Chronic Hypoxia Increases Intracellular Ca2+ Concentration via Enhanced Ca2+ Entry Through Receptor-Operated Ca2+ Channels in Pulmonary Venous Smooth Muscle Cells

Background: Hypoxic pulmonary hypertension (HPH) is characterized by pulmonary vascular remodeling. Intracellular Ca2+ concentration ([Ca2+]i) is an essential signal for myocyte proliferation. Whether chronic hypoxia (CH) affects the basal [Ca2+] i and Ca2+ entry through store- and/or receptor-operated calcium channels (SOCC, ROCC), and whether canonical transient receptor potential (TRPC) proteins are involved in CH-induced Ca2+ influx and proliferation in pulmonary venous smooth muscle cells (PVSMCs) is examined. Methods and Results: Rats were exposed to CH. PVSMCs were isolated from distal pulmonary veins. In freshly isolated PVSMCs, CH increased the basal [Ca2+] i; removal of Ca2+ or application of SKF-96365 reversed the elevated [Ca2+] i, whereas nifedipine had no effect. Receptor-operated Ca2+ entry (ROCE) was expressed in PVSMCs. In freshly isolated PVSMCs from CH rats, ROCE was enhanced, whereas store- operated Ca2+ entry had no alteration. Furthermore, real-time polymerase chain reaction and western blotting showed that mRNA and protein expression level of TRPC6, but neither TRPC1 nor TRPC3, in pulmonary venous smooth muscle (PV) from CH rats and PVSMCs exposed to CH was greater than in normal PV and PVSMCs. The knockdown of TRPC6 in hypoxic PVSMCs with siRNA inhibited the enhanced ROCE and attenuated CH-induced PVSMCs proliferation. Conclusions: The enhanced Ca2+ entry through ROCC, due to upregulated TRPC6, is a novel pathogenic mechanism contributing to the increased basal [Ca2+] i in PVSMCs and excessive PVSMC proliferation during the development of HPH.