Characterization of Angiotensin-Converting Enzyme before and after Cryopreservation of Gir Semen

Abstract

The aim of this study was to characterize the angiotensin-converting enzyme (ACE) in Gir semen before and after cryopreservation. The ejaculate of five sexually mature bulls was used. After collection, one 1-mL aliquot of fresh semen was analyzed immediately, and the rest of the semen was cryopreserved in liquid nitrogen for subsequent analysis. Freshly collected semen and thawed cryopreserved semen were centrifuged twice with Tyrode's albumin lactate pyruvate medium (TALP) to remove plasma and extender, respectively. Samples were then subjected to western blotting, immunocytochemistry, and enzymatic activity techniques. At least one 100 kDa band was observed in every bull analyzed using western blotting with an anti-ACE monoclonal antibody, and band intensity decreased by 70% (p < 0.05) after cryopreservation. Immunocytochemistry showed periacrosomal ACE localization, and the area stained by the fluorescent antibody significantly decreased (p < 0.05) after cryopreservation. Enzyme activity was evaluated using FAPGG substrate hydrolysis, which was significantly lower (p < 0.05) in cryopreserved semen than in fresh semen. Therefore, the process of cryopreservation decreases ACE band intensity and enzyme activity in Gir bull semen, and reduces the stained area in immunocytochemistry.

Protamine and other proteins in sperm and seminal plasma as molecular markers of bull fertility

“…Spermadhesin 2 (spermadhesinZ13) which is a protein with a coding gene located on the 26 th chromosome in cattle is reported to have a negative impact on fertility, reduces sperm motility, and is expressed in many low fertility cattle [35,49]. Štiavnická et al [50] found that a high ubiquitin protein in cow sperm is closely related to quality and low fertility in bulls.…”