Hypoxia Activates Extracellular Signal‐Regulated Kinase (ERK) via EGFR Activation in Human Pulmonary Microvascular Endothelial Cells

Abstract

We have previously shown that hypoxia‐induced proliferation of human pulmonary microvascular endothelial cells (hPMVEC) depends on epidermal growth factor receptor (EGFR)‐mediated arginase II induction. We hypothesized that hypoxia‐induced EGFR activation leads to phosphorylation of extracellular signal‐regulated kinase (ERK) which then leads to increased arginase II expression in hPMVEC. Cells were incubated in hypoxia (5% CO2, 1% O2) for 0, 2, or 24 hours and western blotting was performed for phospho‐ERK and total ERK. We found that hypoxia resulted in greater levels of phospho‐ERK protein with little effect on total ERK protein levels at both 2 and 24 hours. hPMVECs were transfected with an siRNA against EGFR or scramble for 24 hours, allowed to recover for 24 hours, and then incubated in hypoxia or normoxia (5% CO2, 20% O2) for 24 hours. Cells incubated in hypoxia and treated with EGFR siRNA had lower phospho‐ERK levels than hypoxic cells treated with scramble. Furthermore, hypoxic hPMVECs treated with the EGFR siRNA had lower levels of arginase II protein than did hypoxic cells treated with scramble. In the normoxia exposed cells there were lower levels of arginase II protein than in hypoxia exposed cells and there was little effect of the EGFR siRNA on arginase II protein levels. Our findings suggest that in hPMVEC hypoxia activates EGFR which in turn phosphorylates ERK eventually leading to increased levels of arginase II. We speculate that ERK may be a potential therapeutic target to prevent or reverse the vascular remodeling seen in pulmonary hypertension associated with hypoxia.