Mesenchymal Phenotype of Cholangiocytes Plays an Integral Role in the Regulation of Biliary Senescence and Liver Fibrosis in Cholestatic Liver Diseases
Abstract
Cholangiocytes are the epithelial cells lining the bile ducts in the liver and the target of cholestatic liver diseases, such as primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC). Cholangiopathies are characterized by hepatic inflammation, ductular reaction, liver fibrosis and, eventually, carcinogenesis. Epithelial-mesenchymal transition (EMT) is an event by which epithelial cells lose their native characteristics and display functional properties of mesenchymal cells. Vimentin is a type III intermediate filament, and induction of vimentin in epithelial cells results in several important features of EMT, including the adoption of a mesenchymal shape, loss of desmosomes and increased focal adhesion and cell motility. Secretin receptor (SR) is only expressed by cholangiocytes, which plays a key role in the regulation of biliary damage and liver fibrosis.
The overall goal of this study was to evaluate whether EMT occurs in cholangiocytes and contributes to the progression of biliary senescence and liver fibrosis in cholestatic liver diseases. Male 12-week-old Mdr2-/- mice with or without vimentin Vivo-Morpholino treatment, SR-/-mice, SR-/- /Mdr2-/-mice and their corresponding control groups were used for the in vivo studies. Staining was performed in liver sections for evaluation of (i) histological features using hematoxylin and eosin (H&E) staining; (ii) EMT by vimentin and E-cadherin; (iii) ductular reaction by CK-19; (iv) liver fibrosis by Sirius Red staining and Col1a1, and (v) biliary senescence by SA-β-gal and p16. TGF-β1 levels were measured in serum and cholangiocyte supernatants using ELISA kits. Liver specimens from human PSC patients, human intrahepatic biliary epithelial cells (HIBEpiCs), and human hepatic stellate cell lines (HHSteCs) were also used to evaluate changes in EMT.
There was increased expression of a mesenchymal phenotype of cholangiocytes in Mdr2-/- mice, which was reduced by treatment with vimentin Vivo-Morpholino and genetic depletion of SR. Concomitant with reduced vimentin expression, there was decreased liver damage, ductular reaction, biliary senescence, liver fibrosis, and TGF-β1 secretion in Mdr2-/- mice treated with vimentin Vivo-Morpholino and SR-/- /Mdr2-/- mice. Human PSC patients and derived cell lines had increased expression of vimentin and other mesenchymal markers compared to healthy controls and HIBEpiCs, respectively. In vitro silencing of vimentin in HIBEpiCs suppressed TGF-β1-induced EMT and fibrotic reaction. HHSteCs had decreased fibrotic response and increased cellular senescence after stimulation with cholangiocyte supernatant with reduced vimentin levels. Our study demonstrated that loss of vimentin or SR reduces the mesenchymal phenotype of cholangiocytes, which leads to decreased biliary senescence and liver fibrosis. Inhibition of vimentin or SR may be a key therapeutic target in the treatment of cholangiopathies including PSC.
Citation
Zhou, Tianhao (2020). Mesenchymal Phenotype of Cholangiocytes Plays an Integral Role in the Regulation of Biliary Senescence and Liver Fibrosis in Cholestatic Liver Diseases. Doctoral dissertation, Texas A&M University. Available electronically from https : / /hdl .handle .net /1969 .1 /191950.