Characterization of Testicular Angiotensin-Converting Enzyme before and after Semen Cryopreservation and in the Acrosome Reaction of Spermatozoids of Nelore Bulls

Abstract

The aim of this study was to characterize the testicular isoform of angio-tensin-converting enzyme (tACE) before and after semen cryopreservation, and in the acrosome reaction of sperm from Nelore bulls in vitro. Ejaculates of 10 sexually mature Nelore bulls were used. After semen was collected, 1.0 mL of the ejaculate was used for the analysis and the rest was subjected to cryopreservation. Fresh semen before freezing, and frozen/thawed semen were centrifuged twice and the pellet was resuspended intyrode’s albumin lactate pyruvate (TALP). Thereafter, 100 μL aliquots containing 100 × 10 6 spermatozoa were prepared. Aliquots of samples were used for western blot analysis, subjected to capacitation, and thereafter, acrosome reaction assays were performed </>in vitro</i>. With the help of an anti-ACE monoclonal antibody, a 100 kDa protein band was identified in the spermatozoa of Nelore bulls. Cryopreservation reduced the intensity of the protein bands obtained by western blot assay to less than half of that observed prior to freezing ( P < 0.05). Inhibition of ACE by captopril (10 μM), decreased the percentage of capacitated spermatozoa with a positive acrosome reaction ( P < 0.05), indicating the involvement of ACE in these processes. It is concluded that tACE can be found in the spermatozoa of Nelore bulls, and cryopreservation process decreases the intensity of bands of this enzyme; and that the inactivation of tACE reduces the capacity of spermatozoa to undergo the acrosome reaction.