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B cell activating factor (BAFF) signals through BAFF-R to promote mature B cell survival. Recent analyses of BAFF-induced signaling revealed direct association between augmented B cell metabolic fitness and activation of Akt, one of the key regulators of cell survival. The strongest and most reproducible induction of Akt occurs with significant delay (24 h) after BAFF treatment, where it precedes activation of anabolism. It was also recently shown that BAFF induces sustained Erk activation and increased turnover of the proapoptotic molecule Bim. Here we show that these BAFF-induced signaling pathways are mediated by BAFF-R and represent previously unknown arms of I kappa B kinase (IKK)1-dependent signaling. In combination with the known role of IKK1 in regulating transcription of prosurvival genes, our data underscore the central role of IKK1 in coordinating multiple BAFF-R-mediated signaling pathways controlling mature B cell homeostasis.

Inflammatory destruction of intrahepatic bile ducts is a common cause of vanishing bile duct syndrome and cholestasis, often progressing to biliary cirrhosis and liver failure. However, the molecular mechanisms underlying the pathogenesis of inflammatory biliary disease are poorly understood. Here, we show that the two I[kappa]B kinases, IKK1/IKK[alpha] and IKK2/IKK[beta], display distinct collaborative and specific functions that are essential to protect the liver from cytokine toxicity and bile duct disease. Combined conditional ablation of IKK1 and IKK2, but not of each kinase alone, sensitized the liver to in vivo LPS challenge, uncovering a redundant function of the two I[kappa]B kinases in mediating canonical NF-[kappa]B signaling in hepatocytes and protecting the liver from TNF-induced failure. Unexpectedly, mice with combined ablation of IKK1 and IKK2 or IKK1 and NEMO spontaneously developed severe jaundice and fatal cholangitis characterized by inflammatory destruction of small portal bile ducts. This bile duct disease was caused by the combined impairment of canonical NF-[kappa]B signaling together with inhibition of IKK1-specific functions affecting the bile--blood barrier. These results reveal a novel function of the two I[kappa]B kinases in cooperatively regulating liver immune homeostasis and bile duct integrity and suggest that IKK signaling may be implicated in human biliary diseases. cholangitis | inflammation | liver disease | mouse models | signal transduction

Development of obesity-associated insulin resistance and diabetes mellitus type 2 has been linked to activation of proinflammatory pathways in the liver, leading to impaired insulin signal transduction. To further define the role of hepatic NF-[kappa]B activation in this process, we have analyzed glucose metabolism in mice with liver-specific inactivation of the NF-[kappa]B essential modulator gene ([NEMO.sup.L-KO] mice) exposed to a high-fat diet (HFD). These animals are protected from the development of obesity-associated insulin resistance, highlighting the importance of hepatic NF-[kappa]B activation in this context. However, hepatic NEMO deficiency synergizes with HFD in the development of liver steatosis as a consequence of decreased peroxisome proliferator-activated receptor (PPAR-[alpha]) and increased PPAR-[gamma], expression. Steatosis interacts with increased inflammation, causing elevated apoptosis in the livers of these mice under HFD. These changes result in liver tumorigenesis of [NEMO.sup.L-KO] mice under normal diet, a process that is largely aggravated when these mice are exposed to HFD. These data directly demonstrate the interaction of hepatic inflammation, dietary composition, and metabolism in the development of liver tumorigenesis. insulin sensitivity | spontaneous hepatic carcinoma | steatosis | hepatic NEMO deficiency

Natural killer-like (NK) T, regulatory T ([T.sub.R]), and memory type T cells display surface phenotypes reminiscent of activated T cells. Previously, we reported that the generation of [T.sub.R] cells and, to a lesser extent, of memory type T cells, depends on I[kappa]B kinase 2. Here, we show that T cell-specific ablation of I[kappa]B kinase 2, in addition, completely precludes NKT cell development. T cell antigen receptor (TCR)-induced signals to activate NF-[kappa]B are essential for mature T cell activation, leading us to hypothesize that this pathway could play an important role in the generation of the antigen-driven T cell subsets comprising [T.sub.R], memory type T, and NKT cells. TCR-mediated NF-[kappa]B activation critically depends on Bcl10 and PKC[theta]. By using mice deficient for these proteins, we demonstrate that the generation of [T.sub.R] and, to a lesser extent, of memory type T cells, depends on Bcl10 and PKC[theta], and therefore, most likely on NF-[kappa]B activation initiated by TCR engagement. NKT cells, on the other hand, require PKC[theta] for thymic development, whereas absence of Bcl10 leads primarily to the reduction of peripheral NKT cell numbers.

Targeted inactivation of genes in the tumor necrosis factor (TNF)/lymphotoxin (LT) ligand and receptor system has recently revealed essential roles for these molecules in lymphoid tissue development and organization. Lymphotoxin-[Alpha][Beta] (LT[Alpha][Beta])/lymphotoxin-[Beta] receptor (LT[Beta]-R) signaling is critical for the organogenesis of lymph nodes and Peyer's patches and for the structural compartmentalization of the splenic white pulp into distinct B and T cell areas and marginal zones. Moreover, an essential role has been demonstrated for TNF/p55 tumor necrosis factor receptor (p55TNF-R) signaling in the formation of splenic B lymphocyte follicles, follicular dendritic cell networks, and germinal centers. In contrast to a previously described essential role for the p55TNF-R in Peyer's patch organogenesis, we show in this report that Peyer's patches are present in both TNF and p55TNF-R knockout mice, demonstrating that these molecules are not essential for the organogenesis of this lymphoid organ. Furthermore, we show that in the absence of TNF/p55TNF-R signaling, lymphocytes segregate normally into T and B cell areas and a normal content and localization of dendritic cells is observed in both lymph nodes and Peyer's patches. However, although B cells are found to home normally within Peyer's patches and in the outer cortex area of lymph nodes, organized follicular structures and follicular dendritic cell networks fail to form. These results show that in contrast to LT[Alpha][Beta] signaling, TNF signaling through the p55TNF-R is not essential for lymphoid organogenesis but rather for interactions that determine the cellular and structural organization of B cell follicles in all secondary lymphoid tissues.

Cytokines are now recognized to play important roles in the physiology of the central nervous system (CNS) during health and disease. Tumor necrosis factor a (TNF-[alpha]) has been implicated in the pathogenesis of several human CNS disorders including multiple sclerosis, AIDS dementia, and cerebral malaria. We have generated transgenic mice that constitutively express a murine TNF-[alpha] transgene, under the control of its own promoter, specifically in their CNS and that spontaneously develop a chronic inflammatory demyelinating disease with 100% penetrance from around 3-8 weeks of age. High-level expression of the transgene was seen in neurons distributed throughout the brain. Disease is manifested by ataxia, seizures, and paresis and leads to early death. Histopathological analysis revealed infiltration of the meninges and CNS parenchyma by [CD4.sup.+] and [CD8.sup.+] T lymphocytes, widespread reactive astrocytosis and microgliosis, and focal demyelination. The direct action of TNF-[alpha] in the pathogenesis of this disease was confirmed by peripheral administration of a neutralizing anti-murine TNF-[alpha] antibody. This treatment completely prevented the development of neurological symptoms, T-cell infiltration into the CNS parenchyma, astrocytosis, and demyelination, and greatly reduced the severity of reactive microgliosis. These results demonstrate that overexpression of TNF-[alpha] in the CNS can cause abnormalities in nervous system structure and function. The disease induced in TNF-[alpha] transgenic mice shows clinical and histopathological features characteristic of inflammatory demyelinating CNS disorders in humans, and these mice represent a relevant in vivo model for their further study.

Apoptosis and necroptosis are two regulated cell deathmechanisms; however, the interaction between these cell death pathways in vivo is unclear. Here we used cerebral ischemia/reperfusion as a model to investigate the interaction between apoptosis and necroptosis. We show that the activation of RIPK1 sequentially promotes necroptosis followed by apoptosis in a temporally specific manner. Cerebral ischemia/reperfusion insult rapidly activates necroptosis to promote cerebral hemorrhage and neuroinflammation. Ripk3 deficiency reduces cerebral hemorrhage and delays the onset of neural damage mediated by inflammation. Reduced cerebral perfusion resulting from arterial occlusion promotes the degradation of TAK1, a suppressor of RIPK1, and the transition from necroptosis to apoptosis. Conditional knockout of TAK1 in microglial/infiltrated macrophages and neuronal lineages sensitizes to ischemic infarction by promoting apoptosis. Taken together, our results demonstrate the critical role of necroptosis in mediating neurovascular damage and hypoperfusion-induced TAK1 loss, which subsequently promotes apoptosis and cerebral pathology in stroke and neurodegeneration. [ABSTRACT FROM AUTHOR]

Receptor-interacting protein kinase 3 (RIPK3)-mediated necroptosis is thought to be the pathophysiologically predominant pathway that leads to regulated necrosis of parenchymal cells in ischemia-reperfusion injury (IRI), and loss of either Fas-associated protein with death domain (FADD) or caspase-8 is known to sensitize tissues to undergo spontaneous necroptosis. Here, we demonstrate that renal tubules do not undergo sensitization to necroptosis upon genetic ablation of either FADD or caspase-8 and that the RIPK1 inhibitor necrostatin-1 (Nec-1) does not protect freshly isolated tubules from hypoxic injury. In contrast, iron-dependent ferroptosis directly causes synchronized necrosis of renal tubules, as demonstrated by intravital microscopy in models of IRI and oxalate crystal-induced acute kidney injury. To suppress ferroptosis in vivo, we generated a novel third-generation ferrostatin (termed 16-86), which we demonstrate to be more stable, to metabolism and plasma, and more potent, compared with the first-in-class compound ferrostatin-1 (Fer-1). Even in conditions with extraordinarily severe IRI, 16-86 exerts strong protection to an extent which has not previously allowed survival in any murine setting. In addition, 16-86 further potentiates the strong protective effect on IRI mediated by combination therapy with necrostatins and compounds that inhibit mitochondrial permeability transition. Renal tubules thus represent a tissue that is not sensitized to necroptosis by loss of FADD or caspase-8. Finally, ferroptosis mediates postischemic and toxic renal necrosis, which may be therapeutically targeted by ferrostatins and by combination therapy. [ABSTRACT FROM AUTHOR]

Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) is recruited to the TNF receptor 1 to mediate proinflammatory signaling and to regulate TNF-induced cell death. RIPK1 deficiency results in postnatal lethality, but precisely why Ripk1-/- mice die remains unclear. To identify the lineages and cell types that depend on RIPK1 for survival, we generated conditional Ripkl mice. Tamoxifen administration to adult RosaCreERT2Ripk1f1/f1 mice results in lethality caused by cell death in the intestinal and hematopoietic lineages. Similarly, Ripkl deletion in cells of the hematopoietic lineage stimulates proinflammatory cytokine and che-mokine production and hematopoietic cell death, resulting in bone marrow failure. The cell death reflected cell-intrinsic survival roles for RIPK1 in hematopoietic stem and progenitor cells, because Vav-iCre Ripk1f1/f1 fetal liver cells failed to reconstitute hematopoi-esis in lethally irradiated recipients. We demonstrate that RIPK3 deficiency partially rescues hematopoiesis in Vav-iCre Ripk1ml mice, showing that RIPK1-deficient hematopoietic cells undergo RIPK3-mediated necroptosis. However, the Vav-iCre Ripk1f1/f1 Ripk-/- progenitors remain TNF sensitive in vitro and fail to repo-pulate irradiated mice. These genetic studies reveal that hematopoietic RIPK1 deficiency triggers both apoptotic and necroptotic death that is partially prevented by RIPK3 deficiency. Therefore, RIPK1 regulates hematopoiesis and prevents inflammation by suppressing RIPK3 activation. [ABSTRACT FROM AUTHOR]

Development of obesity-associated insulin resistance and diabetes mellitus type 2 has been linked to activation of proinflammatory pathways in the liver, leading to impaired insulin signal transduction. To further define the role of hepatic NF-κB activation in this process, we have analyzed glucose metabolism in mice with liver-specific inactivation of the NF-κB essential modulator gene (NEMOL-KO mice) exposed to a high-fat diet (HFD). These animals are protected from the development of obesity-associated insulin resistance, highlighting the importance of hepatic NF-κB activation in this context. However, hepatic NEMO deficiency synergizes with HFD in the development of liver steatosis as a consequence of decreased peroxisome proliferator-activated receptor (PPAR-α) and increased PPAR-γ expression. Steatosis interacts with increased inflammation, causing elevated apoptosis in the livers of these mice under HFD. These changes result in liver tumorigenesis of NEMOL-KO mice under normal diet, a process that is largely aggravated when these mice are exposed to HFD. These data directly demonstrate the interaction of hepatic inflammation, dietary composition, and metabolism in the development of liver tumorigenesis. [ABSTRACT FROM AUTHOR]

Development of obesity-associated insulin resistance and diabetes mellitus type 2 has been linked to activation of proinflammatory pathways in the liver, leading to impaired insulin signal transduction. To further define the role of hepatic NF-kappaB activation in this process, we have analyzed glucose metabolism in mice with liver-specific inactivation of the NF-kappaB essential modulator gene (NEMO(L-KO) mice) exposed to a high-fat diet (HFD). These animals are protected from the development of obesity-associated insulin resistance, highlighting the importance of hepatic NF-kappaB activation in this context. However, hepatic NEMO deficiency synergizes with HFD in the development of liver steatosis as a consequence of decreased peroxisome proliferator-activated receptor (PPAR-alpha) and increased PPAR-gamma expression. Steatosis interacts with increased inflammation, causing elevated apoptosis in the livers of these mice under HFD. These changes result in liver tumorigenesis of NEMO(L-KO) mice under normal diet, a process that is largely aggravated when these mice are exposed to HFD. These data directly demonstrate the interaction of hepatic inflammation, dietary composition, and metabolism in the development of liver tumorigenesis.