The effects of apelin on IGF1/FSH-induced steroidogenesis, proliferation, Bax expression, and total antioxidant capacity in granulosa cells of buffalo ovarian follicles

Abstract

Abstract Apelin (APLN) was believed to be an adipokine secreted from adipose tissue. However, studies demonstrate that it is a pleiotropic peptide and has several effects on the female reproductive system. In this study, the effect of different doses of IGF1 and FSH in the presence of APLN-13 on the production of estradiol and progesterone was evaluated in the follicular granulosa cells of buffalo ovaries, in addition, the effects of different doses of APLN isoforms (APLN-13 and APLN-17) on proliferation, the expression of Bax protein and total antioxidant capacity activity of the same cells were investigated. Granulosa cells of buffalo ovaries were cultured in the presence of different doses of IGF1 and FSH with or without APLN-13 (10− 9M) to evaluate its effect on the secretion of estradiol and progesterone that tested by ELISA assay. WST-1 method was used to survey the effect of APLN on granulosa cell proliferation and cytotoxicity. In addition, the antioxidant capacity of the cells in the presence of APLN was assessed using FRAP method. mRNA and Bax protein levels were measured in granulosa cells treated with APLN using real-time PCR and western blot techniques. APLN-13 stimulated the effect of IGF1 on the production of estradiol and progesterone, and the progesterone production levels were affected by APLN-13 dose-dependently. However, it did not significantly stimulate the effect of FSH on the secretion of estradiol or progesterone. APLN-13 (all doses) and − 17 (10− 8 and 10− 9 M) improved the proliferation of granulosa cells. Moreover, preincubation of the cells for an hour by APLN receptor antagonist (ML221, 10 µM) did not significantly affect the proliferation of cells. Neither APLN-13 nor APLN-17 were not cytotoxic for the cells compared to the control treatment. APLN-13 at the doses of 10− 6 and 10− 8 M substantially up and down-regulated Bax protein expression; however, such effects were not observed when the cells were preincubated with ML221. In addition, APLN-17 did not influence the expression amount of Bax. Furthermore, both APLN-13 and − 17 improved the total antioxidant capacity of the ovarian granulosa cells, but such effects were not seen when the cells were preincubated with ML221. These findings indicate that APLN enhanced the IGF1 induced steroidogenesis and improved the cell proliferation and antioxidant capacity of follicular granulosa cells of buffalo ovaries; however, its effect on Bax expression was divergent.