The aging center has been and will be
engaged in research activity encompassing the following topics:
1. A major area of interest in our
center is the immune response to vaccines among the elderly.
The risks associated with viral influenza are growing in industrialized
societies, a result of the increasing numbers of elderly and
other chronically ill patients who have excess morbidity and
mortality from influenza. Each winter brings an excess influenza-related
mortality rate of approximately 800 patients, 90% of whom are
elderly persons. The increased morbidity and mortality rate
from influenza in the elderly has been attributed to a decline
in lymphocyte function that occurs with age. Specifically, memory
T lymphocytes that have lost the ability to be costimulated
to produce IL-2 accumulate with age, leading to an abortive
immune response to the virus and to the vaccines that have been
developed against it. The World Health Organization and the
Israeli Ministry of Health recommend yearly vaccination of elderly
persons, but only about 40-60% of elderly vaccinees in the community
develop protective immunity.
Liposomes are phospholipid particles that have been shown to
act as an efficient delivery system to reticuloendothelial and
antigen presenting cells. We have previously found that young
adult mice and humans, who were immunized with the hemagglutinin
antigen that has been entrapped in liposomes containing IL-2,
developed a significantly higher titer of protecting antibodies
than did those (mice or persons) immunized with a free antigen,
or with the liposomal antigen without IL-2.We are currently
examining the efficacy of the liposomal vaccine and its ability
to elicit protective anti-influenza antibodies in old subjects.
An enhanced immune response, resulting in a higher titer of
protective antibodies, may reduce the morbidity and mortality
from influenza, and may prove to be cost effective and cost
saving.
2. Another subject of research is
the role of Microsatellite instability (MIN) and telomere attrition
in the development of immune dysfunction during old age. There
is extensive correlative evidence that DNA damage and mutations
increase with age. Specifically, a decrease in DNA repair has
been observed which may be responsible for increased DNA instability.
Recent findings have shown the role of DNA instability in carcinogenesis.
Six human genes involved in DNA mismatch repair were recently
cloned and characterized; hMsh3,hMLHi, hPMS2, hMsh6, hMsh2 and
hPMS1. Germline mutations in these genes cause, the "Mutator
phenotype", and MIN. Recent studies have shown a tight
association between the failure of lymphocytes from old donors
to build up an adequate proliferativ response to an antigen,
and the accumulation of CD8+CD28 - lymphocyte clones. Lymphocyte
senescence in tissue cultures is also characterized by the disappearance
of the CD28 surface receptor as their number of population doublings
increase and their telomeres shorten. We seek for a possible
relationship between age-associated T cell dysfunction and the
phenomenon of decreased DNA repairability.
Recently we have shown that human lymphocyte DNA
samples obtained in 1987 and ten years later from the
same individuals that have been tested at eight microsatellite
loci developed significant MIN with age. This may indicate an
overall genomic instability in the elderly. We are currently
investigating the relationship between telomere attrition and
MIN in senescing lymphocytes, and the association of such changes
with the characteristic T cell dysfunction of old age.
3. Alzheimer's disease (AD) is an
irreversible neurodegenerative brain disease associated with
the death of brain cells. AD is the leading cause of dementia
among the elderly world - wide and is the fourth leading cause
of death in the western countries. The pathology in the brain
includes the presence of both neurofibrillary tangles and neuritic
plaques. The major component of the neurotic plaques is the
bamyloid peptide (Ab), which derives from the cleavage of the
amyloid precursor protein (APP), presumably via proteolytic
cleavage by the a or b secretase enzymes. The ability to direct
the processing of the protein APP to the "good" pathway
is an essential step towards prevention and treatment of AD
Apolipoprotein E (apoE) is the major apolipoprotein in the central
nervous system. It has been postulated to be involved in growth
and repair after brain injury. It seems that apo E appears to
be necessary for the functioning of neurons. The
e4 allele of
the apo E gene is the chief known genetic risk factor for late
onset sporadic AD During the past five years we investigated
in our laboratory various aspects of Alzheimer's Disease, and
explored the association of apo Ee4 with late sporadic AD. In
another study we showed that persons who had suffered brain
injury and carried the apoEe4, allele sustained neurologic deficits
to much longer periods than persons with brain injury carrying
the apoEe3 allele. Together, these reports and many other indicate
that apo Ee4 allele may be implicated in the pathogenesis of
AD. The role of apo Ee4 in promoting the pathological processes
is still unknown and it may perturb repair functions in the
brain. It may also play a role in the deposition of beta amyloid
in the brain as well as in the hyperphosphorilation of the tau
protein, which leads to the development of the neurofibrillary
tangles. We intend to conduct two parallel studies in order
to examine the role of apo E in AD progression, and the mechanism
by which it contributes to such progression.
A. clinical study
in which we will enroll 100 patients who fulfill the American
Psychiatric Society criteria for mild AD. In addition,
we will enroll 100 more persons who suffer from mild cognitive
impairment. Each enrollee will undergo a thorough work
up concerning the evaluation of disease severity and neuroimaging
(brain PET) as well as apo E genotyping. We will follow
all enrollees up periodically for parameters indicating
disease progression, using clinical and neuropsychological
scales like the clinical dementia rating, the global deterioration
scale and the ADAS-COG as well as functional brain imaging
(PET). We will compare persons with mild AD. Bearing various
apo E genotypes and determine whether the apo Ee4 allele
is a risk factor for increased clinical deterioration
rate as compared with persons with mild AD bearing the
apo Ee3 alleles. We will also be able to determine whether
the apo Ee4 allele is a risk factor for developing AD
among persons with mild cognitive impairment. Such information
may help decide which patient may benefit from early therapeutic
interventions.
B. During the past
two years we developed several methods that enabled us
to study the production and processing of Amyloid Precursor
Protein (APP) by neurons. Using human neuroblastoma cell
lines, we developed a western blot analysis by which we
can assay newly secreted APP in supernatants. We can also
assay alpha (a) and beta (b) secreted APP, the result
of differential processing of APP, as well as the toxic
Ab(42) peptide using a specific enzyme link immunosorbet
assay (ELISA). Altogether, these tests allow us to determine
how much APP was produced, and how itwas preferentially
processed by neurons in response to various stimuli. For
example, we found that heparin increased the secretion
of APP into the culture medium, but enhanced the production
of bAPP, the result of APP cleavage at the beta position.
However low molecular weight heparin enhanced alpha cleavage
of APP as was demonstrated by the increased level of
aAPP
with almost no change in total APP level in the medium.
This pattern was confirmed by the Ab42 levels, as measured
by the ELISA. In this application, we suggest to study
the effect of various apoE proteins on the cleavage pattern
of APP, and on the production of the toxic substance Ab
(42). Using neuroblastoma cell line and primary cultures
we will apply apoE3 and apoE4 at various concentrations
, to the cells and measure at several time points the
effect on APP production and secretion, as well as the
concentration of a and
b secreted APP and Ab(42) in the
medium. We will also examine the effect of apoE3 and apoE4
on the level of the specific mRNA of the enzymes involved
in APP processing. We will use a semiquantative RT-PCR
method to compare the mRNA level of
b secretase and the
presenilins genes in neurons in response to the addition
of either apoE3 or apoE4 to the culture medium. An increase
in the mRNA level of the b secretase or
g secretase (presenilin)
may result in the accumulation of Ab (42) and imply that
the apoE proteins influence APP processing and may act
on the transcriptional level.
