title: N-acetylcysteine attenuates TNF-alpha-induced human vascular endothelial cell apoptosis and restores eNOS expression.
abstract: The circulatory inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) is increased in pathological conditions, such as diabetes, which initiate or exacerbate vascular endothelial injury. Both nitric oxide (NO) and reactive oxygen species may play a dual role (i.e., inhibiting or promoting) in TNF-alpha-induced endothelial cell apoptosis. We investigated the effects of the antioxidant N-acetylcysteine on TNF-alpha-induced apoptosis in human vascular endothelial cell (cell line ECV304) apoptosis, NO production and lipid peroxidation. Cultured vascular endothelial cell (ECV304) were either not treated (control), or treated with TNF-alpha (40 ng/ml) alone or TNF-alpha in the presence of N-acetylcysteine at 30 mmol/l or 1 mmol/l, respectively, for 24 h. Cell viability was measured by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay. Cell apoptosis was assessed by flow cytometry. TNF-alpha-induced endothelial cell apoptosis was associated with increased inducible NO synthase but reduced endothelial NO synthase (eNOS) protein expression. NO production and the levels of the lipid peroxidation product malondialdehyde were concomitantly increased. Treatment with NAC at 30 mmol/l restored eNOS expression and further increased NO production as compared to TNF-alpha alone, resulting in improved cell viability and reduced apoptosis. This was accompanied by increased superoxide dismutase activity, increased glutathione peroxidase production and reduced malondialdehyde levels. N-acetylcysteine at 1 mmol/l, however, did not have significant effects on TNF-alpha-induced endothelial cell apoptosis and cell viability despite it slightly enhanced glutathione peroxidase production. N-acetylcysteine attenuation of TNF-alpha-induced human vascular endothelial cell apoptosis is associated with the restoration of eNOS expression.
authors.affiliation: Anesthesiology Research Laboratory, Department of Anesthesiology, Renmin Hospital of Wuhan University, Wuhan, 430060, China. zhengyuan_xia@yahoo.com
authors.forename: Zhengyuan
authors.initials: Z
authors.lastname: Xia
authors.forename: Min
authors.initials: M
authors.lastname: Liu
authors.forename: Yong
authors.initials: Y
authors.lastname: Wu
authors.forename: Vijay
authors.initials: V
authors.lastname: Sharma
authors.forename: Tao
authors.initials: T
authors.lastname: Luo
authors.forename: Jingping
authors.initials: J
authors.lastname: Ouyang
authors.forename: John H
authors.initials: JH
authors.lastname: McNeill
mesh_terms: D000111:Acetylcysteine / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000201:enzymology; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology; D005434:Flow Cytometry; D005978:Glutathione / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008315:Malondialdehyde / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000096:biosynthesis*; D013482:Superoxide Dismutase / Q000378:metabolism; D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors* / Q000494:pharmacology*
chemical_list: D013778:Tetrazolium Salts; D013844:Thiazoles; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D008315:Malondialdehyde; D007770:L-Lactate Dehydrogenase; D005979:Glutathione Peroxidase; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D013482:Superoxide Dismutase; C022616:thiazolyl blue; D005978:Glutathione; D000111:Acetylcysteine