USA, as represented by the
Secretary, Department of Health and Human Services (Bethesda,
MD)
|
Family
ID: |
51842809 |
Appl.
No.: |
15/026,313 |
Filed: |
October 1, 2014 |
PCT
Filed: |
October 01, 2014 |
PCT No.: |
PCT/US2014/058583 |
371(c)(1),(2),(4) Date: |
March 31, 2016 |
PCT
Pub. No.: |
WO2015/050963 |
PCT
Pub. Date: |
April 09, 2015 |
Prior Publication Data
|
|
|
|
Document
Identifier |
Publication Date |
|
US 20160222348 A1 |
Aug 4, 2016 |
|
Related U.S. Patent Documents
|
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
Issue Date |
|
|
61885209 |
Oct 1, 2013 |
|
|
|
|
Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C12N
5/0607 (20130101); C12N 5/0662 (20130101); C12N
5/0018 (20130101); C12N 5/0661 (20130101); C12N
5/0696 (20130101); C12N 5/0647 (20130101); C12N
2501/115 (20130101); C12N 2501/26 (20130101); C12N
2506/45 (20130101); C12N 2500/44 (20130101); C12N
2500/90 (20130101); C12N 2501/165 (20130101); C12N
2501/155 (20130101); C12N 2506/03 (20130101); C12N
2500/05 (20130101); C12N 2500/42 (20130101); C12N
2500/30 (20130101); C12N 2500/36 (20130101) |
Current International
Class: |
C12N
5/00 (20060101); C12N 5/074 (20100101); C12N
5/077 (20100101); C12N 5/0775 (20100101); C12N
5/0789 (20100101) |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
|
|
|
|
|
|
|
WO 2010/099539 |
|
Sep 2010 |
|
WO |
|
WO 2014/100779 |
|
Jun 2014 |
|
WO |
|
Other References
International Search Report and Written Opinion prepared by the
European Patent Office dated Nov. 27, 2014, for International
Application No. PCT/US2014/058583. cited by applicant .
B.W. Smith et al: "The aryl hydrocarbon receptor directs
hematopoietic progenitor cell expansion and differentiation",
Blood, vol. 122, No. 3, May 30, 2013, pp. 376-385. cited by
applicant .
Wagey et al. "Isolation, Enumeration, and Expansion of Human
Mesenchymal Stem Cells in Culture," Basic Cell Culture Protocols,
Methods in Molecular Biology, 2013, vol. 946, Chapter 20, pp.
315-334. cited by applicant .
Wang et al. "Derivation of Smooth Muscle Cells with Neural Crest
Origin from Human Induced Pluripotent Stem Cells," Cells Tissues
Organs, 2012, vol. 195, pp. 5-14. cited by applicant .
Official Action for Canada Patent Application No. 2,925,774, dated
Jan. 24, 2017 5 pages. cited by applicant .
Official Action for Canada Patent Application No. 2,925,774, dated
Dec. 6, 2017 7 pages. cited by applicant .
Official Action for European Patent Application No. 14790863.6,
dated Jan. 23, 2017 5 pages. cited by applicant .
Official Action for European Patent Application No. 14790863.6,
dated Nov. 16, 2017 4 pages. cited by applicant .
Zanetta et al. "Expression of von Willebrand factor, an endothelial
cell marker, is up-regulated by angiogenesis factors: A potential
method for objective assessment of tumor angiogenesis,"
International Journal of Cancer, Jan. 2000, vol. 85, No. 2, pp.
281-288. cited by applicant .
Official Action for Canada Patent Application No. 2,925,774, dated
Aug. 20, 2018 6 pages. cited by applicant .
Official Action for European Patent Application No. 14790863.6,
dated Sep. 13, 2018 4 pages. cited by applicant.
|
Primary Examiner: Lankford; Blaine
Attorney, Agent or Firm: Sheridan Ross PC
Parent Case Text
CROSS-REFERENCE TO RELATED APPLICATIONS
This application is a national stage application under 35 U.S.C.
371 and claims the benefit of PCT Application No. PCT/US2014/058583
having an international filing date of Oct. 1, 2014, which designed
the United States, which PCT application claimed the benefit of
U.S. Provisional Appl. No. 61/885,209, filed Oct. 1, 2013, the
disclosures of each of which are incorporated by reference herein
in their entirety.
Claims
What is claimed is:
1. A cell culture medium consisting essentially of: Iscove's
modified Dulbecco's medium (IMDM), Ham's F-12 Nutrient Mix, with
L-alanyl-L-glutamine additive, Albumin, a-monothioglycerol,
protein-free hybridoma mixture II, L-ascorbic acid 2-phosphate,
L-alanyl-L-glutamine, Antibiotic,
insulin-transferrin-selenium-ethanolamine supplement, bone
morphogenic protein 4, vascular endothelial growth factor, and
basic fibroblast growth factor.
2. The cell culture medium of claim 1, further comprising
cholesterol lipids.
3. The cell culture medium of claim 1, wherein the antibiotic is
selected form the group consisting of penicillin, streptomycin and
a mixture of penicillin and streptomycin.
4. The cell culture medium of claim 1, wherein the concentration of
albumin, is about 5 mg/ml.
5. The cell culture medium of claim 1, wherein the concentration of
monothioglycerol is from about 350 to about 450 .mu.M.
6. The cell culture medium of claim 1, wherein the concentration of
L-ascorbic acid 2-phosphate is about 50 .mu.g/ml.
7. The cell culture medium of claim 1, wherein the concentration of
L-alanyl-L-glutamine is about 1 mM to about 2 mM.
8. The cell culture medium of claim 1, wherein the concentration of
bone morphogenic protein 4 is about 10 ng/ml.
9. The cell culture medium of claim 1, wherein the concentration of
vascular endothelial growth factor is about 10 ng/ml.
10. The cell culture medium of claim 1, wherein the concentration
of basic fibroblast growth factor is about 10 ng/ml.
Description
BACKGROUND
The use of human stem cells for clinical purposes has become a
subject of increasing interest in recent years. This interest has
only intensified in the wake of the more recent discoveries that
human somatic cells can be induced to form pluripotent stem cells
when certain transcription factors are overexpressed. Human induced
pluripotent stem cells (hiPSCs) can be generated in a variety of
ways, such as reprogramming somatic cells by the expression of four
transcription factors. The hiPSCs exhibit similar properties to
human embryonic stem cells (hESCs), including the ability to
self-renew and differentiate into all three embryonic germ layers:
ectoderm, endoderm, or mesoderm. Additionally, hiPSCs overcome
ethical concerns, relative to generating hESCs from human embryos,
because no embryonic cells are needed to form hiPSCs. Human iPSCs
can be induced into any cell type and, since they can be maintained
over many passages, they can serve as an almost unlimited source to
generate cells from any given person. These properties make
iPSC-derived cells a valuable product for cell therapies and
toxicology or pharmaceutical high throughput screens. However,
therapeutic and commercial uses of iPSC-derived cell products are
hampered by low quantities and cell culture impurity due to
limitations with current methods for producing and maintaining
these cells.
SUMMARY
Described herein are cells, cell culture methods, and cell culture
media compositions useful for producing and maintaining
iPSC-derived cell lines that are of higher purity and maintain cell
type integrity better than current iPSC-derived cell lines. Also
disclosed are methods of using the described cells and media.
One aspect of the present disclosure is an iPSC-derived mesodermal
precursor cell (MPC) line, positive for CD34 and CD31 expression,
that may be used to produce at least four different cell types.
When cultured under appropriate conditions, these mesodermal
precursor cells can be used to produce hematopoietic stem cells
(HSC), mesenchymal stem cells (MSC), smooth muscle cells (SMC), or
unlimited functional endothelial cells (UFEC). One characteristic
that makes the mesodermal precursor cells described herein
desirable is that these cells can be maintained in culture for a
number of days, or passages, without changing phenotype through
differentiation.
The HSCs described herein can be produced by culturing the
described MPCs in medium and under conditions known to cause cells
to differentiate into HSCs. The described HSCs may be characterized
by the expression of CD34, CD31, and CD45. Another characteristic
of the described HSCs is that they have the ability to reconstitute
the hematopoietic system of an irradiated subject, such as a mouse.
The described HSCs also have the ability to maintain their
phenotype for extended periods without differentiating, when
maintained under appropriate conditions.
The described MPCs are also capable of giving rise to UFECs when
cultured under conditions known to allow for differentiation into
cells of an endothelial lineage. The described UFECs can be
characterized by the expression of CD31, vWF, and CD144. In
addition, these cells can mediate the uptake of acetylated low
density lipoproteins (LDL). Furthermore, the UFECs produced using
the methods and cells described herein have the ability to form
vascular-like structures in vitro, a hallmark of endothelial cell
progenitors.
Another cell type capable of being produced by the MPCs described
herein are MSCs. The MSCs described herein can be characterized by
the expression of CD90, CD73, and CD105 in the absence of CD31 and
CD45. These cells can also differentiate in vivo or in vitro into a
number of different cell types, including adipocytes, osteoblasts,
myocytes, or chondrocytes, when cultured under conditions known to
cause progenitor cells to differentiate into the respective cell
type. The described MSCs also have the ability to maintain their
phenotype for extended periods without differentiating, when
maintained under appropriate conditions.
The described MPCs may also be used to generate smooth muscle cells
according to the methods described herein. For example, the
described MPCs can differentiate into smooth muscle cells when
cultured under conditions known to cause progenitor cells to
differentiate into SMCs. The described SMCs are characterized by
the expression of .alpha.-SMA, calponin, and SM22. The described
SMCs also have the ability to maintain their phenotype for extended
periods without differentiating, when maintained under appropriate
conditions.
In some embodiments the cell types described herein may be
generated using the cells of a subject to produce autologous cells
using the cell production methods described herein. The
differentiated autologous cells can then be administered to the
subject for therapeutic purposes.
Described herein are various tissue culture media that may be used
to produce the cells characterized in the present disclosure. In
some embodiments the medium formulation includes a mixture of
Iscove's modified Dulbecco's medium (IMDM), Ham's F-12 Nutrient
Mix, with L-alanyl-L-glutamine additive, albumin,
.alpha.-monothioglycerol, protein-free hybridoma mixture II,
L-ascorbic acid 2-phosphate, L-alanyl-L-glutamine, antibiotic,
cholesterol lipids, insulin-transferrin-selenium-ethanolamine
supplement, bone morphogenic protein 4, vascular endothelial growth
factor, and basic fibroblast growth factor. In some embodiments the
medium formulation includes a mixture of Iscove's modified
Dulbecco's medium (IMDM), Ham's F-12 Nutrient Mix, with
L-alanyl-L-glutamine additive, albumin, .alpha.-monothioglycerol,
protein-free hybridoma mixture II, L-ascorbic acid 2-phosphate,
L-alanyl-L-glutamine, antibiotic,
insulin-transferrin-selenium-ethanolamine supplement, bone
morphogenic protein 4, vascular endothelial growth factor, and
basic fibroblast growth factor. Where the described components are,
or include, proteins, such as albumin, bone morphogenic protein 4,
vascular endothelial growth factor, or basic fibroblast growth
factor, the additive may have an amino acid sequence corresponding
to the human form of the protein. The media described herein may
further include additives such as stem cell factor, Flt-3 ligand,
or thrombopoietin, any of which may be derived from, or correspond
to, the human form of the protein. While any of the media additives
described herein may be derived from, or correspond to, the human
form, this is not necessarily required and additives that are
derived from, or correspond to, those of other mammals may also be
acceptable.
The cells, media, methods of producing the described cells, and
related methods of use are more fully discussed herein.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1. Characterization of hiPSC. Cells were derived and expanded
in feeder free and defined medium. FIG. 1A shows analysis of
pluripotency markers (SSEA-4, Tra-1-60, Oct4, and Nanog) on iPSCs
derived from fibroblasts by immunofluorescence staining. FIG. 1B
shows analysis of pluripotency markers (Oct3/4, Nanog, and Sox2) on
iPSCs derived from fibroblasts by real time RT-PCR. FIG. 1C
illustrates a karyotyping assay on hiPSCs by G-band. FIG. 1D
illustrates bisulfate sequencing analysis of methylation in Nanog
and Oct4 promoter. FIG. 1E illustrates histologic detection in
teratoma formation by Hematoxylin and eosin staining shown that
hiPSCs are able to successfully differentiate into all three germ
layers in vitro.
FIG. 2. Depiction of a step-wise protocol to drive hiPSCs into
mesoderm precursor cells using feeder-free and chemically defined
cell culture media, following by specifically lineage commitment
and maturation.
FIG. 3. Analysis of gene expression level in cells of
undifferentiated and differentiating cells by RT-PCR.
FIG. 4. Kinetic analysis of mesoderm precursor cells from
pluripotent stem cells by FACS, using CD31 and CD34 as markers.
FIG. 4A provides a representative FACS diagram of CD34 and CD31
expression in cells differentiated from normal hiPSCs. FIG. 4B
illustrates the timing of appearance of mesoderm precursors.
FIG. 5. Validation of hiPSC-derived endothelial cells (UFECs)
through mesoderm precursors. FIG. 5A demonstrates that
hiPSC-derived UFECs generated through mesoderm precursors show
typical endothelial cell morphology and express multiple EC markers
(CD31, vWF, and CD144). FIG. 5B illustrates an in vitro functional
angiogenesis assay for hiPSC-derived UFECs. hiPSC-derived UFECs
formed vascular tube-like structures on Matrigel.TM..
FIG. 6. Generation of Mesenchymal stem cells (iMSC) and smooth
muscle cells (iSMCs) from hiPSCs through mesoderm precursor cells.
FIG. 6A. The representative phenotype of iMSC analyzed by FACS.
FIG. 6B. iMSC derived from hiPSCs through mesoderm precursor cells
after osteogenic differentiation in vitro. FIG. 6C. Differentiation
of hiPSC-derived mesoderm precursors into smooth muscle cells. The
expression of SMC markers .alpha.-SMA, calponin, and SM22 was
analyzed by FACS (left) and immunofluorescence staining (the cell
nuclei were stained with DAPI (blue)).
FIG. 7. Generation of hematopoietic lineage cells in suspension
from hiPSCs through mesoderm precursor cells.
FIG. 8. Generation of hematopoietic lineage cells in suspension
from normal hiPSCs. FIG. 8A. The maximum number of CD45-CD34+CD31+
cells generated from iPSCs peaks at day 12 of culture (left); FIG.
8B. Timing of appearance of human CD34 and CD45. Cumulative number
of CD45+CD34+CD31+ cells generated from the differentiation of
iPSCs (right). FIG. 8C. Generation of colony-forming units (CFU) 14
days after the incubation of day 12 hiPSC-derived suspension cells
in semisolid clonogenic culture. Left: number of CFU per
1.times.10.sup.3 cells plated. Right: Representative erythroid
(BFU-E), myeloid (CFU-GM) and mixed (CFU-GEMM) colonies.
FIG. 9. Comparison of the total number of supernatant cells
generated during iPSC differentiation.
FIG. 10. MDM1 produces more CD45+ HSCs at day 10 of culture
compared to MDM.
FIG. 11. Modified iPSC differentiation protocol to favor
hematopoietic differentiation.
DETAILED DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS
Described herein are cells, cell culture methods, and cell culture
media compositions useful for producing and maintaining
iPSC-derived cell lines that are of higher purity and maintain cell
type integrity better than current iPSC-derived cell lines. Also
disclosed are methods of using the described cells and media.
Various terms relating to aspects of the description are used
throughout the specification and claims. Such terms are to be given
their ordinary meaning in the art unless otherwise indicated. Other
specifically defined terms are to be construed in a manner
consistent with the definitions provided herein.
As used in this specification and the appended claims, the singular
forms "a," "an," and "the" include plural referents unless the
content clearly dictates otherwise. Thus, for example, reference to
"a cell" includes a combination of two or more cells, and the
like.
The term "about" as used herein when referring to a measurable
value such as an amount, a temporal duration, and the like, is
meant to encompass variations of up to .+-.10% from the specified
value, as such variations are appropriate to perform the disclosed
methods. Unless otherwise indicated, all numbers expressing
quantities of ingredients, properties such as molecular weight,
reaction conditions, and so forth used in the specification and
claims are to be understood as being modified in all instances by
the term "about." Accordingly, unless indicated to the contrary,
the numerical parameters set forth in the following specification
and attached claims are approximations that may vary depending upon
the desired properties sought to be obtained by the present
invention. At the very least, and not as an attempt to limit the
application of the doctrine of equivalents to the scope of the
claims, each numerical parameter should at least be construed in
light of the number of reported significant digits and by applying
ordinary rounding techniques.
Notwithstanding that the numerical ranges and parameters setting
forth the broad scope of the invention are approximations, the
numerical values set forth in the specific examples are reported as
precisely as possible. Any numerical values, however, inherently
contain certain errors necessarily resulting from the standard
deviation found in their respective testing measurements.
The terms "treating," "treatment," and "therapy" refer to any
success or indicia of success in the attenuation or amelioration of
an injury, pathology or condition, including any objective or
subjective parameter such as abatement, remission, diminishing of
symptoms or making the condition more tolerable to the patient,
slowing in the rate of degeneration or decline, making the final
point of degeneration less debilitating, improving a subject's
physical or mental well-being, or prolonging the length of
survival. The treatment may be assessed by objective or subjective
parameters; including the results of a physical examination,
neurological examination, or psychiatric evaluations.
The term "subject" refers to human and non-human animals, including
all vertebrates, e.g., mammals and non-mammals, such as non-human
primates, mice, rabbits, sheep, dogs, cats, horses, cows, chickens,
amphibians, and reptiles. In many embodiments of the described
methods, the subject is a human.
The terms "induced pluripotent stem cell," "iPSC," and variants
thereof (e.g., hiPSC) refer to pluripotent stem cells made
recombinantly from a somatic cell by expressing certain
transcription factors in the somatic cell, such that it becomes
pluripotent. The transcription factors expressed in the somatic
cell to induce this change are well known in the art; therefore,
these terms are not limited to the particular embodiments of such
cells described herein. Nor is the scope of these terms limited by
the method used to overexpress these factors, be it transduction,
transformation, or another means of exogenous gene expression.
The terms "MDM2" and "MDM+" are used interchangeably through this
and previous applications.
Cell Culture Media
Provided herein are various cell culture media for culturing cells
described herein and, in some cases, for promoting or allowing for
cell differentiation. One type of medium described herein is basic
mesoderm differentiation medium (MDM). In some embodiments MDM can
serve as a cell culture medium. In other embodiments MDM can be
used as a cell differentiation medium. In still further embodiments
MDM may be both a culture medium and a cell differentiation medium.
As those skilled in the art will understand, MDM can have many
embodiments depending on the concentration of the components used
in a given formulation. The primary components of MDM are provided
in Table 1, while Table 2 provides a particular embodiment of the
medium.
TABLE-US-00001 TABLE 1 Primary components of MDM Ingredient Amount
Iscove's Modified Dulbecco's Medium (IMDM) About a 1:1 mixed with
Ham's F-12 Nutrient Mix, with mixture L-alanyl-L-glutamine
(GlutaMax .TM.) additive albumin about 5 mg/ml
.alpha.-monothioglycerol 350-450 .mu.M protein-free hybridoma
mixture II 5% L-ascorbic acid 2-phosphate (GlutaMax .TM.) about 50
.mu.g/ml L-alanyl-L-glutamine about 1 mM to about 2 mM Antibiotic
(penicillin/streptomycin mix) 50 units pen. 50 mg strep.
cholesterol lipids about 1 .mu.g/ml to about 4 .mu.g/ml
insulin-transferrin-selenium-ethanolamine about 0.5% to supplement
(Table 3) about 3% of total mixture bone morphogenic protein 4
about 10 ng/ml vascular endothelial growth factor about 10 ng/ml
basic fibroblast growth factor About 10 to about 25 ng/ml
TABLE-US-00002 TABLE 2 A single embodiment of MDM media, as
exemplified in the Examples, below. Ingredient Amount Iscove's
Modified Dulbecco's Medium (IMDM, 1:1 mixture Invitrogen, Catalog#:
21056-023) mixed with Ham's F-12 Nutrient Mix, with
L-alanyl-L-glutamine (GlutaMax .TM.) additive (Invitrogen,
Catalog#, 31765- 035) Albucult .TM. 5 mg/ml
.alpha.-monothioglycerol (Sigma-Aldrich, Catalog#: M6145) 3.9 .mu.l
per 100 ml protein-free hybridoma mixture II (Invitrogen Catalog#:
5% of total 12040-077) volume L-ascorbic acid 2-phosphate
(Sigma-Aldrich, Catalog#: 50 .mu.g/ml A 8960) L-alanyl-L-glutamine
(GlutaMax .TM.) (2 mM, Invitrogen, 2 mM Catalog#: 35050061)
Antibiotic (Invitrogen, Catalog#: 15140122) 50 units pen. 50 mg
strep. cholesterol lipids (Invitrogen, Catalog#: 12531018) 2.2
.mu.g/ml insulin-transferrin-selenium-ethanolamine supplement 1% of
total (Invitrogen, Catalog#: 515000560) volume bone morphogenic
protein 4 (R&D systems, Catalog#: 10 ng/ml 314-BP-050) vascular
endothelial growth factor (Invitrogen, Catalog#: 10 ng/ml PHC9394)
basic fibroblast growth factor (Pepro Tech, Catalog#: 10 ng/ml
100-18B)
TABLE-US-00003 TABLE 3 Insulin-Transferrin-Selenium-Ethanolamine
media supplement (ITS-X) (100X) Molecular Component Weight (kD)
Concentration (mg/L) mM Insulin 5.8 1,000 172.41379 Transferrin 80
550 6.875 Sodium Selenite 173 0.67 0.003873 Ethanolamine 61 200
3.278688
Also provided herein are various cell culture media for culturing
cells described herein and, in some cases, for promoting or
allowing for cell differentiation. Another type of medium described
herein is basic mesoderm differentiation medium 1 (MDM1). In some
embodiments MDM1 can serve as a cell culture medium. In other
embodiments MDM1 can be used as a cell differentiation medium. In
still further embodiments MDM1 may be both a culture medium and a
cell differentiation medium. As those skilled in the art will
understand, MDM1 can have many embodiments depending on the
concentration of the components used in a given formulation. The
primary components of MDM1 are provided in Table 4, while Table 5
provides a particular embodiment of the medium.
TABLE-US-00004 TABLE 4 Primary components of MDM1 Ingredient Amount
Iscove's Modified Dulbecco's Medium (IMDM) About a 1:1 mixture
mixed with Ham's F-12 Nutrient Mix, with L-alanyl-L-glutamine
(GlutaMax .TM.) additive Albumin About 5 mg/ml
.alpha.-monothioglycerol 350-450 .mu.M Protein-free hybridoma
mixture II 5% L-ascorbic acid 2-phosphate (GlutaMax .TM.) About 50
.mu.g/ml L-alanyl-L-glutamine About 1 mM to 2 mM Antibiotic
(penicillin/streptomycin mix) 50 units pen/50 mg strep
Insulin-transferrin-selenium-ethanolamine About 0.5% to 3%
supplement Bone morphogenic protein 4 About 10 ng/ml Vascular
endothelial growth factor About 10 ng/ml Basic fibroblast growth
factor About 10 to 25 ng/ml
Those skilled in the art will understand that the MDM and MDM1
media described herein can be varied in a variety of ways. For
example, one could perhaps add the individual components of IMDM to
Ham's F-12 Nutrient Mix, with L-alanyl-L-glutamine additive to
arrive at the first ingredient of the medium. Such variations are
contemplated by the inventors and would only be available to the
skilled artisan in view of the detailed description and data
provided by the present application.
A variety of sources of the ingredients listed for MDM and MDM1 may
also be used. In some embodiments albumin may be naturally produced
human albumin. In another embodiment the listed albumin may be
recombinant human albumin, such as Albucult.RTM.. In some
embodiments albumin may be naturally produced bovine albumin. In
another embodiment the listed albumin may be recombinant bovine
albumin. In other embodiments the albumin used in MDM and MDM1 may
be derived from another biological source. For example, the albumin
used in MDM and MDM1 may be natural or recombinant albumin from a
rodent, reptile, avian, canine, feline, primate, lagamorphs,
didelphimorphs, insectivores, carnivores, and the like.
A variety of sources of the cholesterol lipids in MDM may also be
used. In some embodiments the cholesterol lipids may be human
cholesterol lipids. Alternatively, the cholesterol lipids may be
murine cholesterol lipids. In other embodiments the cholesterol
lipids used in MDM may be derived from another biological source.
For example, the cholesterol lipids may be from a rodent, reptile,
avian, canine, feline, primate, lagamorphs, didelphimorphs,
insectivores, carnivores, and the like.
The insulin ingredient used for MDM and MDM1 may also be derived
from a variety of sources. In some embodiments insulin may be
naturally produced human insulin. In another embodiment the listed
insulin may be recombinant human insulin. In other embodiments the
insulin used in MDM and MDM1 may be derived from another biological
source. For example, the insulin may be natural or recombinant
insulin from a rodent, reptile, avian, canine, feline, primate,
lagamorphs, didelphimorphs, insectivores, carnivores, and the
like.
The transferrin ingredient used for MDM and MDM1 may also be
derived from a variety of sources. In some embodiments transferrin
may be naturally produced human transferrin. In another embodiment
the listed transferrin may be recombinant human transferrin. In
other embodiments the transferrin used in MDM and MDM1 may be
derived from another biological source. For example, the
transferrin may be natural or recombinant transferrin from a
rodent, reptile, avian, canine, feline, primate, lagamorphs,
didelphimorphs, insectivores, carnivores, and the like.
The bone morphogenic protein 4 (BMP4) ingredient used for MDM and
MDM1 may also be derived from a variety of sources. In some
embodiments BMP4 may be naturally produced human BMP4. In another
embodiment the listed BMP4 may be recombinant human BMP4. In other
embodiments the BMP4 used in MDM and MDM1 may be derived from
another biological source. For example, the BMP4 may be natural or
recombinant BMP4 from a rodent, reptile, avian, canine, feline,
primate, lagamorphs, didelphimorphs, insectivores, carnivores, and
the like. Other BMPs may be used in place of, or in conjunction
with BMP4. For example, BMP1, BMP2, BMP3, BMP5, BMP7, BMP8a, and
BMP15 are all known to be involved in various aspects of tissue
development or differentiation. Thus, those skilled in the art will
understand, in view of the present disclosure, that the MDM and
MDM1 described herein could also be supplemented with these
proteins, depending on the cells being cultured or the
differentiation path desired. As described above, these BMPs could
also be naturally produced human BMP, recombinant human BMP, or a
natural or recombinant BMP from a rodent, reptile, avian, canine,
feline, primate, lagamorphs, didelphimorphs, insectivores,
carnivores, or other such animal.
The vascular endothelial growth factor (VEGF) ingredient used for
MDM and MDM1 may also be derived from a variety of sources. In some
embodiments VEGF may be naturally produced human VEGF. In another
embodiment the listed VEGF may be recombinant human VEGF. In other
embodiments the VEGF used in MDM and MDM1 may be derived from
another biological source. For example, the VEGF may be natural or
recombinant VEGF from a rodent, reptile, avian, canine, feline,
primate, lagamorphs, didelphimorphs, insectivores, carnivores, and
the like.
The basic fibroblast growth factor (bFGF) ingredient used for MDM
and MDM1 may also be derived from a variety of sources. In some
embodiments bFGF may be naturally produced human bFGF. In another
embodiment the listed bFGF may be recombinant human bFGF. In other
embodiments the bFGF used in MDM and MDM1 may be derived from
another biological source. For example, the bFGF may be natural or
recombinant bFGF from a rodent, reptile, avian, canine, feline,
primate, lagamorphs, didelphimorphs, insectivores, carnivores, and
the like.
The MDM and MDM1 described herein can also be supplemented with
additional ingredients to alter or enhance the function of the
media. For example, in other embodiments the MDM and MDM1 can be
modified to hematopoietic differentiation medium (MDM+ and MDM1+,
respectively) by adding stem cell factor (SCF), Flt-3 ligand, and
thrombopoietin to the medium. In other embodiments the MDM and MDM1
can be modified by adding SCF and thrombopoietin to the medium. In
other embodiments the MDM and MDM1 can be modified by adding SCF
and Flt-3 ligand to the medium. In other embodiments, the MDM and
MDM1 can be modified by adding thrombopoietin and Flt-3 ligand to
the medium. Alternatively, the MDM and MDM1 can be modified by
adding only SCF to the medium. In other embodiments, the MDM and
MDM1 can be modified by adding only Flt-3 ligand to the medium. In
other embodiments, the MDM and MDM1 can be modified by adding only
thrombopoietin to the medium. In particular embodiments, MDM and
MDM1 can be supplemented with recombinant human SCF (Stemcell
Technologies Inc.) at 100 ng/ml, recombinant human Flt-3 ligand
(Stemcell Technologies Inc.) at 100 ng/ml, and recombinant human
thrombopoietin (Stemcell Technologies Inc.) at 100 ng/ml. The MDM+
medium referred to in the examples section provided herein is
formulated by supplementing the MDM from Table 2 with recombinant
human SCF (Stemcell Technologies Inc.) at 100 ng/ml, recombinant
human Flt-3 ligand (Stemcell Technologies Inc.) at 100 ng/ml, and
recombinant human thrombopoietin (Stemcell Technologies Inc.) at
100 ng/ml. The MDM1+ medium referred to in the examples section
provided herein is formulated by supplementing the MDM1 from Table
4 with recombinant human SCF (Stemcell Technologies Inc.) at 50
ng/ml, recombinant human Flt-3 ligand (Stemcell Technologies Inc.)
at 50 ng/ml, and recombinant human thrombopoietin (Stemcell
Technologies Inc.) at 50 ng/ml.
The SCF ingredient used herein may be derived from a variety of
sources. In some embodiments SCF may be naturally produced human
SCF. In another embodiment the listed SCF may be recombinant human
SCF. In some embodiments SCF may be naturally produced murine SCF.
In another embodiment the listed SCF may be recombinant murine SCF.
In other embodiments the SCF used in MDM and MDM1 may be derived
from another biological source. For example, the SCF may be natural
or recombinant SCF from a rodent, reptile, avian, canine, feline,
primate, lagamorphs, didelphimorphs, insectivores, carnivores, and
the like.
The thrombopoietin ingredient described herein may be derived from
a variety of sources. In some embodiments thrombopoietin may be
naturally produced human thrombopoietin. In another embodiment the
listed thrombopoietin may be recombinant human thrombopoietin. In
some embodiments thrombopoietin may be naturally produced murine
thrombopoietin. In another embodiment the listed thrombopoietin may
be recombinant murine thrombopoietin. In other embodiments the
thrombopoietin used in MDM and MDM1 may be derived from another
biological source. For example, the thrombopoietin may be natural
or recombinant thrombopoietin from a rodent, reptile, avian,
canine, feline, primate, lagamorphs, didelphimorphs, insectivores,
carnivores, and the like.
The Flt-3 ligand ingredient used herein may be derived from a
variety of sources. In some embodiments Flt-3 ligand may be
naturally produced human Flt-3 ligand. In another embodiment the
listed Flt-3 ligand may be recombinant human Flt-3 ligand. In some
embodiments Flt-3 ligand may be naturally produced murine Flt-3
ligand. In another embodiment the listed Flt-3 ligand may be
recombinant murine Flt-3 ligand. In other embodiments the Flt-3
ligand used in MDM and MDM1 may be derived from another biological
source. For example, the Flt-3 ligand may be natural or recombinant
Flt-3 ligand from a rodent, reptile, avian, canine, feline,
primate, lagamorphs, didelphimorphs, insectivores, carnivores, and
the like.
Despite the possibilities for having various sources for the
ingredients listed for MDM and MDM1 and MDM- and MDM1-derived
media, there is no requirement that all of the MDM MDM1 protein
components, for example, be derived from the same source. Thus, one
MDM or MDM1 formulation might have transferrin that is obtained
from a natural human source, recombinant human insulin, murine
BMP4, and canine VEGF. This is not to say, however, that all of
these ingredients could not be from the same source in a different
MDM or MDM1 formulation.
One ingredient that may be used in the media described herein is
Protein Free Hybridoma Medium II (PFHM II), which is a serum-free,
protein-free medium that contains no polypeptide growth or
attachment factors, or mediators that may complicate downstream
processing and final product purification.
The media described herein can be supplemented with an antibiotic
to prevent contamination by bacteria. Suitable antibiotics for
tissue culture applications are known in the art. For example,
penicillin and streptomycin, or a combination thereof (pen/strep)
are commonly used. Anti-fungal agents may also be used to prevent
fungal contamination. Suitable anti-fungal agents for tissue
culture applications are known in the art.
The media ingredients listed herein may be used within a range of
concentrations described herein without negatively affecting the
performance of the media. The mixture of Iscove's Modified
Dulbecco's Medium (IMDM) mixed with Ham's F-12 Nutrient Mix with
GlutaMax additive is listed above as being combined in about a 1:1
ratio (i.e., about a 50% to 50% mixture); however, these
ingredients can be mixed in other ratios as well. In some
embodiments, the mixture of Iscove's Modified Dulbecco's Medium
(IMDM) mixed with Ham's F-12 Nutrient Mix with GlutaMax additive is
about 30% to 70% mixture. In some embodiments, the mixture of
Iscove's Modified Dulbecco's Medium (IMDM) mixed with Ham's F-12
Nutrient Mix with GlutaMax additive is about 35% to 65% mixture. In
some embodiments, the mixture of Iscove's Modified Dulbecco's
Medium (IMDM) mixed with Ham's F-12 Nutrient Mix with GlutaMax
additive is about 40% to 60% mixture. In some embodiments, the
mixture of Iscove's Modified Dulbecco's Medium (IMDM) mixed with
Ham's F-12 Nutrient Mix with GlutaMax additive is about 45% to 55%
mixture. In some embodiments, the mixture of Iscove's Modified
Dulbecco's Medium (IMDM) mixed with Ham's F-12 Nutrient Mix with
GlutaMax additive is about 70% to 30% mixture. In some embodiments,
the mixture of Iscove's Modified Dulbecco's Medium (IMDM) mixed
with Ham's F-12 Nutrient Mix with GlutaMax additive is about 65% to
35% mixture. In some embodiments, the mixture of Iscove's Modified
Dulbecco's Medium (IMDM) mixed with Ham's F-12 Nutrient Mix with
GlutaMax additive is about 60% to 40% mixture. In some embodiments,
the mixture of Iscove's Modified Dulbecco's Medium (IMDM) mixed
with Ham's F-12 Nutrient Mix with GlutaMax additive is about 55% to
45% mixture. Any of these concentrations may be combined with the
other ingredients provided herein at any of their listed
concentrations as well and may be used in any MDM, or MDM+, MDM1,
or MDM1+ media described herein or any medium derived
therefrom.
The concentration of albumin listed Table 1 and 4 is about 5 mg/ml;
however, this ingredient may be used at other concentrations
without negatively affecting the performance of the medium. In one
embodiment the concentration of albumin in MDM, MDM+, MDM1, or
MDM1+ is about 5 mg/ml. In one embodiment the concentration of
albumin in MDM, MDM+, MDM1, or MDM1+ is about 1 mg/ml. In one
embodiment the concentration of albumin in MDM, MDM+, MDM1, or
MDM1+ is about 2 mg/ml. In one embodiment the concentration of
albumin in MDM, MDM+, MDM1, or MDM1+ is about 3 mg/ml. In one
embodiment the concentration of albumin in MDM, MDM+, MDM1, or
MDM1+ is about 4 mg/ml. In one embodiment the concentration of
albumin in MDM, MDM+, MDM1, or MDM1+ is about 6 mg/ml. In one
embodiment the concentration of albumin in MDM, MDM+, MDM1, or
MDM1+ is about 7 mg/ml. In one embodiment the concentration of
albumin in MDM, MDM+, MDM1, or MDM1+ is about 8 mg/ml. In one
embodiment the concentration of albumin in MDM, MDM+, MDM1, or
MDM1+ is about 9 mg/ml. In one embodiment the concentration of
albumin in MDM, MDM+, MDM1, or MDM1+ is about 10 mg/ml. Any of
these concentrations may be combined with the other ingredients
provided herein at any of their listed concentrations as well and
may be used in any MDM, MDM+, MDM1, or MDM1+ media described herein
or any medium derived therefrom.
The concentration of .alpha.-monothioglycerol listed Table 1 and
Table 4 is from about 350 .mu.M to about 450 .mu.M; however, this
ingredient may be used at other concentrations without negatively
affecting the performance of the medium. In one embodiment the
concentration of .alpha.-monothioglycerol in MDM, MDM+, MDM1, or
MDM1+ is about 350 .mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, MDM+, MDM1, or MDM1+ is about 360
.mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, MDM+, MDM1, or MDM1+ is about 370
.mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, MDM+, MDM1, or MDM1+ is about 380
.mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol MDM, MDM+, MDM1, or MDM1+ is about 390
.mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, MDM+, MDM1, or MDM1+ is about 400
.mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, MDM+, MDM1, or MDM1+ is about 410
.mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, or MDM+, MDM1, or MDM1+ is about
420 .mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, or MDM+, MDM1, or MDM1+ is about
430 .mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, or MDM+, MDM1, or MDM1+ is about
440 .mu.M. In one embodiment the concentration of
.alpha.-monothioglycerol in MDM, or MDM+, MDM1, or MDM1+ is about
450 .mu.M. Any of these concentrations may be combined with the
other ingredients provide herein at any of their listed
concentrations as well and may be used in any MDM, or MDM+, MDM1,
or MDM1+ media described herein or any medium derived
therefrom.
The concentration of L-alanyl-L-glutamine listed Table 1 and Table
4 is from about 1 mM to about 2 mM; however, this ingredient may be
used at other concentrations without negatively affecting the
performance of the medium. In one embodiment the concentration of
L-alanyl-L-glutamine in MDM, or MDM+, MDM1, or MDM1+ is about 0.3
mM. In one embodiment the concentration of L-alanyl-L-glutamine in
MDM, or MDM+, MDM1, or MDM1+ is about 0.6 mM. In one embodiment the
concentration of L-alanyl-L-glutamine in MDM, or MDM+, MDM1, or
MDM1+ is about 1 mM. In one embodiment the concentration of
L-alanyl-L-glutamine in MDM, or MDM+, MDM1, or MDM1+ is about 1.3
mM. In one embodiment the concentration of L-alanyl-L-glutamine
MDM, or MDM+, MDM1, or MDM1+ is about 1.6 mM. In one embodiment the
concentration of L-alanyl-L-glutamine MDM, or MDM+, MDM1, or MDM1+
is about 2 mM. Any of these concentrations may be combined with the
other ingredients provided herein at any of their listed
concentrations as well and may be used in any MDM, or MDM+, MDM1,
or MDM1+ media described herein or any medium derived
therefrom.
The concentration of cholesterol lipids listed Table 1 is from
about about 1 .mu.g/ml to about 4 .mu.g/ml; however, this
ingredient may be used at other concentrations without negatively
affecting the performance of the medium. In one embodiment the
concentration of cholesterol lipids in MDM, or MDM+ is about 1
.mu.g/ml. In one embodiment the concentration of cholesterol lipids
in MDM, or MDM+ is about 2 .mu.g/ml. In one embodiment the
concentration of cholesterol lipids in MDM, or MDM+, is about 3
.mu.g/ml. In one embodiment the concentration of cholesterol lipids
in MDM, or MDM+ is about 4 .mu.g/ml. In one embodiment the
concentration of cholesterol lipids MDM, or MDM+, MDM1, or MDM1+ is
about 2.2 .mu.g/ml. Any of these concentrations may be combined
with the other ingredients provided herein at any of their listed
concentrations as well and may be used in any MDM, or MDM+, MDM1,
or MDM1+ media described herein or any medium derived
therefrom.
The concentration of L-ascorbic acid 2-phosphate listed Table 1 or
Table 4 is about 50 .mu.g/ml; however, this ingredient may be used
at other concentrations without negatively affecting the
performance of the medium. In one embodiment the concentration of
L-ascorbic acid 2-phosphate in MDM, or MDM+, MDM1, or MDM1+ is
about 30 .mu.g/ml. In one embodiment the concentration of
L-ascorbic acid 2-phosphate in MDM, or MDM+, MDM1, or MDM1+ is
about 40 .mu.g/ml. In one embodiment the concentration of
L-ascorbic acid 2-phosphate in MDM, or MDM+, MDM1, or MDM1+ is
about 45 .mu.g/ml. In one embodiment the concentration of
L-ascorbic acid 2-phosphate in MDM, or MDM+, MDM1, or MDM1+ is
about 50 .mu.g/ml. In one embodiment the concentration of
L-ascorbic acid 2-phosphate MDM, or MDM+, MDM1, or MDM1+ is about
55 .mu.g/ml. In one embodiment the concentration of L-ascorbic acid
2-phosphate MDM, or MDM+, MDM1, or MDM1+ is about 60 .mu.g/ml. In
one embodiment the concentration of L-ascorbic acid 2-phosphate
MDM, or MDM+, MDM1, or MDM1+ is about 65 .mu.g/ml. In one
embodiment the concentration of L-ascorbic acid 2-phosphate MDM, or
MDM+, MDM1, or MDM1+ is about 70 .mu.g/ml. Any of these
concentrations may be combined with the other ingredients provided
herein at any of their listed concentrations as well and may be
used in any MDM, or MDM+, MDM1, or MDM1+ media described herein or
any medium derived therefrom.
The concentration of BMP4 listed Table 1 or Table 4 is about 10
ng/ml; however, this ingredient may be used at other concentrations
without negatively affecting the performance of the medium. In one
embodiment the concentration of BMP4 in MDM, or MDM+, MDM1, or
MDM1+ is about 3 ng/ml. In one embodiment the concentration of BMP4
in MDM, or MDM+, MDM1, or MDM1+ is about 6 ng/ml. In one embodiment
the concentration of BMP4 in MDM, or MDM+, MDM1, or MDM1+ is about
10 ng/ml. In one embodiment the concentration of BMP4 in MDM, or
MDM+, MDM1, or MDM1+ is about 13 ng/ml. In one embodiment the
concentration of BMP4 MDM, or MDM+, MDM1, or MDM1+ is about 16
ng/ml. In one embodiment the concentration of BMP4 MDM, or MDM+,
MDM1, or MDM1+ is about 20 ng/ml. Any of these concentrations may
be combined with the other ingredients provided herein at any of
their listed concentrations as well and may be used in any MDM, or
MDM+, MDM1, or MDM1+ media described herein or any medium derived
therefrom.
The concentration of VEGF listed Table 1 or Table 4 is about 10
ng/ml; however, this ingredient may be used at other concentrations
without negatively affecting the performance of the medium. In one
embodiment the concentration of VEGF in MDM, or MDM+, MDM1, or
MDM1+ is about 3 ng/ml. In one embodiment the concentration of VEGF
in MDM, or MDM+, MDM1, or MDM1+ is about 6 ng/ml. In one embodiment
the concentration of VEGF in MDM, or MDM+, MDM1, or MDM1+ is about
10 ng/ml. In one embodiment the concentration of VEGF in MDM, or
MDM+, MDM1, or MDM1+ is about 13 ng/ml. In one embodiment the
concentration of VEGF MDM, or MDM+, MDM1, or MDM1+ is about 16
ng/ml. In one embodiment the concentration of VEGF MDM, or MDM+,
MDM1, or MDM1+ is about 20 ng/ml. Any of these concentrations may
be combined with the other ingredients provided herein at any of
their listed concentrations as well and may be used in any MDM, or
MDM+, MDM1, or MDM1+ media described herein or any medium derived
therefrom.
The concentration of bFGF listed Table 1 or Table 4 is from about
10 ng/ml to about 25 ng/ml; however, this ingredient may be used at
other concentrations without negatively affecting the performance
of the medium. In one embodiment the concentration of bFGF in MDM,
or MDM+, MDM1, or MDM1+ is about 3 ng/ml. In one embodiment the
concentration of bFGF in MDM, or MDM+, MDM1, or MDM1+ is about 6
ng/ml. In one embodiment the concentration of bFGF in MDM, or MDM+,
MDM1, or MDM1+ is about 7 ng/ml. In one embodiment the
concentration of bFGF in MDM, or MDM+, MDM1, or MDM1+ is about 10
ng/ml. In one embodiment the concentration of bFGF MDM, or MDM+,
MDM1, or MDM1+ is about 15 ng/ml. In one embodiment the
concentration of bFGF MDM, or MDM+, MDM1, or MDM1+ is about 20
ng/ml. In one embodiment the concentration of bFGF MDM, or MDM+,
MDM1, or MDM1+ is about 25 ng/ml. In one embodiment the
concentration of bFGF MDM, or MDM+, MDM1, or MDM1+ is about 30
ng/ml. In one embodiment the concentration of bFGF MDM, or MDM+,
MDM1, or MDM1+ is about 35 ng/ml. Any of these concentrations may
be combined with the other ingredients provided herein at any of
their listed concentrations as well and may be used in any MDM, or
MDM+, MDM1, or MDM1+ media described herein or any medium derived
therefrom.
The concentration of SCF used in MDM+ is about 100 ng/ml. The
concentration of SCF used in MDM1+ is about 50 ng/ml. However, this
ingredient may be used at other concentrations without negatively
affecting the performance of the medium. In addition, SCF may be
used to supplement MDM and MDM1, alone or in combination with other
additives, as discussed herein. In one embodiment the concentration
of SCF in MDM, or MDM+, MDM1, or MDM1+ is about 75 ng/ml. In one
embodiment the concentration of SCF in MDM, or MDM+, MDM1, or MDM1+
is about 80 ng/ml. In one embodiment the concentration of SCF in
MDM, or MDM+, MDM1, or MDM1+ is about 85 ng/ml. In one embodiment
the concentration of SCF in MDM, or MDM+, MDM1, or MDM1+ is about
90 ng/ml. In one embodiment the concentration of SCF MDM, or MDM+,
MDM1, or MDM1+ is about 95 ng/ml. In one embodiment the
concentration of SCF MDM, or MDM+, MDM1, or MDM1+ is about 100
ng/ml. In one embodiment the concentration of SCF MDM, or MDM+,
MDM1, or MDM1+ is about 105 ng/ml. In one embodiment the
concentration of SCF MDM, or MDM+, MDM1, or MDM1+ is about 110
ng/ml. In one embodiment the concentration of SCF MDM, or MDM+,
MDM1, or MDM1+ is about 115 ng/ml. In one embodiment the
concentration of SCF MDM, or MDM+, MDM1, or MDM1+ is about 120
ng/ml. In one embodiment the concentration of SCF MDM, or MDM+,
MDM1, or MDM1+ is about 125 ng/ml. Any of these concentrations may
be combined with the other ingredients provided herein at any of
their listed concentrations as well and may be used in any MDM, or
MDM+, MDM1, or MDM1+ media described herein or any medium derived
therefrom.
The concentration of Flt-3 ligand used in MDM+ is about 100 ng/ml.
The concentration of Flt-3 ligand used in MDM1+ is about 50 ng/ml
however, this ingredient may be used at other concentrations
without negatively affecting the performance of the medium. In
addition, Flt-3 ligand may be used to supplement MDM and MDM1,
alone or in combination with other additives, as discussed herein.
In one embodiment the concentration of Flt-3 ligand in MDM, or
MDM+, MDM1, or MDM1+ is about 75 ng/ml. In one embodiment the
concentration of Flt-3 ligand in MDM, or MDM+, MDM1, or MDM1+ is
about 80 ng/ml. In one embodiment the concentration of Flt-3 ligand
in MDM, or MDM+, MDM1, or MDM1+ is about 85 ng/ml. In one
embodiment the concentration of Flt-3 ligand in MDM, or MDM+, MDM1,
or MDM1+ is about 90 ng/ml. In one embodiment the concentration of
Flt-3 ligand MDM, or MDM+, MDM1, or MDM1+ is about 95 ng/ml. In one
embodiment the concentration of Flt-3 ligand MDM, or MDM+, MDM1, or
MDM1+ is about 100 ng/ml. In one embodiment the concentration of
Flt-3 ligand MDM, or MDM+, MDM1, or MDM1+ is about 105 ng/ml. In
one embodiment the concentration of Flt-3 ligand MDM, or MDM+,
MDM1, or MDM1+ is about 110 ng/ml. In one embodiment the
concentration of Flt-3 ligand MDM, or MDM+, MDM1, or MDM1+ is about
115 ng/ml. In one embodiment the concentration of Flt-3 ligand MDM,
or MDM+, MDM1, or MDM1+ is about 120 ng/ml. In one embodiment the
concentration of Flt-3 ligand MDM, or MDM+, MDM1, or MDM1+ is about
125 ng/ml. Any of these concentrations may be combined with the
other ingredients provide herein at any of their listed
concentrations as well and may be used in any MDM, or MDM+, MDM1,
or MDM1+ media described herein or any medium derived
therefrom.
The concentration of thrombopoietin in MDM+ is about 100 ng/ml. The
concentration of thrombopoietin in MDM1+ is about 50 ng/ml.
However, this ingredient may be used at other concentrations
without negatively affecting the performance of the medium. In
addition, thrombopoietin may be used to supplement MDM, and MDM1,
alone or in combination with other additives, as discussed herein.
In one embodiment the concentration of thrombopoietin in MDM, or
MDM+, MDM1, or MDM1+ is about 75 ng/ml. In one embodiment the
concentration of thrombopoietin in MDM, or MDM+, MDM1, or MDM1+ is
about 80 ng/ml. In one embodiment the concentration of
thrombopoietin in MDM, or MDM+, MDM1, or MDM1+ is about 85 ng/ml.
In one embodiment the concentration of thrombopoietin in MDM, or
MDM+, MDM1, or MDM1+ is about 90 ng/ml. In one embodiment the
concentration of thrombopoietin MDM, or MDM+, MDM1, or MDM1+ is
about 95 ng/ml. In one embodiment the concentration of
thrombopoietin MDM, or MDM+, MDM1, or MDM1+ is about 100 ng/ml. In
one embodiment the concentration of thrombopoietin MDM, or MDM+,
MDM1, or MDM1+ is about 105 ng/ml. In one embodiment the
concentration of thrombopoietin MDM, or MDM+, MDM1, or MDM1+ is
about 110 ng/ml. In one embodiment the concentration of
thrombopoietin MDM, or MDM+, MDM1, or MDM1+ is about 115 ng/ml. In
one embodiment the concentration of thrombopoietin MDM, or MDM+,
MDM1, or MDM1+ is about 120 ng/ml. In one embodiment the
concentration of thrombopoietin MDM, or MDM+, MDM1, or MDM1+ is
about 125 ng/ml. Any of these concentrations may be combined with
the other ingredients provided herein at any of their listed
concentrations as well and may be used in any MDM, or MDM+, MDM1,
or MDM1+ media described herein or any medium derived
therefrom.
The media described herein may have an amount of protein-free
hybridoma mixture II. For example, the final MDM, or MDM+, MDM1, or
MDM1+ medium, or a derivative medium, may have protein-free
hybridoma mixture II is about 5% of the total medium formulation.
Other concentrations may be used without negatively affecting the
performance of the medium. In one embodiment protein-free hybridoma
mixture II is about 2% of the total medium formulation. In one
embodiment protein-free hybridoma mixture II is about 3% of the
total medium formulation. In another embodiment protein-free
hybridoma mixture II is about 4% of the total medium formulation.
In one embodiment protein-free hybridoma mixture II is about 5% of
the total medium formulation. In another embodiment protein-free
hybridoma mixture II is about 6% of the total medium. In one
embodiment protein-free hybridoma mixture II is about 7% of the
total medium formulation. In one embodiment protein-free hybridoma
mixture II is about 8% of the total medium formulation. In one
embodiment protein-free hybridoma mixture II is about 9% of the
total medium formulation. In another embodiment protein-free
hybridoma mixture II is about 10% of the total medium formulation.
Any of these concentrations may be combined with the other
ingredients provided herein at any of their listed concentrations
as well and may be used in any MDM, or MDM+, MDM1, or MDM1+ media
described herein or any medium derived therefrom.
Methods of Culturing Cells and Promoting Cell Differentiation
The media compositions and formulations described herein may be
used for culturing cells. In some embodiments the described media
compositions and formulations may be used to maintain or expand
cells in culture. In other embodiments the described media
compositions and formulations may be used to culture cells in a
manner that promotes their differentiation into a different cell
type. In another embodiment the described media compositions and
formulations may be used to culture cells in a manner that promotes
their differentiation into a different cell type and then the same
medium, or a similar medium derivative, may be used to culture the
differentiated cell. Furthermore, the described media compositions
and formulations may be used to culture cells in a manner that
promotes their differentiation into a different cell type and then
the same medium, or a similar medium derivative, may be used to
culture the differentiated cell in a manner that allows the cell to
differentiate further. Methods for carrying out these culture
techniques are described herein. In view of the description of
these culture methods, certain modifications, based on existing
culture techniques, will be readily apparent to those skilled in
the art, such variations of the described methods are considered to
be within the scope of this disclosure.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 5 days, in the absence of
feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 5 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 5 days, in the presence of a basement membrane
matrix, in the absence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 5
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 5 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 5 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 6 days, in the absence of
feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 6 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 6 days, in the presence of a basement membrane
matrix, in the absence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 6
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 6 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 6 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 7 days, in the absence of
feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 7 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 7 days, in the presence of a basement membrane
matrix, in the absence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 7
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 7 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 7 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 8 days, in the absence of
feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 8 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 8 days, in the presence of a basement membrane
matrix, in the absence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 8
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 8 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 8 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 9 days, in the absence of
feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 9 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 9 days, in the presence of a basement membrane
matrix, in the absence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 9
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 9 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 9 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 10 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 10 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 10 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 10
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 10 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 10 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 11 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 11 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 11 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 11
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 11 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 11 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are method for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 12 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 12 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 12 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 12
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 12 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 12 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 13 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 13 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 13 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 13
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 13 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 13 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 14 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 14 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 14 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 14
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 14 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 14 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 15 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 15 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 15 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 15
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 15 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 15 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 16 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 16 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 16 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 16
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 16 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 16 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 17 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 17 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 17 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 17
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 17 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 17 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 18 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 18 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 18 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 18
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 18 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 18 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 19 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 19 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 19 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 19
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 19 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 19 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 20 days, in the absence
of feeder cells. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 20 days, in the presence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 20 days, in the presence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the mesodermal precursor cell is produced by culturing an iPSC in
the MDM or MDM1 medium described herein for a period of at least 20
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 20 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of at least 20 days, in the absence of a basement membrane
matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 2 passages, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
at least 2 passages, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 2
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 2 passages, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 2
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 2 passages, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 3 passages, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
at least 3 passages, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 3
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 3 passages, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 3
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 3 passages, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 4 passages, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
at least 4 passages, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 4
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 4 passages, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 4
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 4 passages, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 5 passages, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
at least 5 passages, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 5
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 5 passages, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 5
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 5 passages, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 6 passages, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
at least 6 passages, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 6
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 6 passages, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 6
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 6 passages, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 7 passages, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
at least 7 passages, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 7
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 7 passages, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 7
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 7 passages, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 5 days, in the absence of
feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 5
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 5 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 5 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 5 days, in the absence of
a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 5 days, in the absence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 6 days, in the absence of
feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 6
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45, but
not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 6 days, in the presence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 6 days, in the presence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for a period of at least 6 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 6
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 7 days, in the absence of
feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 7
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 7 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 7 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 7 days, in the absence of
a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 7 days, in the absence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 8 days, in the absence of
feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 8
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 8 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 8 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 8 days, in the absence of
a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 8 days, in the absence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 9 days, in the absence of
feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 9
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 9 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 9 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 9 days, in the absence of
a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 9 days, in the absence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 10 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 10
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 10 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 10 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 10 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 10 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 11 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 11
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 11 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 11 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 11 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 11 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 12 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 12
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 12 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 12 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 12 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 12 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided hereins are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 13 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 13
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 13 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 13 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 13 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 13 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 14 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 14
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 14 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 14 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 14 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 14 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 15 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 15
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 15 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 15 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 15 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 15 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 16 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 16
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 16 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 16 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 16 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 16 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 17 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 17
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 17 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 17 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 17 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 17 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 18 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 18
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 18 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 18 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 18 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 18 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 19 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 19
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 19 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 19 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 19 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 19 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 20 days, in the absence
of feeder cells, such that the differentiated mesodermal precursor
cells express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of at least 20
days, in the presence of feeder cells, such that the differentiated
mesodermal precursor cells express CD31 and CD34, but not CD45. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for a
period of at least 20 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for a period of at least 20 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 20 days, in the absence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of at least 20 days, in the absence
of a basement membrane matrix, in the presence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 2 passages, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for at least 2 passages, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 2
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 2
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 2
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 2
passages, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 3 passages, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for at least 3 passages, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 3
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 3
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 3
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 3
passages, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 4 passages, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for at least 4 passages, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 4
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 4
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 4
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 4
passages, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 5 passages, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for at least 5 passages, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 5
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 5
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 5
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 5
passages, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 6 passages, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for at least 6 passages, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 6
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 6
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 6
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 6
passages, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for at least 7 passages, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for at least 7 passages, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 7
passages, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 7
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 7
passages, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for at least 7
passages, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 5 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 5 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
5 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 5 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
5 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 5 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 6 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 6 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
6 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 6 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
6 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 6 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 7 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 7 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
7 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 7 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
7 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 7 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 8 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 8 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
8 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 8 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
8 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 8 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 9 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 9 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
9 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 9 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
9 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 9 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 10 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 10 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
10 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 10 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
10 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 10 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 11 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 11 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
11 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 11 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
11 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 11 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 12 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 12 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
12 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 12 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
12 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 12 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 13 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 13 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
13 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 13 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
13 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 13 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 14 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 14 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
14 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 14 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
14 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 14 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 15 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 15 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
15 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 15 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
15 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 15 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 16 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 16 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
16 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 16 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
16 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 16 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 17 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 17 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
17 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 17 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
17 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 17 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 18 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 18 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
18 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 18 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
18 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 18 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 19 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 19 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
19 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 19 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
19 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 19 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 20 days, in the absence of feeder
cells. In one embodiment the mesodermal precursor cell is produced
by culturing an iPSC in the MDM or MDM1 medium described herein for
a period of 20 days, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
20 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 20 days, in the presence of a
basement membrane matrix, in the presence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
20 days, in the absence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the mesodermal precursor
cell is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 20 days, in the absence of a
basement membrane matrix, in the presence of feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 2 passages, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 2
passages, in the presence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 2 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 2
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 2 passages, in the absence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for 2 passages,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 3 passages, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 3
passages, in the presence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 3 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 3
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 3 passages, in the absence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for 3 passages,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 4 passages, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 4
passages, in the presence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 4 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 4
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 4 passages, in the absence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for 4 passages,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 5 passages, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 5
passages, in the presence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 5 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 5
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 5 passages, in the absence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for 5 passages,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 6 passages, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 6
passages, in the presence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 6 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 6
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 6 passages, in the absence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for 6 passages,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 7 passages, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 7
passages, in the presence of feeder cells. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 7 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells. In
one embodiment the mesodermal precursor cell is produced by
culturing an iPSC in the MDM or MDM1 medium described herein for 7
passages, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 7 passages, in the absence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for 7 passages,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 5 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 5 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
5 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
5 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
5 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
5 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 6 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 6 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
6 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
6 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
6 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
6 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 7 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 7 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
7 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
7 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
7 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
7 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 8 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 8 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
8 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
8 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
8 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
8 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 9 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 9 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
9 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
9 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
9 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
9 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 10 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 10 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
10 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
10 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
10 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
10 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 11 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 11 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
11 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
11 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
11 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
11 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 12 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 12 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
12 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
12 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
12 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
12 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 13 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 13 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
13 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
13 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
13 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
13 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 14 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 14 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
14 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
14 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
14 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
14 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 15 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 15 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
15 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
15 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
15 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
15 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 16 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 16 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
16 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
16 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
16 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
16 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 17 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 17 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
17 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
17 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
17 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
17 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 18 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 18 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
18 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
18 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
18 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
18 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 19 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 19 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
19 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
19 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
19 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
19 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for a period of 20 days, in the absence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for a period of 20 days, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
20 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
20 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
20 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45. In one
embodiment the mesodermal precursor cell is produced by culturing
an iPSC in the MDM or MDM1 medium described herein for a period of
20 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated mesodermal
precursor cells express CD31 and CD34, but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 2 passages, in the absence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 2 passages, in the presence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 2 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 2 passages, in the presence of a basement
membrane matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 2 passages, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 2 passages, in the absence of a basement membrane
matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 3 passages, in the absence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 3 passages, in the presence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 3 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 3 passages, in the presence of a basement
membrane matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 3 passages, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 3 passages, in the absence of a basement membrane
matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 4 passages, in the absence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 4 passages, in the presence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 4 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 4 passages, in the presence of a basement
membrane matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 4 passages, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 4 passages, in the absence of a basement membrane
matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 5 passages, in the absence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 5 passages, in the presence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 5 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 5 passages, in the presence of a basement
membrane matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 5 passages, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 5 passages, in the absence of a basement membrane
matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 6 passages, in the absence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 6 passages, in the presence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 6 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 6 passages, in the presence of a basement
membrane matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 6 passages, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 6 passages, in the absence of a basement membrane
matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45.
Provided herein are methods for producing a mesodermal precursor
cell from an iPSC. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 7 passages, in the absence of feeder cells,
such that the differentiated mesodermal precursor cells express
CD31 and CD34, but not CD45. In one embodiment the mesodermal
precursor cell is produced by culturing an iPSC in the MDM or MDM1
medium described herein for 7 passages, in the presence of feeder
cells, such that the differentiated mesodermal precursor cells
express CD31 and CD34, but not CD45. In one embodiment the
mesodermal precursor cell is produced by culturing an iPSC in the
MDM or MDM1 medium described herein for 7 passages, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated mesodermal precursor cells express CD31 and
CD34, but not CD45. In one embodiment the mesodermal precursor cell
is produced by culturing an iPSC in the MDM or MDM1 medium
described herein for 7 passages, in the presence of a basement
membrane matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 7 passages, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45. In one embodiment the mesodermal precursor cell is
produced by culturing an iPSC in the MDM or MDM1 medium described
herein for 7 passages, in the absence of a basement membrane
matrix, in the presence of feeder cells, such that the
differentiated mesodermal precursor cells express CD31 and CD34,
but not CD45.
Once produced from iPSCs, the mesodermal precursor cells may be
cultured to differentiate into one of at least four different
lineages: hematopoietic stem cells (HSC), mesenchymal stem cells
(MSC), smooth muscle cells (SMC), or unlimited functional
endothelial cells (UFEC) (FIG. 2). All of these cell types are
"induced" cell types because they are not produced naturally by the
processes described herein; therefore, these cells may also be
referred to as induced hematopoietic stem cells (iHSC), induced
mesenchymal stem cells (iMSC), induced smooth muscle cells (iSMC),
or induced unlimited functional endothelial cells (iUFEC). Despite
the possible nomenclature variation, the cells, as discussed herein
are the same.
Hematopoietic Stem Cells (HSC)
Provided herein is a method for producing a hematopoietic stem cell
from a mesodermal precursor cell, where the method involves first
producing a mesodermal precursor cell from an iPSC and then
incubating the mesodermal precursor cell in the MDM+ or MDM1+
medium described herein. In some embodiments, the mesodermal
precursor cells can be incubated in MDM or MDM1 medium to allow for
the production of hematopoietic stem cells. The produced
hematopoietic stem cells can be characterized as expressing CD31,
CD34, and CD45, but not CD38 (CD31.sup.+, CD34.sup.+, CD45.sup.+,
CD38.sup.-). In some embodiments of the method, the mesodermal
precursor cells cultured to give rise to the hematopoietic stem
cells are purified, for example by flow cytometry, prior to being
further cultured to differentiate into hematopoietic stem cells.
Alternatively, in some embodiments of the method, the mesodermal
precursor cells cultured to give rise to the hematopoietic stem
cells are not purified from other cells in the initial iPSC culture
prior to being further cultured to differentiate into hematopoietic
stem cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 3 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM+ medium described herein for a period of at least 3 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
3 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 3
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 3
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 3 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 4 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 4 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
4 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 4
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 4
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 4 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 5 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 5 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
5 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 5
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 5
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 5 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 6 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 6 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
6 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 6
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 6
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 6 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 7 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 7 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
7 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 7
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 7
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 7 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 8 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 8 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
8 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 8
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 8
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 8 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 9 days, in the absence of
feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 9 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
9 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 9
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 9
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 9 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 10 days, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 10 days, in
the presence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
10 days, in the presence of a basement membrane matrix, in the
absence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 10
days, in the presence of a basement membrane matrix, in the
presence of feeder cells. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of at least 10
days, in the absence of a basement membrane matrix, in the absence
of feeder cells. In one embodiment the hematopoietic stem cell is
produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 10 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 3 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 3 days, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 3 days, in the presence
of a basement membrane matrix, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
3 days, in the presence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 3 days, in the absence of
a basement membrane matrix, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
3 days, in the absence of a basement membrane matrix, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 4 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 4 days, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
4 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 4 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
4 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 4 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 5 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 5 days, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
5 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 5 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
5 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 5 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 6 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 6 days, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
6 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 6 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
6 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 6 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 7 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 7 days, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
7 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 7 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
7 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 7 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 8 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 8 days, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
8 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 8 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
8 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 8 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 9 days, in the absence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 9 days, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
9 days, in the presence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 9 days, in the presence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of at least
9 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 9 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of at least 10 days, in the absence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of at least 10 days, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 10 days, in the presence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 10 days, in
the presence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of at least 10 days, in the absence of a basement membrane
matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of at least 10 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 3 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 3 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 3 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 3 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 3 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 3 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 4 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 4 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 4 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 4 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 4 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 4 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 5 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 5 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 5 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 5 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 5 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 5 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 6 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 6 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 6 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 6 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 6 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 6 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 7 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 7 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 7 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 7 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 7 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 7 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 8 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 8 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 8 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 8 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 8 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 8 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 9 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 9 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 9 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 9 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 9 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 9 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 10 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 10 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 10 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of at least 10 days, in the presence of a basement membrane
matrix, in the presence of feeder cells. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 10 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of at least 10 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 3 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 3 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 3 days, in the presence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 3 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 3 days, in the absence of a basement membrane matrix, in
the absence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 3 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 4 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 4 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 4 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 4 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 4 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 4 days, in the
absence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 5 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 5 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 5 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 5 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 5 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 5 days, in the
absence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 6 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 6 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 6 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 6 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 6 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 6 days, in the
absence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 7 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 7 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 7 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 7 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 7 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 7 days, in the
absence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 8 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 8 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 8 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 8 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 8 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 8 days, in the
absence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 9 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 9 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 9 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 9 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 9 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 9 days, in the
absence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 10 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 10 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of at least 10 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
at least 10 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of at least 10 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM medium described herein for a period of at least 10 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 3 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 3 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 3 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 3 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 3 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 3 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 4 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 4 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 4 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 4 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 4 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 4 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 5 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 5 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 5 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 5 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 5 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 5 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 6 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 6 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 6 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 6 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 6 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 6 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 7 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 7 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 7 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 7 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 7 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 7 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 8 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 8 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 8 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 8 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 8 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 8 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 9 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 9 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 9 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 9 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 9 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 9 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 10 days, in the absence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 10 days, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 10 days, in the presence of a
basement membrane matrix, in the absence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM+ or MDM1+ medium described
herein for a period of 10 days, in the presence of a basement
membrane matrix, in the presence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 10 days, in the absence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM+ or MDM1+ medium described herein for a period of 10 days,
in the absence of a basement membrane matrix, in the presence of
feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 3 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 3 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 3 days, in the presence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 3 days, in
the presence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 3 days, in the absence of a basement membrane matrix, in
the absence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 3 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 4 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 4 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 4 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 4 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 4 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 4 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 5 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 5 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 5 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 5 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 5 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 5 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 6 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 6 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 6 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 6 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 6 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 6 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 7 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 7 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 7 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 7 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 7 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 7 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 8 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 8 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 8 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 8 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 8 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 8 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 9 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 9 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 9 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 9 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 9 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 9 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 10 days, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 10 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM+ or
MDM1+ medium described herein for a period of 10 days, in the
presence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM+ or MDM1+ medium described herein for a
period of 10 days, in the presence of a basement membrane matrix,
in the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM+ or MDM1+ medium
described herein for a period of 10 days, in the absence of a
basement membrane matrix, in the absence of feeder cells, such that
the differentiated hematopoietic stem cell expresses CD31, CD34,
and CD45, but not CD38. In one embodiment the hematopoietic stem
cell is produced by culturing a mesodermal precursor cell in the
MDM+ or MDM1+ medium described herein for a period of 10 days, in
the absence of a basement membrane matrix, in the presence of
feeder cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 3 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 3 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 3 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 3 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 3 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 3
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 4 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 4 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 4 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 4 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 4 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 4
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 5 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 5 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 5 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 5 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 5 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 5
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 6 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 6 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 6 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 6 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 6 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 6
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 7 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 7 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 7 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 7 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 7 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 7
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 8 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 8 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 8 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 8 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 8 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 8
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 9 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 9 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 9 days, in the presence of a basement membrane matrix, in
the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 9 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 9 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 9
days, in the absence of a basement membrane matrix, in the presence
of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 10 days, in the absence of feeder cells. In
one embodiment the hematopoietic stem cell is produced by culturing
a mesodermal precursor cell in the MDM medium described herein for
a period of 10 days, in the presence of feeder cells. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 10 days, in the presence of a basement membrane matrix,
in the absence of feeder cells. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 10 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 10 days, in the absence of a basement
membrane matrix, in the absence of feeder cells. In one embodiment
the hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
10 days, in the absence of a basement membrane matrix, in the
presence of feeder cells.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 3 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 3 days, in the presence of feeder cells,
such that the differentiated hematopoietic stem cell expresses
CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 3
days, in the presence of a basement membrane matrix, in the absence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 3 days, in the presence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 3 days, in the absence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 3 days, in the absence of a
basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 4 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 4 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 4 days, in the presence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 4 days, in the presence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 4 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 4
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 5 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 5 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 5 days, in the presence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 5 days, in the presence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 5 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 5
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 6 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 6 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 6 days, in the presence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 6 days, in the presence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 6 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 6
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 7 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 7 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 7 days, in the presence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 7 days, in the presence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 7 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 7
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 8 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 8 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 8 days, in the presence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 8 days, in the presence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 8 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 8
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 9 days, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 9 days, in the
presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38. In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 9 days, in the presence of a basement
membrane matrix, in the absence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 9 days, in the presence of
a basement membrane matrix, in the presence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 9 days, in the
absence of a basement membrane matrix, in the absence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of 9
days, in the absence of a basement membrane matrix, in the presence
of feeder cells, such that the differentiated hematopoietic stem
cell expresses CD31, CD34, and CD45, but not CD38.
In one embodiment the hematopoietic stem cell is produced by
culturing a mesodermal precursor cell in the MDM medium described
herein for a period of 10 days, in the absence of feeder cells,
such that the differentiated hematopoietic stem cell expresses
CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
10 days, in the presence of feeder cells, such that the
differentiated hematopoietic stem cell expresses CD31, CD34, and
CD45, but not CD38. In one embodiment the hematopoietic stem cell
is produced by culturing a mesodermal precursor cell in the MDM
medium described herein for a period of 10 days, in the presence of
a basement membrane matrix, in the absence of feeder cells, such
that the differentiated hematopoietic stem cell expresses CD31,
CD34, and CD45, but not CD38. In one embodiment the hematopoietic
stem cell is produced by culturing a mesodermal precursor cell in
the MDM medium described herein for a period of 10 days, in the
presence of a basement membrane matrix, in the presence of feeder
cells, such that the differentiated hematopoietic stem cell
expresses CD31, CD34, and CD45, but not CD38. In one embodiment the
hematopoietic stem cell is produced by culturing a mesodermal
precursor cell in the MDM medium described herein for a period of
10 days, in the absence of a basement membrane matrix, in the
absence of feeder cells, such that the differentiated hematopoietic
stem cell expresses CD31, CD34, and CD45, but not CD38. In one
embodiment the hematopoietic stem cell is produced by culturing a
mesodermal precursor cell in the MDM medium described herein for a
period of 10 days, in the absence of a basement membrane matrix, in
the presence of feeder cells, such that the differentiated
hematopoietic stem cell expresses CD31, CD34, and CD45, but not
CD38.
Mesenchymal Stem Cells (MSC)
Provided herein is a method for producing a mesenchymal stem cell
from the mesodermal precursor cell described herein, wherein the
method involves first producing a mesodermal precursor cell from an
iPSC, as described by the related methods provided herein, and then
incubating the mesodermal precursor cell under conditions known to
promote differentiation of precursor cells into mesenchymal stem
cells. The produced mesenchymal stem cells can be characterized as
expressing CD90, CD73, and CD105, but not CD31 and CD45. In some
embodiments of the method, the mesodermal precursor cells cultured
to give rise to the mesenchymal stem cells are purified, for
example by flow cytometry, prior to being further cultured to
differentiate into mesenchymal stem cells. Alternatively, in some
embodiments of the method, the mesodermal precursor cells cultured
to give rise to the mesenchymal stem cells are not purified from
other cells in the initial iPSC culture prior to being further
cultured to differentiate into mesenchymal stem cells. The
described iMSCs may be further characterized by their multipotency.
For example the described iMSCs may further differentiate into
adipocytes, osteoblasts, myocytes, or chondrocytes in vitro, when
cultured under proper conditions known in the art, or in vivo, for
example following transplantation.
Smooth Muscle Cells (SMC)
Provided herein is a method for producing smooth muscle cells from
the mesodermal precursor cells described herein, wherein the method
involves first producing a mesodermal precursor cell from an iPSC,
as described by the related methods provided herein, and then
incubating the mesodermal precursor cell under conditions known to
promote differentiation of precursor cells into smooth muscle
cells. The produced smooth muscle cells can be characterized as
expressing .alpha.-SMA, calponin, and SM22. Furthermore, the iSMCs
exhibit the formation of spindles when grown in culture for an
amount of time adequate to allow for cell expansion to levels
sufficient to allow for spindle formation. In some embodiments of
the method, the mesodermal precursor cells cultured to give rise to
the smooth muscle cells are purified, for example by flow
cytometry, prior to being further cultured to differentiate into
smooth muscle cells. Alternatively, in some embodiments of the
method, the mesodermal precursor cells cultured to give rise to the
smooth muscle cells are not purified from other cells in the
initial iPSC culture prior to being further cultured to
differentiate into smooth muscle cells.
Unlimited Functional Endothelial Cells (UFEC)
Provided herein is a method for producing unlimited functional
endothelial cells (UFECs) (also referred to as "iECs") from the
mesodermal precursor cells described herein, wherein the method
involves first producing a mesodermal precursor cell from an iPSC,
as described by the related methods provided herein, and then
incubating the mesodermal precursor cell under conditions known to
promote differentiation of precursor cells into unlimited
functional endothelial cells. The produced UFECs can be
characterized as expressing CD31, vWF and CD144. Other
characteristics of these cells include the ability to form
vascular-like structures on Matrigel.TM. and the uptake of
acetylated-LDL when cultured. In some embodiments of the method,
the mesodermal precursor cells cultured to give rise to the
unlimited functional endothelial cells are purified, for example by
flow cytometry, prior to being further cultured to differentiate
into unlimited functional endothelial cells. Alternatively, in some
embodiments of the method, the mesodermal precursor cells cultured
to give rise to the unlimited functional endothelial cells are not
purified from other cells in the initial iPSC culture prior to
being further cultured to differentiate into unlimited functional
endothelial cells.
Cells for Carrying Out the Described Cell Production Methods
The cell production methods described herein may be carried out
using a variety of cell types to give rise to the initial iPSCs
that serve as the starting point for these methods and cells. For
example, it may be desirable to obtain cells from a subject in need
of a cell transplant, stem cell therapy, of precursor cell therapy,
and use the subject's own autologous cells, such as fibroblasts for
example, to generate autologous iPSCs that may be cultured to give
rise to an autologous form of one or more of the cell types
described herein. In other embodiments, heterologous cells may be
used for producing the cells described herein, even though these
cells may be intended for therapeutic use in a subject. In still
another embodiment, the methods described herein can be carried out
using iPSCs without regard to origin. For instance, the iPSCs could
be human cells, primate cells, or other mammalian cells converted
to iPSCs by conventional methods.
Cells Produced by the Described Methods
Described herein are a variety of cells capable of being produced
by the methods described herein. These cells include iPSC-derived
mesodermal precursor cells (MPC), positive for CD34 and CD31
expression that may be used to produce at least four different cell
types. When cultured under appropriate conditions, the MPCs can be
used to produce hematopoietic stem cells, mesenchymal stem cells,
smooth muscle cells, or unlimited functional endothelial cells. One
characteristic that makes the mesodermal precursor cells described
herein desirable is that these cells can be maintained in culture
for a number of days, or passages, without changing phenotype
through differentiation. In some embodiments the described MPCs can
maintain their phenotype in culture for at least 3 days. In some
embodiments the described MPCs can maintain their phenotype in
culture for at least 5 days. In some embodiments the described MPCs
can maintain their phenotype in culture for at least 7 days. In
some embodiments the described MPCs can maintain their phenotype in
culture for at least 9 days. In some embodiments the described MPCs
can maintain their phenotype in culture for at least 10 days. In
some embodiments the described MPCs can maintain their phenotype in
culture for at least 13 days. In some embodiments the described
MPCs can maintain their phenotype in culture for at least 16 days.
In some embodiments the described MPCs can maintain their phenotype
in culture for at least 20 days. In some embodiments the described
MPCs can maintain their phenotype in culture for at least 24 days,
or more. In some embodiments the described MPCs will have one or
more genes encoding Oct4, Sox2, klf4, or c-MYC incorporated into
the genome as a consequence of being produced from an iPSC that was
made by overexpression of Oct4, Sox2, klf4, and c-MYC. When the
iPSC is produced using a retroviral vector to deliver at least one
gene encoding Oct4, Sox2, klf4, or c-MYC, that gene can become
integrated into the genome of the iPSC and will subsequently be a
part of the genome of the resulting MPC.
Hematopoietic stem cells can be produced by culturing the MPCs
described herein. The described HSCs may be characterized by the
expression of CD34, CD31, and CD45, but not CD38. Another
characteristic of the described HSCs is that they have the ability
to reconstitute the hematopoietic system of an irradiated subject,
such as a mouse. The described HSCs also have the ability to
maintain their phenotype for extended periods without
differentiating, when maintained under appropriate conditions, such
as being cultured using the MDM medium described herein. In another
embodiment the described HSCs can maintain their phenotype for
extended periods without differentiating, when cultured using the
MDM+ or MDM1+ medium described herein. In some embodiments the
described HSCs can maintain their phenotype in culture for at least
3 days. In some embodiments the described HSCs can maintain their
phenotype in culture for at least 5 days. In some embodiments the
described HSCs can maintain their phenotype in culture for at least
7 days. In some embodiments the described HSCs can maintain their
phenotype in culture for at least 9 days. In some embodiments the
described HSCs can maintain their phenotype in culture for at least
10 days. In some embodiments the described HSCs can maintain their
phenotype in culture for at least 13 days. In some embodiments the
described HSCs can maintain their phenotype in culture for at least
16 days. In some embodiments the described HSCs can maintain their
phenotype in culture for at least 20 days. In some embodiments the
described HSCs can maintain their phenotype in culture for at least
24 days, or more. In some embodiments the described HSCs will have
one or more genes encoding Oct4, Sox2, klf4, or c-MYC incorporated
into the genome as a consequence of being produced from an iPSC
that was made by overexpression of Oct4, Sox2, klf4, and c-MYC.
When the iPSC is produced using a retroviral vector to deliver at
least one gene encoding Oct4, Sox2, klf4, or c-MYC, that gene can
become integrated into the genome of the iPSC and will subsequently
be a part of the genome of the resulting HSCs.
Another cell type described herein are unlimited functional
endothelial cells that may be obtained by differentiating the
described MPCs when cultured under conditions known to allow for
differentiation into cells of an endothelial lineage. The described
UFECs can be characterized by the expression of CD31, vWF, and
CD144. In addition, these cells can mediate the uptake of
acetylated low density lipoproteins (LDL). Furthermore, the UFECs
produced using the methods and cells described herein have the
ability to form vascular-like structures when cultured in vitro, a
hallmark of endothelia cell progenitors. One characteristic that
makes the UFECs described herein desirable is that these cells can
be maintained in culture for a number of days, or passages, without
changing phenotype through differentiation. In some embodiments the
described UFECs can maintain their phenotype in culture for at
least 3 days. In some embodiments the described UFECs can maintain
their phenotype in culture for at least 5 days. In some embodiments
the described UFECs can maintain their phenotype in culture for at
least 7 days. In some embodiments the described UFECs can maintain
their phenotype in culture for at least 9 days. In some embodiments
the described UFECs can maintain their phenotype in culture for at
least 10 days. In some embodiments the described UFECs can maintain
their phenotype in culture for at least 13 days. In some
embodiments the described UFECs can maintain their phenotype in
culture for at least 16 days. In some embodiments the described
UFECs can maintain their phenotype in culture for at least 20 days.
In some embodiments the described UFECs can maintain their
phenotype in culture for at least 24 days, or more. In some
embodiments the described UFECs will have one or more genes
encoding Oct4, Sox2, klf4, or c-MYC incorporated into the genome as
a consequence of being produced from an iPSC that was made by
overexpression of Oct4, Sox2, klf4, and c-MYC. When the iPSC is
produced using a retroviral vector to deliver at least one gene
encoding Oct4, Sox2, klf4, or c-MYC, that gene can become
integrated into the genome of the iPSC and will subsequently be a
part of the genome of the resulting UFECs, which are indirectly
derived from the iPSCs.
Also described herein are mesenchymal stem cells (MSCs) that may be
obtained by differentiating the described MPCs under conditions
known to allow for differentiation into cells of a mesenchymal
lineage. The MSCs described herein can be characterized by the
expression of CD90, CD73, and CD105 in the absence of CD31 and
CD45. These cells can also differentiate in vivo or in vitro into a
number of different cell types, including adipocytes, osteoblasts,
myocytes, or chondrocytes, when cultured under conditions known to
cause progenitor cells to differentiate into one of these cell
types. The described MSCs also have the ability to maintain their
phenotype for extended periods without differentiating, when
maintained under appropriate conditions, such as conditions known
to allow for differentiation into cells of an mesenchymal lineage.
In some embodiments the described MSCs can maintain their phenotype
in culture for at least 3 days. In some embodiments the described
MSCs can maintain their phenotype in culture for at least 5 days.
In some embodiments the described MSCs can maintain their phenotype
in culture for at least 7 days. In some embodiments the described
MSCs can maintain their phenotype in culture for at least 9 days.
In some embodiments the described MSCs can maintain their phenotype
in culture for at least 10 days. In some embodiments the described
MSCs can maintain their phenotype in culture for at least 13 days.
In some embodiments the described MSCs can maintain their phenotype
in culture for at least 16 days. In some embodiments the described
MSCs can maintain their phenotype in culture for at least 20 days.
In some embodiments the described MSCs can maintain their phenotype
in culture for at least 24 days, or more. In some embodiments the
described MSCs will have one or more genes encoding Oct4, Sox2,
klf4, or c-MYC incorporated into the genome as a consequence of
being produced from an iPSC that was made by overexpression of
Oct4, Sox2, klf4, and c-MYC. When the iPSC is produced using a
retroviral vector to deliver at least one gene encoding Oct4, Sox2,
klf4, or c-MYC, that gene can become integrated into the genome of
the iPSC and will subsequently be a part of the genome of the
resulting MSCs, which are indirectly derived from the iPSCs.
Smooth muscle cells can be produced by culturing the MPCs described
herein. For example, the described MPCs can differentiate into
smooth muscle cells when cultured under conditions known to cause
progenitor cells to differentiate into SMCs. The described SMCs are
characterized by the expression of .alpha.-SMA, calponin, and SM22.
The described SMCs also have the ability to maintain their
phenotype for extended periods without differentiating, when
maintained under appropriate conditions, such as conditions known
to allow for differentiation into smooth muscle cells. In some
embodiments the described SMCs can maintain their phenotype in
culture for at least 3 days. In some embodiments the described SMCs
can maintain their phenotype in culture for at least 5 days. In
some embodiments the described SMCs can maintain their phenotype in
culture for at least 7 days. In some embodiments the described SMCs
can maintain their phenotype in culture for at least 9 days. In
some embodiments the described SMCs can maintain their phenotype in
culture for at least 10 days. In some embodiments the described
SMCs can maintain their phenotype in culture for at least 13 days.
In some embodiments the described SMCs can maintain their phenotype
in culture for at least 16 days. In some embodiments the described
SMCs can maintain their phenotype in culture for at least 20 days.
In some embodiments the described SMCs can maintain their phenotype
in culture for at least 24 days, or more. In some embodiments the
described SMCs will have one or more genes encoding Oct4, Sox2,
klf4, or c-MYC incorporated into the genome as a consequence of
being produced from an iPSC that was made by overexpression of
Oct4, Sox2, klf4, and c-MYC. When the iPSC is produced using a
retroviral vector to deliver at least one gene encoding Oct4, Sox2,
klf4, or c-MYC, that gene can become integrated into the genome of
the iPSC and will subsequently be a part of the genome of the
resulting SMCs, which are indirectly derived from the iPSCs.
The cells described herein may be made using a variety of cell
types to give rise to the initial iPSCs that serve as the starting
point for producing these cells. For example, it may be desirable
to obtain cells from a subject in need of a cell transplant, stem
cell therapy, of precursor cell therapy, and use the subject's own
autologous cells, such as fibroblasts for example, to generate
autologous iPSCs that may be cultured to give rise to an autologous
form of one or more of the cell types described herein. In other
embodiments, heterologous cells may be used for producing the cells
described herein, even though these cells may be intended for
therapeutic use in a subject. In still another embodiment, the
methods described herein can be carried out using iPSCs without
regard to origin. For instance, the iPSCs could be human cells,
primate cells, or other mammalian cells converted to PSCs by
conventional methods.
Methods of Treatment
Stem cells and lineage precursor cells have been shown to have
therapeutic applications to a variety of diseases. Accordingly, the
stem cells and progenitor cells described herein, whether
autologous or heterologous in nature, may also be used of this
purpose. For example, the HSCs described herein may be used to
treat a subject with a disorder of the hematopoietic system. In
some embodiments the HSCs may be administered to a subject having a
congenital bone marrow disorder. Examples of such disorders include
congenital aplastic anemia (Fanconi anemia), congenital hypoplastic
anemia (Diamond-Blackfan anemia), congenital neutropenias (Kostmann
syndrome, cyclic neutropenia, Shwachman-Diamond syndrome and
others), and congenital thrombocytopenias (TAR syndrome,
amegacaryocytic thrombocytopenia). The described HCSs may be
administered to an individual to treat any one of these disorders.
In some embodiments the described HSCs may be administered to a
subject to treat congenital aplastic anemia (Fanconi anemia). In
some embodiments the described HSCs may be administered to a
subject to treat congenital hypoplastic anemia (Diamond-Blackfan
anemia). In some embodiments the described HSCs may be administered
to a subject to treat Kostmann syndrome. In some embodiments the
described HSCs may be administered to a subject to treat cyclic
neutropenia. In some embodiments the described HSCs may be
administered to a subject to treat Shwachman-Diamond syndrome. In
some embodiments the described HSCs may be administered to a
subject to treat TAR syndrome. In some embodiments the described
HSCs may be administered to a subject to treat amegacaryocytic
thrombocytopenia. In some embodiments the described HSCs
administered to a subject to treat ongenital aplastic anemia
(Fanconi anemia) are autologous. In some embodiments the described
HSCs administered to a subject to treat congenital hypoplastic
anemia (Diamond-Blackfan anemia) are autologous. In some
embodiments the described HSCs administered to a subject to treat
Kostmann syndrome are autologous. In some embodiments the described
HSCs administered to a subject to treat cyclic neutropenia are
autologous. In some embodiments the described HSCs administered to
a subject to treat Shwachman-Diamond syndrome are autologous. In
some embodiments the described HSCs administered to a subject to
treat TAR syndrome are autologous. In some embodiments the
described HSCs administered to a subject to treat amegacaryocytic
thrombocytopenia are autologous. In some embodiments the described
HSCs administered to a subject to treat congenital aplastic anemia
(Fanconi anemia) are heterologous. In some embodiments the
described HSCs administered to a subject to treat congenital
hypoplastic anemia (Diamond-Blackfan anemia) are heterologous. In
some embodiments the described HSCs administered to a subject to
treat Kostmann syndrome are heterologous. In some embodiments the
described HSCs administered to a subject to treat cyclic
neutropenia are heterologous. In some embodiments the described
HSCs administered to a subject to treat Shwachman-Diamond syndrome
are heterologous. In some embodiments the described HSCs
administered to a subject to treat TAR syndrome are heterologous.
In some embodiments the described HSCs administered to a subject to
treat amegacaryocytic thrombocytopenia are heterologous. Those
skilled in the art will understand that the forgoing disclosure
provides only a small listing of disorders of the hematopoietic
system that may be treated using the described HSCs; therefore,
treatment of such disorders known to be susceptible to stem cell
therapy should be considered to be within the scope of this
disclosure. HSCs may also be used to treat defects in angiogenesis
and bone marrow failure. In some embodiments the HSCs described
herein can be administered to a subject to treat angiogenesis and
bone marrow failure. In some embodiments the described HSCs
administered to a subject to treat angiogenesis and bone marrow
failure are autologous. In some embodiments the described HSCs
administered to a subject to treat angiogenesis and bone marrow
failure are heterologous.
The MSCs described herein may be used to treat a subject with a
disorder of the hematopoietic system. In some embodiments the MSCs
may be administered to a subject having an inflammatory,
autoimmune, or degenerative disorder. Examples of such disorders
include repair of infarcted myocardium, diabetes, Crohn's disease,
multiple sclerosis, graft-versus-host disease, hepatitis, and many
bone diseases. In some embodiments the described MSCs may be
administered to a subject to repair of infarcted myocardium. In
some embodiments the described MSCs may be administered to a
subject to treat diabetes. In some embodiments the described MSCs
may be administered to a subject to treat Crohn's disease. In some
embodiments the described MSCs may be administered to a subject to
treat multiple sclerosis. In some embodiments the described MSCs
may be administered to a subject to treat graft-versus-host
disease. In some embodiments the described MSCs may be administered
to a subject to treat hepatitis. In some embodiments the described
MSCs may be administered to a subject to treat a bone disease. In
some embodiments the described MSCs administered to a subject to
repair of infarcted myocardium are autologous. In some embodiments
the described MSCs administered to a subject to treat diabetes are
autologous. In some embodiments the described MSCs administered to
a subject to treat Crohn's disease are autologous. In some
embodiments the described MSCs administered to a subject to treat
multiple sclerosis are autologous. In some embodiments the
described MSCs administered to a subject to treat graft-versus-host
disease are autologous. In some embodiments the described MSCs
administered to a subject to treat hepatitis are autologous. In
some embodiments the described MSCs administered to a subject to
treat a bone disease are autologous. In some embodiments the
described MSCs administered to a subject to repair of infarcted
myocardium are heterologous. In some embodiments the described MSCs
administered to a subject to treat diabetes are heterologous. In
some embodiments the described MSCs administered to a subject to
treat Crohn's disease are heterologous. In some embodiments the
described MSCs administered to a subject to treat multiple
sclerosis are heterologous. In some embodiments the described MSCs
administered to a subject to treat graft-versus-host disease are
heterologous. In some embodiments the described MSCs administered
to a subject to treat hepatitis are heterologous. In some
embodiments the described MSCs administered to a subject to treat a
bone disease are heterologous. Those skilled in the art will
understand that the forgoing disclosure provides only a small
listing of inflammatory, autoimmune, or degenerative disorders that
may be treated using the described MSCs; therefore, treatment of
such disorders known to be susceptible to stem cell therapy should
be considered to be within the scope of this disclosure.
The smooth muscle cells described herein may be used to treat a
subject with a disorder of the cardiac or circulatory system. In
some embodiments the SMCs may be administered to a subject having
myocardial tissue damage, blood vessel damage, arterial disease due
to lack of contractility. In some embodiments the described SMCs
may be administered to a subject to treat myocardial tissue damage.
In some embodiments the described SMCs may be administered to a
subject to treat blood vessel damage. In some embodiments the
described SMCs may be administered to a subject to treat arterial
disease due to lack of contractility. In some embodiments the
described SMCs administered to a subject to treat myocardial tissue
damage are autologous. In some embodiments the described SMCs
administered to a subject to treat blood vessel damage are
autologous. In some embodiments the described SMCs administered to
a subject to treat arterial disease due to lack of contractility
are autologous. In some embodiments the described SMCs administered
to a subject to treat myocardial tissue damage are heterologous. In
some embodiments the described SMCs administered to a subject to
treat blood vessel damage are heterologous. In some embodiments the
described SMCs administered to a subject to treat arterial disease
due to lack of contractility are heterologous. Those skilled in the
art will understand that the forgoing disclosure provides only a
small listing of the disorders that may be treated using the
described SMCs; therefore, treatment of such disorders known to be
susceptible to such therapy should be considered to be within the
scope of this disclosure.
The unlimited functional endothelial cells described herein may
also be used to treat a subject with a disorder of the cardiac or
circulatory system. In some embodiments the UFECs may be
administered to a subject to treat circulatory or cardiac damage
following heart attack, such as poor contractility. In some
embodiments the UFECs may be administered to a subject to treat
pulmonary arterial hypertension. In some embodiments the UFECs may
be administered to a subject to treat ischemic conditions such as,
diabetes, where neovascularization may be beneficial. In
administering the treatments described herein the UFECs
administered may be autologous. In other embodiments, however, the
UFECs administered may be heterologous. Those skilled in the art
will understand that the forgoing disclosure provides only a small
listing of the disorders that may be treated using the described
UFECs; therefore, treatment of such disorders known to be
susceptible to such therapy should be considered to be within the
scope of this disclosure.
Methods and compositions for therapeutic administration of the
described cells to a subject are commonly known in the art and
would be readily apparent to a skilled person in the field. For
example, the described cells may be suspended in a pharmaceutically
acceptable carrier, buffer, or other solution that is suitable for
use with living cells to allow the cells to be administered to a
subject. Routes of administration may include injection,
catheter-based delivery, infusion, and the like. Other suitable
means and routes of administration will be appreciated by those
skilled in the related art and are considered to be within the
scope of this disclosure.
The following examples are provided to supplement the prior
disclosure and to provide a better understanding of the subject
matter described herein. These examples should not be considered to
limit the described subject matter. It is understood that the
examples and embodiments described herein are for illustrative
purposes only and that various modifications or changes in light
thereof will be apparent to persons skilled in the art and are to
be included within, and can be made without departing from, the
true scope of the invention.
Example 1--Production of Induced Pluripotent Stem Cells
Multiple human induced pluripotent stem cells (hiPSCs) were
successfully generated from human umbilical vein endothelial cells
(HUVECs) and fibroblasts by mediated expression of four
transcription factors (Oct4, Sox2, klf4, and c-MYC). Those cells
were continually cultured and propagated in a feeder free
environment under chemically defined conditions. The hiPSCs
maintained normal karyotype, and exhibited similar properties to
human embryonic stem cells (hESCs) including self-renewing and
differentiation into all three embryonic germ layers (FIG. 1).
Example 2--Production of Cells of a Mesodermal Lineage by Culturing
hiPSCs in MDM
Studies were conducted to assess the impact of certain defined
media on more than 10 hiPSC lines derived from both human umbilical
vein endothelial cells (HUVECs) and fibroblasts. The gene
expression profile of differentiation cells was analyzed by FACS
and biologic function assays. Over a 12-day period of culturing
hiPSCs in MDM (as set forth in Table 2) the cells were observed to
be prone to commitment to the mesoderm lineage (FIG. 3) and
efficiently generated mesoderm precursors (FIG. 4). In 10
independent experiments, cultures of 1.times.10.sup.5 hiPSCs
generated large numbers of mesoderm precursors peaking at
30.+-.7.times.10.sup.6 (CD31.sup.+ cells: 51%.+-.4.5, and
CD34.sup.+ cells: 37.%.+-.3.6%) on day 10, and declining
thereafter. The results indicate this culture method produced an
approximate 100-fold increase in human CD34/CD31 positive cells
compared to previously published protocols (see e.g. Wang et al.,
Nat. Biotechnol., 25:317 (2007); Goldman et al., Stem Cells 27:1750
(2009); Lancrin et al., Nature, 457:892 (2009); Morishima, et al.,
J. Cell Physiol., 226:1283 (2011); Rufaihah, et al, Arterioscler.
Thromb. Vasc. Biol., 31:e72 (2011); Tolar J et al., Blood, 117:839
(2011); Niwa, et al. PLoS ONE, 6:e22261(2011); Xu Y, et al. PLoS
ONE, 7:e34321 (2012); White, et al., Stem Cells, 31:92 (2013).
Example 3--Mesoderm Precursor Cells are Multipotent
In order to assess the differentiation potential of mesoderm
precursors (CD31/CD34+ cells) produced from hiPSCs, the cells were
incubated under culture conditions known to induce endothelial cell
formation by precursor cells. Various culture conditions induced
the MPCs to develop into: unlimited functional endothelium
(iUFECs), mesenchymal stem cells (iMSC), smooth muscle cells
(iSMCs), and hematopoietic stem cells (iHSCs).
The mesoderm precursors were cultured under conditions promoting
endothelial cell propagation and maturation, standard culture
conditions (37.degree. C. incubation at 5% CO2) using EGM.TM. 2
medium (Lanza). The cells expanded rapidly with typical
cobblestone-like morphology, and expressed endothelial markers
(CD31, vWF, and VE-cadherin (CD144)) as characterized by
immunohistochemistry (FIG. 5A). The induced endothelial cells
(iECs) exhibited further functional features of endothelial cells,
as confirmed through the formation of vascular-like structures on
Matrigel.TM. and the uptake of acetylated-LDL (FIG. 5B). Notably,
these cells were able to be propagated more than 20 passages while
sustaining an endothelial phenotype, based on cobblestone-like
morphology, expression of endothelial markers, and biologic
functions.
Mesoderm precursor cells were also cultured under conditions
suitable for maintaining MSCs in culture (standard culture
conditions (37.degree. C. incubator at 5% CO2) with MSC growth
medium (15.about.20% fetal-bovine serum and 1%
penicillin--streptomycin in alpha minimal essential medium)), which
revealed the mesoderm precursors can be induced to form MSC-like
cells. The MSC phenotype of these cells was validated by observing
the expression of CD90, CD73, and CD105, but not CD31 or CD45, as
detected by FACS analysis (FIG. 6A). In addition to surface marker
analysis, the most common and reliable way to identify a population
of MSC is to verify their ability to differentiate into adipocytes,
osteoblasts, myocytes, and chondrocytes in vivo and in vitro. To
assess this function potential, the iMSCs derived from mesoderm
precursors were cultured under conditions known to promote
differentiation into osteoblasts (standard culture conditions
(37.degree. C. incubator at 5% CO2) with osteoblasts induction
medium containing: 0.1 .mu.M dexamethasone, 50 .mu.M ascorbic
acid-2-phosphate, 10 mM (3-glycerol phosphate, 10% fetal-bovine
serum, and 1% penicillin--streptomycin in alpha minimal essential
medium) for 21 days, changing media every 4 to 5 days). The
resulting cells were histochemically stained to determine their
specific marker profile. After a one-week induction period, the
cells featured very high phosphatase activity and a vast
extracellular calcium deposit confirmed as Alizarin Red S staining
followed by additional two weeks induction. These results indicate
the iMSCs have the ability to differentiate into osteoblasts and
mediate in vitro bone-formation (FIG. 6B).
The mesoderm precursors were cultured under conditions promoting
SMC propagation and maturation (standard culture conditions
(37.degree. C. incubator at 5% CO2) with SMC growth medium
(SmGM-2.TM., Lonza)). This caused the mesodermal precursor cells to
display SMC-like properties, such as a spindle-like morphology and
the strong expression of smooth muscle-specific markers, including
.alpha.-SMA, calponin, and SM22, as confirmed by FACS analysis and
immunohistochemistry (FIG. 5C). These results indicate that
mesoderm precursors derived from hiPSCs have great potential to
produce SMCs.
To test the hematopoietic lineage differentiation potential, the
hiPSCs were cultured in Matrigel.TM. coated plates in MDM for up to
three weeks. Some of the attached mesoderm precursor cells began to
float starting on day 3 in the culture, and the numbers gradually
increased with time. The suspension cells exhibited morphology
reminiscent of primitive human hematopoietic stem/progenitors (FIG.
7). The suspension cell population expressed hematopoietic
stem/progenitors markers CD34, CD31, and CD45 as confirmed by FACS
(FIG. 8A). In 10 independent experiments, cultures of
1.times.10.sup.5 hiPSCs resulted in remarkably large numbers of
suspension cells peaking at 17.+-.5.times.10.sup.6 on day 12, and
rapidly declining thereafter, consistent with a precise timing of
appearance of hematopoietic cells in the system. The addition of
hematopoietic stem/progenitors growth factors (SCF, Flt-3 ligand,
and thrombopoietin) (i.e., MDM+ medium) in the culture promoted
significant generation of hematopoietic stem/progenitors (FIG. 8A).
Thus, up to 4-6.times.10.sup.6 human CD34.sup.+ cells were
generated at day 12 from an initial culture of 1.times.10.sup.5
hiPSCs, representing an about 500-1000 fold increase in human
CD34.sup.+ cells as compared to previously published protocols
based on EB formation or co-culture on stromal cells.
Interestingly, the majority of CD34.sup.+ cells derived from this
procedure hold a CD38 negative phenotype, consistent with an
immature hematopoietic stem/progenitor cell population. To further
validate their hematopoietic differentiation ability a clonogenic
progenitor assay was performed. The suspension cells generated a
large number of erythroid (burst-forming unit-erythroid (BFU-E)),
myeloid (colony-forming unit-granulocytic, monocytic (CFU-GM)), and
mixed (CFU-granulocytic, erythrocytic, monocytic, megakaryocytic
(CFU-GEMM)) colonies (FIG. 8B). The erythroid and myeloid nature of
BFU-E and CFU-GM colonies was confirmed by expression of the
glycophorin and CD33 markers in these colonies, respectively.
Example 4--Modified Composition of Culture Medium
Table 5 shows a particular embodiment of MDM1.
TABLE-US-00005 TABLE 5 A particular embodiment of MDM1. Ingredient
Amount Iscove's Modified Dulbecco's Medium (IMDM, 1:1 mixture
Invitrogen, Catalog#: 21056-023) mixed with Ham's F-12 Nutrient
Mix, with L-alanyl-L-glutamine (GlutaMax .TM.) additive
(Invitrogen, Catalog#, 31765-035) Albucult .TM. 5 mg/ml
.alpha.-monothioglycerol (350 .mu.M-450 .mu.M) 3.9 .mu.l per 100 ml
protein-free hybridoma mixture II (Invitrogen Catalog#: 5% of total
12040-077) volume L-ascorbic acid 2-phosphate (Sigma-Aldrich,
Catalog#: 50 .mu.g/ml A 8960) L-alanyl-L-glutamine (GlutaMax .TM.)
(2 mM, Invitrogen, 2 mM Catalog#: 35050061) Antibiotic (Invitrogen,
Catalog#: 15140122) 50 units pen. 50 mg strep.
insulin-transferrin-selenium-ethanolamine supplement 1% of total
(Invitrogen, Catalog#: 515000560) volume bone morphogenic protein 4
(R&D systems, Catalog#: 10 ng/ml 314-BP-050) vascular
endothelial growth factor (Invitrogen, Catalog#: 10 ng/ml PHC9394)
basic fibroblast growth factor (Pepro Tech, Catalog#: 10 ng/ml
100-18B)
MDM supplemented with hematopoietic cytokines (SCF, Flt-3 ligand
and TPO) is named MDM+. MDM1 supplemented with hematopoietic
cytokines (SCF, Flt-3 ligand and TPO) is referred to as MDM1+.
Table 6 shows the composition of MDM1+.
TABLE-US-00006 TABLE 6 Composition of MDM1 supplemented with
hematopoietic cytokines (MDM1+). Ingredient Amount MDM1 See Table 4
Recombinant human Stem Cell Factor (rhSCF) 50 ng/mL Recombinant
human Flt-3 ligand (rhFlt-3L) 50 ng/mL Recombinant human
Thrombopoietin (rhTPO) 50 ng/mL
Compared to MDM or MDM+, iPSC differentiation with MDM1 or MDM1+
increases significantly the total number of supernatant cells (FIG.
9). This increase in the total number of supernatant cells resulted
in a 8.7-fold increase in CD45+ cells hematopoietic stem cells
(HSCs) at day 10 when MDM1 was employed during the differentiation
process compared to MDM (FIG. 10).
Example 5--Modified Differentiation Protocol to Favor Hematopoietic
Differentiation
Human iPSC-derived supernatant cells were differentiated with MDM1
and MDM1+ using the otherwise unmodified protocol described in
Examples 1 to 3. At day 10 of differentiation, the differentiation
protocol was modified to further support hematopoietic
differentiation. The protocol is unchanged for differentiation of
the other 3 cell types described in Examples 1 to 3, namely,
mesenchymal stem cells (MSC), smooth muscle cells (SMC), and
unlimited functional endothelial cells (UFEC).
For hematopoietic differentiation, supernatant cells obtained from
day 10 of differentiation were cultured in the upper chamber of a
Transwell.TM. insert (Corning Inc.) in commercially available
hematopoietic culture medium containing: 90% RPMI 1640 Medium
(Gibco, Cat#11875-119), 10% Fetal Bovine Serum (Atlanta
Biologicals, Cat# S10250), recombinant human SCF (Stemcell
Technologies Inc.) at 100 ng/ml, recombinant human Flt-3 ligand
(Stemcell Technologies Inc.) at 100 ng/ml, and recombinant human
thrombopoietin (Stemcell Technologies Inc.) at 100 ng/ml, and
recombinant human granulocyte/macrophage colony-stimulating
factorat at 100 ng/ml (PeproTech, Cat# AF-300-03). The lower
chamber of the Transwell insert was occupied by human umbilical
vein endothelial cells (HUVEC) cultured in commercially available
endothelial cell culture medium EGM.TM. BulletKit.TM. (Lonza, Cat#
cc-3162). (FIG. 11).
Under these co-culture conditions (in an incubator at 37.degree. C.
under 5% CO.sub.2), the majority of the cells within the Transwell
(85.47%.+-.9.5) gradually matured into hematopoietic
stem/progenitor cells (HSPC), displaying the hematopoietic lineage
markers CD34 and CD45, as confirmed by flow cytometry. After 7 days
of co-culture in a Transwell insert, these cells are further
differentiated to the monocytic lineage using a medium containing:
90% RPMI 1640 Medium (Gibco, Cat#11875-119), 10% Fetal Bovine Serum
(FBS, Atlanta Biologicals, Cat# S10250), recombinant human SCF
(Stemcell Technologies Inc.) at 100 ng/ml, recombinant human Flt-3
ligand (Stemcell Technologies Inc.) at 100 ng/ml, and recombinant
human thrombopoietin (Stemcell Technologies Inc.) at 100 ng/ml,
recombinant human granulocyte/macrophage colony-stimulating
factorat at 100 ng/ml (PeproTech, Cat# AF-300-03), and recombinant
human macrophage colony-stimulating factor at 100 ng/ml (PeproTech,
Cat# AF-300-25). Phenotypic expression of monocytic markers (CD14,
CD11b, and CD115) is confirmed by flow cytometry. These monocytes
can subsequently be directed to differentiate into functional
macrophages by using attachment cell culture conditions followed by
RPMI 1640 (Gibco) supplemented with 10% FBS, 2 mmol/L L-glutamine,
penicillin/streptomycin, and recombinant human
granulocyte/macrophage colony-stimulating factor at 20 ng/ml
(PeproTech), as confirmed by the in vitro phagocytosis assay.
(Joachim Weischenfeldt and Bo Porse, Bone Marrow-Derived
Macrophages (BMM): Isolation and Applications. Cold Spring Harbor
Protocol, 2008; 3:1) (FIG. 11).
* * * * *