Hypoxia/reoxygenation up-regulates death receptor expression and enhances apoptosis in human biliary epithelial cells.
OBJECTIVE
To investigate whether ischemia/reperfusion (I/R)-induced apoptosis in the bile duct epithelium could be mediated by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors in biliary epithelial cells, we examined the effects of hypoxia/reoxygenation (H/R) on TRAIL cytotoxicity.


METHODS
Using an H/R model, normal primary human intrahepatic biliary epithelial cells were exposed to hypoxia for 1 h, and then reoxygenated. Expressions of death receptor 4 (DR4) and DR5 mRNA and protein were measured. After 1 h of hypoxia, biliary epithelial cells were treated with TRAIL in different concentrations for 4 h. The death of biliary epithelial cells was confirmed by analysis of apoptosis and methylthiazolyl tetrazolium. The activities of caspase-3 and caspase-8 were determined by fluorometric assay.


RESULTS
Compared with normoxic-cultured cells, the mRNA expressions of DR4 and DR5 were up-regulated from 0 min after reoxygenation, reaching a peak value at 60 min after reoxygenation. The protein expression of DR4 was most intense at 90 min after reoxygenation; the most intense expression of DR5 came at 120 min after reoxygenation. The apoptosis rate increased in the TRAIL treatment group and further increased in the TRAIL plus H/R group, and the effect of concentration-dependent TRAIL-mediated cell killing was more pronounced. Caspase-3 and caspase-8 enzymatic activities after H/R also increased with increased TRAIL concentration.


CONCLUSIONS
H/R up-regulated the expression of DR4 and DR5, and enhanced TRAIL-mediated apoptosis in normal human intrahepatic biliary epithelial cells.
Feng
Li
L
Key Laboratory of Transplant Engineering and Immunology of Health Ministry of China, West China Hospital, Sichuan University, Chengdu, 610041, Sichuan Province, PR China.
Pang
Lili
L
Guo
Yingjia
Y
Ke
Nengwen
N
Li
Shengfu
S
Wei
Liang
L
Li
Quansheng
Q
Li
Youping
Y
D017209:Apoptosis / Q000502:physiology*; D001659:Biliary Tract / Q000166:cytology / Q000503:physiopathology; D053148:Caspase 3 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D000860:Hypoxia / Q000503:physiopathology*; D010100:Oxygen / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism*; D015854:Up-Regulation
D012333:RNA, Messenger; D053220:Receptors, TNF-Related Apoptosis-Inducing Ligand; D053148:Caspase 3; D053181:Caspase 8; D010100:Oxygen