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BACKGROUND: Biliary cancers are more common in fe males, and previous studies have suggested that Helicobacter pylori(H. pylori) exists in the biliary system. However, the effects of H. pylori infection and estrogen on the biological behaviors of human biliary epithelium mucosa remain un known. The present study aimed to clarify their effects on the proliferation, apoptosis, migration and oxidative DNA dam age of a human intrahepatic biliary epithelial cell(HIBEC)line in vitro.METHODS: HIBECs were co-cultured with 17β-estradiol(at 10~(-9) mol/L, 10~(-7) mol/L, and 10~(-5) mol/L) and H. pylori(at MOI=0.5:1, 1:1, and 2:1) and continuously passaged until the15 th generation(approximately 45 days). Then, the following assays were performed. HIBEC proliferation was measured using the CCK-8 assay, plate clone-formation assay and by de termining Ki-67 expression with immunocytochemistry; cell apoptosis and migration were investigated using Annexin-V/PI and transwell assays, respectively; and reactive oxygen species(ROS) and 8-hydroxy-2’-deoxyguanosine(8-OHd G)production were detected by flow cytometry and immuno fluorescence staining combined with confocal laser scanning microscopy, respectively. The results were the basis for evalu ating the level of oxidative stress and the related DNA damage in HIBECs.RESULTS: HIBECs maintained a normal morphology and vitality when treated with 17β-estradiol(at 10~(-9) mol/L) and H. pylori(at MOI=0.5:1 and 1:1). 17β-estradiol at 10~(-7) mol/L and 10~(-5) mol/L and H. pylori at MOI=2:1, by contrast, caused cell death. Compared with controls, HIBECs treated with 17β-estradiol(10~(-9) mol/L) and H. pylori(MOI=1:1) had a higher up-regulation of proliferation, Ki-67 expression, clone formation, migration activity and the expression of ROS and 8-OHd G and exhibited a down-regulation of apoptosis. The above effects were further increased when 17β-estradiol and H. pylori were combined(P<0.05).CONCLUSIONS: H. pylori and 17β-estradiol, separately or in combination, promoted cell proliferation and suppressed apoptosis of HIBECs in vitro. The above phenomena might be related to oxidative stress and its subsequent DNA damage with H. pylori and 17β-estradiol.
BACKGROUND: Biliary cancers are more common in fe males, and previous studies have suggested that Helicobacter pylori (H. pylori) exists in the biliary system. However, the effects of H. pylori infection and estrogen on the biological behaviors of human biliary epithelium mucosa The present study aimed to clarify their effects on the proliferation, apoptosis, migration and oxidative DNA dam age of a human intrahepatic biliary epithelial cell (HIBEC) line in vitro. METHODS: HIBECs were co-cultured with 17β-estradiol ( at 10 -9 mol / L, 10 -7 mol / L and 10 -5 mol / L) and H. pylori (at MOI = 0.5: 1, 1: 1, and 2 The following assays were performed. HIBEC proliferation was measured using the CCK-8 assay, plate clone-formation assay and by de termining Ki-67 expression with immunocytochemistry ; cell apoptosis and migration were investigated using Annexin-V / PI and transwell assays, respectively; and reactive oxygen species (ROS) and 8-hydroxy-2’-deoxyguanosine (8-OHd G) production were detected by flow cytometry and immuno fluorescence staining combined with confocal laser scanning microscopy, respectively. The results were the basis for evalu ating the level of oxidative stress and the related DNA damage in HIBECs.RESULTS: HIBECs maintained a normal morphology and vitality when treated with 17β-estradiol (at 10 -9 mol / L) and H. pylori (at MOI = 0.5: 1 and 1: 1). 17β-estradiol at 10 ~ (-7) mol / L and 10 ~ (-5) mol / L and H. pylori at MOI = 2: 1, HIBECs treated with 17β-estradiol (10 -9 mol / L) and H. pylori (MOI = 1: 1) had a higher up-regulation of proliferation, Ki-67 expression, clone formation, migration activity and the expression of ROS and 8-OHd G and exhibited a down-regulation of apoptosis. The above effects were further increased when 17β-estradiol and H. pylori were combined (P <0.05) .CONCLUSIONS: H. pylori and 17β-estradiol, sepa ratelyor in combination, promoted cell proliferation and suppressed apoptosis of HIBECs in vitro. The above phenomena might be related to oxidative stress and its subsequent DNA damage with H. pylori and 17β-estradiol.