Biomedical Sciences ETDs
Publication Date
Summer 8-1-2023
Abstract
Pulmonary arterial constriction and remodeling resulting from chronic hypoxia (CH) lead to increased pulmonary vascular resistance and resultant pulmonary hypertension in patients with chronic obstructive pulmonary diseases, sleep apnea, or in residents at high altitude. Recent studies from our laboratory have demonstrated that CH augments vasoconstrictor reactivity to a variety of stimuli, including pulmonary arterial smooth muscle cell (PASMC) stretch, membrane depolarization, and stimulation of G protein-coupled receptors. This effect of CH is mediated by a switch in PASMC signaling from Ca2+-dependent mechanisms towards a novel epidermal growth factor receptor (EGFR)-NADPH oxidase 2 (NOX2)-Rho kinase Ca2+ sensitization pathway that occurs in lipid rafts. We have further demonstrated a central role for decreased PASMC membrane cholesterol in this response. Since CH has an effect to both increase reactive vi oxygen species (ROS) in PASMC and decrease membrane cholesterol, we hypothesized that elevated ROS production during CH diminishes PASMC membrane cholesterol. In agreement with this hypothesis, the CH-dependent reduction in PASMC membrane cholesterol was acutely reversed following treatment with the ROS scavengers, tiron and EUK. Furthermore, PASMC treatment with SOTS-1 (superoxide donor), diminished membrane cholesterol in control and cholesterol-treated cells. We further examined caveolin-1 (Cav-1) as a potential mechanism by which ROS regulates membrane cholesterol because Cav-1 is a protein required for the formation of caveolae that is capable of binding directly to cholesterol, serving as a cholesterol transporter, and can regulate cholesterol content in lipid microdomains in a ROS dependent manner. Indeed, CH and exogenous ROS have an effect to diminish Cav-1 protein expression. Since CH has an effect to diminish membrane cholesterol, a component of lipid membrane domains that can regulate both EGFR and NOX2 activity in other cell types, we additionally hypothesized that diminished membrane cholesterol following CH unmasks EGFR-signaling through disinhibition of Rac1. We first show that cholesterol supplementation significantly increased PASMC membrane cholesterol in both the control and CH groups. Additionally, treatment with either MCD (a cholesterol chelator) or epicholesterol (a cholesterol enantiomer) significantly decreased membrane cholesterol in arteries from control animals only. Epicholesterol increased NOX2- dependent superoxide production and Rac1 activity in PASMCs from control but not CH rats. Furthermore, epicholesterol treatment does not alter PASMC caveolae number. In agreement with our hypothesis, the epicholesterol-dependent substitution of vii membrane cholesterol revealed concentration-dependent vasoconstriction to EGF in pulmonary arteries from control rats that is absent under vehicle conditions. Treatment with either fasudil (Rho kinase inhibitor) or gp91ds-tat (NOX2 inhibitor) attenuated this effect. We conclude that ROS mediate CH-dependent decreases in PASMC membrane cholesterol and that this depletion confers enhanced vasoconstriction in an EGFRdependent manner due to disinhibition of NOX2 and Rac1. These findings provide an improved mechanistic understanding of vascular smooth muscle signaling regulation in CH-induced PH and may provide a new approach to the treatment of pulmonary hypertension that seeks to normalize PASMC membrane cholesterol homeostasis.
Document Type
Dissertation
First Committee Member (Chair)
Thomas C. Resta
Second Committee Member
Nikki L. Jernigan
Third Committee Member
Laura Gonzalez Bosc
Fourth Committee Member
William S. Garver
Fifth Committee Member
Sarah K. Medrek
Recommended Citation
Ahmadian, Rosstin. "CHOLESTEROL REGULATION OF EGFR-DEPENDENT VASOCONSTRICTION IN CHRONIC HYPOXIA-INDUCED PULMONARY HYPERTENSION." (2023). https://digitalrepository.unm.edu/biom_etds/239
