The role of NADPH oxidase-mediated oxidative stress in Angiotensin II-induced cellular senescence of vascular smooth muscle cells

Autor: Zih-cian Pan, 潘子茜
Rok vydání: 2008
Druh dokumentu: 學位論文 ; thesis
Popis: 97
Cellular senescence plays an important role in age-related cardiovascular diseases (CVD) including atherosclerosis and abdominal aortic aneurysm. Angiotensin II (Ang II)-induced cellular senescence of vascular smooth muscle cells (VSMCs) is involved in the pathogenesis of CVD. We investigated the roles of NADPH oxidase and members of the mitogen-activated protein kinases (MAPK) in Ang II-induced cellular senescence of human aortic VSMCs. Cellular senescence was assessed with senescence-associated β-galactosidase (SA-β-gal) activity and the expression of two cell cycle inhibitors p21 and p16, three established biomarkers for cellular senescence. Ang II (10-7 M) treatment for 48 h induced VSMCs senescence assessed with senescence-associated β-galactosidase activity and the expression of two cell cycle inhibitors p21 and p16. Ang II treatment for short term (between 2 and 10 min) and long term (between 24 and 48 h) also stimulated reactive oxygen species (ROS) production detected with a fluorescent probe carboxy-2’,7’-dichlorodihydrofluorescein diacetate (H2DCFDA). ROS scavenging with an antioxidant N-acetyl-L-cysteine (NAC) or a membrane-permeable catalase and inhibiting NADPH oxidase with an inhibitor, apocynin, both abolished Ang II-stimulated cellular senescence. Furthermore, small interfering RNA for NADPH oxidase catalytic subunit isoform Nox1 (Nox1 siRNA, 200 nM) markedly inhibited Ang II–induced cellular senescence. Ang II treatment for 24, 36 or 48 h induced mitochondrial membrane depolarization detected with a fluorescent probe 3,3′-dihexyloxacarbocyanine iodide. NAC, apocynin and Nox1 siRNA inhibited Ang II–induced mitochondrial membrane depolarization. We next examined the involvement of MAPKs in Ang II-induced cellular senescence. Ang II treatment transiently activated extracellular signal-regulated kinases 1/2 (ERK 1/2) and p38 MAPK. Inhibitors for ERK 1/2 activation(U0126), p38 MAPK (SB203580), and c-jun N-terminal kinase (JNK) (SP600125), markedly attenuated Ang II–stimulated mitochondrial membrane depolarization and cellular senescence of VSMCs. Interestingly, NAC and apocynin inhibited Ang II-induced activation of ERK 1/2 and JNK, but not p38 MAPK. These results suggested that Ang II induced cellular senescence of HASMCs via NADPH oxidase-dependent ERK1/2 and JNK activation and NADPH oxidase-independent p38 MAPK activation with both leading to mitochondrial dysfunction. Furthermore, Nox1-dependent NADPH oxidase mediates Ang II-induced mitochondrial dysfunction and cellular senescence.
Databáze: Networked Digital Library of Theses & Dissertations
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