Switch from Mnt-Max to Myc-Max induces p53 and cyclin D1 expression and apoptosis during cholestasis in mouse and human hepatocytes

Abstract

Background and rationale Toxic bile acids induce hepatocyte apoptosis for which p53 and cyclin D1 have been implicated as underlying mediators. Both p53 and cyclin D1 are targets of c-Myc, which is also up-regulated in cholestasis. Myc and Mnt use Max as a cofactor for DNA binding. Myc-Max typically activates transcription via E-box binding. Mnt-Max also binds E-box sequence but serves as a repressor and inhibits the enhancer activity of Myc-Max. The current work tested the hypothesis that the switch from Mnt-Max to Myc-Max is responsible for p53 and cyclin D1 up-regulation and apoptosis during cholestasis. Results Following common bile duct ligation or left hepatic bile duct ligation, the expression of p53, c-Myc and cyclin D1 increased markedly while Mnt expression decreased. Nuclear binding activity of Myc to E-box element of p53 and cyclin D1 increased, while that of Mnt decreased in a time-dependent fashion. Lithocolic acid (LCA) treatment of primary human hepatocytes and HuH-7 cells induced a similar switch from Mnt to Myc, increased p53 and cyclin D1 promoter activity, endogenous p53 and cyclin D1 expression and apoptosis. Blocking c-Myc induction in HuH-7 cells prevented the LCA-mediated increase in p53 and cyclin D1 expression and reduced apoptosis. Lowering Mnt expression further enhanced LCA’s inductive effect on p53 and cyclin D1. Bile duct ligated mice treated with lentivirus harboring c-myc siRNA were protected from hepatic induction of p53 and cyclin D1, switch in Mnt to Myc nuclear binding to E-box, and hepatocyte apoptosis. Conclusions The switch from Mnt to Myc during BDL and in hepatocytes treated with LCA is responsible for the induction in p53 and cyclin D1 expression and contributes to apoptosis.

Activation of a Novel c-Myc-miR27-Prohibitin 1 Circuitry in Cholestatic Liver Injury Inhibits Glutathione Synthesis in Mice

“…In the same model, c-Myc protein levels increased by 8 h and remained elevated for approximately 48 h (32). We first examined whether these changes also occur after in vivo LCA treatment.…”

“…The in vitro model recapitulates BDL-induced changes in c-Myc, GCL subunits, Nrf2 ARE binding, and effects of UDCA and SAMe treatments (31)(32)(33) and allows us to dissect the molecular mechanisms responsible more easily. The in vivo models serve to confirm changes that are due to higher levels of toxic bile acids rather than due to increased ductal pressure from ligating the bile duct.…”

“…However, another potential mechanism not yet investigated is the role of c-Myc, which is also induced after BDL (32). Although c-Myc has been shown to activate both GCL subunits at the transcriptional level (3), recent work from Levy and Forman showed it interacts with Nrf2 and negatively regulates AREdependent genes by destabilizing Nrf2 (16).…”

MicroRNA-mediated regulation of glutathione and methionine metabolism and its relevance for liver disease

“…This contradictory finding led us to identify the participation of other proteins in this process using biotinylated ARE in pull-down assays, followed by proteomic analysis. C-Myc, which we had shown previously to be induced during BDL [54] and prohibitin 1 (PHB1), a well-known mitochondrial chaperone protein [55], also play important roles in modulating Nrf2-mediated ARE-dependent gene expression. Interestingly, c-Myc is thought to induce PHB1 at the transcriptional level but we found PHB1 expression fell despite the induction of c-Myc during cholestasis [53].…”

Bile Acids Induce Inflammatory Genes in Hepatocytes

“…9 -11 In these studies, deoxycholic acid (DCA), lithocholic acid, and their metabolites are frequently used. [11][12][13] One caveat of these studies, however, is that plasma and liver concentrations of secondary bile acids (eg, lithocholic acid and DCA) do not dramatically increase in humans with cholestasis or in animal models of cholestasis, 14 -16 because these bile acids are formed in the intestine from primary bile acids, and intestinal concentrations of primary bile acids decrease during cholestasis because of diminished biliary excretion. 14,15 On the other hand, serum levels of the most abundant primary bile acids, such as cholic acid and muricholic acid, increase to 200 -250 mol/L in mice subjected to bile duct ligation (BDL) for 7 days.…”