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Abstract

The placenta is a transient, multifunctional organ that forms during pregnancy to support fetal growth and survival. The placenta facilitates nutrient and gas exchange, protects the fetus from pathogens, and secretes hormones that adapt maternal physiology. These diverse functions are executed by specialized epithelial cells called trophoblasts, which dynamically change the placental and uterine architecture to meet the evolving needs of the developing fetus. Progenitor cells of the placenta, called cytotrophoblasts (CTB), are capable of long-term self-renewal and differentiation into extravillous trophoblasts (EVT) and syncytiotrophoblasts (SCT) that carry out the essential functions of the placenta. Despite their importance, the mechanisms maintaining the balance of CTB progenitor self-renewal, differentiation, and senescence remain poorly understood. Progenitor identity and function are in part shaped by the extracellular matrix (ECM) that provides structural support and a rich source of growth factors. The turnover of ECM by metalloproteases liberates these growth factors, thereby modulating the processes that maintain progenitor populations. Consistent with this, single-cell transcriptomic profiling of human first-trimester placentas identified that 11 metalloproteases are expressed by CTB, suggesting an important role for metalloproteases in maintaining the CTB population. Thus, I hypothesize that metzincin metalloproteases are essential for maintaining the human trophoblast progenitor cell population. This thesis investigates the importance of metalloprotease activity in human trophoblast stem cell (hTSC) organoid growth using the broad-spectrum inhibitor batimastat. Metalloprotease inhibition significantly reduced organoid size without affecting organoid formation frequency, proliferation, or apoptosis. Bulk RNA sequencing revealed 1,989 differentially expressed genes in response to batimastat, with transcriptomic signatures suggesting a loss of CTB progenitor potential and a shift toward a more differentiated state. Overall, this work demonstrates that metzincin metalloproteases may modulate early trophoblast progenitor differentiation. This project advances the understanding of molecular processes governing early human placental development.