Effects Of ErbB4 On Growth And Metastasis Of Human Cholangiocarcinoma QBC939 Cell Line

Part 1 Expression of ErbB4 in human cholangiocarcinoma QBC939 cell lineObjective: To investigate expression of ErbB4 receptor in human cholangiocarcinoma QBC939 cell line.Methods: Human cholangiocarcinoma QBC939 cells were cultured in vitro, immumofluorescence staining were used to detect ErbB4 expressed in cholangiocarcinoma cells.Results: Human cholangiocarcinoma QBC939 cells expressed ErbB4 protein.Conclusions: Human cholangiocarcinoma QBC939 cells express ErBb4, provide feasibility that this study have chosen QBC939 cell line for in vitro subsequent experiments.Part 2 Effects of blockade of Erb B4 on proliferation, migration and invesiveness in human cholangiocarcinoma QBC939 cell lineObjective: To investigate the effects of ErbB4 inhibitor on proliferation, migration and invasiveness of cholangiocarcinoma cells in vitro.Methods: 0μmol/L, 1μmol/L, 5μmol/L, 10μmol/L, 25μmol/L, 50μmol/L, 75μmol/L, 100μmol/L AG1478 were blocked ErbB4 receptor, the proliferation of cholangiocarcinoma cells were measured by CCK-8 assay, moreover, median inhibitory concentration( IC50) of AG1478 in cholangiocarcinoma cells were calculate with SPPS software. After treating with 0μmol/L, 25μmol/L, 50μmol/L, 75μmol/L AG1478, wound-scratch assay was used to detect the ability of migration, Transwell chamber assay was used to detectd the ability of invesiveness of cholangiocarcinoma cells.Results: AG1478(0μmol/L, 1μmol/L, 5μmol/L, 10μmol/L, 25μmol/L, 50μmol/L, 75μmol/L,100μmol/L) inhibited QBC939 cell proliferation, in a dose-dependent manner, IC50 value is 49.14μmol/L After inhibiting ErbB4 receptor, the migration and invesiveness of QBC939 cells were attenuated.Conclusions: ErbB4 inhibitor reduces iNOS expressed in QBC939 cells, furthermore, downstream NO and VEGF reduce.Part 3 Molecular mechanisms that ErbB4 effects on proliferation and metastasis in cholangiocarcinoma cellObjective: To explore the relevance molecular mechanisms that ErbB4 receptor effects on oncobiology.Methods: Western blotting assay was used to analyze iNOS in cholangiocarcinoma cells treated with 50μmol/L(IC50)AG1478. After treating with 0μmol/L, 25μmol/L, 50μmol/L, 75μmol/L AG1478, Concentration of NO in cholangiocarcinoma cells conditioned medium was detected by Giess assay, VEGF was detected by ELISA assay.Results: After inhibiting ErbB4 receptor, the expressed of iNOS was lower. After inhibiting ErbB4 receptor, the concentration of NO and VEGF were decreased.Conclusions: ErbB4 inhibitor induce migration and invasion in QBC939 cells, which is involved in downregulation of iNOS protein expression in QBC939 cells.