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Primary biliary cholangitis (PBC) is a chronic autoimmune liver disease caused by the combined effect of genetic and environmental factors and characterized by the apoptotic necrosis of biliary epithelial cells (BEC) in the small intrahepatic bile ducts. This article describes the effect of B cells, macrophages, natural killer cells, NKT, and T cells on the immune injury of BEC during PBC, so as to provide some guidance for the targeted immune therapy for PBC.
ABSTRACT
Primary biliary cholangitis (PBC) is an autoimmune disease. Although PBC has the features of autoimmune disease, it has poor response to immunosuppressants and good response to the drugs participating in bile acid metabolism, such as ursodeoxycholic acid. Studies have shown that the bicarbonate secretion of biliary epithelial cells is impaired in PBC patients, and bile acid not blocked by HCO3- umbrella enters biliary epithelial cells and mediates their damage and apoptosis, leading to the expression of autoantibodies in apoptotic cells and immunologic injury. In order to explore the role of HCO3- umbrella secreted by biliary epithelial cells in the pathogenesis of PBC, this article briefly introduces the physiological function and production mechanism of HCO3- umbrella and the influencing factors for HCO3- secretion, and it is pointed out that reduced HCO3- secretion may be a key link in the pathogenesis of PBC and a potential therapeutic target.
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Objective: To investigate the toxicity and mechanisms of celastrol (CEL) on human biliary epithelial cells. Methods: The effects of CEL on cell morphology and cell viability changes were observed by CCK-8 experiment and microscope. Cell scratch experiment was used to detect the effect of CEL on cell migration. The effects of CEL on cell cycle and cell apoptosis were detected by flow cytometry. The mRNA and protein expression of apoptosis-related genes Caspase-3, Bax and Bcl-2 were detected by qRT-PCR and Western blotting. Results: CEL inhibited cell proliferation and changed cell morphology at 400-2 000 nmol/L. At 200-800 nmol/L, cell migration was inhibited. At 800-1 200 nmol/L, G0/G1 phase was arrested. At 400-1 200 nmol/L, cell apoptosis was induced and the expression of apoptosis-related genes was increased. Conclusion: CEL showed cholangiocyte toxicity through affecting cell viability, cell migration, preventing cell cycle and promoting cell apoptosis of human biliary epithelial cells.
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Objective To study the effect of bone marrow mesenchymal stem cells (BMMSCs) combined with normothermic mechanical perfusion (NMP) on biliary epithelial cells (BEC) after DCD donor liver transplantation in rats.Methods The third generation of BMMSCs and the BMMSCs modified by Ad/HO-1 (Ad/HO-1/BMMSCs) were cultured,identified and expanded in vitro.To establish a stable NMP system device in vitro.The DCD liver transplantation models were constructed in rats after cardiac ischemia for 30 minutes,220 SD recipient rats were randomly divided into sham operation group (S group,n=44) static cold storage (SCS group,n =44) group,and simple NMP group (P group,n =44),BMMSCs combined with NMP group (BP group,n =44) and BMMSCs modified by Ad/HO-1 combine with NMP group (HBP group,n =44),NMP group,BP group and HBP group were subjected to vitro perfusion for 4h.The group were taken at 0,1,7 and 14 days after transplantation and the relevant indicators were detected,n =6 in each group.The survival rate of the recipient rats,liver function and pathological changes of the bile duct were observed.The expression of cytokeratin 19 (CK19) protein in BEC was detected by immunohistochemistry and Western blot.Apoptotic biliary epithelial cells were detected by TUNEL staining and the expression of apoptosis-related protein caspase-3 was detected by immunohistochemistry.Results The survival time of HBP group was significantly prolonged for (5.6 ±0.8) d in SCS group vs.(18.4 ±2.0) d in NMP group,(20.5 ± 1.5) d in BP group,(82.5 ±3.2) d in HBP group,the differences were statistically significant (all P ＜ O.05).Compared with other groups,the HBP group and the BP group were significantly improved in liver function and biliary pathology,and the expression of CK19 protein in BEC was significantly increased [(0.81 ±0.02) in S group vs.(0.35 ±0.03) in SCS group,(0.47 ±0.02) in NMP group,(0.63 ± 0.02) in BP group,(0.77 ± 0.01) in HBP group on postoperative day (POD) 14],the differences were statistically significant (all P ＜ 0.05).The number of apoptosis and the expression of apoptosis-related protein caspase-3 in HBP group were significantly decreased [(10.0 ± 1.2) in S group vs.(57.3 ±5.5) in SCS group,(40.1 ±4.6) in NMP group,(32.0 ± 2.2) in BP group,(13.7 ± 3.1) in HBP group on POD 14],the difference was statistically significant (all P ＜ 0.05).Compared with the BP group,the protective effect of the HBP group was more obvious,and the difference was statistically significant (P ＜ 0.05).Conclusion By the method of the BMMSCs modified by Ad/HO-1 combined with NMP in vitro preservation of rat,DCD donor liver can significantly improve the effect of BEC on rats and the survival rate after liver transplantation.
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Objective To investigate the mechanism of interlekin-6 (IL-6) in wound healing of human biliary epithelial cells ( BECs ).Methods BECs were cultured in IL-6 at different concentrations:0 ng/L(0 ng/L group),10 ng/L (10 ng/L group),50 ng/L (50 ng/L group),100 ng/L (100 ng/L group),1000 ng/L ( 1000 ng/L group).The effects of IL-6 on the phosphorylation of signal transducer and activator of transcription 3( STAT3 ) and the expression of trefoil family factors 3 (TFF3) were detected.BECs were divided into untreated group,STAT3-RNAi group (BECs transfected with STAT3 RNAi adenovirus) and Control-RNAi group (BECs transfected with vacant RNAi adenovirus).The effects of IL-6 on the expression of TFF3 were detected after RNAi of STAT3.In vitro wound models were constructed for the untreated group,STAT3-RNAi group and Control-RNAi group,and the effects of IL-6 and TFF3 on BECs of the 3 groups were detected.All data were analyzed by using the Student's t test,analysis of variance or Sidak test.Results The expressions of phosphorylated STAT3 in the 50 ng/L group,100 ng/L group and 1000 ng/L group were 0.240 ± 0.052,0.714 ± 0.124,0.327 ± 0.069,respectively,which were significantly higher than 0.033 ± 0.011 of the 0 ng/L group (q =5.246,17.260,7.451,P ＜ 0.05 ).The contents of mRNA and protein of TFF3 increased as the increase of IL-6 concentration (q =12.045,9.889,P ＜ 0.05).After RNAi of STAT3 of the BECs,the expression of TFF3 decreased when the concentration of IL-6 was 1000 ng/L.The expression of TFF3 of the STAT3-RNAi group was 0.037 ± 0.005,which was significantly lower than 0.267 ± 0.038 of the Control-RNAi group and 0.301 ± 0.042 in the untreated group ( q =12.135,13.929,P ＜ 0.05 ).In the in vitro wound model,the speed of BECs migration in the STAT3-RNAi group was (9.1 ± 1.5 ) μm/h,which was slower than (25.1 ± 3.8 ) μm/h of the Control-RNAi group after 12 hours of interference with IL-6 (q =7.737,P ＜ 0.05 ).The speed of BECs migration of STAT3-RNAi group was (39.2 ± 4.7) μm/h after adding 1 g/L of recombinant TFF,which was significantly faster than that of the Control-RNAi group (q =14.507,P ＜0.05).Conclusion IL-6 promotes cell migration and wound healing by activating STAT3 and up-regulating TFF3 expression.
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Objective: To investigate the possible effect of mouse intrahepatic biliary epithelial cell line (mIBEC) on co-cultured human hepatoma cell lines HepG2. Methods: HepG2 and mIBEC cells were co-cultured in a membrane-separated Transwell system. CellTiter 96 ® AQueous One Solution Cell Proliferation Assay (Promega Corporation, Madison, WI) was used to examine HepG2 cell proliferation in the system with or without co-cultured mIBEC. Real-time PCR(RT-PCR) was used to determine Ki67 and caspase3 mRNA expression in HepG2 cells in a system with or without co-cultured mIBEC. Western blotting analysis was used to determine caspase3 protein level in HepG2 cells. Results: The proliferation of the co-cultured HepG2 cells was significantly lower than those cultured alone(P<0.01). Expression of Ki67, a cell proliferation marker, was also significantly down-regulated in mIBEC co-cultured HepG2 cells (P<0.05). The levels of caspase3 mRNA and protein were significantly up-regulated in mIBEC co-cultured HepG2 cells compared with HepG2 cells cultured alone (P < 0.05). Conclusion: mIBEC can inhibit the proliferation of co-cultured HepG2, and caspase3 activation might be one of the reasons for the inhibitory effect of mIBEC against HepG2 cells.