Comparative analysis of the ability of precursor germ cells and epididymal spermatozoa to generate reactive oxygen metabolites

Abstract

Male germ cells at various stages of differentiation from pachytene spermatocytes to mature caudal epididymal spermatozoa were examined for their ability to generate reactive oxygen species (ROS) using sensitive chemiluminescence techniques. In general, spermatozoa were found to spontaneously generate hydrogen peroxide as they progressed through the epididymis, maximal activity being observed on the release of mature cells from the caudal region into a modified Krebs-Ringer's solution. The spontaneous production of hydrogen peroxide rose rapidly during the first 10 min after the spermatozoa had been diluted into culture medium and thereafter stabilized, neither phorbol esters nor A23187 subsequently influencing this activity. Low levels of superoxide generation were also detected in suspensions of epididymal spermatozoa, but did not correlate with maturation status. However, superoxide production could be dramatically enhanced by the addition of exogenous NADPH, in a manner that was closely correlated with the stage of epididymal development being maximal for immature cells recovered from the caput epididymis in all species. Precursor germ cells (pachytene spermatocytes, round and elongate spermatids) similarly generated chemiluminescent signals compatible with the low level generation of ROS. Superoxide generation in these cells could again be stimulated by NADPH, via mechanisms that were inversely related to the stage of germ cell differentiation, the greatest activity being observed in pachytene spermatocytes. These results demonstrate that differentiating male germ cells have the potential to generate ROS, and have implications for the redox regulation of gonadal function and the development of reproductive pathologies involving oxidative stress.