Investigate the effects of rare damaging mutations in TGFB latent complex proteins found in preeclamptic placentas

Project Summary/Abstract Preeclampsia (PE) is a severe pregnancy complication that affects 8% of pregnancies globally, contributing to over 50,000 maternal and 500,000 fetal deaths annually1. Characterized by hypertension, PE arises from poor placental development, partially due to impaired extravillous trophoblast (EVT) differentiation and invasion. Transforming growth factor-beta (TGFβ) signaling is essential for trophoblast differentiation and function, regulating cell migration and extracellular matrix (ECM) remodeling4. Dysregulated TGFβ signaling, including elevated TGFβ levels in preeclamptic placentas, disrupts these processes, leading to poor placental function and adverse outcomes1. Our research focuses on rare damaging mutations in key TGFβ latent complex proteins, including Fibronectin 1 (FN1) and Latent Transforming Growth Factor Beta Binding Protein 1 (LTBP1), that we identified in placentas and umbilical cord blood mesenchymal stem cells (MSCs) from PE-affected pregnancies. We hypothesize that these mutations impair TGFβ latency, enhancing active TGFβ signaling, and disrupting ECM integrity which could ultimately affect EVT migration, invasion, and differentiation. Using patient-derived cells and CRISPR-engineered models, we aim to investigate the functional consequences of these mutations on TGFβ production, signaling, and downstream molecular pathways. In Aim 1, we will study the effects of TGFβ activation and signaling in UC-MSC and iPSC-derived trophoblast stem cells (TSCs) from PE-affected and healthy placentas. We will analyze TGFβ levels, TGFβ down stream signaling, EVT differentiation, and perform functional assays to measure migration and invasion. Bulk RNA sequencing will uncover dysregulated pathways contributing to PE. In Aim 2, we will use CRISPR/Cas9 technology to introduce specific mutations into iPSCs, differentiate them into TSCs and EVTs, and test their effects on ECM integrity, TGFβ latency, and EVT function. RNA sequencing will further identify mutation-specific molecular disruptions. The results have the potential to inform new diagnostic tools, therapeutic targets, and preventive strategies for PE, ultimately improving maternal and fetal health outcomes. Additionally, this research may extend to other placental disorders involving TGFβ signaling and ECM dysfunction, contributing to a broader understanding of placental development and disease.