Culture of preantral ovarian follicles of Bos taurus indicus with alpha-lipoic acid.

Abstract

The aim of this study was to evaluate follicular development, morphological integrity, and antioxidant potential of preantral ovarian follicles from Bos taurus indicus females grown in vitro with alpha-lipoic acid. Ovaries (n = 24) of Bos taurus indicus (n = 12) females were collected during slaughter and fragmented. A randomly obtained fragment from each pair of ovaries was fixed in Bouin (non-cultivated control; D0). These fragments were intended for classical histology (morphology and evaluation of follicular growth), and a fragment from each pair of ovaries was frozen at -80°C (non-cultivated control; D0), and assigned for analysis of oxidative stress [thiobarbituric acid reactive substances (TBARS), nitroblue tetrazolium (NBT), and ferric reducing antioxidant power (FRAP)]. The remaining fragments were cultured in vitro for 6 (D6) or 12 (D12) days, containing only minimum essential medium (MEM) or MEM supplemented with alpha-lipoic acid (50, 100, or 250 ng/ml), on an extracellular matrix of agarose gel, in an oven at 38.5ºC. Every 2 days, 100% of the culture medium was replaced. Supplementation with 100 ng/ml was effective for maintaining follicular integrity after 6 days of culture (primordial: 51.28%; development: 36.88%; P < 0.0001). There was no difference (P > 0.05) between treatments compared with the non-cultivated control treatment (D0), using the NBT and TBARS assays. Therefore, supplementation of the in vitro culture medium of bovine preantral ovarian follicles with a concentration of 100 ng/ml of alpha-lipoic acid at 6 days of culture was effective for maintaining follicular integrity and, after 6 days, maintaining stable levels of reactive oxygen species.