EFFECTS OF IN VITRO PREMATURATION OF OOCYTES WITH cAMP MODULATORS AND THE USE OF EPIGENETIC MODIFIERS ON OOCYTE COMPETENCE AND EMBRYONIC DEVELOPMENT IN BOVINE

Abstract

The biotechnologies applied to animal reproduction have caused great impacts, especially in recent decades. However, even the most complex biotechnologies such as transgenesis and nuclear transfer, for example, are dependent on the technique of in vitro production of embryos (IVPE). The oocyte maturation is a key procedure in IVPE, however, the efficiency of in vitro maturation (IVM) is fundamentally limited by the oocyte quality, i.e. by the oocyte's intrinsic developmental potential. The isolation of immature oocytes from the follicular environment may interrupt the transfer of nutrients and metabolic support provided by the cumulus cells, resulting in premature activation of nuclear maturation, without that the oocyte has undergone these changes, and compromising the oocyte competence and subsequent development. For this reason, maintaining the integrity of intercellular gap junction communications (GJC) during IVM is essential for the completion of meiotic and cytoplasmic maturation. A recent promising approach is the use of enzyme adenylate cyclase (AC) activators and phosphodiesterases (PDEs) inhibitors aiming the maintaining of high cAMP levels during the so-called prematuration. Moreover, among the events that must occur in the nucleus and cytoplasm in order to have full oocyte meiotic competence, the histone acetylation is one of the most important. Considering also reports that trichostatin, an histone deacetylases (HDACs) inhibitor that causes global histone hyperacetylation, leads to a delay of meiotic progression and increases the levels of oocyte competence, this study intends to investigate and optimize the events involved in the oocyte competence acquisition and reflected on subsequent embryonic development after induction of prematuration with cAMP modulators and epigenetic changes through the use of HDACs inhibitors. For this, five experiments were designed, where the meiotic progression and redistribution of cell organelles will be assessed (Exp. I), the status of communication between the oocyte and the cumulus cells (Exp. II), and the relative expression of genes related to competence (Exp. IV) in oocytes obtained in vivo and from slaughterhouse ovaries, immediately after the follicular aspiration (immature) and after prematuration and/or IVM in medium containing cAMP modulators, TSA or a combination of both. We will also study the histone H3 (K9) acetylation levels in treated oocytes and embryos generated by the different treatments (Exp. III), and in the last experiment (Exp. V) the embryo development and quality will be assessed, and there even the pregnancy and birth rates.