Busulfan induces oxidative stress- and Bcl-2 family gene-related apoptosis in epididymal sperm and testis of adult male mice

Busulfan as a chemotherapeutic agent causes testicular germinal epithelium depletion and cytotoxicity in germ cells. The aim of this study was to assess antioxidant status, reactive oxygen species (ROS) generation and apoptosis-related genetic markers of adult male mouse sperm following busulfan treatment. Forty adult NMRI mice (30 ± 5 g) were divided into two groups. Control and busulfan treated group were administered with 100 μL dimethyl sulfoxide and 3.2 mg/kg/day busulfan for 4 days, respectively. The superoxide dismutase and glutathione peroxidase assays were used for analyzing antioxidant status. Then, the levels of Bcl-2 family gene expression, lipid peroxidation and cytotoxicity were evaluated by Real-Time PCR, thiobarbituric and lactate dehydrogenase assays, respectively. The results showed significant decrease on antioxidant status, increase on lipid peroxidation and lactate dehydrogenase in epididymal sperm and testis of busulfan treated mice in comparison with control (P< 0.05). Real Time PCR demonstrated significantly increased-Bax gene expression and decreased-Bcl-2 gene expression in epididymal sperm of treated group (P< 0.05). The high levels of lipid peroxidation and lactate dehydrogenase revealed increased-ROS and severe cytotoxicity in epididymal sperm and testis tissue following busulfan treatment at clinical dose. The oxidative stress and increased-ROS may induce Bcl-2 family gene expression-related apoptosis following busulfan therapy in normal cells.