Antioxidants supplementation improves the quality of in vitro produced ovine embryos with amendments in key development gene expressions

Abstract

The present study assessed the effects of supplementation of different antioxidants on oocyte maturation, embryo production, reactive oxygen species (ROS) production and expression of key developmental genes. In this study, using ovine as an animal model, we tested the hypothesis that antioxidant supplementation enhanced the developmental competence of oocytes. Ovine oocytes aspirated from local abattoir-derived ovaries were subjected to IVM with different concentrations of antioxidants [(Melatonin, Ascorbic acid (Vit C), alpha-tocopherol (Vit E), Sodium selenite (SS)]. Oocytes matured without any antioxidant supplementation were used as controls. The oocytes were assessed for maturation rates and ROS levels. Further, embryo production rates in terms of cleavage, blastocysts and total cell numbers were evaluated after performing in vitro fertilization. Real-Time PCR analysis was used to evaluate the expression of stress related gene (SOD-1), growth related (GDF-9, BMP-15), and apoptosis-related genes (BCL-2 and BAX). We observed that maturation rates were significantly higher in alpha-tocopherol (100 μM; 92.4%) groups followed by melatonin (30 μM; 89.1%) group. However, blastocyst rates in ascorbic acid (100 μM; 19.5%), melatonin (30 μM; 18.4%), alpha-tocopherol (100 μM; 18.2%), and sodium selenite (20 μM; 16.9%) groups were significantly higher (P 0.05) than that observed in the control groups. Total cell numbers in blastocysts in the melatonin, ascorbic acid and alpha-tocopherol groups were significantly higher than those observed in sodium selenite and control groups. ROS production was reduced in groups treated with melatonin (30 μM), vitamin C (100 μM), sodium selenite (20 μM) and α-tocopherol (200 μM) compared with that observed in the control group. Supplementation of antioxidants caused the alterations in mRNA expression of growth, stress, and apoptosis related gene expression in matured oocytes. The results recommend that antioxidants alpha-tocopherol (200 μM), sodium selenite (40 μM), melatonin (30 μM) and ascorbic acid (100 μM) during IVM reduced the oxidative stress by decreasing ROS levels in oocytes, thus improving embryo quantity and quality.