Expressão gênica e viabilidade de folículos pré-antrais submetidos à vitrificação do córtex ovariano de Sapajus apella (macacas-prego)

Abstract

Vitrification is a biotech that has been increasingly showing promise in several species, among them domestic ruminants (such as sheep, goats and cows) and in nonhuman primates (NHP) (as cynomologus and rhesus), and consists in reducing ultra-rapid temperature by the presence of high concentrations of cryoprotective agents (PCA's) in liquid nitrogen. Thus the aim of this study was to develop a methodology for cryopreservation by vitrification of preantral follicles (PF's) in Sapajus apella. For this purpose, we used females (n = 9) adult Sapajus apella squad belonging to the National Primate Center (CENP). Samples were taken from the ovarian cortex by laparotomy so as not to destabilize the animals reproducibly. The fragments exposed to 8 different treatments: Ethylene glycol (EG) + 40% Sucrose (Sac) dissolved in 0.5 M TCM-199, added to Selenium (2,5, 5 or 10 ng / ml) or Trolox (25, 50 or 100 mM). Following exposure to cryoprotective agents was analyzed follicular viability before and after vitrification, from the follicular morphology, expression of Hsp70 genes, Erp29, Erp60 SOD1 and through the analysis by Polymerase Chain Reaction (PCR), in addition to performing measurement of oxidative stress thru the TEAC (English Total Equivalent Antioxidant Activity). The analyzes showed that vitrification allowed the maintenance of follicular viability by previous exposure to concentrations of ACP's alevadas, especially when supplemented with 50 mM trolox (which resulted in high follicular survival rates) and increased expression of the antioxidant enzyme SOD1. While in the absence of it was observed an increase in rates of follicles degenerate and vacuolated, and reduced expression of the antioxidant enzyme SOD1 and increased expression of chaperone Erp29.