[Show abstract][Hide abstract] ABSTRACT: NAFLD is the most common liver disease worldwide but it is the potential evolution to NASH and eventually to hepatocellular carcinoma (HCC), even in the absence of cirrhosis, that makes NAFLD of such clinical importance. Aim: we aimed to create a mouse model reproducing the pathological spectrum of NAFLD and to investigate the role of possible co-factors in promoting HCC. Methods: mice were treated with a choline-deficient L-amino-acid-defined-diet (CDAA) or its control (CSAA diet) and subjected to a low-dose i.p. injection of CCl4 or vehicle. Insulin resistance was measured by the euglycemic-hyperinsulinemic clamp method. Steatosis, fibrosis and HCC were evaluated by histological and molecular analysis. Results: CDAA-treated mice showed peripheral insulin resistance at 1 month. At 1-3 months, extensive steatosis and fibrosis were observed in CDAA and CDAA+CCl4 groups. At 6 months, equal increase in steatosis and fibrosis was observed between the two groups, together with the appearance of tumor. At 9 months of treatment, the 100% of CDAA+CCl4 treated mice revealed tumor versus 40% of CDAA mice. Insulin-like Growth Factor-2 (IGF-2) and Osteopontin (SPP-1) were increased in CDAA mice versus CSAA. Furthermore, Immunostaining for p-AKT, p-c-Myc and Glypican-3 revealed increased positivity in the tumors. Conclusions: the CDAA model promotes the development of HCC from NAFLD-NASH in the presence of insulin resistance but in the absence of cirrhosis. Since this condition is increasingly recognized in humans, our study provides a model that may help understanding mechanisms of carcinogenesis in NAFLD.
PLoS ONE 05/2014; 9(5):e97136. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cholangiocarcinoma (CCA) is a rare malignancy of the liver, arising from bile ducts. The incidence is increasing worldwide, but the prognosis has remained dismal and virtually unchanged in the past 30 years. Although several risk factors have been associated with the development of this cancer, none of them are normally identified in most patients. Diagnosis in advanced stages of the disease and limited therapeutic options contribute to poor survival rates. The recent analysis of genetic and epigenetic alterations occurring in CCA has shed new light in the understanding of the molecular mechanisms leading to the malignant transformation of biliary cells. Further studies in this direction may foster new diagnostic, prognostic and therapeutic approaches. This review provides a global overview of recent advances in CCA and describes the most important genetic mutations and epigenetic alterations so far reported in CCA.
Annals of translational medicine. 10/2013; 1(3):28.
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND/AIMS: Cholangiocyte proliferation plays a role in the progression of cholangiopathies, in particular in primary sclerosing cholangitis. The mechanisms regulating cholangiocyte proliferation are still undefined. Pancreatic Duodenal Homeobox protein 1 (PDX-1) is expressed by reactive cholangiocytes. In the adult pancreas PDX-1 regulates the proliferative response to injury of ductal cells. Its effects can be counteracted by Hairy and enhancer of split 1 (Hes-1). We aimed to verify whether PDX-1/Hes-1 interactions regulate cholangiocyte proliferation in response to injury. METHODS: The effect of the loss of PDX-1 on cholangiocyte proliferation was studied in vitro. In vivo PDX-1-heterozygous (+/-) mice were subjected to either DDC feeding (a model of sclerosing cholangitis) or to bile duct ligation (BDL). PDX-1/Hes-1 interactions on cell proliferation were determined by exposure to All-trans Retinoic Acid (At-RA), an inductor of Hes-1. RESULTS: In vitro, cholangiocyte proliferation was undetectable in cells pre-treated with PDX-1 siRNA. In vivo, increases in bile duct mass and collagen deposition observed after DDC feeding or BDL were significantly reduced in PDX-1(+/-) mice. Hes-1 expression is reduced in proliferating cholangiocytes; At-RA induced a dose-dependent increase in Hes-1 and a decrease in PDX-1 expression. At-RA neutralized the increases in PDX-1 expression and cell proliferation, both in vitro and in vivo in DDC mice. PDX-1 is overexpressed and Hes-1 downregulated in cholangiocytes isolated from PSC livers. CONCLUSION: Hes-1 downregulation allows PDX-1 to act as a major determinant of cholangiocyte proliferation in response to cholestatic injury. These findings provide novel mechanistic insights into the pathophysiology of cholangiopathies.
Journal of Hepatology 11/2012; · 9.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The role of insulin resistance in predicting virological response to therapy of chronic hepatitis C is debated. We assessed the association between basal (defined as homeostasis model assessment of insulin resistance (HOMA-IR)>2) and post-load insulin resistance (as oral glucose insulin sensitivity index<9.8 mg/kg/min) with the rapid and sustained virological responses in chronic hepatitis C.
Observational prospective study of 124 treatment-naïve patients with chronic hepatitis C not fulfilling the metabolic syndrome criteria, adherent to a standard treatment with pegylated interferon alpha plus ribavirin.
Insulin resistance was detected in 50% (by HOMA-IR) and 29% (by oral glucose insulin sensitivity index) of patients. Independent predictors of rapid virologic response were hepatitis C virus (HCV) genotype 2 (odds ratio 5.66; 95% confidence interval 1.88-17.01), HCV genotype 3 (odds ratio 5.23; 95% confidence interval 1.84-14.84) and lower basal ferritin levels (odds ratio 0.99; 95% confidence interval 0.993-0.998). Independent predictors of sustained virologic response were HCV genotype 2 (odds ratio 19.54; 95% confidence interval 2.29-166.41) and HCV genotype 3 (odds ratio 3.24; 95% confidence interval 1.10-9.58). Rapid virologic response was by itself predictive of sustained virologic response (odds ratio 40.90; 95% confidence interval 5.37-311.53).
Insulin resistance, measured by both static and dynamic methods, does not predict rapid or sustained virologic response in chronic hepatitis C patients without the metabolic syndrome.
Digestive and Liver Disease 01/2012; 44(5):419-25. · 2.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cholangiocarcinoma cells over-express oestrogen receptor-β, which displays anti-proliferative and pro-apoptotic effects.
To evaluate the effects of a newly developed and highly selective oestrogen receptor-β agonist (KB9520) on experimental intrahepatic cholangiocarcinoma.
In vitro, the effects of KB9520 on apoptosis and proliferation of HuH-28 cells, HuH-28 cells with selective oestrogen receptor-β silencing (by small interfering RNA), HepG2 cells (oestrogen receptor-α and oestrogen receptor-β negative) and HepER3 cells (HepG2 cells transformed to stably express oestrogen receptor-α) were evaluated. In vivo, the effects of KB9520 on experimental intrahepatic cholangiocarcinoma, induced by thioacetamide administration were tested.
In vitro, KB9520 induced apoptosis and inhibited proliferation of HuH-28 cells. KB9520 effects were absent in cells lacking oestrogen receptor-α and β (HepG2) and in cells expressing only oestrogen receptor-α (HepER3); its pro-apoptotic effect was impaired in cells where oestrogen receptor-β expression was decreased by specific small interfering RNA. In vivo, KB9520 inhibited experimental intrahepatic cholangiocarcinoma development in thioacetamide-treated rats and promoted tumour regression in rats where tumour was already established. In treated animals, tumour areas showed reduced proliferation but increased apoptosis.
KB9520 induced apoptosis in cholangiocarcinoma by selectively acting on oestrogen receptor-β, suggesting that oestrogen receptor-β selective agonists may be a novel and effective therapeutic option for the medical treatment of intrahepatic cholangiocarcinoma.
Digestive and Liver Disease 07/2011; 44(2):134-42. · 2.89 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To study the mechanism(s) linking insulin resistance (IR) to hepatic fibrosis and the role of the epithelial component in tissue repair and fibrosis in chronic hepatitis C (CHC).
Prospective observational study.
Tertiary care academic centre.
78 consecutive patients with CHC.
IR, calculated by the oral glucose insulin sensitivity during oral glucose tolerance test; necroinflammatory activity and fibrosis, defined according to Ishak's score; steatosis, graded as 0 (<5% of hepatocytes), 1 (5-33%), 2 (33-66%) and 3 (>66%). To evaluate the role of the epithelial component in tissue repair and fibrosis, the expansion of the ductular reaction (DR) was calculated by keratin-7 (CK7) morphometry. Nuclear expression of Snail, downregulation of E-cadherin and expression of fibroblast specific protein-1 (FSP1) and vimentin by CK7-positive cells were used as markers of epithelial-mesenchymal transition in DR elements.
IR, the degree of necroinflammation and expansion of the DR (stratified as reactive ductular cells (RDCs), hepatic progenitor cells and intermediate hepatobiliary cells according to morphological criteria) were all associated with the stage of fibrosis. Nuclear Snail expression, E-cadherin downregulation and vimentin upregulation were observed in RDCs. By dual immunofluorescence for CK7 and FSP1, the number of RDCs undergoing epithelial-mesenchymal transition progressively increased together with the necroinflammatory score. By multivariate analysis, total inflammation and insulin resistance were the only factors significantly predicting the presence of advanced fibrosis (Ishak score ≥3) and the expansion of RDCs.
This study indicates that IR is associated with the degree of necroinflammatory injury in CHC and contributes to hepatic fibrosis by stimulating the expansion of RDCs that express epithelial-mesenchymal transition markers.
[Show abstract][Hide abstract] ABSTRACT: Reactive cholangiocytes acquire a neuroendocrine-like phenotype, with synthesis and local release of neuropeptides and hormones. The mechanism that drives such phenotypical changes is still undefined. Pancreatic Duodenal Homeobox-1 (PDX-1) is a transcription factor required for pancreatic development, that sustains pancreatic beta-cell response to injury and insulin synthesis. PDX-1 induces neuroendocrine-like transition of pancreatic ductal cells. Cholangiocyte response to injury is modulated by Glucagon-Like Peptide-1 Receptor (GLP-1R), which, in the pancreas, activates PDX-1. We wanted to verify whether PDX-1 plays any role in cholangiocyte neuroendocrine-like transdifferentiation in response to injury.
PDX-1 expression was assessed in cholangiocytes from normal and one week bile duct ligated (BDL) rats. Changes in PDX-1 expression and activation upon GLP-1R activation were then assayed. The effects of the lack of PDX-1 in cholangiocytes were studied in vitro by siRNA and in vivo by the employment of PDX-1-deficient (+/-) mice.
BDL but not normal cholangiocytes express PDX-1. GLP-1R activation elicits, in a PI3K-dependent fashion, PDX-1 expression, together with its nuclear translocation. In vitro, GLP-1R-induced increases in VEGF and IGF-1 mRNA expression were blunted in cells with PDX-1 siRNA. In vivo, the VEGF and IGF-1 mRNA expression in the liver after one week BDL was markedly reduced in PDX-1-deficient mice, together with reduced bile duct mass.
In response to injury, reactive cholangiocytes de novo express PDX-1, the activation of which allows cholangiocytes to synthesize IGF-1 and VEGF. These findings suggest that PDX-1 drives the acquisition of the neuroendocrine-like phenotype by cholangiocytes in response to cholestatic injury.
Journal of Hepatology 10/2010; 53(4):663-70. · 9.86 Impact Factor