Journal of Pharmacological Sciences
Online ISSN : 1347-8648
Print ISSN : 1347-8613
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Nicotine- and Tar-Free Cigarette Smoke Induces Cell Damage Through Reactive Oxygen Species Newly Generated by PKC-Dependent Activation of NADPH Oxidase
Hiroshi AsanoTakahiro HorinouchiYosuke MaiOsamu SawadaShunsuke FujiiTadashi NishiyaMasabumi MinamiTakahiro KatayamaToshihiko IwanagaKoji TeradaSoichi Miwa
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2012 Volume 118 Issue 2 Pages 275-287

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Abstract

We examined cytotoxic effects of nicotine/tar-free cigarette smoke extract (CSE) on C6 glioma cells. The CSE induced plasma membrane damage (determined by lactate dehydrogenase leakage and propidium iodide uptake) and cell apoptosis {determined by MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] reduction activity and DNA fragmentation}. The cytotoxic activity decayed with a half-life of approximately 2 h at 37°C, and it was abolished by N-acetyl-L-cysteine and reduced glutathione. The membrane damage was prevented by catalase and edaravone (a scavenger of OH) but not by superoxide dismutase, indicating involvement of OH. In contrast, the CSE-induced cell apoptosis was resistant to edaravone and induced by authentic H2O2 or O2 generated by the xanthine/xanthine oxidase system, indicating involvement of H2O2 or O2 in cell apoptosis. Diphenyleneiodonium [NADPH oxidase (NOX) inhibitor] and bisindolylmaleimide I [BIS I, protein kinase C (PKC) inhibitor] abolished membrane damage, whereas they partially inhibited apoptosis. These results demonstrate that 1) a stable component(s) in the CSE activates PKC, which stimulates NOX to generate reactive oxygen species (ROS), causing membrane damage and apoptosis; 2) different ROS are responsible for membrane damage and apoptosis; and 3) part of the apoptosis is caused by oxidants independently of PKC and NOX.

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© The Japanese Pharmacological Society 2012
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