Overexpression of CYP2E1 enhances sensitivity of hepG2 cells to fas-mediated cytotoxicity.
Hepatocellular carcinoma (HCC) has been reported to be resistant to Fas-mediated apoptosis. In present study, experiments were conducted to investigate the potential effects of CYP2E1 overexpression on susceptibility of HCC to Fas-mediated cytotoxicity. HCC cell line HepG2 was infected with Ad-CYP2E1 to enhance the expression of CYP2E1, followed by treatment with low toxic dose of recombinant human Fas ligand (FasL, 0.5 ng/ml) in the presence of Actinomycin D (Act D, 125 ng/ml). High level of Fas expression was found in HepG2 cells. Its protein level and distribution kept unchanged after different treatments. Compared with control, CYP2E1 expressed HepG2 cells were more sensitive to FasL plus Act D. The sensitivity was elevated in a multiplicity of infection (m.o.i)-dependent manner, which was dramatically suppressed by CYP2E1 inhibitor diallyl disulfide (DAS) (p < 0.01). The percentage of apoptotic cells caused by FasL/Act D was increased from 18.7 to 75% after infection with Ad-CYP2E1 (p < 0.01). DAS treatment resulted in 60% reduction of apoptotic ratio (p < 0.01). Antioxidants GSH ethyl ester, Vitamin C and Vitamin E efficiently protected against cytotoxicity induced by FasL plus Act D in CYP2E1-expressed HepG2 cells. After adding FasL/Act D, increased caspases activities, lipid preoxidation and reduced GSH level, as well as mitochondrial release of cytochrome c were found in Ad-CYP2E1 infected cells (all p < 0.01); these changes were significantly attenuated by DAS (all p < 0.05). These results suggested that CYP2E1 potentiates Fas-mediated HepG2 cells toxicity via the induction of oxidative stress to promote apoptosis. Adenovirus-mediated overexpresson of CYP2E1 may have an important role in the elimination of hepatoma cells mediated by immune effector cells in the liver.
Yan
Qing-Guo
QG
State Key Laboratory of Cancer Biology, Department of Pathology, Xijing Hospital, the Fourth Military Medical University, ShaanXi, Xi'an, China.
Shi
Jian-Guo
JG
Zhang
Feng
F
Zhao
Qing-Tao
QT
Pang
Xiao-Wen
XW
Chen
Rui
R
Hu
Pei-Zhen
PZ
Li
Qin-Long
QL
Wang
Zhe
Z
Huang
Gao-Sheng
GS
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D019392:Cytochrome P-450 CYP2E1 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D053222:Fas Ligand Protein / Q000235:genetics / Q000494:pharmacology*; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology*; D018384:Oxidative Stress / Q000187:drug effects / Q000235:genetics; D019014:fas Receptor / Q000235:genetics / Q000378:metabolism*
D053222:Fas Ligand Protein; D019014:fas Receptor; D019392:Cytochrome P-450 CYP2E1