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Current
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 -Cellular
and Molecular Biology of Aging
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BASIC BIOMEDICAL RESEARCH: CELLULAR
AND MOLECULAR BIOLOGY OF AGING
Table
of Contents
BASIC BIOMEDICAL RESEARCH: CELLULAR
AND MOLECULAR BIOLOGY OF AGING
The Division of Basic Research at the Reynolds Department
of Geriatrics (RDG) is under the direction of Dr. Sue Griffin,
the 1999 UAMS College of Medicine Distinguished Faculty Scholar.
In addition to her success in obtaining a competitive renewal
of her NIA-sponsored program project, Dr. Griffin holds an
important position that is highly complementary to her role
in the RDG, serving as the Director of Research at the Central
Arkansas Veterans Heathcare System (CAVHS) Geriatric Research,
Education & Clinical Center (GRECC), where she is deeply
involved with Alzheimer’s disease (AD) research, seeking
to reveal the fundamental causes of AD and to apply this knowledge
to the treatment of patients and the eradication of the disease.
The focus of the basic biomedical research program at the
Reynolds Department of Geriatrics and Reynolds Institute on Aging
is the cellular and molecular biology of aging. The research
encompasses Alzheimer's Disease pathogenesis, mechanisms of
gene action in the regulation of cellular aging, protein degradation
in immune senescence, mitochondrial dysfunction in normal
and dystrophic muscle, and the genetics of cellular senescence,
among other topics.
Table of Contents
Alzheimer's Disease Pathogenesis
Sue T. Griffin, PhD, Professor and Vice Chair for Basic
Research
In the mid 1980s, Dr. Griffin proposed that immune responses
engendered by neuronal dysfunction and death contributed to
the progressive accumulation of neuropathologic features in
AD, and thus to its clinical symptoms. Two risk factors for
the development of sporadic AD have been identified-aging
and head trauma-although the "cause" of AD is unknown
in the majority of cases. However, duplication of chromosome
21 (Down syndrome), mutations of the ß-amyloid precursor
protein (ß-APP) genes, and genetic variations (e.g.,
presenilin genes) are causative. As envisioned by Dr. Griffin,
the immune responses mounted as a defense against neuronal
degeneration in AD are, on a smaller scale, the same as those
that occur in response to minor insults such as those accrued
with the wear and tear of time, except that the responses
become chronic, presumably in cycle with chronic neuronal
degeneration.
Attempting to understand the possible relationship between
the chronic immune response and neuronal degeneration, Dr.
Griffin proposed (and research in her laboratory supported
the existence of) a neurodegenerative cascade driven by chronically
overexpressed, immune response-generated cytokines synthesized
and released by glia in the brain. Two consequences of overexpression
of one such cytokine-interleukin-1 (IL-1)-suggested that it
was a prime candidate for this driving force. IL-1 stimulates
excessive expression and processing of a normal membrane protein,
ß-APP, of the neurotoxic ß-amyloid found in AD
plaques. In addition, although IL-1 at low doses supports
neuronal survival, at high doses it is toxic to neurons. IL-1
also activates astrocytes to enlarge and to synthesize and
release several proteins, including apolipoprotein E, complement
factors, and S100ß, a neurite extension factor. Each
of these proteins is present in ß-amyloid plaques in
AD, and S100ß in particular has been implicated in the
growth of dystrophic neuronal processes in these plaques.
Dr. Griffin's work has given credence to the idea that chronic
overexpression of IL-1 sets in motion a self-propagating cascade
of neurodegeneration that further activates microglia, with
increased expression of IL-1, and keeps the cytokine cycle
in motion. Her work with postmortem brain tissue from AD patients
demonstrated overexpression of IL-1 in activated microglia,
overexpression of biologically active S100ß in activated
astrocytes, and overgrowth of dystrophic neuronal processes
(neurites) overexpressing ß-APP in ß-amyloid plaques.
This past year Dr. Griffin's group has shown that S100ß,
like IL-1, regulates the expression of the proinflammatory
protein IL-6.
To further investigate the potential importance of these
findings, Dr. Griffin's laboratory studied the expression
of IL-1 and S100ß in fatally head-injured patients as
early as 12 hours after acute brain injury and found a dramatic
increase in the number of activated microglia and astrocytes
overexpressing IL-1 and S100 , respectively. This laboratory's
work also showed that with normal human aging there is a gradual,
significant rise in IL-1 and S100 levels, suggesting that
the wear and tear of time is injurious and elicits expression
of these potentially neurodegenerative cytokines. This work
and that of other laboratories have given rise to a novel
treatment strategy for decreasing the risk or delaying the
onset of clinical signs of AD. In particular, Dr. Griffin's
work provides an explanation at the molecular level of why
anti-inflammatory strategies may reduce the risk and or delay
the onset of AD. In addition, it provides the molecular framework
for future, more precise intervention strategies. In collaboration
with Eli Lilly and Company (Indianapolis, Indiana), Dr. Griffin's
group has shown that in a transgenic mouse model of AD, S100ß
is overexpressed months before the neuropathologic changes
characteristic of AD appear.
This past year Dr. Griffin and her collaborators reported
in three scientific papers that homozygosity for a specific
polymorphism in the IL-1  -encoding
IL-1A gene at least triples the risk of AD (odds ratios [ORs]
= 3, 4.5, and 7.1, respectively) and decreases the age of
onset by 7-9 years. Moreover, homozygosity for this IL-1A
polymorphism plus homozygosity for a specific polymorphism
in the IL-1B gene in the IL-1 ß-encoding region confers
even greater risk of AD (OR = 11). The first two findings
were reported in the Annals of Neurology in March 2000
and the third appeared in Neurology in July 2000. These
findings have now been confirmed by studies at another center.
Dr. Griffin's AD work is supported by a program project grant
(NIA AG10208, 1995-2007), and for her studies in Down's syndrome
(invariably associated with AD pathologic changes), she has
just received NIH funding for the years 2000-2005. These funds
provide support for Dr. Griffin, Robert E. Mrak, MD, PhD,
six full-time technical personnel, five postdoctoral fellows,
and two research assistant professors at the Little Rock CAVHS,
as well as three faculty and four technical personnel in the
United Kingdom.
In close collaboration with Dr. Cornelia Beck, Dr. Griffin
submitted a successful grant application to establish an Alzheimer's
Disease Center (ADC) at UAMS. She was also instrumental in
recruiting a senior clinical AD specialist as director of
the clinical core and codirector of the ADCC.
Dr. Griffin's master's degree candidate, Paul Edwards, successfully
defended his thesis and received his Master of Science degree
in physiology in August 2001. Dr. Griffin currently sponsors
two undergraduate students and has been chosen as mentor by
Mona Gupta, an Honors in Research UAMS medical student.
Helen Deng, MD, Research Assistant Professor
Dr. Deng's principal contribution this past year has been
in establishing a genotyping facility, which has been greatly
expanded with the funding of Charlotte A. Peterson, PhD's
Microarray Core (described below). The Joan Taylor Foundation
funds Dr. Deng's position. She and David D. Liu, MD have applied
for funding from the UAMS Foundation for Institutional Review
Board-approved studies of patients in the RDG's Geriatrics
Clinic to determine the prevalence of the specific IL-1 polymorphisms
that Dr. Griffin has shown are related to the development
of AD. Through collaborative efforts with colleagues in China,
Dr. Deng and Dr. Liu continue to determine the prevalence
of these polymorphisms in Chinese populations.
Table of Contents
Molecular Signaling
in Alzheimer's Disease Pathogenesis
Steven W. Barger, PhD, Assistant Professor
Dr. Barger's expertise in molecular biology and his strong
background in the area of age-related changes relevant to
the pathogenesis of AD have enabled him to make several important
discoveries regarding the mechanisms involved in brain cell-cell
interactions and signaling that result in neurodegeneration
and neuron cell loss in AD. His outstanding achievements were
recognized by the Inglewood Foundation in 1996 by his appointment
as the RCOA's Alzheimer Scholar for 3 years with full salary
and technical support. His research is supported by an NIA-sponsored
researcher-initiated (R01) grant (2001-2005), his project
in Dr. Griffin's NIA-sponsored program project (1995-2007),
and as a coinvestigator on the research project "Cytokines,
Neurodegeneration, and Down's Syndrome" (2000-2005).
These funds provide partial support for Dr. Barger's salary
and for salaries for two postdoctoral fellows, a graduate
student, and a technician. Dr. Barger sponsored an additional
graduate student, Mao Xianrong, who successfully completed
his doctorate in May 2001.
Dr. Barger originated much of the work on the effects of
glutamatergic stress on neurons to induce expression of -APP
and release of its secreted fragment. He has shown that this
secreted fragment is an important distress signal from the
neuron that activates microglia and induces their synthesis
of IL-1. This past year he reported that this activation of
microglia results in the release of glutamate, which exhibits
fulminant neurotoxicity and further perpetuates the degenerative
cycle. Dr. Barger published four research papers, a review,
and a book chapter this year.
Gene Action in the Regulation
of Immune Cell Aging
Mark D. Crew, PhD, Assistant Professor
Major histocompatibility complex (MHC) class I proteins display
a high degree of interspecies and allelic variation. It is
well appreciated that genetic variability in the extracellular
portion of class I molecules has significance in autoimmunity
and disease resistance. The biologic importance of sequence
and length variation in the transmembrane (TM) domain, however,
is unknown. Thus, the effect of TM domain variation on overall
MHC class I function is being examined by transfecting human
MHC class I genes with altered TM domains into B cells and
analyzing the transfected gene products with respect to biosynthesis,
cell-surface expression, and recognition by cytolytic T lymphocytes.
This past year Dr. Crew continued his focus on the role of
such genes in transplantation biology. In addition, he has
been involved in developing immunotherapies for ovarian cancer,
which increases in incidence with advancing age. His research
is funded through collaborative efforts on NIH and Department
of Defense grants.
Table of Contents
Changes in Bone Marrow Stem
Cells during Aging
Beata Lecka-Czernik, PhD, Research Assistant Professor
The fate of bone marrow stem cells involved in bone remodeling
and maintenance of bone mineral density has been a focus of
Dr. Lecka-Czernik's research. She has demonstrated that aging
changes these cells so that instead of forming osteoblasts
and contributing to bone formation, they are more likely to
differentiate into adipocytes, increasing the fat content
in bone. She has also characterized the essential role of
the transcription factor peroxisome proliferator-activated
receptor (PPAR)- in activating the bone marrow adipocyte gene
program and suppressing osteoblast gene expression in aged
bone marrow. Her work is particularly exciting because it
yields a better understanding of stem cell biology during
aging and may contribute to the development of pharmacologic
intervention that would preserve the osteoblastic potential
of these cells, preventing their "escape" toward
the adipocyte phenotype. This work is currently funded by
a researcher-initiated (R01) NIH grant.
Membrane Lipids, Aging,
and Lymphocyte and Neutrophil Signal Transduction
Usha Ponnappan, PhD, Assistant Professor, and David
A. Lipschitz, MD, PhD, Professor and Chair
The focus of this VA Merit Review award, with Drs. Ponnappan
and Lipschitz as co-principal investigators, is understanding
of the role that lipids play in signal transduction in neutrophil
populations during aging. In the membrane of T lymphocytes,
myristate concentration significantly decreases with age,
and returning the concentration to normal is accompanied by
a reversal of the age-related reduction in T-lymphocyte proliferative
potential. This past year Dr. Ponnappan showed that specific
age-related functional changes in a mitogen-activated protein
kinase (erk2) accompanied the activation of neutrophils.
Dr. Ponnappan also received a competitive renewal of her
NIH-funded research project "Role of Transcription Factor
NFKB in Immunosenescence," also
with Dr. Lipschitz. The goal of this study is to delineate
the involvement of nuclear factor (NF)KB
in immune dysregulation associated with aging in T lymphocytes.
Having demonstrated a significant role for the proteasome
in aging, she will attempt to localize the specific defect
within the subunits of the proteasome responsible for this
dysregulation.
Table of Contents
Genetics of Cellular Senescence
Robert J. Shmookler Reis, PhD, Professor
This past year Dr. Reis continued to make marked progress
on three fronts. For his Merit Review award for the project
"Mechanisms and Agents Inducing Homologous Recombination
in Human Cells" (1998-2003), Dr. Reis studies the frequent
chromosomal abnormalities characteristic of transformed and
tumorigenic cell lines that may be generated by increased
homologous recombination. He and his group have found markedly
increased plasmid recombination in five immortal human cell
lines, relative to normal controls, and elevated chromosomal
recombination in human fibroblasts following stable transformation
with simian virus 40 large T antigen. Current studies include
mutational mapping of the domains of the T-antigen oncogene
responsible for this increased recombinogenicity.
On a second front, Dr. Reis studies "Polymorphic Genes
Modulating Lifespan in Caenorhabditis elegans"
(funded through 2001), using a rapid chromosome-mapping procedure
to analyze polymorphic loci in very long-lived members of
recombinant-inbred populations of the nematode C. elegans.
He has localized five genes that determine lifespan and shown
that these genes act essentially independently. He is currently
developing high-resolution mapping and isolation procedures
to isolate and characterize genes governing longevity. Dr.
Reis and his group have applied to the NIA for competitive
continuation of funding for this project.
The third front of Dr. Reis's research efforts involves his
participation in the NIA-sponsored program project of Stavros
Manolagas, MD, PhD, "Molecular and Cellular Mechanisms
of Osteoporosis" (1997-2001). Focusing on the relation
of immunosenescence to osteoporosis, Dr. Reis has identified
seven candidate genetic loci that may be associated with the
risk for osteoporosis in a mutant mouse model of osteoporosis.
He and his colleagues are expanding these studies to patients
to determine whether this or other gene sequences influence
the risk of osteoporosis in humans.
Table of Contents
Mitochondrial Function and
Aging
Joan McEwen, PhD, Associate Professor
The mechanisms of mitochondrial synthesis, assembly, and function
are the focus of Dr. McEwen's research efforts. Under a grant
from the American Heart Association, she has focused on the
mechanisms of assembly of the subunits and prosthetic groups
of cytochrome c oxidase.
In the second year of a Merit Review award (1999-2004) for
the research project "Role of Mitochondrial Metabolism
in Histoplasma capsulatum Virulence," she is exploring
the role of mitochondrial pathways in disease virulence. The
human pathogenic fungus H. capsulatum enters hosts
via inhalation into the lungs. In healthy, immunocompetent
individuals, infection is most often asymptomatic or mild
and resolves without clinical intervention. However, in patients
with impaired immune systems, including the elderly, who have
a greater susceptibility to a variety of infectious diseases,
the incidence of disease penetrance is higher, and serious
pulmonary or disseminated histoplasmosis may develop.
Data collected over the past year fully support Dr. McEwen's
hypotheses concerning mitochondrial electron transport activities
in fungal infection. In her view, the fungus adapts to adverse
conditions by up-regulating expression of genes for mitochondrial
respiratory system proteins. These findings highlight as a
rational drug target an alternative oxidase that is resistant
to nitric oxide but is rapidly induced by it, as it is likely
to play an important virulence role by allowing fungal respiratory
metabolism to continue in the face of inhibition of the cytochrome
oxidase system. Drugs specific for alternative oxidase may
be effective in curing a variety of human fungal diseases,
including the more common candidal infections. In addition,
such drugs would be of particular importance in treating geriatric
patients because of these patients' increased susceptibility
to fungal infections.
Maintenance of Muscle Mass during
Aging
Charlotte A. Peterson, PhD, Associate Professor
The focus of Dr. Peterson's work, funded by NIH and VA Merit
awards, has been on defining the mechanisms underlying the
loss of muscle during aging, which often leads to the loss
of functional independence. Her research efforts have led
to the characterizing of changes in gene expression, neuromuscular
interactions, the inflammatory response, the response to activity,
and the response to specific growth factors.
This past year Dr. Peterson participated as project leader
of one component of Dr. Sue Griffin's program project, funding
for which is slated to begin in June 2002. Dr. Peterson will
extend the analysis of the role of IL-1 in muscle and determine
whether IL-1 gene polymorphisms modulate muscle responses
to damage and are predictive of specific gene expression patterns
and the inflammatory response after an acute bout of resistance
exercise. The long-term goal of the work is to test the hypothesis
that IL-1 expression and the inflammatory response after an
acute exercise bout are predictive of the hypertrophic response
of muscle to chronic resistance training by elderly individuals.
Dr. Peterson also submitted a revision of her application
to be considered for joint funding by the NIH and the National
Aeronautics and Space Administration (NASA) beginning in April
2002. The original application narrowly missed the funding
cutoff. This work is a continuation of studies in the rat
identifying underlying molecular mechanisms controlling muscle
size and strength. These and other ongoing studies in Dr.
Peterson's laboratory provide evidence that the gradual decline
in muscle mass and strength that often accompanies aging may
be reversed, even at the molecular level, by exercise.
Dr. Peterson has published four papers, and two papers are
under review. Recently, she was named Director of the Microarray
Core Facility on campus and is establishing a Genomics/Bioinformatics
Group. Through gene expression analysis, she has found that
inherent changes in gene expression in cells responsible for
muscle repair may occur with age so that the cells display
an adipocyte-like phenotype. Thus, she is the first to show
that common mechanisms may contribute to loss of bone and
muscle with age.
Table of Contents
Macrophages, Erythropoiesis,
and Aging
Kodetthoor B. Udupa, PhD, Research Professor
Macrophages appear to play a role in regulating normal erythropoiesis.
Employing an in vitro culture system of the burst-forming
unit-erythroid (BFU-E), Dr. Udupa has shown that depletion
of macrophages from murine marrow significantly increases
the number of BFU-E. The addition of macrophages or macrophage-conditioned
medium neutralizes this reaction, indicating that soluble
factors mediate suppression. In fact, cytokines produced by
macrophages that are likely to affect erythroid growth, including
IL-1,
tumor necrosis factor (TNF)-,
granulocyte-macrophage colony-stimulating factor, and interferon-y,
exert suppressive effects on BFU-E. This past year Dr. Udupa
and his group demonstrated an inverse relationship between
marrow macrophage number and rate of erythropoiesis. In addition,
intramedullary concentrations of IL-1
and TNF-
are changed, depending on erythropoietic status. Marrow macrophage
number also increases with age and may contribute to age-related
diminution of the ability of the erythron to respond to increased
stimulation. His work is supported by Ortho-McNeil Pharmaceutical
Corporation and Aventis Pharmaceuticals (formerly Hoechst
Marion Roussel, Inc.).
Mechanisms of Muscle Atrophy
Esther E. Dupont-Versteegden, PhD, Research Assistant
Professor
Dr. Dupont-Versteegden joined the UAMS faculty 2 years ago.
As a part of Dr. Charlotte Peterson's group, she continues
to explore the molecular mechanisms underlying muscle atrophy.
In a paper in the American Journal of Physiology, she
reported that the number of dead or dying nuclei increases
after spinal cord transection and that exercise attenuates
this decline in muscle cell numbers. These studies are important
because they indicate that exercise can maintain muscle mass
in extreme situations such as spinal cord injury, providing
hope in the geriatric population for the maintenance of muscle
mass by exercise. In the last year, she has been working on
identifying pathways acting in the muscle cells that control
muscle mass in both young and old. Her work is supported by
a research grant from the American Federation for Aging Research.
A grant application to the NIA will likely be funded in 2002.
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