MicroRNAs are endogenous small non-coding RNAs that regulate gene expression and cancer development. A rare population of hepatocellular cancer stem cells (HSCs) holds the extensive proliferative and self-renewal potential necessary to form a liver tumour. We postulated that specific transcriptional factors might regulate the expression of microRNAs and subsequently modulate the expression of gene products involved in phenotypic characteristics of HSCs. We evaluated the expression of microRNA in human HSCs by microarray profiling, and defined the target genes and functional effects of two groups of microRNA regulated by IL-6 and transcriptional factor Twist. A subset of highly chemoresistant and invasive HSCs was screened with aberrant expressions of cytokine IL-6 and Twist. We demonstrated that conserved let-7 and miR-181 family members were up-regulated in HSCs by global microarray-based microRNA profiling followed by validation with real-time polymerase chain reaction. Importantly, inhibition of let-7 increases the chemosensitivity of HSCs to sorafenib and doxorubicin whereas silencing of miR-181 led to a reduction in HSCs motility and invasion. Knocking down IL-6 and Twist in HSCs significantly reduced let-7 and miR-181 expression and subsequently inhibited chemoresistance and cell invasion. We showed that let-7 directly targets SOCS-1 and caspase-3, whereas miR-181 directly targets RASSF1A, TIMP3 as well as nemo-like kinase (NLK). In conclusion, alterations of IL-6- and Twist-regulated microRNA expression in HSCs play a part in tumour spreading and responsiveness to chemotherapy. Our results define a novel regulatory mechanism of let-7/miR-181s suggesting that let-7 and miR-181 may be molecular targets for eradication of hepatocellular malignancies.

Background-Simvastatin is a cholesterol-lowering drug that is widely used to prevent and treat atherosclerotic cardiovascular disease. Simvastatin exhibits numerous pleiotropic effects including anti-cancer activity. However, the effect of simvastatin on cholangiocarcinoma has not been evaluated.

Cholangiocytes are the epithelial cells that line the biliary tree. In the adult liver, they are a mitotically dormant cell population, unless ductular reaction is triggered by injury. The ability of cholangiocytes to proliferate is important in many different human pathological liver conditions that target this cell type, which are termed cholangiopathies (i.e. primary biliary cirrhosis, primary sclerosing cholangitis and biliary atresia). In our article, we provide background information on the morphological and functional heterogeneity of cholangiocytes, summarize what is currently known about their proliferative processes, and briefly describe the diseases that target these cells. In addition, we address recent findings that suggest cholangiocyte involvement in epithelialtomesenchymal transformation and liver fibrosis, and propose directions for future studies.

S. Castration inhibits biliary proliferation induced by bile duct obstruction: novel role for the autocrine trophic effect of testosterone. Am J Physiol Gastrointest Liver Physiol 301: G981-G991, 2011. First published September 8, 2011; doi:10.1152/ajpgi.00061.2011.-Increased cholangiocyte growth is critical for the maintenance of biliary mass during liver injury by bile duct ligation (BDL). Circulating levels of testosterone decline following castration and during cholestasis. Cholangiocytes secrete sex hormones sustaining cholangiocyte growth by autocrine mechanisms. We tested the hypothesis that testosterone is an autocrine trophic factor stimulating biliary growth. The expression of androgen receptor (AR) was determined in liver sections, male cholangiocytes, and cholangiocyte cultures [normal rat intrahepatic cholangiocyte cultures (NRICC)]. Normal or BDL (immediately after surgery) rats were treated with testosterone or antitestosterone antibody or underwent surgical castration (followed by administration of testosterone) for 1 wk. We evaluated testosterone serum levels; intrahepatic bile duct mass (IBDM) in liver sections of female and male rats following the administration of testosterone; and secretin-stimulated cAMP levels and bile secretion. We evaluated the expression of 17␤-hydroxysteroid dehydrogenase 3 (17␤-HSD3, the enzyme regulating testosterone synthesis) in cholangiocytes. We evaluated the effect of testosterone on the proliferation of NRICC in the absence/presence of flutamide (AR antagonist) and antitestosterone antibody and the expression of 17␤-HSD3. Proliferation of NRICC was evaluated following stable knock down of 17␤-HSD3. We found that cholangiocytes and NRICC expressed AR. Testosterone serum levels decreased in castrated rats (prevented by the administration of testosterone) and rats receiving antitestosterone antibody. Castration decreased IBDM and secretinstimulated cAMP levels and ductal secretion of BDL rats. Testosterone increased 17␤-HSD3 expression and proliferation in NRICC that was blocked by flutamide and antitestosterone antibody. Knock down of 17␤-

The proliferation of cholangiocytes occurs during the progression of cholestatic liver diseases and is critical for the maintenance and/or restoration of biliary mass during bile duct damage. The ability of cholangiocytes to proliferate is important in many different human pathologic conditions. Recent studies have brought to light the concept that proliferating cholangiocytes serve as a unique neuroendocrine compartment in the liver. During extrahepatic cholestasis and other pathologic conditions that trigger ductular reaction, proliferating cholangiocytes acquire a neuroendocrine phenotype. Cholangiocytes have the capacity to secrete and respond to a variety of hormones, neuropeptides, and neurotransmitters, regulating their surrounding cell functions and proliferative activity. In this review, we discuss the regulation of cholangiocyte growth by neuroendocrine factors in animal models of cholestasis and liver injury, which includes a discussion of the acquisition of neuroendocrine phenotypes by proliferating cholangiocytes and how this relates to cholangiopathies. We also review what is currently known about the neuroendocrine phenotypes of cholangiocytes in human cholestatic liver diseases (ie, cholangiopathies) that are characterized by ductular reaction.