Shear Stress Markedly Alters the Proteomic Response to Hypoxia in Human Pulmonary Endothelial Cells.

Blood flow produces shear stress that homeostatically regulates the phenotype of pulmonary endothelial cells exerting anti-inflammatory and anti-thrombotic actions and maintaining normal barrier function. Hypoxia due to diseases, such as COPD, causes vasoconstriction, increased vascular resistance and pulmonary hypertension. Hypoxia-induced changes in endothelial function play a central role in the development of pulmonary hypertension. However, the interactive effects of hypoxia and shear stress on the pulmonary endothelial phenotype have not been studied. Human pulmonary microvascular endothelial cells were cultured in normoxia or hypoxia while subjected to physiological shear stress or in static conditions. Unbiased proteomics was used to identify hypoxia-induced changes in protein expression. Using publicly available scRNAseq datasets, differences in gene expression between the alveolar endothelial cells from COPD and healthy lungs were identified. 60 proteins were identified whose expression changed in response to hypoxia in conditions of physiological shear stress but not in static conditions. These included proteins that are crucial for endothelial homeostasis e.g. JAM-A, ERG or implicated in pulmonary hypertension e.g. thrombospondin-1. 55 of these 60 have not been previously implicated in the development of hypoxic lung diseases. mRNA for five of the 60 (ERG, MCRIP1, EIF4A2, HSP90AA1 and DNAJA1) showed similar changes in the alveolar endothelial cells of COPD compared to healthy lungs in females but not in males. These data show that the proteomic responses of the pulmonary microvascular endothelium to hypoxia are significantly altered by shear stress and suggest that these shear-hypoxia interactions are important in the development of hypoxic pulmonary vascular disease.