Estudio de la interacción de factores ambientales y genéticos durante el desarrollo embrionario de ratón

Programa de Doctorado en Biotecnología y Tecnología QuímicaDuring embryogenesis the developing embryo is not isolated from its environment, this being a fundamental part of organism¿s life cycle and plays an important role in determining the phenotype. The embryogenesis is tightly regulated by different signalling pathways too. Any variation of the environmental or genetic factors can alter the development and leads to embryonic malformations, which can be enhanced if combined. In this work we have focused on maternal diabetes as an environmental risk factor. Congenital malformations in diabetic pregnancy are still a major health concern despite the improvements in glycaemic control strategies. Embryopathies produced as a consequence of maternal diabetes have been studied intensely in both experimental and clinical scenarios. Despite the numerous studies made in diabetic mice, is necessary to stablish the best strain as well as the most appropriate diabetes induction protocol. In our study we tested different protocols in females from different strains, and we stablished the most effective protocol for each one. As it is well known, abnormalities in maternal folate and diabetes during pregnancy have both been shown to induce neural tube defects (NTDs) in humans and animal models. However, the possible interaction between these two factors in the development of NTDs remains unclear. Folr1 is a membrane receptor that mediates the high affinity internalization and delivery of folate to the cytoplasm of the cell. Functional defect in this gene produce severe abnormalities in embryos, including cranial NTDs. It has been demonstrated that Folr1 embryonic mRNA was decreased under maternal diabetes. The metiltetrahidrofolate reductase (MTHFR) is a key enzyme involved in folate metabolism that converts 5.10-methylenetetrahydrofolate to 5-methyltetrahydrofolate. Mutation in the 677C¿T in the Mthfr gene is the most common genetic cause of hyperhomocysteinemia and is associated with NTDs and pregnancy complications. We analysed the combination of maternal diabetes and mutations of Folr1 and of Mthfr using mouse embryo culture and mouse models, and we observed that hyperglycaemia in combination with the alteration of folic acid metabolism seems to increase the incidence of congenital malformations in the embryos with majority of them being NTDs. Moreover, hyperglycaemia has been also shown to downregulate the expression of elements in the non-canonical Wnt-PCP pathway, such as the Dishevelled-associated activator of morphogenesis 1 (Daam1) and Vangl2. Daam1 is a formin that is essential for actin polymerization and for cytoskeletal reorganization, and it is expressed strongly in certain organs during mouse development, including the eye, neural tube and heart. Daam1gt/gt and Daam1gt/+ embryos develop ocular defects (anophthalmia or micropthalmia) that are similar to those detected as a result of hyperglycaemia. Indeed, studying the effects of maternal diabetes on the Wnt-PCP pathway demonstrated that there was strong association with the Daam1 genotype, whereby the embryopathies observed in Daam1gt/+ mutant embryos of diabetic dams was more severe. There was evidence that embryonic exposure to glucose in vitro diminishes the expression of genes in the Wnt-PCP pathway, leading to altered cytoskeletal organization, cell shape and cell polarity in the optic vesicle. Hence, the Wnt-PCP pathway appears to influence cell morphology and cell polarity, events that drive the cellular movements required for optic vesicle formation and that, in turn, are required to maintain the fate determination. We also observed a number of Daam1+/gt from diabetic mothers that presented NTDs. These finding shows, although without a statistical significance, a possible interaction between the diabetes and the Wnt-PCP signalling pathway in the development of NTDs. However, to confirm this hypothesis it would be necessary to make more studies.Universidad Pablo de Olavide. Departamento de Fisiología, Anatomía y Biología CelularPostprin