Mechanisms of lipid accumulation in oocytes under metabolic altered environments: in vitro maturation and obesity

Abstract

Developing oocytes stock pile lipids within the ooplasm and use it as energy source. This physiological event can be detrimental when over occur, leading to excessive intraoocytic lipid accumulation. Results obtained in a previous research grant (2014/21034-3) shown, in bovine, that the in vitro maturation (IVM) of oocytes, when compared to in vivo maturation, lead to increased intraoocytic lipid accumulation, associated with a massive deregulation of cumulus cell metabolic pathways. Also, in this previous grant we demonstrated, for the first time, a presumptive mechanism of fatty acid transport from cumulus cell to the oocyte via Fatty Acid Binding Protein 3 (FABP3) and transzonal projections (TZPs). We also demonstrated that such transport is exacerbated during IVM. These previous results shown that IVM generate a metabolic altered environment that reflects in abnormal cumulus cells metabolism and lipid accumulation in oocytes. While IVM represents an in vitro model to study oocytes' lipid accumulation, the obesity could be a model to study such accumulation in a metabolic altered environment in vivo. Preliminary results attached to this submitted grant, demonstrate, in murine, that obesity also lead to excessive lipid accumulation in oocytes. Taken these previous results this research proposal aim to investigate causes and mechanisms of oocyte lipid accumulation in metabolic altered environment in vitro (IVM) and in vivo (obesity), in bovines and murines, respectively. Despite demonstrated that the IVM induce oocyte lipid accumulation, it is neither unknown the effects of IVM on cumulus cell regulation of metabolic and signaling pathways, nor how specific components of IVM media interfere on the accumulation process. Similarly, it is unknown the mechanisms by which oocytes from obese females excessively accumulate lipids. To deeper our understanding on such points we designed three main studies. In Study 1 the transcriptome of cumulus cell from immature, in vivo-matured or in vitro-matured cumulus-oocyte complexes (COCs) will be generated in order to detect alteration on metabolic and signaling pathways associated with the oocyte maturation. The results of this experiment will allow to generated novel metabolic pathways to be investigated. In Study 2 experiments will be done to determine the effects of extracellular vesicles (EVs) from fetal calf serum and of FSH supplementation levels in IVM media on the oocyte's lipid accumulation and presumptive fatty acid transport via FABP3 and TZPs. To these aims immature oocytes, oocytes in vitro-matured for 9 hours, and oocytes in vitro-matured for 24 hours (in MII) will be analyzed in terms of lipid content and FAPB3 levels. The corresponding cumulus cells from these COCs will be used to quantify PLIN2 and mRNA of FABP3 and PLIN2. In Study 3 a murine model of obesity will be used to investigate the mechanisms of lipid accumulation within the COC. In this study, the first experiment will aim to determine and compare lipid stocks in cumulus and oocytes during the oocyte maturation in lean and obese mice. In the second group of experiments four potential mechanisms of lipid accumulation will be investigated in pre-ovulatory follicles and ovulated oocytes of lean and obese mice, as follows: (1) increased synthesis of lipids within the COC; (2) reduced beta-oxidation; (3) fatty acids transport from cumulus to the oocyte via FABPs and TZPs; and (4) impaired lipophagy within the COC. The results that will be obtained from this research proposal will allow to comprehend the mechanism associated with intraoocytic lipid accumulation in metabolic altered conditions, and may lead to the development of strategies to attenuated the negative effects of oocytes' lipid accumulation on fertility. (AU)