Cholestasis impairs hepatic lipid storage via AMPK and CREB signaling in hepatitis B virus surface protein transgenic mice.

Clinical studies demonstrated that nonalcoholic steatohepatitis is associated with liver-related outcomes in chronic hepatitis B. Furthermore, primary biliary fibrosis and biliary atresia occurred in patients with HBV infection. Interestingly, hepatitis B virus surface protein (HBs) transgenic mice spontaneously develop hepatic steatosis. Our aim is to investigate the effect of Abcb4 knockout-induced cholestasis on liver steatosis in HBs transgenic mice. Hybrids of HBs transgenic and Abcb4-/- mice were bred on the BALB/c genetic background. Lipid synthesis, storage, and catabolism as well as proteins and genes that control lipid metabolism were analyzed using HPTLC, qPCR, western blot, electrophoretic mobility shift assay (EMSA), lipid staining, and immunohistochemistry. Hepatic neutral lipid depots were increased in HBs transgenic mice and remarkably reduced in Abcb4-/- and HBs/Abcb4-/- mice. Similarly, HPTLC-based quantification analyses of total hepatic lipid extracts revealed a significant reduction in the amount of triacylglycerols (TAG), while the amount of free fatty acids (FFA) was increased in Abcb4-/- and HBs/Abcb4-/- in comparison to wild-type and HBs mice. PLIN2, a lipid droplet-associated protein, was less expressed in Abcb4-/- and HBs/Abcb4-/-. The expression of genes-encoding proteins involved in TAG synthesis and de novo lipogenesis (Agpat1, Gpat1, Mgat1, Dgat1, Dgat2, Fasn, Hmgcs1, Acc1, Srebp1-c, and Pparγ) was suppressed, and AMPK and CREB were activated in Abcb4-/- and HBs/Abcb4-/- compared to wild-type and HBs mice. Simulating cholestatic conditions in cell culture resulted in AMPK and CREB activation while FASN and PLIN2 were reduced. A concurrent inhibition of AMPK signaling revealed normal expression level of FASN and PLIN2, suggesting that activation of AMPK-CREB signaling regulates hepatic lipid metabolism, i.e. synthesis and storage, under cholestatic condition. In conclusions, in vivo and mechanistic in vitro data suggest that cholestasis reduces hepatic lipid storage via AMPK and CREB signaling. The results of the current study could be the basis for novel therapeutic strategies as NASH is a crucial factor that can aggravate chronic liver diseases.

Hepatitis B virus surface proteins accelerate cholestatic injury and tumor progression in Abcb4-knockout mice.

Understanding of the pathophysiology of cholestasis associated carcinogenesis could challenge the development of new personalized therapeutic approaches and thus improve prognosis. Simultaneous damage might aggravate hepatic injury, induce chronic liver disease and even promote carcinogenesis. We aimed to study the effect of Hepatitis B virus surface protein (HBsAg) on cholestatic liver disease and associated carcinogenesis in a mouse model combining both impairments. Hybrids of Abcb4-/- and HBsAg transgenic mice were bred on fibrosis susceptible background BALB/c. Liver injury, serum bile acid concentration, hepatic fibrosis, and carcinogenesis were enhanced by the combination of simultaneous damage in line with activation of c-Jun N-terminal kinase (JNK), proto-oncogene c-Jun, and Signal transducer and activator of transcription 3 (STAT3). Activation of Protein Kinase RNA-like Endoplasmic Reticulum Kinase (PERK) and Eukaryotic translation initiation factor 2A (eIF2α) indicated unfolded protein response (UPR) in HBsAg-expressing mice and even in Abcb4-/- without HBsAg-expression. CONCLUSION: Cholestasis-induced STAT3- and JNK-pathways may predispose HBsAg-associated tumorigenesis. Since STAT3- and JNK-activation are well characterized critical regulators for tumor promotion, the potentiation of their activation in hybrids suggests an additive mechanism enhancing tumor incidence.