Hypoxia interacts with NFAT signaling in rat pulmonary arterial smooth muscle cells (PASMCs)

Nuclear factors of activated T‐cells (NFATs) are calcium sensitive transcription factors regulated by calcineurin. They play critical roles in inflammation and cell proliferation. Recent studies showed that calcineurin/NFAT pathway is involved in the pathogenesis of pulmonary hypertension. Here we exposed rats to hypoxia for 3 weeks with daily injection of the calcineurin inhibitor cyclosporin (CsA, 35 mg/kg/day). Right ventricular systolic pressure (CsA treated: 34.3±2.0 mmHg; untreated: 53.9±2.6 mmHg, p<0.05), and right heart hypertrophy were significantly reduced in CsA treated rats. Real‐time RT‐PCR showed that NFATc3 mRNA was predominantly expressed in PA of rats and mice. It was 3‐5 times the amount of other NFAT isoforms, except that it was similar to NFATc1 in mouse PA. NFATc3 and NFATc1 proteins were also detected by western blot. NFATc3 nuclear translocation was examined by expressing hNFATc3‐EGFP fusion protein in rat PASMCs. Confocal images showed that nuclear accumulation of hNFATc3‐EGFP occurred within 5 min and peaked at about 15‐20 min after PASMCs were stimulated by ionomycin and PMA. NFAT‐dependent transcription activity, monitored with a 4xNFAT‐luciferase reporter, was increased 4 folds by ionomycin+PMA. Moreover, the stimulatory effect of ionomycin+PMA was further potentiated in PASMCs exposed to 24 hr hypoxia (2% O 2 ). Since many agonists stimulate PLC to activate IP 3 ‐induced Ca 2+ release and PKC similar to ionomycin+PMA, this observation suggests that hypoxia may interact synergistically with agonists to enhance NFAT‐dependent gene transcription in PASMCs.