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Aab Home Page The Center for Aging and Developmental Biology Department of Biomedical Genetics The Center for Cardiovascular Research The Center for Oral Biology Center for Pediatric Biomedical Research The Center for Vaccine Biology and Immunology New Research Building Research at the URMC Rochester Alzheimer's Disease Center |
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Center for Aging and Developmental Biology
This is an unprecedented opportunity to apply recent advances in our understanding of basic mechanisms to problems affecting the development, maturation and dysfunction of the nervous system. This includes the study in diverse experimental organisms of neurogenesis, differentiation, cell death, synaptic specification, behavioral development, critical periods and normal changes in neural organization and function that accompany senescence. Additional emphasis will be placed on molecular-pathological integration and its potential for translation opportunities in therapy of neurological disorders. The Center will build upon existing programs in systems, toxicological, and cognitive Neurosciences at the University of Rochester with the long term goal of developing effective therapies for neurological dysfunction associated both with disease and normative aging. Current Faculty of the Center for Aging and Developmental Biology
Research: Neurobiology of Aging and Alzheimer's Disease; Profiling expressions of multiple messages from single defined neurons in human and animal brains. The study of molecular biology of the brain has been limited by methods that take sizeable pieces of tissue, grind them up and perform biological studies. However, we know that within any, even very small sample of tissue are cells of many different types, and in the case of neurodegenerative disease, in many different stages of disease progression. These facts argued for examining single cells in aging and neurodegenerative disease. To do this we use methods of immunochemistry, insitu hybridization, and combinations of these two. In addition, we have recently developed methods for extracting messenger RNA from single identified cells, amplifying this message a million fold and determining the levels of expression of multiple genes from each single cell. The resulting masses of data are analyzed using advanced methods of multi-variate statistics and bio-informatics. Studies using these methods have demonstrated the re-expression of some cell cycle genes in neurons from the Alzheimer's diseased brain. Research: Genetic analysis of regulatory network controlling the commitment and differentiation of retinal neurons inmammalian development. Transcriptional factors play an important role in the neuronal cell specification, differentiation, and maturation. In our earlier studies, we have shown that the targeted deletions of he close-related POU-domain genes, brn-3a, brn-3b and brn-3c, result in the loss of neurons in the somatosensory, retina and inner ear, respectively. To further investigate the mechanisms of the neuronal cell loss in these mutants, we have used the lacZ and human placental alkaline phosphatase knocked into brn-3b locus to study the fate and axonal projection process of the brn-3b mutant retinal ganglion cells in vivo and in explant culture. Our results indicated that brn-3b gene was not required for the initial commitment of neuronal progenitor cells to the retinal ganglion cell fate or for the migration of these cells to the retinal ganglion cell layer. However, brn-3b gene was imperative for the terminal differentiation of retinal ganglion cells. Without brn-3b gene, the retina ganglion cells failed to project axonal process during development and degenerated by apoptosis. Our results suggest that brn-3b gene controls the activity of genes which play essential roles in the formation of retinal ganglion cells. Similarly, we use the targeted mutation approach to study the role of mouse math5 gene, a bHLH transcriptional factor gene homologous to Drosophila atonal gene. Expression of the math5 were first detected in cells destined for the retinal ganglion cell fate and prior to the activation of brn-3b gene. The expression profile of math5 gene suggests its role in the initial commitment of retinal progenitor cells to retinal ganglion cell fate. Deletion of math5 in mouse resulted in the severe loss of retinal ganglion cells during embryonic development. Our results indicated that math5 may indeed function in determining the retinal ganglion cell fate. Research: Effects of HIV-1 infection on neuronal function, signaling, and fate in the CNS Our laboratory has focused on the effects of chronic inflammation caused by human immunodeficiency virus type 1 (HIV-1) infection in the central nervous system (CNS). In particular, we are interested in the ability of the HV-1 regulatory gene product Tat to down-regulate the production of dopamine in the nigrostriatal pathway, as a possible mechanism for the Parkinsonism associated with HIV-1 infection of the CNS. We are also actively exploring crosstalk between neurons and glial cells by molecules with immunologic functions, including the novel chemokine fractalkine, and the intercellular adhesion molecule, ICAM-5. Both of these proteins are expressed in certain subsets of telencephalic neurons and may directly mediate communication between neurons and microglia (which have receptors for both proteins) in normal and pathologic settings. Fractalkine may have both autocrine and paracrine functions to protect glutamatergic neurons in HIV-1 encephalitis. We are now investigating whether fractalkine species have a neuroprotective function in other neurodegenerative conditions. We have recently demonstrated that the frequency of ICAM-5 that can be detected in serum of patients with HIV-1 is markedly elevated compared to normal volunteers. Because ICAM-5 is exclusively located in the CNS, these findings suggest that ICAM-5 may be a potential marker of blood-brain barrier dysfunction in HIV-1 infection of the CNS. Based on this observation, we are also investigating how ICAM-5 is transported across the blood-brain barrier into the serum during acute damage vulnerable populations of neurons from hypoxic-ischemic injury and chronic damage from lentiviral and retroviral infections of the CNS. These findings in turn may help us to understand how communication between neurons and microglia can result in neurodegeneration of the dendritic arbor and loss of higher cortical functions in diseases that result in inflammation of the CNS.
Research: Nervous system development and neuronal regeneration following traumatic injury There is growing evidence that the molecular mechanisms that underlie the sculpting of neuronal networks during development also apply to aspects of adult neuronal plasticity and repair following injury. Much progress has been made in identifying molecules that regulate axonal pathfinding but still little is known about how these molecules directly contribute to the wiring and maintenance of complex neuronal connectivity in vivo. Using a genetic approach, our research aims to elucidate the mechanisms that govern the assembly of specific neural circuitry. Understanding the principles of network assembly will have important implications for the development of strategies aimed to promote neural repair following trauma or disease. Ongoing research projects are aimed to understand the role played by semaphorins and their receptors in the developing, mature, and regenerating mammalian nervous system. The Semaphorins are a large family of secreted and membrane bound proteins, several of which have been shown to participate in the patterning of the developing nervous system. More than 25 members of the semaphorin gene family have been identified and grouped into subclasses based on structural similarities. Class 3 semaphorins, which include mammalian Sema3A, Sema3C, and Sema3F, are secreted proteins that function as potent chemorepellents for specific classes of neurons. Sema3A is essential for neural development, since mice lacking this secreted semaphorin exhibit severe defects in projections of cranial and spinal nerves. Recently, the neuropilins and plexins have been identified as cell surface receptors for semaphorins. Neuropilins are a small family of type 1 transmembrane proteins that bind secreted semaphorins with high affinity. Neuropilins are essential components of the semaphorin receptor complex and impart functional specificity toward different secreted semaphorin family members. We have found that neuropilin-2 is a receptor for the secreted semaphorin Sema3F in cultured sympathetic neurons. To study neuropilin-2 signaling in vivo, we have recently generated a null mutation at the mouse neuropilin-2 locus. Our initial analysis of neuropilin-2 null mice revealed specific defects in cranial nerves and several major central nervous system fiber tracts that normally express neuropilin-2. Interestingly, despite of many major defects in the nervous system, many of the neuropilin-2 null mice survive into adulthood, providing a unique opportunity to determine the role played by semaphorins and their receptors throughout development and in the mature nervous system. Long-term goals of ongoing research projects include: (1) to elucidate the molecular mechanisms by which axons are guided to their targets during neural development; (2) to apply this knowledge to systems that model traumatic injury of the adult nervous system in order to develop means and strategies to promote neuronal regeneration.
Research: The Role and Control of Neurogenesis in the Development and Modification of the Cerebral Cortex The formation of our cerebral cortex during embryonic development depends on the generation of the appropriate number of neurons and their proper assembly into functional circuits. Modifications of this developmental program can generate differences in cortical size across species (such as the evolutionary expansion of the primate cerebral cortex), produce cortical pathology within a species, and even allow neuronal production to continue into adulthood in some brain regions but not others. In my laboratory, we are interested in how the neurons of the cerebral cortex are produced and organized into neural circuits. To address this issue we are focusing on the process of neurogenesis - the means by which cortical neurons are generated from dividing progenitor cells. Our long-term objectives are to understand: (1) the role of neurogenesis in building cortical circuits and how it is regulated, (2) how specific changes in neurogenesis contributed to the evolutionary expansion of the cortex and may underlie the pathogenesis of particular cortical abnormalities in humans, and (3) why neurogenesis in some brain regions persists beyond early developmental periods, into adulthood. This research involves comparing cell proliferation parameters across diverse species to determine how changes in neurogenesis generate differences in cortical size and organization. Moreover, the use of experimental in vivo and in vitro models will allow us to decipher the mechanisms that govern these proliferation parameters during normal development and, in some brain regions, throughout life. Research: Use of yeast and mouse models for the study of childrens neurodegenerative diseases. Dr. Pearce uses yeast and mouse models for the study of childrens neurodegenerative diseases. Batten disease is one of the more common childhood neurodegenerative diseases. Children with this disease usually suffer visual failure, psychomotor deterioration, seizures and premature death. Although the underlying defect to this disease, a defective CLN3 gene, was first identified in 1995, very little progress has been made in understanding Cln3p function and how a defective Cln3p causes Batten disease. Individuals with Batten disease exhibit accumulation of lipopigments in the lysosome, predominantly the proteolipid, mitochondrial ATPase subunit c, leading people to believe that the turnover of this protein is somehow affected. Our lab has cloned the yeast homolog to Cln3p, designated Btn1p, and has established that this protein is not involved in the degradation of mitochondrial proteins. We have established that vaculolar/lysosomal pH regulation is altered in yeast strains lacking Btn1p. We are using the numerous genetic techniques available to a yeast molecular biologist to study the function of Btn1p, which can then be applied to understanding Batten disease. Similarly this approach is used for investigating other childhhod disorders, such as Niemann Pick Type C disease. We are also studying a mouse that lacks the mouse equivalent of CLN3. The process and progression of CNS-degeneration which is specific to lacking the CLN3-protein is being examined using a variety of histological and molecular biological techniques. In addition a molecular pattern is being established by looking at differences in gene expression between normal and cln3-knockout mice using microarray technology.
Research: Transgenic animal modeling and the development of heteroplasmic transmitochondrial animal models of human disease Dr. Pinkert's research involves gene transfer, expression and regulation
using transgenic animal modeling. His work has illustrated the potential
of modifying the immune system, enhancing growth performance and the
feasibility of biopharmaceutical production (molecular farming). Additionally,
a number of enabling technologies and procedures have been developed
for the genetic engineering of both nuclear and mitochondrial genomes.
Most recently, Dr. Pinkert's laboratory has embarked on pioneering
studies revolving around mitochondrial transfer techniques and the
development of animals harboring foreign mitochondrial genomes. In
humans, severely debilitating and lethal metabolic and cellular disorders
exist due to mutations arising exclusively within the mitochondrial
genome. At present, there are scores of mutations of the mitochondrial
genome that are known to be the underlying causes of various degenerative
disorders. These mtDNA mutations, many of which exist in a heteroplasmic
state (where both normal and mutant mitochondrial genomes coexist
in varying proportions within the same individual), mainly affect
tissues with high cellular energy requirements such as brain, optic
nerve, cardiac muscle, skeletal muscle, kidney and endocrine organs.
In contrast to nuclear genes, mitochondrial gene replication and function
differ markedly in a number of ways - from exclusive matrilineal inheritance
of mitochondria and mitochondrial DNA (mtDNA), to the presence of
hundreds or thousands of mitochondria (each with one to ten copies
of the mitochondrial genome) within a given cell. Various human diseases
have been associated with specific mtDNA point mutations, deletions
and duplications including diabetes mellitus, myocardiopathy, retinitis
pigmentosa, MERRF and MELAS diseases, as well age-associated changes
in the functional integrity of mitochondria (as seen in Parkinson's,
Alzheimer's and Huntington's diseases). Therefore, the ability to
manipulate the mitochondrial genome and to regulate the expression
of mitochondrial genes would provide one possible mode of gene therapy
for many of these disease states. The creation of transmitochondrial
mice represents a new model system that will provide a greater understanding
of mitochondrial dynamics, leading to therapeutic strategies for human
metabolic diseases affected by aberrations in mitochondrial function
or mutation.
Research: Neuronal development in the nematode C. elegans The extraordinary diversity of cell type in the nervous system arises from complex genetic networks that act during development. Understanding the pathways that specify neural competence, pan-neuronal characteristics and neural-subtype identity is essential for understanding and treating neurological disease. The relatively simple nervous system of the nematode C. elegans, combined with the organism's fully-described cell lineage, sequenced genome, and conservation of developmental pathways, make it an ideal system for approaching these issues. Our work focuses on three-celled sensory organs in the C. elegans male tail called rays. Each ray contains a structural cell and two distinct sensory neurons, all three of which descend from a single precursor cell. Ray development requires the functions of two evolutionarily-conserved basic-helix-loop-helix (bHLH) transcription factors: LIN-32, the C. elegans atonal/MATH ortholog, and HLH-2, the E/daughterless ortholog. We have previously shown that mutations in these factors disrupt the ray developmental lineage at multiple points, suggesting that LIN-32 and HLH-2 activate multiple targets important for different steps of ray cell fate specification. To find additional components of the ray developmental pathway, we have used DNA microarrays to compare gene expression between males with extra rays and males lacking rays. Using this approach, we have identified a number of new ray differentiation genes (downstream targets of the ray developmental pathway) as well as regulatory factors that could be components of these pathways. We are currently taking a multidisciplinary approach (using forward and reverse genetics, biochemistry, genomics and bioinformatics) to characterize these components, identify new ones, and understand how their functions are integrated as part of a complex genetic network. We expect that these relationships will lend insight into similar, less experimentally-accessible processes in higher organisms.
Research: Cellular and Molecular Mechanisms of Neuroreceptor Interactions and Plasticity in the CNS My research interests revolve around issues related to the actions of neurotransmitters and the consequences of neurotransmitter receptor interactions on adult as well as developing neurons in the central nervous system. Within this context, my laboratory takes the view that neurons undergo constant adaptations in response to acute or chronic environmental changes and that this ultimately leads to altered patterns or efficiency of neurotransmission. Such plastic adaptations may occur in development, in response to neurodegenerative insults and may be manifest in alterations in neuronal morphology and/or functional properties, as well as in the molecular or genetic profiles of the neuron. Thus, my laboratory adopts a multidisciplinary approach, incorporating neuroanatomical, patch clamp electrophysiological and molecular biological techniques. My laboratory has also developed molecular biological techniques that can be combined with electrophysiology to analyze gene expression profiles and function in the same cell. The analysis at the level of single cells is now being extended to include the detection of proteins. This combined strategy is being employed in ongoing research projects to investigate GABA, glutamate and cholinergic receptors in development, including the influence of neurotrophins, and in an animal model of chronic alcoholism. These projects have in common the theme of revealing coordinated changes in function, the expression of genes and, ultimately, the encoded proteins.
Research: Development and Implementation of HSV Amplicon-based Gene Transfer Our research relates to the development of novel gene transfer technologies based on the plasmid-based Herpes Simplex Virus (HSV) amplicon vector system. The ultimate goal is to develop optimized amplicon vectors to safely express neuroprotective factors to treat human neurodegenerative disorders, as well as to express novel antigens that impart protection against infectious and oncologic diseases. Disease targets include Parkinson's and Alzheimer's diseases, prostate cancer, chronic lymphocytic leukemia, and AIDS. Another avenue of research relates to examination of immune responses elicited as a result of amplicon administration in the CNS. Understanding the fundamental efficacy and safety issues regarding therapeutic gene delivery will allow us to properly implement potential clinical applications employing HSV amplicon vectors.
Research: Response of effected and neighboring neurons to cytoskeletal disruption in late-onset neuro-degenerative diseases, particularly Alzheimer's Disease, Amyotrophic Lateral Sclerosis, and Parkinson's Disease. Late-onset neurodegenerative diseases, including Alzheimer's disease
(AD), Amyotrophic Lateral Sclerosis (ALS), and Parkinson's Disease
(PD), are the result of abnormal function of specific populations
of neurons within the central nervous system. The deficits leading
to clinical symptomology include a progressive synapse loss within
specific neuronal networks. The synapse loss is likely mediated in
part by cell loss but also by synapse dysfunction. Numerous factors
have been identified as synaptic degenerators, including disruption
of the cytoskeletal network, particularly of the cytoskeletal proteins
mediating axonal transport. Our primary research interests focus on
axonal transport disruption mediated by cytoskeletal protein alterations.
We are particularly interested in the initial response of both effected
and neighboring uneffected cells to events of cytoskeletal disruption.
Ongoing research focuses on the response of neurons undergoing microtubule
disruption. To this end, we use primary hippocampal neurons in co-culture
to examine microtubule disruption due to nocodazole (a microtubule
inhibitor drug) or pathogenic tau proteins, including P301L and R406W.
We are also investigating effects of overexpression of normal four-repeat
tau on microtubule patterns. We are embarking on the development of
late-onset models which will express tau pathogenitors at the CNS
location and time of choosing of the investigator. Our analyses are
multi-faceted combining histological, immunocytochemical, ultrastructural,
quantitative, and molecular biological techniques to analyze alterations
resulting from cytoskeletal disruption. Long-term research goals include:
1) to determine the initial response of cell defense systems (lysosomal,
ubiquitin-proteasomal-system (UPS)) to disruption of cytoskeletal
proteins (microtubules and neurofilaments in particular), 2) to determine
the response of cell defense systems of near neighbor neurons (apparently
uneffected), 3) to determine if similarities exist between the late-onset
neurodegenerative diseases of Alzheimer's Disease, Amyotrophic Lateral
Sclerosis, and Parkinson's Disease, in regards to the response of
neurons undergoing cytoskeletal disruption, and 4) to create models
which allow control over the CNS location and age of expression of
cytoskeletal disruptors. The overall goal of our research is to mimic
molecular cytoskeletal disruption, and to mechanistically dissect
the response of both effected and neighboring uneffected neurons with
an aim towards providing therapeutic targets. Research: Alzheimer's disease (AD) is the most prevalent form of dementia associated with aging. Immune responses have been proposed as being integral for initiating and/or propagating AD pathogenesis within the brain. With recent immunotherapeutic approach targeting Aß for treating AD in humans, it has become more evident that immune responses play a critical role in the pathogenesis of AD. However, the real immunological mechanisms underlying the beneficial effects (decreased Aß plaques and behavior improvement) and the adverse effects (brain inflammation) induced by Aß vaccination remain largely unknown. My research in Dr. Federoff's lab focus on the T cell responses induced by Aß. Aß is delivered to mice by the herpes simplex virus (HSV) amplicon. Cellular immune responses against Aß induced by HSV-Aß vaccination are evaluated by ELISPOT and CTL assays. We are currently testing the roles of CD4+ and CD8+ T cell subsets in Aß-mediated immune responses. The other direction of our research focus on the role of molecular adjuvants in modulating the Th1 and Th2 immune responses to HSV-Aß. The goal of our research is to design safer anti-Aß vaccines by either specifically blocking or activating certain subsets of T cells by the HSV-Aß vaccine, or by incorporating proper molecular adjuvants with the HSV-Aß vaccine if desired.
Research: Synaptic transmission in the central nervous system Synaptic transmission in the central nervous system governs behavioral functions of individuals. Many multi-gene based neurodegenerative disorders, such as Alzheimer's disease, and Parkinson's disease, have a delayed onset of obvious behavioral deficits. However, telltale behavioral signs are detected earlier with a more sensitive battery of behavioral now. These more subtle behavioral deficits may be a reflection of early pathologic changes in synaptic transmission in the affected forebrain structures due to the gain or loss of function of the mutated genes. My research investigates the function of neurons in the forebrain structures, including neostriatum, cortex, and hippocampus, in normal and pathological conditions. Specifically, I am interesting in studying how lipid metabolites, cytokines, chemokines, and apolipoprotein metabolites contribute to the abnormal function of affected neurons in these neurodegenerative disorders, with respect to synaptic transmission and intrinsic membrane properties in in vitro and in vivo models.
Research: Study of Environmental and Genetic Interactions in Parkinson's Disease Parkinson's disease affects over one million Americans and is the second leading neurodegenerative disease. The etiology of Parkinson's disease is currently under investigation but is thought to involve both genetic and environmental triggers. Despite multiple initiating factors we and others envisage a common pathobiologic model of Parkinson's disease. Common to both sporadic and genetic forms of Parkinson's disease is the profound loss of nigrostriatal dopamine neurons and the hallmark pathological feature of intracytoplasmic inclusions called Lewy bodies. Lewy bodies are comprised of a number of proteins including alpha-synuclein. We are investigating the role of dopamine and synuclein in the pathogenesis of Parkinson's disease. Utilizing both in vitro and in vivo models of Parkinson's disease we are studying the role of protein aggregation in this neurodegenerative disease. Furthermore, we are exploring the utility of single-chain antibodies as a possible therapeutic model for Parkinson's disease. |
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Howard
J. Federoff, M.D., Ph.D.
Paul
D. Coleman, Ph.D. 
Lin
Gan, Ph.D.
Harris
Gelbard, M.D.,Ph.D.
Roman
Giger, Ph.D.
David
Kornack, Ph.D.
David
Pearce, Ph.D.
Carl
A. Pinkert, Ph.D.
Douglas
Portman, Ph.D.
Hermes
Yeh, Ph.D.
William
J. Bowers, Ph.D.
Linda Callahan, Ph.D.
Ya-Hui GraceChiu, Ph.D.
Shao-Ming (Patrick) Lu, Ph.D.
Kathleen
A. Maguire-Zeiss, Ph.D.