Effect of retinol as antioxidant on the post-thaw viability and the expression of apoptosis and developmental competence-related genes of vitrified preantral follicles in buffalo (Bubalus bubalis).

Abstract

The present study evaluated the effect of supplementation of retinol in the vitrification solution on the viability, apoptosis and development-related gene expression in vitrified buffalo preantral follicles. Preantral follicles isolated from cortical slices of ovaries were randomly assigned into three groups: Group1-Control fresh preantral follicles; Group 2-Vitrification treatment (Vitrification solution 1 (VS1) -TCM-199+25mM HEPES+Foetal bovine serum (FBS) 10%, Ethylene glycol (EG): 10%, Dimethyl sulphoxide (DMSO): 10%, Sucrose-0.3M for 4min; VS2- TCM-199+25mM HEPES+FBS10%, EG:25%, DMSO: 25%, Sucrose:0.3M for 45s); Group3-vitrification treatment +5μM of Retinol. Preantral follicles were placed in corresponding vitrification medium and plunged into liquid nitrogen (-196°C). After a week, the follicles were thawed and analysed for follicular viability and gene expression. There was no significant difference in the viability rates among the Group 1(Fresh preantral follicles) (91.46±2.39%), Group 2 (89.59±2.46%) and Group 3 (87.19±4.05%). There was a significantly (p<.05) higher mRNA expression of BCL2L1, GDF-9 and BMP-15 in the vitrification+retinol group compared with the control group. There was a significantly (p<.05) higher expression of Caspase-3 and Annexin-5 in the vitrification group and Vitrification+retinol group compared with control group of follicles. It is concluded that the supplementation of 5μM of Retinol in Vitrification solution was an efficient vitrification procedure for the vitrification of buffalo preantral follicles.