Endothelin‐1 (ET‐1) induces hypoxia‐inducible factor 1 (HIF‐1) in Pulmonary Arterial Smooth Muscle Cells (PASMCs).

Numerous cellular responses to hypoxia are mediated by the transcription factor, HIF‐1. Recent data suggest that, under certain conditions, HIF‐1 may require feed forward for full expression (Peng, et al. J Physiol, 2006). Recently, ET‐1 was found to increase HIF‐1 levels in tumor cells (Spinella, et al. Cancer Res 2007). Since hypoxia increases ET‐1 levels in the lung, we hypothesized that during moderate prolonged hypoxia ET‐1 might feed forward and amplify activation of HIF‐1 in PASMCs. Primary cultures of rat PASMCs were treated with ET‐1 (10 −8 M; 48 hr) or exposed to hypoxia (4% O 2 ; 60 hrs). Protein levels of the oxygen‐sensitive α subunit of HIF‐1 (HIF‐1α) were markedly increased in nuclear extracts from both hypoxic cells and cells treated with ET‐1 under non‐hypoxic conditions. Time course studies revealed that accumulation of HIF‐1α in response to ET‐1 required greater than 4 hr of exposure. Real‐time PCR revealed that ET‐1 (10 −10 ‐10 −8 M; 48 hr) increased Hif1a mRNA expression. ET‐1 also decreased mRNA and protein expression of prolyl hydroxylase 2 (PHD2), the protein responsible for targeting HIF‐1α for degradation. The induction of HIF‐1α by moderate prolonged hypoxia (4% O 2 ; 60 hr) was blocked by BQ‐123, an ET‐1 receptor subtype A antagonist, whereas BQ‐123 had no effect on the induction of HIF‐1α by severe acute hypoxia (1% O 2 ; 4 hr). These results suggest that ET‐1 induces HIF‐1α by upregulation of HIF‐1α synthesis and downregulation of PHD2‐mediated degradation. Furthermore, the sustained induction of HIF‐1α in PASMCs during moderate prolonged hypoxia may require amplification by ET‐1. Funded by: HL67191