Pulmonary

Pulmonary arterial hypertension (PAH) is a progressive disease defined by increased pulmonary vascular resistance, resulting in right ventricular hypertrophy and untimely death from right heart failure. Heterozygous mutations in BMPR2 are associated with approximately 70% of hereditary PAH cases and 20% of idiopathic PAH cases, respectively. Although patients carrying a BMPR2 mutation are more likely to develop PAH than non-carriers, only 20% will develop while the majority will remain asymptomatic. PAH is characterized by excessive vascular remodeling involving endothelial cell (EC) dysfunction and impaired apoptosis and uncontrolled proliferation of vascular smooth muscle cells (VSMCs). Paracrine signaling between ECs and VSMCs, together with mechanical cues from blood flow, govern vascular remodeling, permeability, and homeostasis. The crosstalk between mechanical stimuli and cell-cell signaling is poorly understood to date. Many therapeutic strategies developed using conventional two-dimensional (2D) systems often fail due to poor recapitulation of in vivo conditions, in terms of cell-cell interactions, signaling, and tissue organization. Animal models do not reflect human disease conditions owing to species differences and distinct pathogenic mechanisms. Despite significant advancements in the use of bioengineered models to study EC-VSMC interactions, none of the previous studies have accurately replicated EC-VSMC interactions akin to that seen in the blood vessel wall. Therefore, new alternative models are needed that significantly overcome the existing in vitro models' limitations and have more translational potential than animal models. This project is organized with two specific aims. Aim 1: We plan to generate ECs, and lineage-specific (neuroectoderm, lateral plate mesoderm, and paraxial mesoderm) VSMCs from induced pluripotent stem cells (iPSCs) carrying BMPR2 mutation and unaffected mutant carrier an and develop 3D artery tissue-on-chip (ATOC), mimicking EC-VSMC niche to study BMPR2 mutation-mediated vascular remodeling under variable and tunable hemodynamic conditions.

The microfluidic device includes two stacked microchannels separated by a porous membrane, enabling paracrine signaling between cells in an in vitro co-culture setting under flow conditions. The model will allow for simultaneous mimicking of EC dysfunction and hyperplasia of VSMCs. A panel of drugs known to modulate the BMP pathway will be used to assess the phenotype changes. Aim 2: We hypothesize that VSMCs-secreted biochemical cues are expected to cause hyperpermeability in the EC layer under flow conditions. Studies are planned to investigate whether flow-derived shear-stress alone or the synergistic effect of EC-VSMCs paracrine signaling and shear-stress causes EC hyperpermeability using RNAi technology. Further, RNA sequencing will be utilized to evaluate changes in signaling pathways related to vascular remodeling and discover new therapeutic targets/signaling pathways. The research outcomes are expected to

1. create an in vitro microfluidics-based tissue model of the artery under hemodynamic conditions to advance understanding of how shear stress within artery vasculature may alter signaling pathways related to disease development and progression;
2. provide mechanistic insights into sremportal the critical role of mechano-biochemical signaling cues in EC-VSMC crosstalk, on vascular remodeling;
3. provide a cost-effective yet efficient platform for characterization of other mutation-linked PAH.