ABSTRACT
Objective To investigate the inhibitory effect of 5-FU on human intrahepatic biliary epithelial cells with TGF-β1-induced mesenchymal transition and potential mechanism.Methods The primary epithelial cells from human intrahepatic bile ducts were cultured,the cells were identified by immunofluorescence microscopy.The cells were randomly divided into 3 groups: normal control group,TGF-β1 group and TGF-β1+5-FU group.The expression of CK-19,E-cadherin,vimentin and α-SMA protein were measured by immunofluorescence microscopy and Western blot assay.Western blot was used to detect the expression levels of TGF-β1.Real-time PCR to detect mRNA expression of TGF-β1,Ⅰand collagen type Ⅲ.Results The primary epithelial cells of intrahepatic bile ducts were successfully cultured.In TGF-β1 group,the protein expression of vimentin,α-SMA and the mRNA expression levels ofⅠand collagen type Ⅲ were significantly higher than that of normal control group(P0.05),TGF-β1 protein and mRNA expressions were significantly lower as compared with that of normal TGF-β1 group(P<0.05).Conclusions 5-FU could inhibit TGF-β1-induced biliary epithelial-mesenchymal transition with down-regulated expression of TGF-β1.
ABSTRACT
Objective To establish the method of combined hepatocyte growth factor (HGF) with epidermal growth factor (EGF) cultured human gallbladder epithelial cells(HGBECs) in vitro .Methods The epithelial layer was peeled away from human gallbladder ,epithelial layer were digested with collagenase Ⅳ and scraped repeatedly .HGBECs were isolated and seeded in cell cul-ture plates containing medium supplemented with or without 10 ng/mL EGF or with 10 ng/mL HGF and 10 ng/mL EGF respec-tively .Then the morphologic changes of the cells were observed and taken photos with inverted phase contrast microscope ,and counted number of cells ,MTT assay detected vigor of cells in different groups .Results The number of the HGBECs of the HGF+EGF group was obviously more than the EGF group ,the duration of the HGBECs of the HGF+ EGF group was obviously longer than the EGF group(19 .3 ± 2 .5)d vs .(14 .2 ± 2 .4)d ,P< 0 .05 .And the HGBECs of the group with HGF+ EGF had better cell vigor .Conclusion HGF combines with EGF added to medium can obviously promote the proliferation of HGBECs and prolong the duration and stabilize morphology of HGBECs in vitro .