Abstract
Abnormal proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs) are hallmark characteristics of vascular remodeling in pulmonary hypertension induced by chronic hypoxia. In this study, we investigated the role of the Na+/H+ exchanger (NHE) and alterations in intracellular pH (pHi) homeostasis in meditating increased proliferation and migration in PASMCs isolated from resistance‐sized pulmonary arteries from chronically hypoxic rats or from normoxic rats that were exposed to hypoxia ex vivo (1% or 4% O2, 24–96 h). We found that PASMCs exposed to either in vivo or ex vivo hypoxia exhibited greater proliferative and migratory capacity, elevated pHi, and enhanced NHE activity. The NHE inhibitor, ethyl isopropyl amiloride (EIPA), normalized pHi in hypoxic PASMCs and reduced migration by 73% and 45% in cells exposed to in vivo and in vitro hypoxia, respectively. Similarly, EIPA reduced proliferation by 97% and 78% in cells exposed to in vivo and in vitro hypoxia, respectively. We previously demonstrated that NHE isoform 1 (NHE1) is the predominant isoform expressed in PASMCs. The development of hypoxia‐induced pulmonary hypertension and alterations in PASMC pH i homeostasis were prevented in mice deficient for NHE1. We found that short‐term (24 h) ex vivo hypoxic exposure did not alter the expression of NHE1, so we tested the role of Rho kinase (ROCK) as a possible means of increasing NHE activity. In the presence of the ROCK inhibitor, Y‐27632, we found that pHi and NHE activity were normalized and migration and proliferation were reduced in PASMCs exposed to either in vivo (by 68% for migration and 22% for proliferation) or ex vivo (by 43% for migration and 17% for proliferation) hypoxia. From these results, we conclude that during hypoxia, activation of ROCK enhances NHE activity and promotes PASMC migration and proliferation.
Highlights
Persistent alveolar hypoxia is associated with many chronic lung diseases, and results in the development of pulmonary hypertension, significantly impacting patient morbidity and mortality
We previously demonstrated that the alkaline shift in pHi observed in pulmonary arterial smooth muscle cells (PASMCs) isolated from chronically hypoxic rats and mice is due to upregulation of Na+/H+ exchanger (NHE) isoform 1 (NHE1) and enhanced NHE activity (Rios et al 2005; Shimoda et al 2006)
In PASMCs isolated from rats exposed to chronic hypoxia (CH) perfused with a bicarbonate-free extracellular solution average baseline pHi was significantly higher compared to that measured in PASMCs from normoxic rats (Fig. 1A), confirming our previous findings in mice
Summary
Persistent alveolar hypoxia is associated with many chronic lung diseases, and results in the development of pulmonary hypertension, significantly impacting patient morbidity and mortality. Several studies have proposed an important role for changes in pulmonary arterial smooth muscle cell (PASMC) intracellular pH (pHi) in the pathogenesis of hypoxic pulmonary hypertension (Huetsch and Shimoda 2015). Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.
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