Influência da n-acetilcisteína (nac) durante o cultivo in vitro de folículos antrais iniciais em bovinos

Abstract

This study investigates the effects of different concentrations of N-acetylcysteine (NAC) on follicular growth and morphology, as well as viability, reactive oxygen species (ROS) levels and meiotic progression of oocytes from bovine early antral follicles cultured in vitro. To this end, isolated early antral follicles (~500 μm) were cultured in TCM-199+ alone or supplemented with 1.0, 5.0 or 25.0 mM NAC at 38.5°C with 5% CO2 per 8 days. Follicle diameters were evaluated on days 0, 4 and 8 of culture. At the end of cultivation, ROS levels, chromatin configuration and viability (calcein-AM and ethidium homodimer-1 staining) were investigated in isolated cumulus oocyte complexes (COCs). Comparisons of follicular diameters between treatments were performed using the Tukey test. Data on percentages of morphologically normal follicles, growth rates, and chromatin configuration were compared using Fisher's test (P < 0.05). The results showed an increase in follicular diameters after culture in all treatments compared to the beginning of culture, except for follicles cultured with 25.0 mM NAC. Fluorescence microscopy showed that oocytes cultured in all treatments stained positively with calcein-AM, and that 5.0 mM NAC reduced the fluorescence for ethidium homodimer-1. The intracellular levels of ROS in oocytes from follicles cultured with 1.0mM NAC showed a significant reduction in relation to the other treatments. The presence of NAC in the culture medium did not influence the rates of oocytes at the germinal vesicle stage. In conclusion, NAC at concentrations of 1.0 reduces ROS levels and 5.0mM reduces staining for ethidium homodimer-1, but NAC 25.0mM reduces follicular growth and the percentages of continuously growing follicles.