Genetic inactivation of ReIA/p65 sensitizes adult mouse hepatocytes to TNF-induced apoptosis in vivo and in vitro

Background & Aims: The transcription factor nuclear factor (NF)-kappa B plays a critical role in mediating survival of hepatocytes in response to tumor necrosis factor (TNF)-alpha during development because mice deficient for the NF-kappa B subunit ReIA/p65 die in utero because of TNF-induced liver apoptosis. For the adult liver, conflicting concepts exist as to whether soluble TNF can trigger apoptosis when NF-kappa B activation is impaired. By creating a mouse model in which the transactivating NF-kappa B subunit ReIA/p65 can be genetically inactivated in hepatocytes using the Cre/lox system, we sought to clarify the role of NF-kappa B in TNF-mediated hepatocyte apoptosis. Methods: Deletion of RelA/p65 in the liver was achieved using an inducible conditional knockout system (rela(F/F)MxCre mice) or, hepatocyte-specifically, using a developmental conditional knockout system (rela(F/F)AlbCre mice). Results: Disruption of RelA/p65 rendered mice sensitive to lethal liver injury upon TNF administration. Primary RelA/p65-deficient hepatocytes showed no NF-kappa B activation and undergo rapid apoptosis after TNF treatment. In contrast, hepatocytes deficient for I kappa B-kinase beta (IKK beta), displayed residual NF-kappa B activity and consecutively only mild apoptosis in response to TNF. TNF-induced apoptosis in RelA/p65-deficient hepatocytes was accompanied by prolonged activation of c-jun activating kinase (JNK) and rapid, largely proteasome-independent elimination of the long splice form of the antiapoptotic cellular FLICE inhibitor protein (c-FLIPL). Gene silencing of caspase-8, caspase-inhibitors, inhibition of JNK, or administration of antioxidants inhibited apoptosis and elimination of c-FLIPL. Conclusions: RelA/p65 is essential for TNF-induced NF-kappa B activation in adult hepatocytes. Genetic deletion of a functional RelA/p65 sensitizes these cells to apoptosis in response to soluble TNF in vivo and in vitro.