59

ACTA2

BACKGROUND: The epithelial-mesenchymal transition (EMT) has been implicated as a key mechanism in the pathogenesis of liver fibrosis. The miR-200 family has been shown to inhibit EMT. METHODS: Liver fibrosis levels were assessed with Masson's trichrome staining of liver samples obtained from biliary atresia (BA) patients. The expressions of cytokeratin-7 (CK-7) and alpha-smooth muscle actin (alpha-SMA) in the liver sections were detected by immunohistochemical and immunofluorescent staining. EMTs were induced by transforming growth factor (TGF)-beta1 in human biliary epithelial cells (BECs) in vitro. RESULTS: We showed that the EMT-related proteins CK-7 and alpha-SMA colocalized to the intrahepatic BECs in the liver sections of patients with BA. The level of alpha-SMA expression was related to liver fibrosis stage in BA. EMT in primary human intrahepatic BECs was induced by TGF-beta1 in vitro. miR-200b is one member of the miR-200 family and significantly inhibited TGF-beta1-mediated EMT in BECs. CONCLUSION: Together, these data suggest that the occurrence of EMT in BECs might contribute to BA fibrosis. miR-200b significantly affects the development and progression of TGF-beta1-dependent EMT and fibrosis in vitro.

PURPOSE: epithelial mesenchymal transition (EMT) plays a central role in the development of fibrotic complications of the lens. The current study is designed to check whether EMT of lens epithelial cells (LECs) is regulated by epigenetic modifications and to evaluate the effect of Trichostatin-A (TSA) on the transforming growth factor-beta (TGF-beta)-induced EMT. METHODS: Fetal human LECs (FHL124) were treated with TGF-beta2 in the presence or absence of TSA. Levels of mRNA, protein, as well as localization of alpha-smooth muscle actin (alphaSMA) were studied along with migration of LECs. Acetylation of histone H4 was analyzed and chromatin immunoprecipitation (ChIP) was carried out to study the level of acetylated histone H4 at the promoter of alphaSMA gene (ACTA2). Student's t-test was used for statistical analysis. RESULTS: TGF-beta2 treatment resulted in myofibroblast-like changes and increased migratory capacity of FHL124. Protein and mRNA expression of alphaSMA increased, and immunofluorescence revealed presence of extensive stress fibers. TSA treatment preserved epithelial morphology, retarded cell migration, and abrogated an increase in alphaSMA levels. TSA led to the accumulation of acetylated histone H4 that was reduced on TGF-beta2 treatment. However, increased level of histone H4 acetylation was found at the ACTA2 promoter region during TGF-beta treatment. CONCLUSIONS: The increased level of alphaSMA, a hallmark of EMT in LECs, is associated with increased level of histone H4 acetylation at its promoter region, and TSA helps in suppressing EMT by epigenetically reducing this level. TSA thus shows promising potential in management of fibrotic conditions of the lens.