Graduate School of Medical Sciences
A partnership with the Sloan Kettering Institute

Gary Gibson

Professor
Niroshana Anandasabapathy

Bio

Signal Transduction and Oxidative Process in Neurogeneration

Dr. Gary Gibson and his research group are trying to discover the underlying cause of age-related neurodegenerative diseases and to develop effective therapies. The most common age-related neurodegenerative diseases are Alzheimer's, Parkinson's and Huntington diseases. The research shows that fundamental mechanisms of damage may be similar in many age-related diseases and stroke. The brain is very dependent on oxygen and glucose (sugar). Our research suggests that a reduced ability to use glucose and oxygen either causes the disease(s) or is a critical clinically relevant change. Dr.Gibson and his team are trying to determine why this reduction occurs and its consequences for brain function. Abnormalities in the use of glucose and oxygen cause abnormal regulation of calcium and production of excess free radicals, and these may lead to the brain dysfunction. Thus, preventing the free radical damage or change in calcium with specific drugs should protect these proteins and thus save the brain. They are using cell from patients, genetically modified cells and animal models to test these possibilities and to evaluate the effectiveness of drugs.

Description of research projects. Our research and be divided into four areas.

KGDHC and neurodegenerative disease. Mitochondrial dysfunction and oxidative stress are consistent features of multiple neurodegenerative diseases including Alzheimer's disease (AD). Reductions in the α-ketoglutarate dehydrogenase complex (KGDHC), oxidative stress, diminished metabolism and neurodegeneration appear closely linked. The activity of KGDHC, a key mitochondrial enzyme complex that consists of three proteins, is markedly reduced in brains in several neurodegenerative diseases. The overall goals of this research is to determine the consequences of the decline, the cause of the reduction in KGDHC in brains from patients with neurodegenerative disease, and how the decline in activity or the consequences might be ameliorated. The reduction in KGDHC activity is not simply secondary to neurodegeneration, since the decline occurs in brain areas vulnerable to degeneration, and in regions without overt neuropathology. Future studies will test the closely associated hypotheses that: (i) the diminished KGDHC activity in AD is due to the sensitivity of KGDHC to post-translational modifications including oxidative stress; (ii) mitochondrial function including the response to oxidant stress and release of pro-apoptotic proteins is sensitive to reductions in the KGDHC activities; (iii) neurons are particularly sensitive to diminished KGDHC (iv) a direct interaction of KGDHC with thioredoxin underlies the sensitivity of KGDHC to oxidative stress and converts diminished KGDHC to abnormal cell function. The experiments will determine if the reductions in brain KGDHC activity and other neurodegenerative diseases are due to post-translational changes by using a combination of antibodies that detect common modifications and mass spectrometry. Experiments to determine the consequences of reduced KGDHC activities on cell function will utilize a specific inhibitor, as well as genetic manipulation of individual KGDHC subunits in multiple cell models and cell types from transgenic mice. The research will test the role of each subunit and determine if neurons are more vulnerable than other brain cells. These experiments will test the relation of diminished KGDHC to mitochondrial membrane potentials, to the stimulation of the release of cytochrome c and to the response to oxidant challenge. Further experiments will test whether thioredoxin mediates the consequences of an inhibition of KGDHC through a linkage with cellular thiol metabolism. In summary, the sensitivity of KGDHC to multiple oxidative stressors and its importance to cellular function suggest that multiple factors converge at KGDHC to cause neurodegeneration. Successful completion of these studies will provide new understanding of the molecular mechanisms underlying AD and other neurodegenerative diseases are likely to suggest new therapeutic strategies.

Enzymes of the tricarboxylic acid cycle in neurodeneration.Diminished brain metabolism and oxidative stress are characteristic features of Alzheimer's Disease (AD). The mechanisms underlying these changes are as yet poorly defined. Our recent studies indicate that a marker of mitochondrial damage, the α ketoglutarate dehydrogenase complex (KGDHC), correlates at least as well with clinical disability as do plaque and tangle counts. KGDHC and several other mitochondrial enzymes are known to be sensitive to reactive oxygen species (ROS). These studies test the hypothesis that: impairment of select mitochondrial enzymes by ROS is an important component of the cascade of events that leads to diminished metabolism and to the cognitive deficits in AD. This hypothesis will be tested on human autopsy brains collected by our collaborators at the Mt Sinai (NY) ADRC, who have collected several hundred samples of brain from patients whose pre terminal neuropsychological status has been determined using the Clinical Dementia Rating (CDR). Quantitative markers of oxidative stress and activities of specific mitochondrial enzymes will be compared to clinical status (CDR) and to markers of AD pathology including plaque and tangle counts, by refined statistical methods described in the proposal. Tissue culture models will also be used, so as to do mechanistic experiments on the effects of specific ROS on the activities of the same mitochondrial enzymes examined in the necessarily correlational studies of human autopsied brain. The models will be: (1) cultured fibroblasts from AD patients, to test the effects of ROS on cells which have the same genetic background as that in which the disease is expressed; (2) culture models of neurons, the most vulnerable cell type in AD brains. These models will also provide systems to test the efficacy of approaches to limit or reverse the changes in mitochondrial enzymes due to ROS. Thus, these studies will not only help clarify mechanisms in AD, but also have direct implications for the development of new therapies.

Thiamine deficiencyThiamine (vitamin B1) deficiency (TD) produces a mild, chronic impairment of oxidative metabolism that models the diminished metabolism and reduced activities of the thiamine-dependent mitochondrial enzymes that occur in brain in several common age-related neurodegenerative disorders. Regionally selective neurodegeneration and activation of astrocytes, microglia and endothelial cells occur in TD and in these diseases. Vascular changes, inflammatory responses, oxidative stress and neuronal death are present in brains from TD mice and in brains from patients who die from common neurodegenerative diseases. A mechanistic sequence of events cannot be discovered in autopsied human brain, but can be studied effectively in experimental animals. Several features of the TD model make it amenable to analysis of the interactions leading to neurodegeneration: 1) The time course of the events leading to neuronal death is prolonged (11 days). 2) The death occurs in a discrete nucleus with a well-defined number of neurons. 3) The model exists in mice so transgenic animals can be used to test mechanism. The proposed experiments will test the following hypothesis: TD-induced abnormalities in metabolism increase neural production of cytokines, which activate microglia and astrocytes and stimulate endothelial cells to promote entry of peripheral immune cells. Cytotoxic compounds that are released from these activated cells combine to produce neurodegeneration. The underlying mechanisms will be tested in vivo by the strategy successfully used in our ongoing experiments. The sequences of responses of cell-specific changes in markers of inflammation, vascular responses and oxidative stress will be established. Specific drug treatments and, where feasible, transgenic animals will be used to test whether each specific step is critical in the cascade leading to neurodegeneration. An understanding of these mechanisms will likely suggest new ways to overcome the consequences of the mild, chronic impairment of oxidative metabolism that accompanies numerous age-related neurodegenerative disorders.

Fibroblasts studies.The continuing goal of our research is to understand the molecular basis of demonstrated abnormalities in calcium regulation and their clinical relevance to Alzheimer's Disease. Calcium regulation is closely linked to the metabolism of reactive oxygen species (ROS) and oxygen-dependent pathways of energy production, both of which are altered in AD brain. However, interpretation of observations in brain has been hampered by the difficulty in determining whether an alteration is causative or secondary to neurodegeneration. Abnormalities in calcium regulation persist in fibroblasts cultured from patients with either familial or non-familial forms of AD compared to controls including patients with other neurodegenerative disorders. Properties that persist in cultured fibroblasts are inherent traits of AD cells that are independent of diet, drugs or neurodegeneration. Our completed studies with fibroblast cell lines from multiple members of one presenilin-1 AD family provide evidence of increased oxidative stress in AD cells. To determine the role of ROS in the production of the abnormalities of calcium regulation and oxygen-dependent pathways of energy production in cells from AD patients, the proposed experiments will use cultured fibroblasts to test a three part hypothesis: (a) cells from AD patients produce excessive ROS and are more sensitive to oxidants than controls, (b) abnormalities in cellular ROS metabolism underlie AD-related changes in cellular calcium regulation and mitochondrial function, (c) AD/control differences in calcium and ROS are diagnostic and predictive for AD. The results of these experiments will reveal whether various AD gene mutations alter the interactions of oxidative stress, oxidative metabolism and calcium. Demonstration that alterations in specific oxidant species or their cognate antioxidant systems are a triggering event that lead to abnormalities in calcium signaling and mitochondrial function would have pathological, therapeutic and diagnostic significance.

Current Areas of Focus

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