U.S. patent application number 10/180669 was filed with the patent office on 2002-12-19 for relation of apolipoprotein e genotype and estrogen deprivation in alzheimer's disease.
Invention is credited to Einstein, Gillian, Schmechel, Donald E., Shaughnessy, Laura W..
Application Number | 20020192725 10/180669 |
Document ID | / |
Family ID | 23687464 |
Filed Date | 2002-12-19 |
United States Patent
Application |
20020192725 |
Kind Code |
A1 |
Einstein, Gillian ; et
al. |
December 19, 2002 |
Relation of apolipoprotein E genotype and estrogen deprivation in
alzheimer's disease
Abstract
A method of screening a subject for risk of developing
Alzheimer's disease comprises determining the presence of at least
one ApoE4 allele in a subject, and determining the presence or
absence of decreased estrogen levels in said subject (e.g., due to
previous or impending menopause or hysterectomy). The presence of
at least one ApoE4 allele (and particularly two ApoE4 alleles) in
combination with decreased estrogen levels in said subject
indicating said subject is at greater risk of developing
Alzheimer's disease (e.g., as compared to subjects with at
corresponding number of ApoE4 alleles, but who do not have
decreased estrogen levels), and that the subject will receive
greater benefit from estrogen replacement therapy in treating
Alzheimer's disease than a subject who does not carry one or two
ApoE4 alleles.
Inventors: |
Einstein, Gillian;
(Bethesda, MD) ; Shaughnessy, Laura W.; (Durham,
NC) ; Schmechel, Donald E.; (Durham, NC) |
Correspondence
Address: |
MYERS BIGEL SIBLEY & SAJOVEC
PO BOX 37428
RALEIGH
NC
27627
US
|
Family ID: |
23687464 |
Appl. No.: |
10/180669 |
Filed: |
June 26, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10180669 |
Jun 26, 2002 |
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09425650 |
Oct 22, 1999 |
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6432643 |
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Current U.S.
Class: |
435/7.21 ;
435/6.11 |
Current CPC
Class: |
C12Q 2600/172 20130101;
C12Q 1/6883 20130101; C12Q 2600/156 20130101; C12Q 2600/106
20130101 |
Class at
Publication: |
435/7.21 ;
435/6 |
International
Class: |
C12Q 001/68; G01N
033/567 |
Goverment Interests
[0001] This invention was made with government support under Grant
Number 5 T32 AG00029 from the National Institutes of Health. The
government has certain rights to this invention.
Claims
We claim:
1. A method of screening a human female subject for risk of
developing Alzheimer's disease, comprising: (a) determining the
presence of at least one ApoE4 allele in a subject, and (b)
determining the presence or absence of decreased estrogen levels in
said subject; the presence of at least one ApoE4 allele in
combination with decreased estrogen levels in said subject
indicating said subject is at greater risk of developing
Alzheimer's disease.
2. A method according to claim 1, wherein said step of determining
the presence of at least one ApoE4 allele is carried out by
collecting a biological sample containing DNA from said subject,
and then detecting the presence or absence of DNA encoding ApoE4 in
said biological sample.
3. A method according to claim 2, wherein said detecting step is
carried out by amplifying DNA encoding ApoE4.
4. A method according to claim 1, wherein said step of determining
the presence of at least one ApoE4 allele is carried out by
collecting an ApoE sample from said subject, and then detecting the
presence or absence of the ApoE4 isoform in said ApoE sample.
5. A method according to claim 1, wherein said subject has
previously been determined to have one or more factors indicating
that such subject is afflicted with Alzheimer's disease.
6. A method according to claim 1, wherein said determining step
comprises detecting whether said subject is homozygous for the gene
encoding ApoE4.
7. A method for screening a human female subject for responsiveness
to estrogen replacement therapy for the treatment of Alzheimer's
disease, comprising: determining the presence of at least on ApoE4
allele in said subject, wherein the presence of at least one apoE4
gene allele indicates said subject will receive greater benefit
from estrogen replacement therapy that a subject who does not carry
at least one ApoE4 allele.
8. A method according to claim 7, wherein said step of determining
the presence of at least one ApoE4 allele is carried out by
collecting a biological sample containing DNA from said subject,
and then detecting the presence or absence of DNA encoding ApoE4 in
said biological sample.
9. A method according to claim 8, wherein said detecting step is
carried out by amplifying DNA encoding ApoE4.
10. A method according to claim 7, wherein said step of determining
the presence of at least one ApoE4 allele is carried out by
collecting an ApoE sample from said subject, and then detecting the
presence or absence of the ApoE4 isoform in said ApoE sample.
11. A method according to claim 7, wherein said subject has
previously been determined to have one or more factors indicating
that such subject is afflicted with Alzheimer's disease.
12. A method according to claim 7, wherein said determining step
comprises detecting whether said subject is homozygous for the gene
encoding ApoE4.
13. A method for treating a human female subject for Alzheimer's
disease, comprising: (a) determining the presence of at least on
ApoE4 allele in said subject; and then (b) administering estrogen
replacement therapy to said subject in an Alzheimer's disease
treatment effective amount.
14. A method according to claim 13, wherein said administering step
is carried out by orally administering an estrogenic agent to said
subject.
15. A method according to claim 13, wherein said administering step
is carried out by transdermally administering an estrogenic agent
to said subject.
16. A method according to claim 13, wherein said estrogen
replacement therapy includes the step of administering a
progestationally active compound to said subject in an amount
effective to inhibit estrogenic endometrial proliferation.
17. A method according to claim 13, wherein said step of
determining the presence of at least one ApoE4 allele is carried
out by collecting a biological sample containing DNA from said
subject, and then detecting the presence or absence of DNA encoding
ApoE4 in said biological sample.
18. A method according to claim 17, wherein said detecting step is
carried out by amplifying DNA encoding ApoE4.
19. A method according to claim 13, wherein said step of
determining the presence of at least one ApoE4 allele is carried
out by collecting an ApoE sample from said subject, and then
detecting the presence or absence of the ApoE4 isoform in said ApoE
sample.
20. A method according to claim 13, wherein said subject has
previously been determined to have one or more factors indicating
that such subject is afflicted with Alzheimer's disease.
21. A method according to claim 13, wherein said determining step
comprises detecting whether said subject is homozygous for the gene
encoding ApoE4.
22. A method of treating a human female subject for Alzheimer's
disease, said method comprising the steps of: selecting a subject
that carries at least on ApoE4 allele, and then administering
estrogen replacement therapy to said subject in an Alzheimer's
disease treatment effective amount.
23. A method according to claim 22, wherein said administering step
is carried out by orally administering an estrogenic agent to said
subject.
24. A method according to claim 22, wherein said administering step
is carried out by transdermally administering an estrogenic agent
to said subject.
25. A method according to claim 22, wherein said estrogen
replacement therapy includes the step of administering a
progestationally active compound to said subject in an amount
effective to inhibit estrogenic endometrial proliferation.
Description
FIELD OF THE INVENTION
[0002] The present invention concerns methods of screening for
Alzheimer's disease susceptibility in subjects, along with methods
of classifying susceptible individuals for treatment and methods of
treating Alzheimer's disease.
BACKGROUND OF THE INVENTION
[0003] Alzheimer's disease (AD) is marked by a devastating decrease
in cognitive ability which is correlated with a decline in the
number of synapses in the hippocampus and neocortex. One of the
risk factors for development of AD is the gene coding for the E4
allele of a lipid carrier protein, apolipoprotein E (APOE, gene).
The APOE E4 allele is a risk factor for late-onset, familial and
sporadic AD while the APOE3 and E2 alleles are either neutral (E3)
or protective (E2). Another risk factor is sex; both the incidence
and prevalence of AD is greater in females than in males.
[0004] U.S. Pat. No. 5,508,167 to A. Roses et al., assigned to Duke
University, discloses methods of diagnosing or prognosing
Alzheimer's disease in a subject. The methods involve directly or
indirectly detecting the presence or absence of an apolipoprotein E
type 4 (ApoE4) isoform or DNA, encoding ApoE4 in the subject. The
presence of ApoE4 indicates the subject is afflicted with
Alzheimer's disease or at risk of developing Alzheimer's disease.
This basic finding has led to a number of developments in the
Alzheimer's disease field.
[0005] For example, U.S. Pat. No. 5,773,220 to S. DeKosy and M.
Kamboh, assigned to the University of Pittsburgh, describes a
method for screening for the risk of developing Alzheimer's disease
in a subject by detecting the presence or absence of the ApoE
allele and the presence or absence of the alphal-antichymotrypsin
(ACT) allele. The presence of two ACT/A alleles, in conjunction
with the presence of one or two ApoE4 alleles, is said to indicate
an increased risk for Alzheimer's disease.
[0006] In addition, U.S. Pat. No. 5,935,781 to J. Poirer, assigned
to McGill University. This patent describes a method for the
identification of human subjects responsive to cholinomimetic
therapy. The method comprises determining the absence of
apolipoprotein E4 (apoE4) alleles in a biological sample of the
patient, where the absence of at least one apoE4 allele indicates a
predisposition to respond to cholinomimetic therapy. Methods of
administering cholinomimetics to such identified subjects are also
described.
[0007] While the identification of ApoE4 as a risk factor for
Alzheimer's disease has led to a number of new developments in the
field, AD remains a complex disease for which treatment is
difficult and the ultimate prognosis is poor. Accordingly, there
remains a need for new ways to screen for AD, classify patients for
appropriate AD treatment, and treat AD.
SUMMARY OF THE INVENTION
[0008] A first aspect of the present invention is a method of
screening (e.g., diagnosing or prognosing) a subject for risk of
developing Alzheimer's disease. The method comprises: (a)
determining the presence of at least one ApoE4 allele in a subject,
and (b) determining the presence or absence of decreased estrogen
levels in said subject. The presence of at least one ApoE4 allele
(and particularly two ApoE4 alleles) in combination with decreased
estrogen levels in said subject indicating said subject is at
greater risk of developing Alzheimer's disease (e.g., as compared
to subjects with at corresponding number of ApoE4 alleles, but who
do not have decreased estrogen levels).
[0009] A second aspect of the present invention is a method for
screening a subject for responsiveness to estrogen replacement
therapy for the treatment of Alzheimer's disease. The method
comprises determining the presence of at least on ApoE4 allele in
the subject. The presence of at least one apoE4 gene allele allele
(and particularly two ApoE4 alleles) indicates that the subject
will receive greater benefit from estrogen replacement therapy that
a subject who does not carry at least one ApoE4 allele allele (and
particularly two ApoE4 alleles). Alternatively stated, the presence
of at least one ApoE4 allele indicates a predisposition, or
potential, of that subject to beneficially respond to estrogen
replacement therapy (e.g., a greater likelihood that that subject
will beneficially respond to estrogen replacement therapy as
compared to a subject that does not carry at least one ApoE4
allele).
[0010] A third aspect of the present invention is a method for
treating a subject for Alzheimer's disease. The method comprises:
(a) determining the presence of at least one ApoE4 allele in said
subject allele (and particularly two ApoE4 alleles); and then (b)
administering estrogen replacement therapy to that subject (i.e., a
subject carrying one or two ApoE4 alleles) in an Alzheimer's
disease treatment effective amount.
[0011] A fourth aspect of the present invention is a method of
treating a human female subject for Alzheimer's disease, where that
subject carries at least one ApoE4 allele allele (and particularly
two ApoE4 alleles). The method comprises administering estrogen
replacement therapy to the subject in an Alzheimer's disease
treatment effective amount.
[0012] In a particularly preferred embodiment, the estrogen
replacement therapy is initiated to a susceptible subject as
described above prior to the onset of menopause, or at least
concurrently with the onset of menopause, or is initiated
concurrently with a hysterectomy. The object in this embodiment is
to reduce, inhibit, or eliminate a gap in estrogen levels, or the
time for which the subject is exposed to decreased estrogen levels,
so that the risk of early neuronal cell death in that patient is
reduced, and the time of onset of Alzheimer's disease is delayed,
and/or the progression of that disease is slowed.
[0013] A fifth aspect of the present invention is the use of an
estrogen replacement therapy active agent for the preparation of a
medicament for the treatment of Alzheimer's disease.
[0014] The foregoing and other objects and aspects of the present
invention are explained in detail in the drawings herein and the
specification set forth below.
BRIEF DESCRIPTION OF THE DRAWINGS
[0015] FIG. 1 shows that there is no difference in spine density in
the dorsal blade of the dentate gyrus in ApoE3 as compared to ApoE4
mice, comparing Sham OVX and OVX mice.
[0016] FIG. 2 illustrates that CA1 apical dendrite spine density is
significantly decreased in female ApoE4 mice deprived of
estrogen.
[0017] FIG. 3 shows that stratum radiatum synapthophysin
immunoreactivity is significantly decreased in ApoE4 mice deprived
of estrogen.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0018] Terms and phrases used herein have their ordinary meaning in
the art, as further explained below.
[0019] "ApoE4" refers to apolipoprotein E4.
[0020] "Concurrently" as used herein means at the same time or
sufficiently close in time that substantially the same
physiological effect is achieved.
[0021] "Estrogen replacement therapy" as used herein refers to a
long-term therapy in which estrogen or estrogenic active agents are
administered to a subject continuously over an extended period of
time (e.g., a month, a year, or more) to maintain sustained blood
levels of the active agent to combat the effects of menopause or
hysterectomy (e.g., loss of calcium from bone and increased
incidience of classical osteoporotic fractures of the forearm and
hip, ischemic heart disease, etc.). The administration may be daily
or periodically.
[0022] "Decreased estrogen levels" as used herein means that the
subject has estrogen levels indicating that the subject is a
candidate for or should receive estrogen replacement therapy (e.g.,
an estrogen level at a post-menopausal rather than a pre-menopausal
level; an estrogen level at a post-hysterectomy rather than a
pre-hysterectomy level in a pre-menopausal woman).
[0023] Subjects suitable for the present invention include those
who have not previously been diagnosed as afflicted with
Alzheimer's disease, those which have previously been determined to
be at risk of developing Alzheimer's disease, and those who have
been initially diagnosed as being afflicted with Alzheimer's
disease where confirming information is desired, or where a focused
treatment option is desired. For example, patients diagnosed or
determined to be afflicted with dementia, particularly patients who
had previously been clinically normal who are determined to be
afflicted with a progressive dementia, are suitable subjects. Thus,
the present invention may be employed in detecting both familial
Alzheimer's disease (late onset and early onset) as well as
sporadic Alzheimer's disease. Many Alzheimer's disease patients
encountered in practice have no obvious family history and have
been classified as sporadic. However, genetic factors in early- and
late-onset of familial Alzheimer's disease (FAD) are well
documented. Late-onset Alzheimer's disease is the classification
usually used if the disease is diagnosed to occur after the age of
65 in humans. The present invention is particularly useful in
identifying patients who would benefit from the early
administration of an estrogen replacement therapy. Subjects
suitable for the present invention are, in general, human subjects,
and are preferably female subjects.
[0024] The step of detecting the presence or absence of ApoE4 or of
DNA encoding such isoform (including the number of alleles for
ApoE4) may be carried out either directly or indirectly by any
suitable means. A variety of techniques are known to those skilled
in the art. All generally involve the step of collecting a sample
of biological material containing either DNA or ApoE from the
subject, and then detecting whether or not the subject possesses
ApoE4 or DNA encoding such isoform from that sample. For example,
the detecting step may be carried out by collecting an ApoE sample
from the subject (for example, from cerebrospinal fluid, or any
other fluid or tissue containing ApoE), and then determining the
presence or absence of an ApoE4 isoform in the ApoE sample (e.g.,
by-isoelectric focusing or immunoassay). In the alternative, the
detecting step may be carried out by collecting a biological sample
containing DNA from the subject, and then determining the presence
or absence of DNA encoding an ApoE4 isoform in the biological
sample. Any biological sample which contains the DNA of that
subject may be employed, including tissue samples and blood
samples, with blood cells being a particularly convenient source.
Numerous techniques for detecting the presence of one or two ApoE4
alleles in a subject are known, including but not limited to those
described in U.S. Pat. No. 5,508,167 to Roses et al., U.S. Pat. No.
5,773,220 to S. DeKosy and M. Kamboh, and U.S. Pat. No. 5,935,781
to Poirer (applicants specifically intend that the disclosures of
all United States patent references cited herein be incorporated
herein by reference).
[0025] Patients may be selected for treatment by the methods of the
present invention based upon knowledge of the ApoE4 profile of that
individual patient (i.e., the presence of one or two ApoE4
alleles). The ApoE profile may be obtained in the manner described
above. Of course, it is not necessary that such screening or
profiling be at the same time or place, or by the same individual,
as making the selection for therapy, so long as the selection is
based upon this information.
[0026] Estrogen replacement therapy may be carried out by any
suitable means. All typically involve administering an active agent
such as estrogen or an estrogen analog (typically a steroid that
has estrogen activity) to the subject in an estrogen replacement
therapy effective amount, which is commensurate with an Alzheimer's
disease treatment effective amount (e.g., an amount effective to
delay the onset of AD, slow the progression of AD, etc.). Any
suitable route of administration may be employed, including, but
not limited to, oral administration, aerosol administration to
airway surfaces, intravenous injection, subcutaneous injection,
intramuscular injection, transdermal administration (e.g., a
patch), etc. Oral and transdermal formulations are currently
preferred. Numerous estrogen replacement therapy preparations and
protocols are known, including but not limited to those described
in U.S. Pat. Nos. 5,922,349; 5,897,539; 5,565,199; 5,468,736;
5,422,119; 5,288,717; and 5,023,084, the disclosures of all of
which are incorporated by reference herein in their entirety. Other
agents, such as progesterone (or progestin) in a hormone
replacement therapy effective amount, may be administered along
with the estrogen to provide a combination therapy, if desired
(typically to reduce undesirable side-effects of estrogen
monotherapy, as such estrogenic endometrial proliferation and
corresponding risk of endometrial cancer).
[0027] Suitable active agents for estrogen replacement therapy
include, but are not limited to, natural and synthetic estrogens
such as conjugated equine estrogen, ethinyl estradiol, micronized
estradiol, 17.beta. estradiol, mestranol, estradiol valerate,
11-nitrato estradiol, 7-.alpha.-methyl-11-nitrato-estradiol,
piperazine estrone sulfate, quinestranol, and 8,9-dehydroestereone
(particularly alkali metal salts and sulfate esters thereof). See,
e.g., U.S. Pat. No. 5,422,119 at column 6; U.S. Pat. No. 5,288,717.
Of course, all active agents may be prepared as a pharmaceutically
acceptable salt or ester, in accordance with known techniques,
[0028] The progestin component may be any progestationally active
compound, including but not limited to progsterone,
17-hydroxyprogesterone, dihydroprogesterone, medroxyprogesterone
acetate, norethindrone, norethindrone acetate, norethynodrel,
ethynodioldiacetate, norgesterel, levo-norgesterel, gestodene,
delta-15-levonorgesterel, norgestimate, 17-deacetyl norgestimate,
nomegestereol, nesterone, desogesterel and 3-keto-desogesteral.
See, e.g., U.S. Pat. No. 5,422,119 at column 6.
[0029] In general, a pharmaceutical formulation or medicament for
estrogen replacement therapy is prepared by bringing an effective
of the active agent into contact with a pharmaceutically acceptable
carrier, such as lactose or talc (for an oral administration),
intimately admixing the two, and forming (when necessary) the
mixture into a suitable unit dosage form such as a patch for
transdermal administration or a tablet, dragee, capsule or pill for
an oral dosage form.
[0030] The amount of active agent administered will depend upon
factors such as the specific active agent, the age, weight and
condition of the subject, the route of administration, etc. For
example, the estrogenic active agent may be administered in an a
amount of from 0.3 to 1.2 mg daily for orally administered
conjugated equine estrogen; or from about 25 .mu.g/day to about 150
.mu.g/day of transdermal .beta.-estradiol.
[0031] Examples of commercially available estrogen preparations
include: Alora.TM. (skin patch), Climara.TM. (skin patch),
Estraderm.TM. (skin patch), FemPatch.TM. (skin patch), Estrace.TM.
(pill or skin patch), Estrab.TM. (pill), Menset.TM. (pill),
Ogen.TM. (pill), Ortho-est.TM. (pill), and Premarin.TM. (pill).
[0032] Examples of commercially available estrogen/progestin
combination formulations include, but are not limited to,
Combipatch.TM. (skin patch) and Prempro.TM. (pill).
[0033] Where necessary, estrogen levels may be directly or
indirectly determined in a subject by measuring serum estradiol
levels in accordance with known techniques, or by measuring vaginal
or urethral pH levels, as described in U.S. Pat. No. 5,916,176 to
Caillouette. Of course, decreased estrogen levels may be inferred
from menopause or hysterectomy.
[0034] The present invention is illustrated in greater detail in
the following non-limiting examples.
EXAMPLE 1
Effect of Ovarectomy on Spine and Hippocampal
Neuron Density in Transgenic ApoE4 and ApoE3 Mice
[0035] This example explored whether the interaction of the ApoE4
risk factor and the estrogen risk factor might exacerbate the
pathology associated with AD in a putative animal model of AD. To
explore this possibility, we compared spine and synapse density in
two populations of hippocampal neurons in 15-month-old human
transgenic APOE4 and APOE3 mice ovariectomized (OVX) or sham OVX
mice at 14 months of age. CA1 pyramidal neurons are one of the most
susceptible populations of neurons in AD, while dentate granule
cells are less vulnerable.
[0036] Animals and Surgery.
[0037] Female transgenic mice created from the C57B1/6J line with
the human isoforms of APOE3 and APOE4 were used (P. -T. Xu et al.,
Neurobiol. Dis. 3, 229-235 (1996). Three sibling pairs were
obtained for each group. At 14 months of age, one member of each
pair was ovariectomized (OVX) while the other member received a
sham OVX surgery.
[0038] Tissue Preparation.
[0039] At 15 months of age, all animals were euthanized, their
brains removed and cut along the midline. Each brain was
immersion-fixed in 4% paraformaldehyde in 0.1 M phosphate buffer
(PB), pH 7.4, at room temperature (RT). After 140 minutes, the left
hemisphere was removed and rinsed in 0.1 M PB. Using a vibratome,
sagittal slices of 300 microns were collected in 0.1 M PB and
stored at 4.degree. C. until intracellular injection with Lucifer
yellow. The right hemisphere was immersion-fixed for 48 hours at
4.degree. C., embedded in paraffin, and sectioned sagittally into 8
micron slices that were mounted on slides in preparation for
synaptophysin immunohistochemistry (SYN-IR).
[0040] Lucifer Yellow Injections and Immunoreactivity.
[0041] Hippocampal CA1 pyramidal cells and dentate granule cells
were injected intracellularly with Lucifer yellow in accordance
with known techniques (G. Einstein, J. Neurosci. Meth. 26, 95-103
(1988)). At least three slices from the dorsal hippocampus were
injected per animal, with 10-15 granule cells and CA1 neurons
injected per slice.
[0042] After injection, slices were rinsed in PB and post-fixed
overnight in 10% formalin. The tissue was then put in 25% sucrose
(in 0.1M phosphate buffered saline-PBS) for cryoprotection and
resectioned at 60 microns on a freezing microtome. Free-floating
sections were rinsed in PBS, blocked, incubated in biotinylated
anti-Lucifer-yellow antibody followed by incubation in ABC and
visualization with DAB. The tissue was mounted on gelatin-subbed
slides and coverslipped. Under a 100.times. objective and using a
drawing tube, four neurons each from CA1 and the dorsal blade of
the dentate gyrus were drawn and the spines counted.
[0043] Synaptophysin Immunoreactivity.
[0044] Slides containing sections through the dorsal hippocampus
were incubated with the mouse monoclonal antibody to synaptophysin
followed by incubation with a biotinylated secondary antibody, an
ABC solution and then visualized using DAB (E. Masliah et al., Exp.
Neurol. 113, 131 (1991)). Following coverslipping, images of the
stratum radiatum and molecular layers of the dentate were grabbed
using an MTI camera and the imaging program Image 1 NIH Image 1.63
was used to analyze the density of synaptophysin staining in each
of these areas.
[0045] Table 1 below shows that there is no difference in the
optical density of SYN-IR for the molecular layers of the dorsal
blade of the dentate gyrus.
1TABLE 1 inner molecular middle molecular outer molecular GROUP
layer layer layer APOE3 SHAM 44.92 .+-. 7.8 37.50 .+-. 6.8 42.95
.+-. 5.1 APOE3 OVX 49.60 .+-. 5.6 42.50 .+-. 5.0 45.90 .+-. 4.5
APOE4 SHAM 46.10 .+-. 5.7 33.40 .+-. 5.0 42.50 .+-. 7.1 APOE4 OVX
48.96 .+-. 4.1 37.20 .+-. 3.0 44.37 .+-. 4.3
[0046] FIG. 1 shows that there is no difference in spine density in
the dorsal blade of the dentate gyrus in ApoE3 as compared to ApoE4
mice, comparing Sham OVX and OVX mice. FIG. 2 illustrates that CA1
apical dendrite spine density is significantly decreased in female
ApoE4 mice deprived of estrogen. FIG. 3 shows that stratum radiatum
synapthophysin immunoreactivity is significantly decreased in ApoE4
mice deprived of estrogen.
[0047] Observations.
[0048] (1) With our paradigm, mice carrying the APOE4 gene had a
significant decrease in spine density on the apical dendrites of
CA1 pyramidal neurons when compared to sham OVX sibs. In contrast,
mice carrying the APOE3 gene had no change in spine density of CA1
pyramidal neurons when compared to sham OVX sibs. (2) Mice carrying
the APOE4 gene had a significant decrease in the density of SYN-IR
in the stratum radiatum when compared to sham OVX sibs. In
contrast, mice carrying the APOE3 gene had no change in the density
of SYN-IR in the stratum radiatum when compared to sham OVX sibs.
(3) Mice carrying the APOE3 and E4 gene had no change in spine
density of dorsal blade dentate granule cells when compared to sham
OVX sibs. (4) Mice carrying the APOE3 and E4 gene had no change in
the density of SYN-IR in the molecular layer of the dentate when
compared to sham OVX sibs.
[0049] These data show that the most vulnerable population of
neurons is the CA1 neurons in the aging APOE4 mice deprived of
estrogens. The decrease in both spine and SYN-IR density indicate a
significant decrease in synapses and in neuronal connectivity. The
interaction of the two risk factors, APOE4 and estrogen
deprivation, exacerbates the pathology associated with AD.
[0050] The foregoing is illustrative of the invention, and is not
to be construed as limiting thereof. The invention is defined by
the following claims, with equivalents of the claims to be included
therein.
* * * * *