Effect of HIF-1 on pulmonary arterial smooth muscle cell (PASMC) function and p38-MAP kinase pathway activation

RATIONALE: Chronic hypoxia (CH) leads to pulmonary hypertension (PH) via sustained vasoconstriction and vascular remodeling, characterized by PASMC hyperproliferation, medial wall thickening, and muscle extension into distal arterioles. Prior studies showed CH increases PASMC proliferative and migratory capacity and resistance to apoptosis, processes dependent on activation of hypoxic inducible factor 1 (HIF-1). Pulmonary arteries from humans with PH and CH have also been shown to exhibit constitutive phosphorylation of p38, and this signaling pathway has been shown to promote PASMC proliferation and vascular thickening. Prior research in cancer cells showed persistent phosphorylation of p38 and its downstream target, MK2, ultimately increased MK2 degradation and decreased total MK2 expression. MK2 plays a role as a nuclear chaperone of caspase-3, the final executioner of apoptosis. In this study we sought to dissect how HIF-1 induction influences activation of p38 and expression of MK2. METHODS: Intralobar pulmonary arteries were isolated from male Wistar rats by dissection. PASMCs were enzymatically isolated and grown in vitro. For experiments, PASMCs were growth arrested and subjected to 1) normoxia or hypoxia (1% and 4% O 2 ) using a humidified modular incubator; 2) treatment with either vehicle or dimethyloxalylglycine (DMOG), a potent inhibitor of prolyl hydroxylases (PHDs) which leads to HIF-1 activation; or 3) treatment with an adenoviral construct containing an O 2 -insensitive, constitutively active HIF-1. Immunoblot was performed to characterize expression of HIF-1, phosphorylated p38, phosphorylated MK2, and total MK2. RT-PCR was performed to determine expression of MK2 mRNA. Sub-cellular fractionation was performed on PASMCs subjected to the experimental conditions above to characterize the intracellular distribution of MK2. RESULTS: Induction of HIF-1 by hypoxia induced rapid phosphorylation of p38 and MK2 at 4 hours, followed by a decrease in total MK2 expression at 6 hours, which was sustained at 48 hours. Similarly, induction of HIF-1 with DMOG resulted in sustained phosphorylation of p38 and decreased total MK2 expression at 48 hours. Forced expression of HIF-1 also resulted in decreased total MK2 expression. Sub-cellular fractionation revealed that treatment with hypoxia and DMOG resulted in an increased amount of total and phosphorylated MK2 in the cytoplasm, and a decreased amount of total MK2 in the nucleus. RT-PCR of PASMCs subjected to hypoxia or treated with DMOG showed no decrease in MK2 mRNA expression at 48 hours, suggesting this process is not due to direct transcriptional regulation by HIF-1. CONCLUSION: Our work demonstrates that induction of HIF-1 by multiple methods in PASMCs was associated with rapid phosphorylation of p38 and MK2, followed by sustained p38 phosphorylation with and a subsequent decrease of total MK2 protein expression. Since MK2 plays an important role in mediating apoptosis by acting as a nuclear chaperone for caspase-3, our findings suggest a mechanism by which CH decreases total MK2 expression and its availability to facilitate apoptosis. We will explore the significance of this effect on PASMC resistance to apoptosis in follow-up studies. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.