GSK3β/NF-κB dependent transcriptional regulation of homeostatic hepatocyte Tnf production.

BACKGROUND & AIMS:Maintenance of hepatocyte homeostasis plays an important role in mediating the pathogenesis of many diseases. A growing body of literature has established a critical role played by TNFα in maintaining hepatocyte homeostasis; however, the transcriptional mechanisms underlying constitutive Tnf expression is unknown. METHODS:Whole liver fractions and primary hepatocytes from adult control C57BL/6 mice and the murine hepatocyte cell line, AML12, was assessed for constitutive Tnf expression. Impacts of GSK3β and NF-κB inhibition on constitutive Tnf expression was assessed in AML12 cells. Finally, AML12 cell proliferation following GSK3β and NF-κB inhibition was evaluated. RESULTS:Constitutive Tnf gene expression is present in whole liver, primary hepatocytes and cultured AML12 hepatocytes. Cytokine induced Tnf gene expression is regulated by NF-κB activation. Pharmacologic inhibition of GSK3β resulted in a time- and dose-dependent inhibition of Tnf gene expression. GSK3β inhibition decreased nuclear levels of the NF-κB subunits p65 and p50. We determined that NF-κB transcription factor subunit p65 binds to consensus sequence elements present in the murine TNFα promoter and inhibition of GSK3β decreases binding and subsequent Tnf expression. Finally, AML12 cell growth was significantly reduced following GSK3β and NF-κB inhibition. CONCLUSION:These results demonstrate that GSK3β and NF-κB are essential for mediating Tnf expression and constitutive hepatocyte cell growth. These findings add to a growing body of literature on TNFα mediated hepatocyte homeostasis and identify novel molecular mechanisms involved in mediating response to various disease states in the liver.