RESUMO
【Objective】 To use hairy enhancer of split 1 (Hes1) to regulate the differentiation of liver epithelial progenitor cells (LEPCs) into cholangiocytes. 【Methods】 The vectors, pTet-on and pTRE2hyg-Hes1, were transfected into LEPCs. The expression of Hes1 was induced by doxycycline (DOX) with different concentrations (0, 0.1, 1, 5, 10, 50, 100 and 500 μg/mL). The expressions of Hes1, molecular markers of hepatocyte and cholangiocyte, glutathione synthetase (Gss), keratin 19 (Krt19) and hepatic nuclear factor 1β (HNF1β) in LEPCs were verified by Western blotting, RT-PCR, Real-time PCR, immunocytochemistry and immunofluorescence. 【Results】 The expression of Hes1 in LEPCs transfected by pTet-on/pTRE2hyg-Hes1 was increased by 11.21 fold when induced by DOX at 10 ug/mL, which drove the LEPCs to differentiate into biliary epithelial cells. With increasing expression of Hes1, cholangiocyte markers, Krt19 and HNF1β, were significantly upregulated, while the hepatocyte marker, Gss, was obviously downregulated. 【Conclusion】 DOX at 10 μg/mL may induce a suitably up-regulated expression of Hes1 in LEPCs double-transfected by pTet-on and pTRE2hyg-Hes1, and the suitable high-expression rather than over-expression of Hes1 can regulate LEPCs to differentiate into cholangiocytes.