Epithelial to mesenchymal transition in development of liver fibrosis in small-for-size fatty liver graft

Abstract

BACKGROUND AND AIMS: Although graft size and chronic liver diseases have been proposed as risk factors for post-transplantation liver fi brosis, the underlying mechanisms remain unclear. Our previous study showed that the distinct regeneration pattern featured by oval cell activation in small-for-size fatty liver graft was accompanied with prominent periportal ductular reaction (DR). As intense proliferation of cholangiocytes is associated with recruitment of fi broblasts and subsequent fi brosis, we aim to investigate whether DR involves the process of epithelial to mesenchymal transition (EMT) in this model. MATERIALS AND METHODS: A rat orthotopic liver transplantation model using either small-for-size fatty (40% fatty change) or normal grafts (as control) in cirrhotic recipients was applied. Liver and blood samples were collected at day 2, 4, 7, 14 and 28 after transplantation. Intragraft gene profi le was mined by cDNA microarray. A panel of markers either indicating or inducing EMT was examined at mRNA or protein levels. Functional studies using PIL2 oval cell line were conducted to confi rm the proliferation and differentiation of oval cells. RESULTS: More extensive ductular proliferation with stronger expression of OV-6 in the cholangiocytes was observed in small-for-size fatty grafts at day 7 and day 14 after transplantation, which was consistent with our previous fi ndings. Gene expression of EMT inhibitor, BMP7 and epithelial markers such as CK19 and E-cadherin was signifi cantly decreased at day 7 (BMP7, 1.2 vs 4.7 folds, p=0.03) or day 14 (CK19, 1.3 vs 2.3 folds, p=0.01; E-cadherin, 1.0 vs 3.5 folds, p=0.00) at mRNA levels in small-for-size fatty grafts, whereas the expression of Notch2 and profi brogenic cytokine, TGF-b1, was strikingly increased at day 4 (Notch2, 2.8 vs. 1.0 fold, p=0.00) or day 14 (TGF-b1, 2178 vs 1146 folds, p=0.03). Co-expression of CK19 and vimentin or CK19 and S100A4 in ductular cells at day 7 and day 14 was more remarkable in small-for-size fatty grafts. CONCLUSION: Reactive ductules identifi ed in small-for-size fatty liver grafts may undergo epithelial-to-mesenchymal transition and further lead to fi brosis in the course of graft regeneration. Elucidating the relationship between EMT and fi brogenesis as well as the integration of Notch/TGF-b signaling in this process may help develop novel therapeutic strategies to prevent and treat post-transplantation liver fi brosis.