title: NADPH oxidase/ROS-dependent PYK2 activation is involved in TNF-α-induced matrix metalloproteinase-9 expression in rat heart-derived H9c2 cells.
abstract: TNF-α plays a mediator role in the pathogenesis of chronic heart failure contributing to cardiac remodeling and peripheral vascular disturbances. The implication of TNF-α in inflammatory responses has been shown to be mediated through up-regulation of matrix metalloproteinase-9 (MMP-9). However, the detailed mechanisms of TNF-α-induced MMP-9 expression in rat embryonic-heart derived H9c2 cells are largely not defined. We demonstrated that in H9c2 cells, TNF-α induced MMP-9 mRNA and protein expression associated with an increase in the secretion of pro-MMP-9. TNF-α-mediated responses were attenuated by pretreatment with the inhibitor of ROS (N-acetyl-l-cysteine, NAC), NADPH oxidase [apocynin (APO) or diphenyleneiodonium chloride (DPI)], MEK1/2 (U0126), p38 MAPK (SB202190), JNK1/2 (SP600125), NF-κB (Bay11-7082), or PYK2 (PF-431396) and transfection with siRNA of TNFR1, p47(phox), p42, p38, JNK1, p65, or PYK2. Moreover, TNF-α markedly induced NADPH oxidase-derived ROS generation in these cells. TNF-α-enhanced p42/p44 MAPK, p38 MAPK, JNK1/2, and NF-κB (p65) phosphorylation and in vivo binding of p65 to the MMP-9 promoter were inhibited by U0126, SB202190, SP600125, NAC, DPI, or APO. In addition, TNF-α-mediated PYK2 phosphorylation was inhibited by NAC, DPI, or APO. PYK2 inhibition could reduce TNF-α-stimulated MAPKs and NF-κB activation. Thus, in H9c2 cells, we are the first to show that TNF-α-induced MMP-9 expression is mediated through a TNFR1/NADPH oxidase/ROS/PYK2/MAPKs/NF-κB cascade. We demonstrated that NADPH oxidase-derived ROS generation is involved in TNF-α-induced PYK2 activation in these cells. Understanding the regulation of MMP-9 expression and NADPH oxidase activation by TNF-α on H9c2 cells may provide potential therapeutic targets of chronic heart failure.
authors.affiliation: Department of Physiology and Pharmacology and Health Aging Research Center, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan; Heart Failure Center, Division of Cardiology, Department of Internal Medicine, Chang Gung Memorial Hospital at Keelung, Keelung, Taiwan. Electronic address: chuenmao@mail.cgu.edu.tw.
authors.forename: Chuen-Mao
authors.initials: CM
authors.lastname: Yang
authors.forename: I-Ta
authors.initials: IT
authors.lastname: Lee
authors.forename: Ru-Chun
authors.initials: RC
authors.lastname: Hsu
authors.forename: Pei-Ling
authors.initials: PL
authors.lastname: Chi
authors.forename: Li-Der
authors.initials: LD
authors.lastname: Hsiao
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D002460:Cell Line; D051418:Focal Adhesion Kinase 2 / Q000378:metabolism*; D020780:Matrix Metalloproteinase 9 / Q000096:biosynthesis*; D009206:Myocardium / Q000201:enzymology / Q000378:metabolism; D019255:NADPH Oxidases / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
chemical_list: D000975:Antioxidants; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D019255:NADPH Oxidases; D051418:Focal Adhesion Kinase 2; C496054:Ptk2b protein, rat; D020780:Matrix Metalloproteinase 9; C572337:Mmp9 protein, rat; D000111:Acetylcysteine
keywords: MAPKs; Matrix metalloproteinase-9; NADPH oxidase; PYK2; ROS