Cytotoxicity and Genotoxicity of Cigarette Smoke Extract on Human Osteoblastic U-2 OS Cells

• Main Authors: Yu-Pong Chen, 陳鈺浲
• Other Authors: Tsu-Shing Wang
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/29842353774267003503

碩士 === 中山醫學大學 === 生物醫學科學學系碩士班 === 101 === Cigarette smoking was first identified as a risk factor for osteoporosis; however, the precise mechanism of smoking in osteoporosis remains unclear. Therefore, the purpose of this study is to investigate whether cigarette smoke extract (CSE) induces cytotoxicity and genotoxicity in Human osteosarcoma U2-OS cells. The results showed that CSE treatment for 24 hours could significantly induce the cytotoxicity and genotoxicity (DNA strand breaks and DNA cross-links) by coment assay and reactive oxygen species by DCFDA - Cellular Reactive Oxygen Species Detection Assay in U2-OS cells.In addition, the pan-caspase inhibitor BOC-D-FMK did not block CSE-induced cytotoxicity ,but N-acetylcysteine, glutathione, c-Jun N-terminal kinases inhibitor (SP600125) , Poly (ADP-ribose) polymerase 1 inhibitor (NU1025),and Necroptosis inhibitor (Necrostatin-1) can block cytotoxicity by crystal violet assay in U2-OS cells. CSE-induced increase in LC3-II protein expression performed autophagy, and can be inhibited by the addition of N-acetylcysteine and glutathione. CSE induced cell necrosis by Annexin-V/Propidium iodide double statining.The summary of these results, CSE-induced cell death of U2-OS cells through reactive oxygen species production increase, induced c-Jun N-terminal kinases directly or causing DNA damage-induced DNA repair enzyme Poly (ADP-ribose) polymerase 1 induction of receptor-interacting protein 1 lead to the activation of c-Jun N-terminal kinases, mitochondrial membrane potential release apoptosis inducing factor leading to cell necrosis, smoking may be induced osteoporosis occurs by this pathway. N-acetylcysteine and glutathione and CSE in combination in vitro by DTNB assay, N-acetylcysteine and glutathione may be used for the prevention of smoke induced osteoporosis.