Reproduction Abstracts

Introduction: Differentiation of the embryoblast and trophoblast cell lineages is an ontogenetic milestone in preimplantation embryo development. Both cell lineages are supplied with nutrients from uterine secretions, reflecting an ultrafiltrate of maternal plasma. We have recently shown that an insulin-dependent diabetes mellitus leads to maternal hyperlipidaemia and to a strong increase in intracellular lipids in trophoblast and, especially, embryoblast cells in rabbit blastocysts (Schindler et al. 2014, Endocrinology 10.1210/en.2013-1760).

Our aim was to determine mechanisms of lipid accumulation in the embryoblast and trophoblast in rabbit blastocysts derived from a diabetic (in vivo) and hyperlipidaemic (in vitro) environment.

Methods: For in vivo analysis blastocysts from diabetic rabbits were flushed from the uterus six days after mating. To evaluate the effect of an increased environmental lipid level in detail, blastocysts from healthy rabbits were cultured in vitro with a specified lipid mixture (Gibco, Chemically Defined Lipid Concentrate). Intracellular lipid accumulation was visualised by Oil Red-O staining. Marker molecules involved in lipid metabolism were analysed by qPCR, Western Blot and immunohistochemistry, separately in the embryoblast and trophoblast.

Results and Discussion: In embryoblast cells intracellular lipid droplets and fatty acid uptake (FATP4) and binding (FABP4), as well as PPARg expression were increased after lipid stimulation. In trophoblast cells intracellular lipid droplets were also increased but accompanied by a down-regulation of fatty acid oxidation (CPT1 and PPARa) and fatty acid synthesis (FASN), indicating that two different signalling pathways were activated in blastocysts. Consistently with these results, embryos from diabetic rabbits revealed also an increased expression of FATP4, FABP4 and PPARg in embryoblast cells and down-regulation of CPT1 and FASN in trophoblast cells.

Our study shows that embryoblast and trophoblast handle hyperlipidaemic conditions in different ways, leading in the result to similar phenotypes. This observation underlies the importance of more differentiated analyses in early embryos.