The long-term goal of this research program is to understand the etiology and pathophysiology of mitochondrial genetic disorders. From this understanding, it may be possible to define the role of mitochondrial respiratory chain dysfunction ina broad range of neurodegenerative disorders. A number of disorders are now known to be caused by mutations in the mitochondrial genome. The pathophysiology most often involves the nervous system although other systems can also be affected, particularly skeletal and cardiac muscle. In addition to the genetically-defined mitochondrial disorders, there is substantial experimental evidence that mitochondrial dysfunction is a pathophysiological component of the late- onset neurodegenerative diseases of Parkinson, Alzheimer, and Huntington. During the previous period of effort, the mitochondrial mutations that are th primary pathogenic factors in Leber hereditary optic neuropathy (LHON), one of a broad group of optic atrophies, have been identified. LHON may now be the neurodegenerative disorder with the best-defined genetic basis. As such, LHON is a good model system by which the mechanisms of neurodegeneration can be elucidated and studied.
One aim of the proposed experiments is to undertake a detailed analysis of the phenotypic effects of primary LHON mutations on cellular energy metabolism. Biochemical and metabolic analyses will be carried out with fibroblast cell cultures to test the effects of primary LHON mutations upon mitochondrial respiratory chain function and citric acid cycle activity. In addition, neuronal cell lines that lack mitochondrial DNA will be developed. These mtDNA-less sublines will then be used for cybrid fusions to enucleated LHON fibroblasts. The resulting cybrids, neuronal cells that carry LHON mutations in their mtDNA, will be used to determine, for the first time, the effects of known pathogenic mitochondrial mutations upon energy metabolism in neuronal cells and upon the expression of neuronal characteristics. The hypothesis will be tested that the pathogenicity of primary LHON mutations is due to an reduction in oxidative phosphorylation. We will also determine whether respiratory chain impairment in these cybrid cells predisposes neuronal cells to neurodegeneration through activation of apoptosis or programmed cell death. Finally, phylogenetic analysis will be used to study the origin and spread of primary LHON mutations throughout the human population during evolution, and the etiological role of secondary LHON mutations.
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