U.S. patent application number 10/024880 was filed with the patent office on 2002-07-18 for cardiovascular components for transplantation and methods of making thereof.
Invention is credited to Bell, Eugene.
Application Number | 20020094573 10/024880 |
Document ID | / |
Family ID | 22950575 |
Filed Date | 2002-07-18 |
United States Patent
Application |
20020094573 |
Kind Code |
A1 |
Bell, Eugene |
July 18, 2002 |
Cardiovascular components for transplantation and methods of making
thereof
Abstract
Cardiovascular components such as biocompatible heart valves and
annular sewing rings are disclosed, as well as, methods for making
the same. The heart valves include biodegradable polymer fiber
scaffolds and collagen. Also disclosed are donor aortic heart
valves processed without the use of crosslinking chemicals.
Inventors: |
Bell, Eugene; (Boston,
MA) |
Correspondence
Address: |
ELLEN LEONNIG
TEI BIOSCIENCES, INC.
7 ELKINS STREET
BOSTON
MA
02127
US
|
Family ID: |
22950575 |
Appl. No.: |
10/024880 |
Filed: |
December 19, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10024880 |
Dec 19, 2001 |
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09420527 |
Oct 19, 1999 |
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60251125 |
Dec 4, 2000 |
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Current U.S.
Class: |
435/398 ;
264/222; 623/2.12; 623/900 |
Current CPC
Class: |
A61L 27/3691 20130101;
A61L 27/507 20130101; A61L 27/362 20130101; A61F 2/2415 20130101;
C12N 2502/02 20130101; A61L 2430/40 20130101; A61L 27/3804
20130101; A61L 27/3604 20130101; C12N 2533/54 20130101; A61K 35/44
20130101; C12N 2502/14 20130101; A61L 27/3687 20130101; C12N 5/0656
20130101; A61K 35/34 20130101; C12N 2533/50 20130101 |
Class at
Publication: |
435/398 ;
264/222; 623/2.12; 623/900 |
International
Class: |
A61F 002/24 |
Claims
What is claimed is:
1. A semilunar heart valve, comprising a biodegradable polymer
fiber scaffold and collagen.
2. The semilunar heart valve of claim 1, wherein the biodegradable
polymer fiber scaffold is a biopolymer fiber scaffold.
3. The semilunar heart valve of claim 1, wherein the collagen is
porcine fetal collagen.
4. The semilunar heart valve of claim 1, wherein the collagen is
fibrillar collagen.
5. The semilunar heart valve of claim 4, wherein the fibrillar
collagen is liquid dense fibrillar collagen.
6. The semilunar heart valve of claim 2, wherein the biopolymer
fiber scaffold is a collagen biopolymer scaffold.
7. The semilunar heart valve of claim 6, wherein the collagen is
selected from the group consisting or collagen type I, collagen
type II, collagen type III, collagen type IV, collagen type V,
collagen type VI, collagen type VII, collagen type VIII, collagen
type IX, collagen type X, collagen type XI, collagen type XII,
collagen type XIII, collagen type XIV, and collagen type XVII.
8. The semilunar heart valve of claim 6, wherein the collagen
biopolymer scaffold is crosslinked.
9. The semilunar heart valve of claim 1, wherein the biodegradable
polymer fiber scaffold is derived from an aortic porcine valve
processed without a crosslinking agent.
10. The semilunar heart valve of claim 1, further comprising
signaling molecules.
11. The semilunar heart valve of claim 1, wherein the polymer
scaffold has a structure determined by a digital program.
12. A method of making a semilunar heart valve, comprising the
steps of: (a) assembling a mold which replicates the structure of a
semilunar heart valve having between two lateral edges a hollow
representing the aortic root and hollows representing a plurality
of leaflets with outer and inner surfaces, the inner surfaces of
the hollows representing the plurality of leaflets connecting with
the hollow representing the aortic root and forming the intimal
surface of the hollow representing the aortic root; (b) covering
the intimal surface of the hollow representing the aortic root and
the outside surface of the hollow representing the plurality of
leaflets with a biodegradable polymer fiber scaffold; (c) filling
the hollow representing the aortic root and the hollows
representing the plurality of leaflets with collagen; and (d)
freeze-drying the polymer fiber scaffold and the collagen forming a
tissue with two lateral edges.
13. The method of making a semilunar heart valve of claim 12,
wherein the biodegradable fiber scaffold is a biopolymer fiber
scaffold.
14. The method of making a semilunar heart valve of claim 12,
wherein the collagen is porcine fetal collagen.
15. The method of making a semilunar heart valve of claim 12,
wherein the collagen is fibrillar collagen.
16. The method of making a semilunar heart valve of claim 12,
wherein the collagen is enriched with signaling molecules.
17. The method of making a semilunar heart valve of claim 16,
wherein the signaling molecules are selected from the group
consisting of sonic hedgehog; NK-2, XNKx-3.3 (tinman), hCsx and Gax
homeobox gene products; TGFbeta, VEGF, FGF, IGF, PDGF, and BMP4
cytokine proteins.
18. The method of making a semilunar heart valve of claim 12,
further comprising the steps of removing the tissue from the mold,
and sewing together the two lateral edges of the tissue.
19. The method of making a semilunar heart valve of claim 12,
further comprising the steps of, seeding the tissue with cells
which normally populate human semilunar valve tissue.
20. The method of making a semilunar heart valve of claim 19,
wherein the cells are selected from the group consisting of
fibrosa, spongiosa, and ventricularis cells.
21. The method of making a semilunar heart valve of claim 21,
further comprising the step of culturing the cells.
22. The method of making a semilunar heart valve of either claim 12
or 19, further comprising the step of seeding the tissue with
endothelial or mesothelial cells.
23. An annular sewing ring for attachment of a heart valve to the
aortic wall of a host, comprising: a biopolymer cloth and a
biopolymer rope shaped in a circle, wherein the biopolymer cloth is
wrapped around and stitched to the biopolymer rope.
24. The annular sewing ring of claim 23, wherein the biopolymer
fiber cloth is collagen cloth and the biopolymer rope is collagen
rope.
25. The annular sewing ring of claim 24, wherein the collagen is
selected from the group consisting of collagen type I, collagen
type II, collagen type III, collagen type IV, collagen type V,
collagen type VI, collagen type VII, collagen type VIII, collagen
type IX, collagen type X, collagen type XI, collagen type XII,
collagen type XIII, collagen type XIV, and collagen type XVII.
26. The annular sewing ring of claim 23, wherein the ring is seeded
with cells which normally populate semilunar valve tissue.
27. The annular sewing ring of claim 23, wherein the ring is
enriched with signaling molecules.
28. The annular sewing ring of claim 23, wherein the signaling
molecules are selected from the group consisting of sonic hedgehog;
NK-2, XNKx-3.3 (tinman), hCsx and Gax homeobox gene products;
TGFbeta, VEGF, FGF, IGF, PDGF, and BMP4 cytokine proteins.
29. A semilunar heart valve made according to the method comprising
the steps of: (a) assembling a mold which replicates the structure
of a semilunar heart valve having between two lateral edges a
hollow representing the aortic root and hollows representing a
plurality of leaflets with outer and inner surfaces, the inner
surfaces connecting with the hollow representing the aortic root
and forming the intimal surface of the hollow representing the
aortic root; (b) covering the intimal surface of the hollow
representing the aortic root and the outside surface of the hollow
representing the plurality of leaflets with a biopolymer fiber
scaffold; (c) filling the hollow representing the aortic root and
the hollows representing the plurality of leaflets with collagen;
and (d) freeze-drying the biopolymer fiber scaffold and the liquid
dense fibrillar collagen forming a tissue with two lateral edges.
Description
BACKGROUND OF THE INVENTION
[0001] The heart includes four natural valves that function to
regulate flow direction as blood is pumped between the lungs and
the various blood vessels. The mitral and tricuspid valves, which
are known as the atrioventricular or intraflow valves operate to
prevent backflow into the atria during ventricular contraction
while permitting blood to flow therethrough during ventricular
relaxation. The aortic and pulmonary valves are known as semilunar
or outflow valves and are located where blood leaves the heart.
[0002] Semilunar valves consist of three membranous cup-like
structures or cusps attached, at the same level, to the wall of a
cylindrical aortic vessel so that the cusps press on each other
when they are filled with blood, preventing backflow in diastole.
The direction of blood flow is upward. On contraction of the
vessel, that is during systole, the cusps are pressed against the
vessel wall by the force of blood flowing past the attached edges
of the cusps toward the free edges of the cusps, allowing the blood
to flow freely.
[0003] Each open pocket of the semilunar valve defines a volume
called the aortic sinus which is filled with blood when the valve
is closed. If the leaflet is cut away from the wall of the aorta it
can be spread out in the form of a flat hemicircular membrane. The
hemicircle is the edge of the leaflet which is attached to the wall
of the aorta while the top more or less linear edge was the free
edge of the leaflet. Each end of the leaflets called a commisure.
The work of A. A. H. J. Sauren (The Mechanical behavior of the
Aortic Valve (PhD thesis) Eindhoven, The Netherlands: Eindhoven
Technical University, 1981), which is incorporated herein by
reference, has shown in whole mounts of leaflets that the
supporting scaffold of the leaflet consists of collagen fibers,
having fractile properties, which extend from one commisure to the
other providing support for the applied load of blood. Equations
which describe the fiber system of the leaflet have been derived by
C. S. Perkin and D. M. McQueen (Mechanical equilibrium determines
the fractile fiber architecture of aortic heart valve leaflets. Am.
J. Physiol. 266, H319-H328, 1994) from their function which is to
support a uniform load when the aortic valve is closed. What they
find is a single parameter family of collagen fibers with fractile
properties which compare closely with the whole mount fiber
preparations. Their work serves as the basis for creating a digital
program which a textile machine, or a sewing machine, could use to
reproduce an approximation of the fiber scaffold of the valve
leaflet.
[0004] Histologically the leaflet consists of three tissue layers,
the fibrosa, the spongiosa and the ventricularis. The fibrosa of
the leaflet faces the aortic wall, enclosing the fiber system
described above; the fiber scaffold is arranged in corrugated
fashion permitting radial expansion of the valve leaflet. Adjacent
to the fibrosa is the spongiosa, a loosely organized connective
tissue with collagen elastin, proteoglycans and
mucopolysaccharides. Furthest away from the aortic wall is the
ventricularis consisting of a sheet of elastin thought to provide
the tensile recoil needed to maintain the corrugated shape of the
fibrosa. The surfaces of the leaflets in contact with the blood are
covered by a layer of endothelial cells.
[0005] Heart valves, e.g., semilunar valves, are deformed by a
variety of pathological processes. In many cases the diseased or
defective valve can be surgically removed and replaced with a
prosthetic valve. Two main types of artificial valves exist: (1)
mechanical valves made from metal or plastic material; and (2)
valves made from animal tissue.
[0006] Artificial valves, whether mechanical or made from animal
tissue, have serious drawbacks. For example, mechanical valves
carry a significant risk of thrombus formation. Also, the stress
associated with the junction between the stent or frame and the
biological portion of the bioprosthetic valve appears to be
involved in structural failure over time. Valves made from animal
tissue are typically crosslinked with chemicals, e.g.,
glutaraldehyde during processing. Treatment of the animal tissue
with glutaraldehyde causes calcification and/or the structural
breakdown of the tissue, thus, reducing the area available as
binding sites for human host cells. In addition, both mechanical
valves and valves constructed from animal tissue do not have the
capacity to grow, i.e., these types of valves can neither be
occupied or remodeled by host cells nor can they be biologically
integrated.
[0007] A need exists, therefore, for an improved prosthetic heart
valve that overcomes or minimizes the above-mentioned problems.
SUMMARY OF THE INVENTION
[0008] The invention features novel biocompatible cardiovascular
components, e.g., semilunar heart valves, for transplantation. The
invention also features methods for constructing these novel
biocompatible cardiovascular components which preserve the nativity
of the biological materials used. In addition, the invention
features a novel annular sewing ring for attachment of a
cardiovascular component to the aortic wall of a host. The
components can be used in vitro, for example, for model systems for
research, or in vivo as prostheses or implants to replace diseased
or defective heart valves. In either case, the valves can be seeded
with cells, e.g., spongiosa cells, fibrosa cells, ventricularis
cells, smooth muscle cells, and/or endothelial and mesothelial
cells.
[0009] In one aspect of the invention, the cardiovascular component
is a semilunar valve which includes a biodegradable polymer fiber
scaffold, e.g., a biopolymer fiber scaffold, and collagen. In a
preferred embodiment, the collagen is fetal porcine collagen. In
another preferred embodiment, the collagen is fibrillar collagen.
In yet another preferred embodiment, the biopolymer fiber scaffold
is a collagen biopolymer scaffold.
[0010] In another aspect of the invention, the cardiovascular
component is a semilunar valve which includes a biodegradable
polymer fiber scaffold, e.g., a biopolymer fiber scaffold, and
collagen wherein the biopolymer scaffold fiber is derived from an
aortic porcine valve processed in the absence of a crosslinking
agent, e.g., glutaraldehyde.
[0011] In yet another aspect of the invention, the cardiovascular
component is a semilunar valve which includes a biodegradable fiber
scaffold, e.g., a biopolymer fiber scaffold, and collagen wherein
the scaffold has a structure determined by a digital program.
[0012] The invention further pertains to a method of making a
semilunar heart valve, comprising the steps of: (a) assembling a
mold which replicates the structure of a semilunar heart valve
having between two lateral edges a hollow representing the aortic
root and hollows representing a plurality of leaflets with outer
and inner surfaces, the inner surfaces connecting with the hollow
representing the aortic root, thus, forming the intimal surface of
the aortic root; (b) covering the intimal surface of the hollow
representing the aortic root, i.e., the surface of the hollow
representing the aortic root which connects, with the hollow
representing the valve leaflets, and the outside surface of the
hollow representing the valve leaflets, i.e., the surface away from
the aortic wall with a biodegradable polymer fiber scaffold; (c)
filling the hollow representing the aortic root and the hollows
representing the plurality of leaflets with collagen, e.g., fetal
porcine collagen, fibrillar collagen e.g., liquid dense fibrillar
collagen; and (d) freeze-drying the polymer fiber scaffold and the
liquid dense collagen forming a tissue with two lateral edges.
[0013] The invention still further pertains to an annular sewing
ring for attachment of a heart valve to the aortic wall of a host
which includes a biopolymer cloth and a biopolymer rope shaped in a
circle, wherein the biopolymer cloth is wrapped around and stitched
to the biopolymer rope.
[0014] The invention yet further pertains to a semilunar heart
valve made according to a method which includes the steps of: (a)
assembling a mold which replicates the structure of a semilunar
heart valve having between two lateral edges a hollow representing
the aortic root and hollows representing a plurality of leaflets
with outer and inner surfaces, the inner surfaces connecting with
the hollow representing the aortic root; (b) covering the intimal
surface of the hollow representing the aortic root and the outside
surface of the hollow representing the plurality of leaflets with a
biodegradable polymer fiber scaffold; (c) filling the hollow
representing the aortic root and the hollows representing the
plurality of leaflets with liquid dense collagen; and (d)
freeze-drying the polymer fiber scaffold and the liquid dense
collagen forming a tissue with two lateral edges.
BRIEF DESCRIPTION OF THE DRAWINGS
[0015] FIG. 1 shows the aorta slit open longitudinally and laid
flat so that the structure of the valve leaflets of the semilunar
valve can be displayed.
[0016] FIG. 2 shows the valve leaflets of the semilunar valve from
below in the closed condition; filled with blood and pressing on
each other thereby preventing backflow.
[0017] FIG. 3 shows the valve leaflets of the semilunar valve
schematically in longitudinal section.
[0018] FIG. 4 shows the mold design for constructing a semilunar
heart valve.
[0019] FIG. 4A shows the back cover of the mold which represents
the outside of the aorta.
[0020] FIG. 4B shows the front cover of the mold which represents
the inside of the aorta displaying the attached leaflet molds.
[0021] FIG. 4C shows the back side of the front cover of the mold
displaying the back side of the leaflet molds as they join with the
aorta.
DETAILED DESCRIPTION OF THE INVENTION
[0022] The features and other details of the invention will now be
more particularly described and pointed out in the claims. It will
be understood that the particular embodiments of the invention are
shown by way of illustration and not as limitations of the
invention. The principle features of this invention can be employed
in various embodiments without departing from the scope of the
invention.
[0023] The present invention features novel biocompatible
cardiovascular components for transplantation, e.g., heart valves,
e.g., semilunar heart valves. The term "biocompatible" as that term
is used herein, means exhibition of essentially no cytotoxicity
while in contact with body fluids or tissues. "Biocompatibility"
also includes essentially only minimal interactions, i.e.,
interactions leading to immune rejection or to persistent
inflammation responses, with recognition proteins, e.g., naturally
occurring antibodies, cell proteins, cells, and other components of
biological systems. The invention also features methods of making
these components which preserves the nativity of the biological
material comprising the components.
[0024] In one aspect of the invention, the cardiovascular component
is a heart valve, e.g., a semilunar heart valve which includes a
biodegradable polymer fiber scaffold, e.g., a biopolymer fiber
scaffold, and collagen. A semilunar heart valve is composed of
three membranous cup-like structures or cusps attached, at the same
level, to the wall of a cylindrical arterial vessel, e.g., the
aorta, so that the cusps press on each other when they are filled
with blood, preventing backflow in diastole. Methods for making the
polymer fibers which comprise the polymer fiber scaffolds are
taught in U.S. Pat. No. 5,851,290, entitled "Apparatus and Method
for Spinning and Processing Collagen Fiber," which is incorporated
herein by reference.
[0025] The term "biodegradable polymers," as that term is used
herein, includes any polymer that naturally degrades or breaks down
over time by hydrolysis, for example, poly-.alpha.-hydroxyesters
such as poly-1-lactic acid and poly-1-glycolic acid, polydioxinone,
polyvinyl alcohol, surgical gut, and combinations thereof, or which
degrades over time by enzymatic action, for example, biopolymer,
e.g., collagen.
[0026] A biopolymer is a naturally occurring polymeric substance
formed from individual molecules in a biological system or
organism. Biopolymers can also be man-made by manipulation of the
individual molecules once obtained outside the biological system or
organism. The biopolymer is suitable for introduction into a living
organism, e.g., a mammal, e.g., a human. The biopolymer is
non-toxic and bioabsorbable when introduced into a living organism
and any degradation products of the biopolymer should also be
non-toxic to the organism. The biopolymers of the invention can be
formed into cardiovascular components, e.g., heart valves, e.g.,
semilunar heart valves, which include biocompatible fibers, e.g.,
collagen fibers, biocompatible fabrics, e.g., collagen fabrics.
Examples of molecules which can form biopolymers and which can be
used in the present invention include collagen, laminin, elastin,
fibronectin, fibrinogen, thrombospondin, gelatin, polysaccharides,
poly-1-amino acids and combinations thereof. In one embodiment, a
combination or mixture of one or more biopolymers can be used to
form the cardiovascular components, e.g., heart valves, e.g.,
semilunar heart valves, of the invention. For example, a
combination of laminin and type IV collagen can be used to form the
biopolymer fibers described herein. A preferred molecule for
biopolymer production is collagen.
[0027] Preferred sources of molecules which form biopolymers
include mammals such as pigs, e.g., near-term fetal pigs, sheep,
fetal sheep, cows, and fetal cows. Other sources of the molecules
which can form biopolymers include both land and marine vertebrates
and invertebrates. In one embodiment, the collagen can be obtained
from skins of near-term, domestic porcine fetuses which are
harvested intact, enclosed in their amniotic membranes. Collagen or
combinations of collagen types can be used in the cardiovascular
components, e.g., heart valves, e.g., semilunar heart valves
described herein. A preferred type of collagen is porcine fetal
collagen. Another preferred type of collagen is fibrillar collagen,
e.g., fibrillar collagen can be produced by processing a solution
of monomeric liquid collagen, e.g., non-polymeric liquid collagen.
Fibrillar collagen is a type of collagen which contains fibrils.
The language "fibrillar collagen" or "collagen microfibril" is art
recognized and is intended to include collagen in the form
described in Williams, B. R. et al. (1978) J. Biol. Chem. 253
(18):6578-6585 and U.S. patent application Ser. No. 08/910,853,
filed Aug. 13, 1997, entitled "Compositions, Devices, and Methods
for Coagulating Blood" by Eugene Bell and Tracy M. Sioussat, the
contents of which are incorporated herein by reference. In a
preferred embodiment, the collagen microfibrils are prepared
according to the methods taught in U.S. patent appln. 60/095,627,
entitled "Bone Precursor Compositions," which are incorporated
herein by reference. Liquid dense fibrillar collagen is fibrillar
collagen in a liquid form which can be dried to a dense fibrillar
tissue, e.g, a matt. Biopolymer and collagen matts are described in
copending patent application Ser. No. 09/042,549, entitled
"Biopolymer Matt for Use in Tissue Repair and Reconstruction," the
contents of which are incorporated herein by reference.
[0028] Examples of collagen or combinations of collagen types
include collagen type I, collagen type II, collagen type III,
collagen type IV, collagen type V, collagen type VI, collagen type
VII, collagen type VIII, collagen type IX, collagen type X,
collagen type XI, collagen type XII, collagen type XIII, collagen
type XIV, and collagen type XVII. A preferred combination of
collagen types includes collagen type I, collagen type III, and
collagen type IV.
[0029] Preferred mammalian tissues from which to extract the
molecules which can form biopolymer include entire mammalian
fetuses, e.g., porcine fetuses, dermis, tendon, muscle and
connective tissue. As a source of collagen, fetal tissues are
advantageous because the collagen in the fetal tissues is not as
heavily crosslinked as in adult tissues. Thus, when the collagen is
extracted using acid extraction, a greater percentage of intact
collagen molecules is obtained from fetal tissues in comparison to
adult tissues. Fetal tissues also include various molecular factors
which are present in normal tissue at different stages of animal
development.
[0030] In a preferred embodiment, the cardiovascular components,
e.g., heart valves, e.g., semilunar heart valves, are collagen
cardiovasular components, e.g., collagen heart valves, e.g.,
collagen semilunar heart valves. Collagen solutions can be produced
by salt extraction, acid extraction, and/or pepsin extraction from
the starting material. In a preferred embodiment, the collagen used
is produced by sequentially purifying two forms of collagen from
the same collagen-containing starting material. First, intact
collagen is acid extracted from the starting material, the extract
is collected and collagen is prepared as a collagen solution, e.g.,
by precipitating the collagen with sodium chloride and solubilizing
the collagen in a medium having an acidic pH. Meanwhile, truncated
collagen, i.e., collagen from which the teleopeptides have been
cleaved or partly cleaved leaving only the helical portion or the
helical portion with some telopeptides, is extracted from the
starting material using enzyme, e.g., an enzyme which is functional
at an acidic pH, e.g., pepsin, extraction. Then, the collagen from
this pepsin extract is purified separately by similar methods as
from the first extract.
[0031] Proteins necessary for cell growth, morphogenesis,
differentiation, and tissue building can also be added to the
biopolymer molecules or to the biopolymer fibrils to further
promote cell ingrowth and tissue development and organization
within the cardiovascular components, e.g., hearts valves, e.g.,
semilunar heart valves. The phrase "proteins necessary for cell
growth, morphogenesis, differentiation, and tissue building" refers
to those molecules, e.g., proteins which participate in the
development of tissue. Such molecules contain biological,
physiological, positional, and structural information for
development or regeneration of the tissue structure and function.
Examples of these macromolecules include, but are not limited to,
sonic hedgehog; NK-2, XNKx-3.3 (tinman), hCsx and Gax homeobox gene
products; TGFbeta, VEGF, FGF, IGF, PDGF, BMP4 cytokine proteins,
growth factors, extracellular matrix proteins, proteoglycans,
glycosaminoglycans and polysaccharides. Alternatively, the
cardiovascular components, e.g., heart valves, e.g., semilunar
heart valves of the invention can include extracellular matrix
macromolecules in particulate form or extracellular matrix
molecules deposited by cells or viable cells or deliberately added
to the valve scaffold. Methods for processing tissues for making
extracellular matrix macromolecules in particulate form are taught
in U.S. Pat. No. 5,800,537, entitled "A Method and Construct for
Producing Graft Tissue From Extracellular Matrix," the contents of
which are incorporated herein by reference.
[0032] The collagen used to create the cardiovascular components,
e.g., heart valves, e.g., semilunar heart valves, may be enriched
with signaling molecules which play a role in vascular development.
Products of three classes of genes are implicated: hedgehog,
homeobox and cytokine. They include but are not limited to the
following proteins: sonic hedgehog; NK-2, XNKx-3.3 (tinman), hCsx
and Gax homeobox gene products; TGFbeta, VEGF, FGF, IGF, PDGF, and
BMP4 cytokine proteins. Differentiation induced by the use of
combinations of the foregoing proteins is promoted by incubation of
the cell laden scaffold in vitro under tissue culture
conditions.
[0033] The term "growth factors" is art recognized and is intended
to include, but is not limited to, one or more of platelet derived
growth factors (PDGF), e.g., PDGF AA, PDGF BB; insulin-like growth
factors (IGF), e.g., IGF-I, IGF-II; fibroblast growth factors
(FGF), e.g., acidic FGF, basic FGF, .beta.-endothelial cell growth
factor, FGF 4, FGF 5, FGF 6, FGF 7, FGF 8, and FGF 9; transforming
growth factors (TGF), e.g., TGF-.beta.1, TGF-.beta.1.2,
TGF-.beta.2, TGF-.beta.3, TGF-.beta.5; vascular endothelial growth
factors (VEGF), e.g., VEGF, epidermal growth factors (EGF), e.g.,
EGF, amphiregulin, betacellulin, heparin binding EGF; interleukins,
e.g., IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10,
IL-11, IL-12, IL-13, IL-14; colony stimulating factors (CSF), e.g.,
CSF-G, CSF-GM, CSF-M, BMP cytokine proteins; nerve growth factor
(NGF); stem cell factor; hepatocyte growth factor, and ciliary
neurotrophic factor. The term encompasses presently unknown growth
factors that may be discovered in the future, since their
characterization as a growth factor will be readily determinable by
persons skilled in the art.
[0034] The term "extracellular matrix proteins" is art recognized
and is intended to include one or more of fibronectin, laminin,
vitronectin, tenascin, entactin, thrombospondin, elastin, gelatin,
collagens, fibrillin, merosin, anchorin, chondronectin, link
protein, bone sialoprotein, epinectin, hyaluronectin, undulin,
epiligrin, and kalinin. The term encompasses presently unknown
extracellular matrix proteins that may be discovered in the future,
since their characterization as an extracellular matrix protein
will be readily determinable by persons skilled in the art.
[0035] The term "proteoglycan" is art recognized and is intended to
include one or more of decorin and dermatan sulfate proteoglycans,
keratin or keratan sulfate proteoglycans, aggrecan or chondroitin
sulfate proteoglycans, heparan sulfate proteoglycans, biglycan,
syndecan, perlecan, or serglycin. The term encompasses presently
unknown proteoglycans that may be discovered in the future, since
their characterization as a proteoglycan will be readily
determinable by persons skilled in the art.
[0036] The term "glycosaminoglycan" is art recognized and is
intended to include one or more of heparan sulfate, chondroitin
sulfate, dermatan sulfate, keratan sulfate, hyaluronic acid. The
term encompasses presently unknown glycosaminoglycans that may be
discovered in the future, since their characterization as a
glycosaminoglycan will be readily determinable by persons skilled
in the art.
[0037] The term "polysaccharide" is art recognized and is intended
to include one or more of heparin, dextran sulfate, chitin, alginic
acid, pectin, and xylan. The term encompasses presently unknown
polysaccharides that may be discovered in the future, since their
characterization as a polysaccharide will be readily determinable
by persons skilled in the art.
[0038] Suitable living cells include, but are not limited to, cells
derived from the layers of tissue comprising the semilunar heart
valve, e.g., spongiosa, fibrosa, and ventricularis cells,
epithelial cells, and mesothelial cells, e.g., keratinocytes,
adipocytes, hepatocytes, neurons, glial cells, astrocytes,
podocytes, mammary epithelial cells, islet cells; endothelial
cells, e.g., aortic, capillary and vein endothelial cells; and
mesenchymal cells, e.g., dermal fibroblasts, mesothelial cells,
stem cells, osteoblasts, smooth muscle cells, striated muscle
cells, ligament fibroblasts, tendon fibroblasts, chondrocytes, and
fibroblasts.
[0039] In one embodiment, when the biopolymer is collagen, the
collagen can be treated with an enzyme, e.g., lysyl oxidase which
primes the collagen for crosslinking. Lysyl oxidase, which can be
purified from a variety of sources including, for example, calf
aorta, human placenta, chicken embryo epiphyseal cartilage, pig
skin, (see Shackleton, D. R. and Hulmes, D. J. S. (1990) Biochem.
J. 266:917-919), and several locations in pig embryos, converts the
68 -amino group of lysine to an aldehyde. This aldehyde is a
reactive functional group which spontaneously binds to other lysine
.epsilon.-amino groups or other aldehydes on other collagen
molecules to form irreversible covalent crosslinks. The result is
that collagen becomes insoluble. Lysyl oxidase can be added to the
collagen solutions under conditions which allow for the aldehyde
conversion of the lysines. The lysyl oxidase is then removed from
the collagen solution and the collagen is processed as described
herein during which the spontaneous crosslinks form. For example,
during the processing of the collagen cardiovascular component,
e.g., during the polymerization step, the crosslinks spontaneously
form as the concentration of collagen per unit volume increases.
The lysyl-oxidase-mediated crosslink is strong, irreversible and is
a linkage naturally found in collagen. Collagen crosslinked in this
manner is insoluble and susceptible only to specific enzymatic
attack during remodeling of tissues. Lysyl oxidase can also be used
to crosslink collagen for use as matt and matt compositions as well
as spun fibers, gels, etc.
[0040] In still another embodiment, the strength of the
cardiovascular component can be increased by standard collagen
crosslinking methods using, e.g., ultraviolet, dehydrothermal, or
chemical crosslinkers. Typical chemical crosslinkers include, for
example, glutaraldehyde, formaldehyde, acrylamide, carbodiimides,
such as those known in the art, e.g.,
1-ethyl-3-(dimethyaminopropyl)carbodiimide, diones known to those
skilled in the art, e.g., 2,5-hexanedione, diimidates, e.g.,
dimethylsuberimidate, or bisacrylamides, e.g.,
N,N'-methylenebisacrylamid- e.
[0041] In still yet another embodiment of the invention, the
cardiovascular components, e.g., heart valves, e.g., semilunar
heart valves, comprise biopolymer fiber scaffold derived from an
aortic porcine valve processed in the absence of a crosslinking
agent, e.g., glutaraldehyde or chemicals similar thereto. By
eliminating the step of treating an aortic porcine valve with
glutaraldehyde or with chemicals similar to glutaraldehyde, the
calcification or structural breakdown of the aortic porcine valve
tissue is eliminated. Accordingly, binding sites for host human
cells and other cells are maintained with the present
invention.
[0042] Moreover another embodiment of the invention, the
cardiovascular components, e.g., heart valve, e.g., semilunar heart
valve, comprise a biodegradable polymer fiber scaffold, e.g., a
biopolymer fiber scaffold, having a structure determined by a
digital program. The work of A. A. H. J. Sauren (The Mechanical
behavior of the Aortic Valve (PhD thesis) Eindhoven, The
Netherlands: Eindhoven Technical University, 1981), which is
incorporated herein by reference, has shown in whole mounts of
leaflets that the supporting scaffold of the leaflet consists of
collagen fibers, having fractile properties, which extend from one
commisure to the other providing support for the applied load of
blood. Equations which describe the fiber system of the leaflet
have been derived by C. S. Perkin and D. M. McQueen (Mechanical
equilibrium determines the fractile fiber architecture of aortic
heart valve leaflets. Am. J. Physiol. 266, H319-H328, 1994) from
their function which is to support a uniform load when the aortic
valve is closed. What they found is a single parameter family of
collagen fibers with fractile properties which compare closely with
the whole mount fiber preparations. Their work serves as the basis
for creating a digital program which a textile machine, or a sewing
machine, could use to reproduce an approximation of the fiber
scaffold of the valve leaflet.
[0043] The present invention also features a novel biocompatible
annular sewing ring for attachment of a heart valve to the aortic
wall of a host. The annular ring is comprised of a biopolymer,
e.g., collagen. In a preferred embodiment, the biopolymer, e.g.,
collagen, is a biopolymer fiber, e.g., collagen fiber.
[0044] Referring to the figures, the semilunar valve of the aorta
consists of three leaflets or cusps which resemble pockets attached
to the wall of the aorta along their inferior edge (FIG. 1) with
the superior edge of each pocket being free. If the wall of the
aorta is slit between two leaflets along a vertical axis and the
aorta is opened, three leaflets resembling pockets, all in a row,
are seen attached to the flattened aortic wall or root. The aorta
or the aortic root and its attached leaflets can be made in a mold;
for example, a mold consisting of three parts which fit together
tightly to make a closed aortic wall to which the three pocket like
structures are attached (FIG. 4).
[0045] The parts of the mold for replicating a semilunar heart
valve can be constructed from inert materials, e.g., stainless
steel or stainless steel coated with Teflon.RTM.. An example of
such a mold is shown in FIG. 4. Referring to FIG. 4, the mold
includes three integral parts, which are shown in views A, B, and
C. The three parts fit together between two lateral edges to form a
box with a hollow interior to form the aortic root and attach to
three valve leaflets by interconnecting U-shaped channels. The
first part (A) is shaped as a rectangular box cover which
represents the back or outside wall of the aorta, e.g., the aortic
root. The second part (B) also employs a rectangular shape and fits
together with the first part A to form the front or inside wall of
the aorta. The second part (B) itself includes a front and back
side which represent the front or inside walls and the back or
outside walls, respectively, of the valve leaflets or cusps which
attach to the front or inside wall of the aorta (A). The front or
inside walls of the valve leaflets are represented on the front
side of the second part (B) as hollow pockets which include the
free edge, the nodulus arantis, the commisures, and the lower edge
of each valve leaflet. The commisures and the lower edge of each
valve leaflet, which form the hollow pockets, communicate with the
aorta and are represented by hollow U-shaped channels which
penetrate the wall of the second part (B and C). The third part
includes a plate (C) which forms the outside face of each valve
leaflet, i.e., the face intimal with the aorta. The third part
completes the hollow channels which form the valve leaflets (B'a
and B'b) and includes a U-shaped curve structure that attaches to
the back of the second part. The bottom of the U-shaped curve of
the third part (B'a) includes a wider diameter than the other areas
of the communicating hollows (B', C) to form the base or floor of
the bottom edge of each valve leaflet.
[0046] With an assembled mold, sheets of a biodegradable polymer
fiber scaffold, e.g., a biopolymer fiber scaffold, e.g., a collagen
fiber scaffold, e.g., a woven (braided, knitted) collagen fiber
scaffold, are laid into the respective hollows (B) of the mold so
that they lie in contact with the intimal surface of the aortic
root and with the outside surface of the valve pockets, i.e., the
surface away from the aortic wall. The sheets of a biodegradable
polymer fiber scaffold, e.g., a biopolymer fiber scaffold, e.g., a
collagen fiber scaffold, e.g., a woven (braided, knitted) collagen
fiber scaffold, extend into the U shaped slit and are bent to
overlap the displaced plate (C) so that they lie in contact with
the intimal surface of the aortic wall. Fiber scaffolds containing
more than one type of biodegradable polymers can also be formed,
for example, by combining a biodegradable polymer fiber scaffold
with a collagen fiber scaffold in the hollows of the mold as
described above.
[0047] In a preferred embodiment, a crosslinked collagen fiber
scaffold is laid into the respective hollows of the mold. In
another preferred embodiment, a biopolymer fiber scaffold comprised
of processed pig heart valves can be laid into the respective
hollows of the mold. In yet another preferred embodiment, sheets of
woven collagen threads constructed from a textile machine can be
laid into the respective hollows of the mold. As described
previously, from equations for the collagen fiber support system
for the valve leaflet or cusp, a digital program can be written for
a textile or sewing machine which can produce a collagen fiber
scaffold in the form of three cusps using the crosslinked collagen
fibers described immediately above. Even without such a program
from information already available the general pattern of the fiber
system can be reproduced. For example each cusp can have the
geometry of the expanded cusp shown in FIG. 3, but cusps would be
connected at the commisures.
[0048] The hollows of the mold are then filled with collagen, e.g.,
fetal porcine, fibrillar collagen, e.g., liquid dense fibrillar
collagen. In a preferred embodiment, the third part of the mold (C,
B'b) is removed and the fiber reinforced collagen which has formed
lining the aortic wall and the valve leaflet is seeded with cells
which would normally populate the respective tissues of the
semilunar valve, e.g., cells derived from the fibrosa, spongiosa,
and ventricularis tissues. The mold can then be introduced into a
cell culturing system to allow attachment, growth, and
differentiation of the seeded cells. In another preferred
embodiment, the valve is seeded with cells which normally populate
the outer tissue covering the semilunar valve, e.g., allogeneic or
autogenous endothelial or mesothelial cells, to provide an
epithelial covering to the valves and aorta. These endothelial or
mesothelial cells are used to populate all tissue surfaces except
the back or outer surface of the aortic wall after the first cells
seeded into the tissue have remodeled the biopolymer structure.
[0049] As described supra, the collagen, e.g., fetal porcine
collagen, e.g., liquid dense fetal porcine collagen, used to fill
the components of the mold may be enriched with signaling molecules
which play a role in vascular development. Cardiovascular complexes
of signaling molecules can be derived from the extracellular matrix
of young or very young porcine fetuses containing the proteins
listed below. Products of three classes of genes are implicated:
hedgehog, homeobox and cytokine. They include but are not limited
to the following proteins: sonic hedgehog; NK-2, XNKx-3.3 (tinman),
hCsx and Gax homeobox gene products; TGFbeta, VEGF, FGF, IGF, PDGF,
and BMP4 cytokine proteins. Differentiation induced by the use of
combinations of the foregoing proteins is promoted by incubation of
the cell laden scaffold in vitro under tissue culture
conditions.
[0050] The tissue which has formed can be lifted out of the mold
and the two lateral edges of the valve are sewn together with a
biopolymer thread, e.g., collagen or synthetic thread, to form a
tubular valve. In a preferred embodiment, additional endothelial or
mesothelial cells may be seeded onto the structure to cover the
intimal surface of the aorta and the surfaces of the valve
leaflets. In another embodiment, the tubular valve can be
incorporated into a pulsatile closed circulatory loop containing a
nutrient fluid having the visco-elastic properties of blood. The
valve can be oriented in the circulation so that during diastole it
back flows to fill the pockets of the valve. Mechanical
conditioning of the valve can be carried out for a period of 2-4
weeks. It can remain in culture ready for delivery on demand.
[0051] As described supra, the heart valves, e.g., semilunar
valves, of animals, e.g., the pig heart, have been used as
substitutes for defective human valves for many years. The usual
procedure for processing the valves after removal from an animal
consists of crosslinking them with chemicals, e.g., glutaraldehyde.
Chemicals such as glutaraldehyde destroy all biological information
associated with the scaffold of the valve. Accordingly, chemicals
such as glutaraldehyde cause the tissue of the removed valve to
calcify and/or breakdown structurally, thus, eliminating binding
sites for cells, e.g., human host cells, which might otherwise be
expected to seed into the leaflets of the valves as well as into
the cuffs of the valves. There is also the loss of binding sites
that would otherwise permit endothelial or mesothelial cells to
populate the surfaces of the valve structures. To overcome these
limitations the following method has been developed to process the
valves after removal from the donor animal. This method eliminates
the chemical crosslinking step, e.g., crosslinking with
glutaraldehyde.
[0052] In a preferred embodiment, the valve, e.g., the semilunar
valve, is removed from the donor animal, e.g., a pig. Using a 10%
solution of NAOH, cellular components are stripped from the
collagen fiber scaffold of the valve leaflets and annulus (the
aortic wall), thus, eliminating viruses, and other microorganisms,
cells and cell surface antigenic determinants retaining the fibrous
scaffold. The processed valve can then be laid into a valve mold,
e.g., the semilunar mold described supra, and be formed into a
heart valve for transplantation.
[0053] In a preferred embodiment, the processed donor valve is laid
into the respective hollows (B) of the mold so that they lie in
contact with the intimal surface of the aortic root and with the
outside surface of the valve pockets, i.e., the surface away from
the aortic wall. The donor valve, e.g., pig valve, e.g., the pig
semilunar valve, extend into the U shaped slit and are bent to
overlap the displaced plate (C) so that they lie in contact with
the intimal surface of the aortic wall. The hollows of the mold are
then filled with collagen, e.g., fetal porcine collagen, or
fibrillar collagen e.g., liquid dense fibrillar collagen. In a
preferred embodiment, the contents of the mold are freeze dried to
form a structure, e.g., a biopolymer foam, e.g., a collagen foam
around the donor valve. Methods for freeze drying collagen and
forming biopolymer foams and collagen foams of varying densities
are taught in U.S. Pat. Nos. 5,891,558 and 5,709,934, each entitled
"Biopolymer Foams for Use in Tissue Repair and Reconstruction," the
contents of which are incorporated herein by reference.
[0054] In a preferred embodiment, as described supra, the third
part of the mold (C, B'b) is removed and the donor valve reinforced
foam which has formed lining the aortic wall and the valve leaflet
is seeded with cells which would normally populate the respective
tissues of the semilunar valve, e.g., cells derived from the
fibrosa, spongiosa, and ventricularis tissues. The mold can then be
introduced into a cell culturing system to allow attachment,
growth, and differentiation of the seeded cells. In another
preferred embodiment, the valve is seeded with cells which normally
populate the outer tissue covering the semilunar valve, e.g.,
allogeneic or autogenous endothelial or mesothelial cells, to
provide an epithelial covering to the valves and aorta. These
endothelial or mesothelial cells are used to populate all tissue
surfaces except the back or outer surface of the aortic wall after
the first cells seeded into the tissue have remodeled the
biopolymer structure.
[0055] As described supra, the collagen, e.g., fetal porcine
collagen, or fibrillar collagen, e.g., liquid dense fibrillar
collagen, used to fill the components of the mold may be enriched
with signaling molecules which play a role in vascular development.
Products of three classes of genes are implicated: hedgehog,
homeobox and cytokine. They include but are not limited to the
following proteins: sonic hedgehog; NK-2, XNKx-3.3 (tinman), hCsx
and Gax homeobox gene products; TGFbeta, VEGF, FGF, IGF, PDGF, and
BMP4 cytokine proteins. Differentiation induced by the use of
combinations of the foregoing proteins is promoted by incubation of
the cell laden scaffold in vitro under tissue culture
conditions.
[0056] The tissue which has formed can be lifted out of the mold
and the two lateral edges of the valve are sewn together with a
biopolymer thread, e.g., collagen or synthetic thread, to form a
tubular valve.
[0057] In a preferred embodiment, the valve is rehydrated in tissue
culture medium and seeded with fibroblasts internally and with
endothelial cells on its surfaces to demonstrate cell attachment,
absence of toxicity, cell proliferation and differentiation. The
toxicity and low information content of glutaraldehyde treated
tissues which prevent the repopulation of processed valves are no
longer a problem when valves are prepared by the methods of this
disclosure.
[0058] In another embodiment, the tubular valve can be incorporated
into a pulsatile closed circulatory loop containing a nutrient
fluid having the visco-elastic properties of blood. The valve can
be oriented in the circulation so that during diastole it back
flows to fill the pockets of the valve. Mechanical conditioning of
the valve can be carried out for a period of 2-4 weeks or longer.
It can remain in culture ready for delivery on demand.
[0059] The invention also features a biocompatible annular sewing
ring which can be used for attachment of a valve, e.g., a heart
valve, e.g., a semilunar valve, to the aorta. In one embodiment, a
biopolymer cloth or biopolymer matt, e.g., a collagen cloth or
collagen matt, is prepared. Biopolymer and collagen matts are
described in copending patent application Ser. No. 09/042,549,
entitled "Biopolymer Matt for Use in Tissue Repair and
Reconstruction," the contents of which are incorporated herein by
reference. The biopolymer cloth or matt, e.g., collagen cloth or
matt, can be wrapped around and stitched to a biopolymer matt in a
rope-like structure, e.g., a collagen matt rope shaped as a circle
and stitched to the rope. Various shaped biopolymer matts, e.g.,
various shaped collagen matts, are described in copending patent
application Ser. No. 09/042,549, entitled "Biopolymer Matt for Use
in Tissue Repair and Reconstruction," the contents of which are
incorporated herein by reference. These, biopolymer matts, e.g.,
collagen matts, can be cast into various shapes, e.g., tubes or
orbs, such as spheres, to produce membranous structures which can
contain material or liquids for specialized functions. Examples of
implants made from matt, matt composite, or matt compositions
include, for example, vessels, ducts, ureters, bladders and bone
implants. Biopolymer matts, e.g., collagen matts, can be cast as
tubes or orbs, such as spheres, to produce membranous structures
which can contain material or liquids for specialized functions.
Examples of implants made from matt, matt composite, or matt
compositions include, for example, vessels, ducts, ureters,
bladders and bone implants from matt cylinders filled with bone
replacement material.
[0060] In a preferred embodiment, the biocompatible annulus, e.g.,
the collagen annulus, can be seeded with cells and enriched with
signaling molecules which play a role in vascular development.
Products of three classes of genes are implicated: hedgehog,
homeobox and cytokine. They include but are not limited to the
following proteins: sonic hedgehog; NK-2, XNKx-3.3 (tinman), hCsx
and Gax homeobox gene products; TGFbeta, VEGF, FGF, IGF, PDGF, and
BMP4 cytokine proteins. Differentiation induced by the use of
combinations of the foregoing proteins is promoted by incubation of
the cell laden scaffold in vitro under tissue culture conditions.
In another preferred embodiment, additional endothelial or
mesothelial cells may be seeded onto the structure to cover the
outer surface of the biocompatible annulus.
[0061] The contents of all cited references including literature
references, issued patents, published patent applications, and
co-pending patent applications cited throughout this application
including the background are hereby expressly incorporated by
reference in their entirety.
[0062] Equivalents
[0063] Those skilled in the art will recognize, or be able to
ascertain using no more than routine experimentation, many
equivalents to specific embodiments of the invention described
specifically herein. Such equivalents are intended to be
encompassed in the scope of the following claims.
* * * * *