Mitochondria are the main sources of energy in the cell. They are unique mammalian organelles because they contain their own DNA (mtDNA), whose genes encode components of the respiratory chain/oxidative phosphorylation system. They are essential for the normal functioning of all cells in the body, and are absolutely critical for the function of those tissues that are highly dependent on aerobic metabolism, including heart, skeletal muscle, and brain. Since 1988, single large-scale mtDNA rearrangements, more than 100 mtDNA point mutations, as well as mendelian-inherited multiple mtDNA deletions have been associated with human diseases. Mitochondrial neuro-gastrointestinal encephalomyopathy (MNGIE) is an autosomal recessive disorder associated with multiple deletions and depletion of mtDNA in skeletal muscle. The major clinical features are: ptosis, external ophthalmoparesis, gastrointestinal dysmotility, cachexia, peripheral neuropathy, and leukodystrophy. We mapped the disease to chromosome 22q13.32-qter and subsequently identified loss-of-function mutations in the thymidine phosphorylase (TP) gene as the cause of the disorder. With the support of a NIH grant, we have continued our investigation of MNGIE. Clinicians from around the world have sent us blood samples to test for defects in thymidine phosphorylase. To date, we have identified 51 MNGIE patients. All patients tested have shown very low or no detectable activity of thymidine phosphorylase in huffy coat samples. In addition, we have identified dramatic increases of thymidine levels in plasma from patients. These findings led us to hypothesize that elevated intracellular levels of thymidine cause alterations of mitochondrial nucleotide pools that, in turn, induce point mutations, multiple deletions, and depletion of mtDNA. To test our hypothesis, we propose to study this disorder in vivo using human autopsy samples and in vitro using fibroblasts from patients. In addition, we have produced thymidine phosphorylase knock-out mice as a model for MNGIE. Our proposed studies of the pathogenesis of MNGIE are likely to enhance our understanding of nucleotide metabolism and will likely lead to more rational therapies for this uncommon, but devastating illness.
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