The effects of 12-week exposure to zidovudine (AZT) at 400, 500, and 600mg/kg/d were examined on expression of 542 mitochondria-related genes and mitochondrial DNA (mtDNA) copy number in the liver of male and female B6C3F1 mice to understand mitochondrial role in sex-related differences in development of lactic acidosis. Plasma lactate levels and hematologic parameters were also examined. Results indicated increased red blood cell (RBC) count in vehicle-treated controls, whereas a dose-related decline in the RBC count was noted in AZT-treated mice compared to the basal levels before treatments began. These decreases were associated with significant dose-related increases in mean corpuscular volume and mean corpuscular hemoglobin levels. This effect was greater in AZT-treated females compared to males. In both sexes, 12-week AZT or vehicle exposure significantly reduced plasma lactate levels compared to the basal levels. Results also showed modest, but significant, changes in the expression of genes associated with apoptosis and lipid metabolism at 600mg/kg/d AZT. Neither drug nor sex influenced hepatic mtDNA copy number. Altogether, 12-week AZT exposure as high as 600mg/kg/d did not impair hepatic mitochondria or induce lactic acidosis in B6C3F1 mice. However, AZT-mediated hematologic toxicity appeared to be greater in females compared to males.