Figure 2: The effect of high-fat diet with metformin treatment on sperm ROS concentrations, oxidative DNA damage, and mitochondrial membrane potential. (a) Intracellular sperm ROS generation (DCFDA). (b) Percentage of spermatozoa positive for MitoSox Red staining (mitochondrial superoxide). (c) Percentage of spermatozoa positive for 8OHdG staining (oxidative DNA damage). (d) The ratio of red-to-green JC-1 fluorescents in spermatozoa (mitochondrial membrane potential). Data are expressed as mean ± s.e.m. Data are representative of n = 9 control diet, n = 8 high-fat diet and n = 8 high-fat diet + metformin males. a and b letters denote significance at P < 0.05. Figures bearing the same superscript letter are not significantly different. ROS: reactive oxygen species; 8OHdG: 8-hydroxy-2'-deoxyguanosine; CD: control diet; HFD: high-fat diet; s.e.m.: standard error of the mean.