Efeito do fator de necrose tumoral alfa (TNF-α) e da interleucina-1 beta (IL-1β) sobre o desenvolvimento in vitro de folículos secundários bovinos isolados

Abstract

Ovarian follicular growth is regulated by various substances, such as hormones, growth factors and cytokines. Tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are proinflammatory cytokines that have several sites of synthesis in the ovary. In this context, the study of the effect of these cytokines during the in vitro growth of preantral follicles is of great importance for understanding the mechanisms that control follicular development in vitro. The objective of the present study was to determine the effect of TNF-α and IL-1β on development and survival of bovine secondary follicles after 18 days of culture in vitro. Secondary follicles (~0.2mm) were isolated from ovarian cortex of cows and individually cultured. The secondary follicles were randomly distributed in the following treatments: (I) TCM-199+ alone (culture control), or supplemented with (II) 10 ng/mL IL-1β, (III) 10 ng/mL TNF-α and (IV) both TNF- α and IL-1β. The follicles were cultured at 38.5°C, with 5% CO2 in air, for 18 days. Thereafter, the effect of these treatments on growth, follicular survival, antrum formation, viability, ultrastructure and mRNA levels for GDF9, C-MOS, H1foo and Cyclin B1 were then evaluated. At the end of 18 days of in vitro culture, TNF-α increased follicular diameter and the rate of antrum formation. On the other hand, IL-1β and both IL-1β and TNF-α did not influence follicle growth. All treatments maintained viability rates after culture between 74.32 and 92.00%. Ultrastructural analysis of showed that follicles cultured in the presence of TNF-α had oocyte citoplasm and granulosa cells well preserved. The presence of TNF-α, IL-1β or both did not influence the expression o mRNAs for GDF9, C-MOS, H1foo and Cyclin B1 after 18 days of culture. In conclusion, TNF-α promotes follicular growth and maintains the ultrastructure of isolated bovine secondary follicles cultured in vitro.