Chronic hypoxia-induced upregulation of store-operated and receptor-operated Ca2+ channels in pulmonary arterial smooth muscle cells: a novel mechanism of hypoxic pulmonary hypertension.
Chronic hypoxic pulmonary hypertension is associated with profound vascular remodeling and alterations in Ca(2+) homeostasis in pulmonary arterial smooth muscle cells (PASMCs). Recent studies show that transient receptor potential (TRPC) genes, which encode store-operated and receptor-operated cation channels, play important roles in Ca(2+) regulation and cell proliferation. However, the influence of chronic hypoxia on TRPC channels has not been determined. Here we compared TRPC expression, and store- and receptor-operated Ca(2+) entries in PASMCs of normoxic and chronic hypoxic rats. Reverse-transcription polymerase chain reaction (RT-PCR), Western blot, and immunostaining showed consistently that TRPC1, TRPC3, and TRPC6 were expressed in intralobar pulmonary arteries (PAs) and PASMCs. Application of 1-oleoyl-2-acetyl-sn-glycerol (OAG) to directly activate receptor-operated channels, or thapsigargin to deplete Ca(2+) stores, caused dramatic increase in cation entry measured by Mn(2+) quenching of fura-2 and by Ca(2+) transients. OAG-induced responses were approximately 700-fold more resistant to La(3+) inhibition than thapsigargin-induced responses. siRNA knockdown of TRPC1 and TRPC6 specifically attenuated thapsigargin- and OAG-induced cation entries, respectively, indicating that TRPC1 mediates store-operated entry and TRPC6 mediates receptor-operated entry. In hypoxic PAs, there were 2- to 3-fold increases in TRPC1 and TRPC6 expression. They were accompanied by significant increases in basal, OAG-induced, and thapsigargin-induced cation entries in hypoxic PASMCs. Moreover, removal of Ca(2+) or inhibition of store-operated Ca(2+) entry with La(3+) and SK&F-96365 reversed the elevated basal [Ca(2+)](i) in PASMCs and vascular tone in PAs of chronic hypoxic animals, but nifedipine had minimal effects. Our results for the first time to our knowledge show that both store- and receptor-operated channels of PASMCs are upregulated by chronic hypoxia and contribute to the enhanced vascular tone in hypoxic pulmonary hypertension.
Lin
Mo-Jun
MJ
Division of Pulmonary and Critical Care Medicine, Johns Hopkins Asthma and Allergy Center, 5501 Hopkins Bayview Circle, Baltimore, MD 21224, USA.
Leung
George P H
GP
Zhang
Wei-Min
WM
Yang
Xiao-Ru
XR
Yip
Kay-Pong
KP
Tse
Chung-Ming
CM
Sham
James S K
JS
D000818:Animals; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000096:biosynthesis* / Q000235:genetics / Q000502:physiology; D002412:Cations / Q000378:metabolism; D015687:Cell Hypoxia; D002478:Cells, Cultured; D004075:Diglycerides / Q000494:pharmacology; D006976:Hypertension, Pulmonary / Q000209:etiology / Q000378:metabolism* / Q000503:physiopathology; D017136:Ion Transport; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D011651:Pulmonary Artery / Q000166:cytology / Q000378:metabolism* / Q000503:physiopathology; D034622:RNA Interference; D051381:Rats; D017208:Rats, Wistar; D050052:TRPC Cation Channels; D019284:Thapsigargin / Q000494:pharmacology; D015854:Up-Regulation
D015220:Calcium Channels; D002412:Cations; D004075:Diglycerides; D050052:TRPC Cation Channels; C492145:Trpc6 protein, rat; C096351:transient receptor potential cation channel, subfamily C, member 1; D019284:Thapsigargin; C038237:1-oleoyl-2-acetylglycerol; D002118:Calcium