The Impact of NADPH Oxidase 5 Activity and Reactive Oxygen Species on Capacitated Human Sperm.

Abstract

Background: Progesterone (P4) activates sperm calcium channels (CatSper), allowing calcium to enter the cell, which activates NADPH Oxidase-5 (NOX5) and produces reactive oxygen species (ROS). While calcium and ROS are essential for sperm capacitation, the role of NOX5 in capacitated sperm is unclear. This study investigated NOX5 activity in capacitated human sperm.

Methods: Forty semen samples from fertile men were processed, with motile sperm separated and divided into nine groups: control (Ham's F-10), solvent (DMSO), progesterone, diphenyleneiodonium chloride (DPI, NOX5 inhibitor), phorbol-12-myristate 13-acetate (PMA, NOX5 activator), P4+DPI, P4+PMA, Trolox, and P4+ Trolox. Sperm kinematics, membrane integrity, survival rate, and ROS production was evaluated. Data were analyzed using ANOVA and Kruskal-Wallis tests, p≤ 0.05 considered statistically significant.

Results: Progressive motility significantly decreased with DPI (56.2±2.1%) and PMA (56.5±2.1%), both alone and combined with progesterone (58.0±2.0% and 57.4±2.2%), compared to the progesterone group (66.0±1.9%). No significant change was observed in the Trolox groups. Progesterone, alone or combined with DPI, PMA, and Trolox, significantly reduced sperm linearity from 0.6±0.01 to 0.5±0.01%. Straight-line velocity decreased in P4+PMA and P4+Trolox groups (88.2±4.4 and 89.7±3.9 μm/s) compared to the control group (105.0±5.5 μm/s). Trolox reduced ROS content, while other treatments had no effect on ROS levels.

Conclusion: NOX5 does not play a prominent role in capacitated sperm. The negative effects of PMA and DPI on sperm motility appear independent of their actions on NOX5 and ROS production. Trolox did not affect sperm motility and survival, indicating that capacitated sperm require little or no ROS.