TNFalpha is a pleiotropic cytokine that induces either cell proliferation or cell death. Inhibition of NF-kappaB activation increases susceptibility to TNFalpha-induced death, concurrent with sustained JNK activation, an important contributor to the death response. Sustained JNK activation in NF-kappaB-deficient cells was suggested to depend on reactive oxygen species (ROS), but how ROS affect JNK activation was unclear. We now show that TNFalpha-induced ROS, whose accumulation is suppressed by mitochondrial superoxide dismutase, cause oxidation and inhibition of JNK-inactivating phosphatases by converting their catalytic cysteine to sulfenic acid. This results in sustained JNK activation, which is required for cytochrome c release and caspase 3 cleavage, as well as necrotic cell death. Treatment of cells or experimental animals with an antioxidant prevents H(2)O(2) accumulation, JNK phosphatase oxidation, sustained JNK activity, and both forms of cell death. Antioxidant treatment also prevents TNFalpha-mediated fulminant liver failure without affecting liver regeneration.

Inflammation may underlie the metabolic disorders of insulin resistance and type 2 diabetes. IkappaB kinase beta (IKK-beta, encoded by Ikbkb) is a central coordinator of inflammatory responses through activation of NF-kappaB. To understand the role of IKK-beta in insulin resistance, we used mice lacking this enzyme in hepatocytes (Ikbkb(Deltahep)) or myeloid cells (Ikbkb(Deltamye)). Ikbkb(Deltahep) mice retain liver insulin responsiveness, but develop insulin resistance in muscle and fat in response to high fat diet, obesity or aging. In contrast, Ikbkb(Deltamye) mice retain global insulin sensitivity and are protected from insulin resistance. Thus, IKK-beta acts locally in liver and systemically in myeloid cells, where NF-kappaB activation induces inflammatory mediators that cause insulin resistance. These findings demonstrate the importance of liver cell IKK-beta in hepatic insulin resistance and the central role of myeloid cells in development of systemic insulin resistance. We suggest that inhibition of IKK-beta, especially in myeloid cells, may be used to treat insulin resistance.

IkappaB kinase beta (IKKbeta), required for NF-kappaB activation, links chronic inflammation with carcinogenesis. We investigated whether IKKbeta is involved in chemically induced liver cancer, a model not involving overt inflammation. Surprisingly, mice lacking IKKbeta only in hepatocytes (Ikkbeta(Deltahep) mice) exhibited a marked increase in hepatocarcinogenesis caused by diethylnitrosamine (DEN). This correlated with enhanced reactive oxygen species (ROS) production, increased JNK activation, and hepatocyte death, giving rise to augmented compensatory proliferation of surviving hepatocytes. Brief oral administration of an antioxidant around the time of DEN exposure blocked prolonged JNK activation and compensatory proliferation and prevented excessive DEN-induced carcinogenesis in Ikkbeta(Deltahep) mice. Decreased hepatocarcinogenesis was also found in mice lacking IKKbeta in both hepatocytes and hematopoietic-derived Kupffer cells. These mice exhibited reduced hepatocyte regeneration and diminished induction of hepatomitogens, which were unaltered in Ikkbeta(Deltahep) mice. IKKbeta, therefore, orchestrates inflammatory crosstalk between hepatocytes and hematopoietic-derived cells that promotes chemical hepatocarcinogenesis.

The proinflammatory cytokine tumor necrosis factor (TNF) alpha signals both cell survival and death. The biological outcome of TNFalpha treatment is determined by the balance between NF-kappaB and Jun kinase (JNK) signaling; NF-kappaB promotes survival, whereas JNK enhances cell death. Critically, identity of a JNK substrate that promotes TNFalpha-induced apoptosis has been outstanding. Here we show that TNFalpha-mediated JNK activation accelerates turnover of the NF-kappaB-induced antiapoptotic protein c-FLIP, an inhibitor of caspase-8. This is not due to direct c-FLIP phosphorylation but depends on JNK-mediated phosphorylation and activation of the E3 ubiquitin ligase Itch, which specifically ubiquitinates c-FLIP and induces its proteasomal degradation. JNK1 or Itch deficiency or treatment with a JNK inhibitor renders mice resistant in three distinct models of TNFalpha-induced acute liver failure, and cells from these mice do not display inducible c-FLIP(L) ubiquitination and degradation. Thus, JNK antagonizes NF-kappaB during TNFalpha signaling by promoting the proteasomal elimination of c-FLIP(L).

Variants of NOD2, an intracellular sensor of bacteria-derived muramyl dipeptide (MDP), increase susceptibility to Crohn's disease (CD). These variants are thought to be defective in activation of nuclear factor kappaB (NF-kappaB) and antibacterial defenses, but CD clinical specimens display elevated NF-kappaB activity. To illuminate the pathophysiological function of NOD2, we introduced such a variant to the mouse Nod2 locus. Mutant mice exhibited elevated NF-kappaB activation in response to MDP and more efficient processing and secretion of the cytokine interleukin-1beta (IL-1beta). These effects are linked to increased susceptibility to bacterial-induced intestinal inflammation and identify NOD2 as a positive regulator of NF-kappaB activation and IL-1beta secretion.