Gallic acid promotes the in vitro development of sheep secondary isolated follicles involving the phosphatidylinositol 3-kinase pathway

Abstract

This study was conducted to evaluate the effect of addition of gallic acid as the single antioxidant to the base medium for in vitro culture of sheep secondary follicles and if the phosphatidylinositol 3-kinase (PI3K) pathway is involved in the action of gallic acid. Secondary follicles were isolated and cultured for 12 days in α-MEM supplemented with bovine serum albumin (BSA), insulin, glutamine, hypoxanthine, transferrin, selenium, and ascorbic acid (control medium: α-MEM⁺) or in α-MEM supplemented with BSA, insulin, glutamine, hypoxanthine and different concentrations of gallic acid (25, 50 or 100 μM), thus replacing transferrin, selenium and ascorbic acid in the medium. Follicle morphology, glutathione (GSH), and mitochondrial activity, and meiotic resumption were evaluated. Furthermore, inhibition of PI3K pathway was performed by pretreatment with LY294002. After 12 days of culture, the follicle survival in a medium containing 100 μM gallic acid was similar (P > 0.05) to α-MEM⁺ and greater (P < 0.05) compared with other gallic acid concentrations. Antrum formation, follicle diameter, GSH, and mitochondrial activity, and meiotic resumption, however, were greater (P < 0.05) when 100 μM gallic acid was included in the α-MEM⁺ culture medium compared with the control medium. Furthermore, LY294002 inhibited (P < 0.05) follicle survival, development, and meiotic resumption stimulated by 100 μM gallic acid. In conclusion, concentration of 100 μM of gallic acid can be a substitute for transferrin, selenium, and ascorbic acid in the base medium during in vitro culture of sheep secondary follicles, inducing follicle development likely through the PI3K pathway.

Keywords: Antioxidant; Oocyte; Ovine; PI3K; Preantral follicle.