Hypoxia/reoxygenation up-regulates death receptor expression and enhances apoptosis in human biliary epithelial cells

Abstract: Aims: 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. Main 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. Key findings: 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. Significance: H/R up-regulated the expression of DR4 and DR5, and enhanced TRAIL-mediated apoptosis in normal human intrahepatic biliary epithelial cells. [Copyright &y& Elsevier]