Modeling Normal and Abnormal Trophoblasts

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PROJECT SUMMARY Preeclampsia is a major source of maternal and fetal morbidity and mortality in pregnancy. Multiple studies have documented abnormalities in placental trophoblasts obtained from preeclamptic pregnancies at term, in development and function of both syncytiotrophoblast (STB) and extravillous trophoblast (EVTB). These cell types arise early in gestation, and it is unclear whether the abnormalities contribute to the pathophysiology of preeclampsia or are only a result of the preeclamptic environment. The proposed project will use three innovative models to address this gap in knowledge: (1) Induced pluripotent stem cells (iPSC), derived from control and preeclamptic pregnancies and differentiated to a mixed population of peri-implantation stage trophoblast by exposure to BMP4, and signaling inhibitors; (2) Induced trophoblast stem cells (TSC), obtained by conversion of the control and EOPE iPSC lines, and differentiated to the first trimester, villous stage of placental development, along either the STB or EVTB lineage; (3) 3D trophoblast organoids, derived from the TSC lines, and differentiated to predominantly STB or EVT. There are three specific aims: Aim 1 will test whether STB differentiation and, as a consequence, function, is defective in EOPE in the various models of first trimester placenta. Aim 1A will test the hypothesis that defects in STB assembly and function will be discernible in EOPE in the models of peri-implantation and villous STB developed in this laboratory, particularly under conditions of oxidative stress. In Aim 1B, single nucleus RNAseq will be used to distinguish whether TB types, especially the two previously identified clusters of 8TB, are differentially affected. Aim 2 will characterize EVTB differentiation and invasive properties in normal pregnancy and EOPE by using the TSC models. Aim 2A will determine whether defective TB invasion in EOPE is characteristic of first trimester-like EVTB, as it is in peri-implantation TB. Aim 28 will utilize organoid cultures to assess EVTB differentiation in 3-dimensional space, in homology to anchoring villi, and the role of cell-matrix interactions in acquisition of specific EVTB subtypes. Aim 3 will determine whether mitochondrial defects affect EVT and STB in EOPE Aim 3A will determine whether diminished ATP production reduces invasion in EOPE under stressful conditions. Aim 3B will test whether EOPE TBs are more susceptible to stress-induced production of reactive oxygen species. Accomplishment of these aims will uncover the mechanisms behind impaired STB and EVTB differentiation and function in patient-derived culture models of first trimester placental development.