Swelling-activated Cl- currents and intracellular CLC-3 are involved in proliferation of human pulmonary artery smooth muscle cells

Wenbin Liang, Lihong Huang, Dongling Zhao, Jeff Z. He, Parveen Sharma, Jie Liu, Anthony O. Gramolini, Michael E. Ward, Hee Cheol Cho, Peter H. Backx

Research output: Contribution to journalArticlepeer-review


Background: Proliferation of pulmonary artery smooth muscle cells (PASMCs) leads to adverse vascular remodeling and contributes to pulmonary arterial hypertension, a condition associated with a 15% annual mortality despite treatment. We previously showed that swelling-activated Cl- currents (ICl,swell) are upregulated in PASMC proliferation and that nonspecific Cl- current blockers inhibit proliferation. However, the specific role of ICl,swell in PASMC proliferation and its molecular underpinning remain unknown. Methods and results: In the present study, we found that the specific ICl,swell blocker, DCPIB (4-[(2-butyl-6,7- dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy] butanoic acid), dose-dependently blocked (IC50=2.7μmol/l) ICl,swell and inhibited (IC50=6.9μmol/l) proliferation in isolated human PASMCs (hPASMCs). To identify the Cl- channel genes underlying I Cl,swell and regulating hPASMC proliferation, we measured the mRNA expression of candidate Cl- channel genes (CLC-1 to CLC-7, CLC-Ka and CLC-Kb, and BEST-1 to BEST-4) in hPASMCs. CLC-2 to CLC-7 and BEST-1 are expressed in hPASMCs, with the most abundant gene being CLC-3, a channel gene previously linked to ICl,swell. Although stable expression of a microRNA-adapted shRNA targeting CLC-3 transcripts in hPASMCs selectively reduced CLC-3 mRNA by more than 80% and inhibited hPASMC proliferation (by >45%) compared with control-shRNA, it did not alter ICl,swell. Consistent with this observation, immunocytostaining studies revealed that CLC-3 protein is primarily located in intracellular areas of cultured proliferative hPASMCs. The intracellular CLC-3 protein levels were profoundly reduced by shRNA targeting CLC-3. The other molecular candidate for ICl,swell (i.e.,CLC-2) also showed a mainly intracellular distribution. Conclusion: Our findings support the conclusion that both ICl,swell and CLC-3 play a role in PASMC proliferation, but CLC-3 channels do not underlie ICl,swell in these cells.

Original languageEnglish (US)
Pages (from-to)318-330
Number of pages13
JournalJournal of hypertension
Issue number2
StatePublished - Feb 2014
Externally publishedYes


  • 3-dihydro-1-oxo-1H-inden-5- yl)oxy] butanoic acid
  • 4-[(2-Butyl-6
  • 7-dichloro-2-cyclopentyl-2
  • CLC-3
  • proliferation
  • pulmonary arterial hypertension
  • RNAi

ASJC Scopus subject areas

  • Internal Medicine
  • Physiology
  • Cardiology and Cardiovascular Medicine


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