Targeting and Modeling the Biomechanical Environment of the Pulmonary Vasculature to Study Pulmonary Arterial Hypertension

Abstract

Endothelial dysfunction is a known consequence of Bone Morphogenetic Protein Receptor Type II (BMPR2) mutations in pulmonary arterial hypertension (PAH). Initial studies aimed to prevent heritable PAH by inhibiting the increased phosphorylation and downstream activity of a key deregulated protein, SRC. After inhibiting SRC failed to prevent PAH, we were motivated to engineer a new, more biomimetic model to study pulmonary vascular diseases. Standard 2D cell culture models fail to mimic the mechanical environment seen in the pulmonary vasculature. Hydrogels have emerged as promising platforms for 3D disease modeling due to their tunable physical and biochemical properties. In order to recreate the mechanical parameters seen in PAH, we created a novel 3D hydrogel-based artificial arteriole platform that reproduced the pulsatile flow rates and pressures seen in the lung vasculature. We probed Bmpr2R899X and wild type (WT) endothelial cells to better understand how the addition of oscillatory flow and physiological pressure altered gene expression, influenced cell morphology, and recapitulated endothelial permeabilities in our in vitro model. The addition of oscillatory flow and pressure resulted in several gene expression changes in both WT and Bmpr2R899X cells. However, for many pathways with relevance to PAH etiology, Bmpr2R899X cells responded differently when compared to the WT cells. Bmpr2R899X cells did not elongate in the direction of flow, and instead remained static in morphology despite mechanical stimuli. The increased permeability seen in PAH was successfully reproduced in our model. Using clinical data, we aimed to correlate findings in our model with single nucleotide polymorphisms (SNP) in metabolic pathways in PAH patients. A SNP in PPARA was found to be associated with increased right ventricular function in PAH patients. A partnered transcription co-activator of PPARA, CITED2, was found to be significantly up-regulated in Bmpr2R899 cells when exposed to pressure and flow in our model, underscoring the model’s potential feasibility to aid clinical PAH research.