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Rho Kinase and Na + /H + Exchanger Mediate Endothelin‐1 Induced Pulmonary Arterial Smooth Muscle Cell Proliferation and Migration
Author(s) -
Huetsch John Christian,
Yun Xin,
Jiang Haiyang,
Shimoda Larissa
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.892.11
Subject(s) - intracellular ph , endothelin 1 , pulmonary hypertension , amiloride , chemistry , hypoxic pulmonary vasoconstriction , rho associated protein kinase , vasoconstriction , protein kinase c , microbiology and biotechnology , sodium–hydrogen antiporter , medicine , endocrinology , kinase , intracellular , biochemistry , biology , sodium , receptor , organic chemistry
Objective Remodeling of the pulmonary vasculature, involving increased proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs), is a key component of the pathology underlying pulmonary hypertension (PH). Endothelin‐1 (ET‐1) is known to be an important contributor to both pulmonary vasoconstriction and remodeling in PH. We aimed to determine whether ET‐1 modulates PASMC function via a pathway involving Rho kinase (ROCK) and the Na + /H + exchanger (NHE). Methods PASMCs were isolated from distal pulmonary arteries of male C57/B6 mice as well as male Wistar rats and expanded in culture. To measure NHE activity, cells were incubated with pH‐sensitive fluorescent dye and the Na + ‐dependent recovery of intracellular pH (pH i ) following acidification by ammonium pulse challenge was measured via fluorescence microscopy. NHE1 mRNA expression was measured via qRT‐PCR and protein expression was measured via immunoblot. Proliferation was assessed via ELISA for BrdU incorporation over 24 hr. Migration was measured over 24 hr using Transwell filters with 8 μm pores. Cells were treated with 10 −8 M ET‐1 versus vehicle for 24–48 hr. NHE was pharmacologically inhibited with 10 μM ethyl‐isopropyl amiloride. ROCK was inhibited with 10 μM Y‐27632. Results ET‐1 treatment, compared to vehicle, resulted in increased resting pH i and NHE activity in PASMCs. ET‐1 did not result in increased NHE1 mRNA or protein expression. ROCK inhibition prevented ET‐1‐induced NHE activation. PASMC proliferation and migration were both increased by ET‐1 treatment, compared to vehicle. ROCK inhibition attenuated PASMC proliferation and migration in the presence of ET‐1. Similarly, NHE inhibition also attenuated ET‐1‐induced PASMC proliferation and migration. Conclusions ET‐1 results in increased PASMC proliferation and migration via a pathway involving ROCK and NHE activation. This suggests a novel pathway which may contribute to vascular remodeling in PH. Support or Funding Information NIH K08HL133475 & R01HL073859 This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .