Transplantation/ImmunologyLPS-Induced Epithelial-Mesenchymal Transition of Intrahepatic Biliary Epithelial Cells
Introduction
Repeated biliary infection is a characteristic feature of a range of chronic inflammatory liver diseases including hepatolithiasis (HL) and biliary stricture or obstruction. Biliary chronic inflammation plays an important role in liver fibrosis or even cirrhosis. The causative organism of biliary infection is often gram-negative bacteria, which could release endotoxin and induce inflammatory reaction. Studies have showed that when the IBECs are exposed to a high level of lipopolysaccharide during bacterial infections, IBECs often show cellular damage and pathologic proliferation. This response of IBECs plays a role in the progression of liver fibrogenesis or even cirrhosis [1].
Although the activation of hepatic stellate cells remains a central event in liver fibrosis, the role of epithelial-mesenchymal transition (EMT) of both hepatocytes and cholangiocytes in liver fibrosis is attracting great attentions. EMT has been implicated in a variety of biological processes such as fibrogenesis, embryonic development, and tumor progression. Recent clinical and animal studies have demonstrated that bile duct epithelial cells can undergo EMT, thereby contributing to hepatic fibrosis 2, 3, 4, 5. Consistent with these studies, our previous retrospective clinical study shown that we observed the stimulation of the TGF-β1/smad2/3 signaling pathway followed by the loss of epithelial markers and the acquirement of mesenchymal markers in bile duct epithelial cells from patients with primary hepatolithiasis.
Transforming growth factor beta1 (TGF-β1) is known to be the most potent inducer of EMT, and it initiates morphological transition of the cells from an epithelial to a fibroblastic appearance, accompanied by a loss of epithelial cell markers such as E-cadherin and a gain of mesenchymal cell markers such as vimentin, fibronectin, and N-cadherin 6, 7 However, how the TGF-β1 is induced in chronic liver diseases remains unclear. As infection is a common pathogenic process of chronic liver disease that mainly involves bile duct injury, and also participates in liver fibrosis, we hypothesized that biliary infection is related to the induction of TGF-β1-mediated EMT and subsequent fibrogenesis. Therefore, in this study, primary IBECs were cultured with LPS to investigate whether these can activate TGFβ/Smads signaling pathway and induce EMT.
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Antibodies and Reagents
Antibodies and reagents included anti-TGF-β1, anti-E-cadherin, anti-S100A4, anti-p-Smad 2/3, and anti-α-SMA (DAKO, Glostrup, Denmark); Smad2/3 siRNA and control siRNA (Santa Cruz Technology, Santa Cruz, CA); paclitaxel and LPS (Sigma Chemical Co., St. Louis, MO, USA); Takara PrimeScript TMRT-PCR Kit (Takara Biotechnology Co., Dalian, China); RIMP1640 (Sigma Chemical Co.); and Trizol (Invitrogen, Carlsbad, CA, USA).
Cell Culture
Human intrahepatic Biliary Epithelial Cells (HIBEpiCs) were purchased from the
Effects of LPS on TGF-β1 mRNA Expression by HIBEpiCs
Twenty-four hours after the administration of LPS, marked TGF-β1 mRNA was detected. The expression of TGF-β1 mRNA reached the peak at 48 h, but the mRNA level decreased after 72 h. These results suggest that the expression of TGF-β1 was induced by LPS and reached a peak value at 48 h (Fig. 1 ∗P < 0.01; ∗∗P < 0.05). Therefore, we detected the expression of proteins involved in the EMT process at 48 and 72 h in the following experiments.
LPS-induced expression of E-cadherin, S100A4, and α-SMA in HIBEpiCs
Resting HIBEpiCs showed epithelioid cobblestone morphology,
Discussion
In this study, stimulation with LPS causes a TGFβ/Smads-mediated EMT of HIBEpiCs. These results indicated that biliary infection played a potential role in inducing EMT of bile duct epithelial cells and subsequent peribiliary fibrosis. Biliary infection is an independent risk factor for liver injury and fibrosis. However, the exact molecular mechanism remains unclear. Bacteria, mainly gram-negative bacteria, retrograde into the biliary tract through cholangiopancreatic ampulla. LPS, endotoxin
Acknowledgments
The authors acknowledge support for this work by the Science and Technology Foundation of Guizhou Province (no. 2009GZ14798).
References (16)
- et al.
Portal tract fibrogenesis in the liver
Lab Invest
(2004) - et al.
Hepatocyte growth factor attenuates liver fibrosis induced by bile duct ligation
Am J Pathol
(2006) - et al.
Cholangiocytes with mesenchymal features contribute to progressive hepatic fibrosis of the polycystic kidney rat
Am J Pathol
(2007) - et al.
Epithelial-mesenchymal transition contributes to portal tract fibrogenesis during human chronic liver disease
Lab Invest
(2008) Epithelial to mesenchymal transition in renal fibrogenesis: Pathologic significance, molecular mechanism, and therapeutic intervention
J Am Soc Nephrol
(2004)- et al.
Paclitaxel interrupts TGF-β 1 signaling between gallbladder epithelial cells and myofibroblasts
J Surg Res
(2007) - et al.
Smad proteins and transforming growth factor-β signaling
Kidney Int Suppl
(2000) - et al.
Chronic allograft nephropathy: Intraepithelial signals generated by transforming growth factor-β and bone morphogenetic protein-7
Am J Transplant
(2006)
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These authors contributed equally to this work.