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

Ricardo G Almeida,Elisvânia F Santos,Fernando H G Furtado,Deiler S Costa,Fabio J C Faria

doi:10.4236/as.2019.104041

Ricardo G Almeida, Elisvânia F Santos + Show 3 more

Open Access

https://doi.org/10.4236/as.2019.104041

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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 × 106 spermatozoa were prepared. Aliquots of samples were used for western blot analysis, subjected to capacitation, and thereafter, acrosome reaction assays were performed in vitro. 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 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.

Highlights

Angiotensin-converting enzyme (ACE) is a zinc-dependent metalloprotease, which is anchored to the plasma membrane and exists in two isoforms
The use of a monoclonal anti-ACE antibody revealed at leastone 100-kDa protein band in the spermatozoid suspension from Nelore bulls
The pattern of the pre- (Figure 1(a)) and post-cryopreservation (Figure 1(b)) protein bands in semen with the anti-ACE antibody showed that the amount of protein was reduced following cryopreservation

Summary

Introduction

Angiotensin-converting enzyme (ACE) is a zinc-dependent metalloprotease, which is anchored to the plasma membrane and exists in two isoforms. SACE converts inactive angiotensin I into angiotensin II, a potent vasoconstrictor that stimulates the release of the hormone aldosterone, which induces sodium retention and increases blood pressure. It enhances this effect by inhibiting the action of the vasodilator bradykinin [1] [2] [3] [4] [5]. The second isoform is of lower molecular weight (110 kDa), transcribed by the same gene, and is found exclusively in male germ cells; it was designated as a testicular isoform (tACE) This isoform is not expressed in animals before puberty, which suggests that hormonal stimulation is required for its synthesis [3]. Hypophysectomized rats do not express tACE before puberty; animals that receive testosterone, follicle stimulate follicle-stimulating hormone (FSH), and luteinizing hormone (LH) soon after pituitary withdrawal can synthesize the enzyme, indicating the need for reproductive hormones in the expression of tACE [6]

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