Search

The stress-responsive serum- and glucocorticoid-inducible kinase Sgk-1 is involved in osmoregulation and cell survival and may contribute to fibrosis and hypertension. However, the function of Sgk-1 in vascular remodeling and thrombosis, 2 major determinants of pulmonary hypertension (PH), has not been elucidated. We investigated the role of Sgk-1 in thrombin signaling and tissue factor (TF) expression and activity in pulmonary artery smooth muscle cells (PASMC). Thrombin increased Sgk-1 activity and mRNA and protein expression. H2O2 similarly induced Sgk-1 expression. Antioxidants, dominant-negative Rac, and depletion of the NADPH oxidase subunit p22phox diminished thrombin-induced Sgk-1 expression. Inhibition of p38 mitogen-activated protein kinase, phosphatidylinositol 3-kinase, and phosphoinositide-dependent kinase-1 prevented thrombin-induced Sgk-1 expression. Thrombin or Sgk-1 overexpression enhanced TF expression and procoagulant activity, whereas TF upregulation by thrombin was diminished by kinase-deficient Sgk-1 and was not detectable in fibroblasts from mice deficient in sgk-1 (sgk1(-/-)). Similarly, dexamethasone treatment failed to induce TF expression and activity in lung tissue from sgk1(-/-) mice. Transcriptional induction of TF by Sgk-1 was mediated through nuclear factor kappaB. Finally, Sgk-1 and TF proteins were detected in the media of remodeled pulmonary vessels associated with PH. These data show that thrombin potently induces Sgk-1 involving NADPH oxidases, phosphatidylinositol 3-kinase, p38 mitogen-activated protein kinase, and phosphoinositide-dependent kinase-1, and that activation of nuclear factor kappaB by Sgk-1 mediates TF expression and activity by thrombin. Because enhanced procoagulant activity can promote pulmonary vascular remodeling, and Sgk-1 and TF were present in the media of remodeled pulmonary vessels, this pathway may play a critical role in vascular remodeling in PH.

Background: Human urotensin II (hU-II) is a potent vasoactive peptide possibly involved in pulmonary hypertension. Because the signaling mechanisms activated by this peptide in the pulmonary vasculature are largely unknown, we investigated the role of hU-II in the activation of NADPH oxidase and the control of redox-sensitive kinase pathways, expression of plasminogen activator inhibitor-1 (PAI-1), and proliferation in pulmonary artery smooth muscle cells (PASMCs)., Methods and Results: hU-II upregulated expression of the NADPH oxidase subunits p22phox and NOX4 and increased the levels of reactive oxygen species (ROS), which were abrogated by transfecting p22phox or NOX4 antisense vectors. p22phox and NOX4 also contributed to hU-II-induced activation of extracellular signal-regulated kinase 1/2, p38 mitogen-activated protein kinase, c-Jun N-terminal kinase, and protein kinase B (Akt). Furthermore, hU-II increased the expression of PAI-1 and enhanced PASMC proliferation in an NADPH oxidase- and kinase-dependent manner., Conclusions: hU-II is a potent activator of ROS generation by NADPH oxidase in PASMCs, leading to redox-sensitive activation of mitogen-activated protein kinases and Akt and subsequently to enhanced PAI-1 expression and increased proliferation. These findings suggest that hU-II may play a novel role in pulmonary hypertension by promoting remodeling processes via activation of NADPH oxidases.

Pulmonary hypertension and vascular remodeling processes are associated with oxidative stress, hypoxia and enhanced levels of thrombin and vascular endothelial growth factor (VEGF). The hypoxia-inducible transcription factor HIF regulates the expression of VEGF under hypoxia. The HIF pathway is also activated by thrombin or CoCl2, likely via reactive oxygen species (ROS). In this study we investigated whether the redox-modifying enzymes superoxide dismutase (SOD), glutathione peroxidase (GPX) and catalase affect HIF levels and the expression of VEGF mRNA in pulmonary artery smooth muscle cells (PASMC). Stimulation of PASMC with thrombin or CoCl2 increased ROS production and enhanced HIF-alpha protein and VEGF mRNA levels as well as HIF-dependent reporter gene activity. These responses were inhibited by vitamin C and by overexpression of GPX and catalase, whereas the opposite effects were observed in SOD-expressing cells. These findings suggest that an 'antioxidant' state with reduced levels of H2O2 limits the activation of the HIF pathway, whereas a 'prooxidant' state allowing elevated H2O2 levels promotes it. Thus, shifting the redox balance to a more reduced environment, thereby limiting VEGF expression, may be beneficial for treating remodeling processes during pulmonary hypertension.