Wnt/β-catenin signaling plays an important role in liver development and regeneration. Its aberrant activation, however, is observed in a subset of primary hepatocellular cancers (HCCs). In the current study, we compare and contrast the tumor characteristics of HCC in the presence or absence of mutations in the β-catenin gene (CTNNB1). Frozen HCCs (n = 32), including five fibrolamellar (FL) variants, and control livers (n = 3) from Health Sciences Tissue Bank and Department of Surgery at the University of Pittsburgh Medical Center, were examined for mutations in CTNNB1, protein levels of β-catenin, tyrosine-654-phosphorylated-β-catenin (Y654-β-catenin), and glutamine synthetase (GS). Missense mutations in the exon-3 of CTNNB1were identified in 9/32 HCCs. Total β-catenin levels were higher than controls in most tumors; however, GS was exclusively increased in HCCs with mutations. Phenotypically, greater percentages of mutated HCCs showed macrovascular and microvascular invasion. Also, the tumor size was greater than double in mutated HCCs. High levels of total β-catenin protein were observed in multinodular tumors independent of β-catenin mutations. In addition, significant cases with mutations showed absence of cirrhosis. Finally, the highest levels of Y654-β-catenin were exclusively observed in fibrolamellar (FL)-HCC cases. Conclusion: Thus, HCCs that harbor missense mutations in exon-3 of CTNNB1 exhibit, histologically, a more aggressive phenotype. Also, CTNNB1 mutations might lead to HCC in the absence of cirrhosis. Finally, FL-HCC cases display a unique up-regulation of tyrosine-phosphorylated-β-catenin, suggesting robust receptor tyrosine kinase signaling in this tumor type. (HEPATOLOGY 2009.)
β‐Catenin, the downstream effector of the Wnt signaling, plays important roles in hepatic development, regeneration, and tumorigenesis. However, its role at hepatocyte adherens junctions (AJ) is relatively poorly understood, chiefly due to spontaneous compensation by γ‐catenin. We simultaneously ablated β‐ and γ‐catenin expression in mouse liver by interbreeding β‐catenin–γ‐catenin double‐floxed mice and Alb‐Cre transgenic mice. Double knockout mice show failure to thrive, impaired hepatocyte differentiation, cholemia, ductular reaction, progressive cholestasis, inflammation, fibrosis, and tumorigenesis, which was associated with deregulation of tight junctions (TJ) and bile acid transporters, leading to early morbidity and mortality, a phenotype reminiscent of progressive familial intrahepatic cholestasis (PFIC). To address the mechanism, we specifically and temporally eliminated both catenins from hepatocytes using adeno‐associated virus 8 carrying Cre‐recombinase under the thyroid‐binding globulin promoter (AAV8‐TBG‐Cre). This led to a time‐dependent breach of the blood–biliary barrier associated with sequential disruption of AJ and TJ verified by ultrastructural imaging and intravital microscopy, which revealed unique paracellular leaks around individual hepatocytes, allowing mixing of blood and bile and leakage of blood from one sinusoid to another. Molecular analysis identified sequential losses of E‐cadherin, occludin, claudin‐3, and claudin‐5 due to enhanced proteasomal degradation, and of claudin‐2, a β‐catenin transcriptional target, which was also validated in vitro. Conclusion: We report partially redundant function of catenins at AJ in regulating TJ and contributing to the blood–biliary barrier. Furthermore, concomitant hepatic loss of β‐ and γ‐catenin disrupts structural and functional integrity of AJ and TJ via transcriptional and posttranslational mechanisms. Mice with dual catenin loss develop progressive intrahepatic cholestasis, providing a unique model to study diseases such as PFIC. (H epatology 2018;67:2320‐2337).
The oxysterol receptor liver X receptor (LXR) is a nuclear receptor best known for its function in the regulation of lipid and cholesterol metabolism. LXRs, both the α and β isoforms, have been suggested as potential therapeutic targets for several cancer types. However, there was a lack of report on whether and how LXRα plays a role in the development of hepatocellular carcinoma (HCC). In the current study, we found that systemic activation of LXRα in the VP‐LXRα knock‐in ( LXRαKI ) mice or hepatocyte‐specific activation of LXRα in the VP‐LXRα transgenic mice sensitized mice to liver tumorigenesis induced by the combined treatment of diethylnitrosamine (DEN) and 3,3',5,5'‐tetrachloro‐1,4‐bis (pyridyloxy) benzene (TCPOBOP). Mechanistically, the LXRα‐ responsive up‐regulation of interleukin‐6 (IL‐6)/signal transducer and activator of transcription 3 (STAT3) signaling pathway and the complement system, and down‐regulation of bile acid metabolism, may have contributed to increased tumorigenesis. Accumulations of secondary bile acids and oxysterols were found in both the serum and liver tissue of LXRα activated mice. We also observed an induction of monocytic myeloid–derived suppressor cells accompanied by down‐regulation of dendritic cells and cytotoxic T cells in DEN/TCPOBOP‐induced liver tumors, indicating that chronic activation of LXRα may have led to the activation of innate immune suppression. The HCC sensitizing effect of LXRα activation was also observed in the c‐MYC driven HCC model. Conclusion: Our results indicated that chronic activation of LXRα promotes HCC, at least in part, by promoting innate immune suppressor as a result of accumulation of oxysterols, as well as up‐regulation of the IL‐6/Janus kinase/STAT3 signaling and complement pathways.
Hepatocyte growth factor (HGF) and β-catenin have both been implicated in the pathogenesis of hepatocellular carcinoma (HCC). Human HGF (pCMV-HGF) gene delivery induces hepatomegaly in mice as is also evident in the β-catenin transgenic mice. The present study was aimed at examining any role of β-catenin in HGF-induced hepatomegaly. Here we report that human HGF gene delivery in a 1 week study in mice leads to hepatomegaly secondary to increased hepatocyte proliferation, which is accompanied by β-catenin activation in the liver. The primary mechanism of β-catenin activation is the loss of c-Met-β-catenin association at the hepatocyte membrane leading to nuclear translocation of β-catenin. In a four-week study, albeit β-catenin activation was observed along with significant hepatomegaly, c-Met-β-catenin complex remained unchanged. In addition, increased E-cadherin-β-catenin association was evident as well. HGF gene delivery failed to induce hepatomegaly in the β-catenin liver conditional knockout mice. These data demonstrate that importance of β-catenin in HGF-induced hepatocyte proliferation in vivo, thus highlighting the cooperation of HGF and β-catenin pathways in liver growth. (Supported by RSG-03-141-01-CNE and NIH - 1RO1DK62277 to SPSM).
Smads serve as intracellular mediators of transforming growth factor beta (TGF-beta) signaling. After phosphorylation by activated type I TGF-beta receptors, Smad proteins translocate to the nucleus, where they serve as transcription factors and increase or decrease expression of TGF-beta target genes. Mice lacking one copy each of Smad2 and Smad3 suffered midgestation lethality due to liver hypoplasia and anemia, suggesting essential dosage requirements of TGF-beta signal components. This is likely due to abnormal adhesive properties of the mutant hepatocytes, which may result from a decrease in the level of the beta1-integrin and abnormal processing and localization of E-cadherin. Culture of mutant livers in vitro revealed the existence of a parallel developmental pathway mediated by hepatocyte growth factor (HGF), which could rescue the mutant phenotype independent of Smad activation. These pathways merge at the beta1-integrin, the level of which was increased by HGF in the cultured mutant livers. HGF treatment reversed the defects in cell proliferation and hepatic architecture in the Smad2(+/-); Smad3(+/-) livers.
