Trophoblast miRNAs Inhibit Invasion and Angiogenesis in Fetal Growth Restriction - PROJECT SUMMARY/ABSTRACT This K08 proposal presents a comprehensive five-year research and career development plan to address critical questions pertaining to impaired placentation associated with fetal growth restriction (FGR). As a pediatric and fetal surgeon, this research project, along with its accompanying didactic efforts will equip me with unique cross- disciplinary skills to facilitate my transition to independence as a physician-scientist in placental biology. The catalyst for this proposal is based on the rigor of previous research and our preliminary data in the field. FGR is a significant problem that carries a high rate of perinatal mortality and a high frequency of lifelong poor health. Pathology of fetal growth restricted placentas demonstrates reduced extravillous trophoblast invasion and poor feto-placental angiogenesis. In addition, microRNAs, which are short non-coding RNAs, have been implicated in the pathogenesis of FGR. They can be exported from one cell to communicate with and induce effect on other cells via small transporters known as extracellular vesicles (EVs) and are critical for normal placentation. Based on these data, our central hypothesis is that anti-angiogenic miRNAs within human extravillous derived EVs inhibit extravillous trophoblast invasion and feto-placental endothelial angiogenesis through paracrine signaling, resulting in FGR. In Aim 1, we will model the metabolic stress associated with FGR by culturing extravillous trophoblasts in a nutrient restricted media. The miRNA cargo within extravillous trophoblast-released EVs will then be sequenced to determine the effect nutrient-restriction has on anti-angiogenic miRNA expression. In Aim 2, nutrient-restricted EVs derived from extravillous trophoblasts will be incubated with extravillous trophoblasts and microvascular endothelial cells (MVECs) to determine if their cargo impairs trophoblast invasion and placental feto-placental angiogenesis, respectively. Lastly, in Aim 3, we will utilize a blastocyst trophectoderm transduction model to upregulate anti-angiogenic miRNAs only within the placenta. We have previously identified miRNAs that are upregulated in mouse FGR placental tissue and will determine their effects on implantation and fetal growth in this novel in vivo model of pregnancy. The proposed experiments will impact multiple clinical disciplines including obstetrics, neonatology, and fetal care by providing mechanistic data relevant to the role of miRNAs in placental pathologic changes evident in FGR. This foundational work will provide an excellent vehicle for my maturation into an independent investigator. Experiments will be performed in an environment with an established history of successful mentorship of junior faculty to independence. With the support of this application, I will 1) advance my technical skills (RNA- sequencing, invasion assays, angiogenic assays, and blastocyst transduction) and 2) learn advanced molecular principles and biostatistics and improve my mentoring and writing skills.
