Relative contribution of leukocytes and of spermatozoa to reactive oxygen species production in human sperm suspensions.

The contribution of leukocytes and of spermatozoa to reactive oxygen species (ROS) production in prepared sperm suspensions from donors and subfertility patients was compared. In both groups, more leukocytes/10(6) spermatozoa were counted in samples which produced detectable ROS than in those that did not: Donors-645 vs. 170 (medians, n = 7; p < 0.01, Kruskal-Wallis), Subfertile group-1785 (n = 18) vs. 11 (n = 8) (p < 0.005, Kruskal-Wallis), respectively. Leukocyte concentrations were correlated with basal (r = 0.826, p < 0.001) and with ROS production stimulated with 50 mumol N-formyl, met, leu, phe l-1 (N-FMLP) (r = 0.835, p < 0.001) and 100 nmol phorbol 12-myristate 13-acetate l-1 (PMA) (r = 0.835, p < 0.001) measured using a chemiluminescence assay. Leukocytes were removed from the sperm suspensions of 6 donors and from 96 ejaculates from 21 subfertility patients and ROS production was determined. Subsequently, in all 6 donors, N-FMLP did not stimulate ROS production indicating that leukocyte removal was complete, though in one case PMA stimulated low levels of ROS production. In 65 ejaculates from subfertile men the N-FMLP response was completely eliminated but in 7 of these samples PMA continued to stimulate ROS production. We conclude that infiltrating leukocytes are the predominant source of ROS production in unpurified sperm preparations. Some purified sperm suspensions could be stimulated to produce ROS by the addition of PMA indicating that spermatozoa themselves may produce ROS, albeit in much smaller amounts.