Hypoxic Pulmonary Endothelial Cells Release Epidermal Growth Factor which Results in Vascular Smooth Muscle Cell Arginase 2 Expression and Proliferation
The hallmark of pulmonary hypertension (PH) is vascular remodeling leading to alterations in hemodynamics and vasoreactivity. Endothelial cells (EC) are central in the response of a vessel to stimuli associated with PH. EC can proliferate contributing to vascular remodeling, but likely a more important role of EC is the release of factors that induce proliferation in the underlying smooth muscle cells (SMC) leading to intimal thickening. Hypoxia is associated with the development of PH in certain human diseases. We have previously shown that human pulmonary microvascular EC (hPMVEC) respond to hypoxia with proliferation that is dependent on arginase 2 up-regulation. This arginase 2 up-regulation is mediated by the release of epidermal growth factor (EGF) and resultant activation of EGF receptors (EGFR) on hPMVEC. We hypothesized that the release of EGF by hPMVEC would also result in proliferation of human pulmonary arterial SMC (hPASMC) via EGFR-mediated arginase 2 up-regulation. To test this hypothesis, we used conditioned media (CM) from hPMVEC grown either in normoxia or hypoxia to study the role of EGF released by hPMVEC on the regulation of arginase 2 and proliferation in hPASMC. When hPASMC were grown with CM from hypoxic hPMVEC there was a 9-fold induction of arginase 2 protein levels and more than a doubling of viable hPASMC numbers compared to hPASMC grown with CM from normoxic hPMVEC. In another experiment, hPVMEC were transfected with a scramble siRNA (scramble) or an siRNA against EGF (siEGF) and then the hPMVEC placed in hypoxia to generate conditioned media. In the hPASMC treated with the CM from siEGF-treated hypoxic hPMVEC there were lower levels of arginase 2 protein than in the hPASMC grown in CM from hPMVEC grown in normoxia; and substantially less arginase 2 than in hPASMC treated with CM from hypoxic hPMVEC treated with scramble. Furthermore, hPASMC treated with hypoxic CM from siEGF-treated hPMVEC had substantially lower viable cell numbers than in hPASMC treated with hypoxic CM from scramble-treated hPMVEC. When hPASMC were treated with hypoxic CM from hPMVEC the addition of an EGF neutralizing antibody to the CM prevented the increase in viable cell numbers seen with hypoxic CM. When hPASMC were transfected with siRNA against EGFR and then treated with hypoxic CM from hPMVEC the hypoxia-induced increase in arginase 2 protein levels and viable cell numbers were prevented. These data demonstrate that hPMVEC release EGF, which can activate EGFR on hPASMC to induce arginase 2 protein expression and increase viable cell numbers. We speculate that EGF neutralizing antibodies or EGFR blocking antibodies are potential therapeutics to prevent and/or attenuate the vascular remodeling seen in PH associated with hypoxia.
This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.