Effect of Quercetin Supplementation in Extender on Sperm Kinematics, Extracellular Enzymes Release, and Oxidative Stress of Egyptian Buffalo Bulls Frozen-Thawed Semen.

Ahmed R M El-Khawagah,Mohamed M M Kandiel,Haney Samir

doi:10.3389/fvets.2020.604460

Ahmed R M El-Khawagah, Mohamed M M Kandiel + Show 1 more

Open Access

https://doi.org/10.3389/fvets.2020.604460

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Abstract

Buffalo spermatozoa are more sensitive for cryopreservation compared to other species. This study aimed to evaluate the consequences of quercetin against cryodamage of buffalo frozen–thawed spermatozoa characteristics. Semen of Egyptian bulls (n = 4) was extended in OptiXcell extender incorporated with quercetin at 0 (control), 2.5, 5.0, 10.0, 20.0, 40.0, and 80.0 μM before cryopreservation. Frozen–thawed semen was evaluated for sperm motility by computer-assisted sperm analyzer (CASA), viability, morphology, membrane, and acrosome integrities. The kinematics parameters including average path velocity (VAP; μm/s), straight linear velocity (VSL; μm/s), curvilinear velocity (VCL; μm/s), amplitude of lateral head displacement (ALH; μm), beat cross frequency (BCF; Hz), linearity [LIN, (VSL/VCL) × 100], and straightness [STR, (VSL/VAP) × 100] were assessed. The sperm-free extender was evaluated for aspartate aminotransferase (AST), alanine aminotransferase (ALT), and H2O2. Homogenized sperm cells were evaluated for oxidative stress biomarkers [superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX)], and lipid peroxidation [malondialdehyde (MDA)]. The highest values of total motility, progressive motility, viability, intact acrosome, and membrane integrity substantially improved with 10 μM of quercetin. STR (%) was substantially low (P < 0.01), and VCL (μm/s) and ALH (μm) were markedly high (P < 0.05) in 10 μM of quercetin. The outflow of ALT enzyme to extracellular fluid was lower with 10 μM of quercetin (P < 0.001) and higher at 2.5 μM of quercetin. The spermatozoa leaked AST was markedly lower at 5.0, 10 (P < 0.001) and 20 μM (P < 0.05) of quercetin. The activity of antioxidant enzymes was eminently low at all quercetin concentrations, and this was accompanied by the decrease in H2O2 in the media. SOD activity at 10–80 μM, CAT at 5.0–40 μM, and GPX at 2.5–80.0 μM of quercetin in spermatozoa were substantially low. MDA level significantly (P < 0.001) decreased at all quercetin concentrations. In conclusion, the incorporation of quercetin at the level of 10 μM is promising in improving buffalo semen characteristics and lower the freezing–thawing oxidative stress.

Highlights

Semen cryopreservation and its use in artificial insemination are beneficial policies for prolonged conservation of the hereditary material of superior bulls
Moderate levels of reactive oxygen species (ROS) are required for successful capacitation, acrosome reaction, and oocyte fusion [5], their overproduction during cryopreservation leads to sperm dysfunction [4]
The present study aimed to evaluate the role of quercetin in preventing the cryodamage to spermatozoa of buffalo bulls in the OptiXcell extender and to verify its influence on post-thawing spermatozoa kinematics, semen characteristics, and oxidative stress biomarkers

Summary

Introduction

Semen cryopreservation and its use in artificial insemination are beneficial policies for prolonged conservation of the hereditary material of superior bulls. Cryopreservation irreversibly damages spermatozoa resulting in their dysfunction. Despite the remarkable success in sperm cryopreservation technology, there is about a 50% decline in sperm motility after freezing and thawing [1]. The loss in sperm functionality is attributed to the cryo-damages resulting from the harsh process of freezing and thawing [2]. Several mechanisms have been claimed to be the main cause of sperm damage during the cryopreservation process including thermal shock (with intracellular and extracellular ice formation), cellular dehydration, and osmotic stress [3]. It has been suggested that freezing and thawing of spermatozoa triggers the production of reactive oxygen species (ROS) [4]. Moderate levels of ROS are required for successful capacitation, acrosome reaction, and oocyte fusion [5], their overproduction during cryopreservation leads to sperm dysfunction [4]

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