Effect of Aminopeptidase N on functions and fertility of mouse spermatozoa in vitro

Aminopeptidase N (APN) is defined as a multifunctional enzyme, which regulate cellular physiology of a wide variety of cells in human. Earlier studies reported that mammalian semen shares this common enzyme as a major protein of seminal plasma that has correlation with male fertility, while the regulatory mechanisms of APN in spermatozoa are still far from being well understood. Present study was designed to investigate the role of APN in biological and chemical functions of spermatozoa using an in vitro antagonistic approach. Results showed that lower APN activity in sperm culture medium significantly increased sperm motility and the percentage of high speed spermatozoa and decreased the percentage of slow speed spermatozoa after a dose dependent inhibitor treatment (10, 100, and 1000 μM leuhistin) on epididymal mouse spermatozoa in a capacitating media for 90 min. Both 100 μM and 1000 μM decreased APN activity, while only 1000 μM decreased cell viability and increased PKA activity significantly compared to control. Nonetheless capacitation status, acrosome reaction status, and lactate dehydrogenase activity were not affected. Intriguingly, the treatment affected embryonic development through decreasing tyrosine phosphorylation of proteins and increasing reactive oxygen species levels. Further in silico analysis revealed associated regulatory proteins, which have critical functional role for male fertility.