Potential health risks associated with the clinical use of nucleoside analogue reverse transcriptase inhibitors (NRTIs) exist because the molecular basis for their effectiveness also actuates their potential toxicities. Current research suggests that one of the manifestations of this toxicity is mitochondrial damage leading to cardiac disease. The purpose of this project is to determine the nature and extent of NRTI-induced damage in mitochondria by evaluating the types and frequencies of mutations induced in mitochondrial tRNA genes of human cells exposed in vitro, and in tissues of mouse and human exposed transplacentally. The impact of these mutations upon markers of mitochondrial function (EM studies of mitochondrial structure, mtDNA depletion studies, and OXPHOS/COX enzyme assays) will be evaluated, using a modified parallelogram approach, in human lymphocytes exposed in vitro, in heart cells and lymphocytes of mice exposed in utero, and in lymphocytes and umbilical cord tissue of human infants exposed in utero. Short-term and long-term studies will be conducted to evaluating the persistence of mitochondrial DNA mutations and mitochondrial dysfunction in rodents and children exposed in utero. Commonly used NTRIs (AZT, 3TC, and d4T), either alone or in combination, will be evaluated. Preliminary studies will be conducted to determine the human-equivalent dose necessary to replicate in vivo in the mouse the plasma levels of AZT found in human children. The long range goals of this project are i) to determine the relationship between the types and frequencies of mtDNA mutations and alterations in mitochondrial function, and ii) to determine the relative potency of individual NRTIs, and/or combinations of commonly used NRTIs, to induce deleterious mitochondrial damage. Elucidation of the mechanisms underlying NRTI-induced mitochondrial damage will benefit HIV-infected patients taking NRTIs to control infection, HIV-exposed individuals who take NRTIs prophylactically to prevent HIV infection, and children receiving perinatal NRTIs to prevent HIV infection.
