Project Summary A fundamental question relevant to clinical IVF is whether and how the peri-conceptional milieu affects embryo implantation and placentation and how clinical manipulations of this environment influence perinatal outcomes. Changes in trophoblast differentiation and function, perturbations of placental vascular development and/or sub-optimal endometrial regeneration during clinical or laboratory manipulations such as trophectoderm biopsy could all contribute to abnormal implantation and placentation and lead to an abnormal fetal growth phenotype. We hypothesize that epigenetic changes in specific placental cellular compartments (embryonic or extraembryonic) and/or the endometrium contribute to the significant adverse outcomes associated with IVF and, specifically, following fresh (hyperstimulated) vs. frozen/thawed embryo transfer or natural conception or following trophectoderm biosy. We suggest that these epigenetic alterations lead to abnormal gene expression at critical times during development, implantation and placentation and result in abnormal growth and other complications. In Specific Aim 1A we will identify site- and gene-specific epigenetic differences in placentas, in isolated trophoblasts, placental vasculature and endothelial cells from pregnancies following fresh/hyperstimulated vs. frozen/thawed vs. control pregnancies. In Specific Aim 1C, we will begin exploring whether there is an endometrial contribution to the observed peri-conceptional milieu growth-related differences as well. In Specific Aim 2 we will identify specific epigenetic differences in placentas and in isolated trophoblast cells, placental vasculature and endothelial cells in pregnancies following trophectoderm biopsy. In this "clinical" project, we focus on identifying the epigenetic signature(s) related to abnormal placental function and an abnormal birth weight phenotype. In Specific Aim 3 we will determine whether the epigenetic signature determined at birth persists into childhood and may be associated with long term health consequences such as obesity. This initiative continues to be discovery-driven, has and will continue to inform mechanistic studies for our mouse model (Project 2) and should contribute to our understanding of optimal human embryo development, implantation and placentation.