PPAR{gamma} regulates hypoxia-induced Nox4 expression in human pulmonary artery smooth muscle cells through NF-{kappa}B.
NADPH oxidases are a major source of superoxide production in the vasculature. The constitutively active Nox4 subunit, which is selectively upregulated in the lungs of human subjects and experimental animals with pulmonary hypertension, is highly expressed in vascular wall cells. We demonstrated that rosiglitazone, a synthetic agonist of the peroxisome proliferator-activated receptor-γ (PPARγ), attenuated hypoxia-induced pulmonary hypertension, vascular remodeling, Nox4 induction, and reactive oxygen species generation in the mouse lung. The current study examined the molecular mechanisms involved in PPARγ-regulated, hypoxia-induced Nox4 expression in human pulmonary artery smooth muscle cells (HPASMC). Exposing HPASMC to 1% oxygen for 72 h increased Nox4 gene expression and H(2)O(2) production, both of which were reduced by treatment with rosiglitazone during the last 24 h of hypoxia exposure or by treatment with small interfering RNA (siRNA) to Nox4. Hypoxia also increased HPASMC proliferation as well as the activity of a Nox4 promoter luciferase reporter, and these increases were attenuated by rosiglitazone. Chromatin immunoprecipitation assays demonstrated that hypoxia increased binding of the NF-κB subunit, p65, to the Nox4 promoter and that binding was attenuated by rosiglitazone treatment. The role of NF-κB in Nox4 regulation was further supported by demonstrating that overexpression of p65 stimulated Nox4 promoter activity, whereas siRNA to p50 or p65 attenuated hypoxic stimulation of Nox4 promoter activity. These results provide novel evidence for NF-κB-mediated stimulation of Nox4 expression in HPASMC that can be negatively regulated by PPARγ. These data provide new insights into potential mechanisms by which PPARγ activation inhibits Nox4 upregulation and the proliferation of cells in the pulmonary vascular wall to ameliorate pulmonary hypertension and vascular remodeling in response to hypoxia.
Lu
Xianghuai
X
Department of Medicine, Atlanta Veterans Affairs, Emory University Medical Centers, Georgia, USA.
Murphy
Tamara C
TC
Nanes
Mark S
MS
Hart
C Michael
CM
D000818:Animals; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000494:pharmacology; D000860:Hypoxia / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000235:genetics* / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D047495:PPAR gamma / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011651:Pulmonary Artery / Q000166:cytology / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D000077154:Rosiglitazone; D045162:Thiazolidinediones / Q000494:pharmacology
D007004:Hypoglycemic Agents; D016328:NF-kappa B; D047495:PPAR gamma; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D045162:Thiazolidinediones; D000077154:Rosiglitazone; D006861:Hydrogen Peroxide; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; D010100:Oxygen