Interferons (IFNs)1 are a family of multifunctional cytokines that block viral infection, inhibit cell proliferation, and modulate cell differentiation. Whereas type I IFNs (IFN-␣, IFN-␤, and IFN-) bind to a common cell-surface receptor, the receptor for type II IFN (IFN-␥) is a distinct entity (1). IFNs transduce signals from the cell surface, resulting in selective gene induction through the activation of JAK tyrosine kinases and STAT transcription factors (1-3). Upon JAK-mediated tyrosine phosphorylation, the STAT proteins (STAT1, STAT2, and STAT3) dimerize and translocate to the nucleus to activate transcription of IFN-stimulated genes (3). IFN also activates the nuclear factor-B (NF-B) transcription factor in a serine/threonine kinase-dependent signaling pathway that protects cells against apoptosis (4, 5).NF-B regulates the expression of genes involved in cell signaling, stress responses, growth, survival, and apoptosis by binding to cis-acting B sites in the promoters and enhancers of these genes. Viruses, cytokines, lipopolysaccharides, and other stimulating agents promote NF-B translocation to the nucleus and DNA binding. Active DNA-binding forms of NF-B are dimeric combinations of Rel proteins (p50, p52, c-Rel, v-Rel, RelA/p65, and RelB). In most cell types, the predominant form of NF-B is the p50⅐p65 heterodimer. NF-B dimers are retained in the cytoplasm of unstimulated cells in an inactive state by the binding of a family of inhibitory IB proteins.Whereas STAT proteins play crucial roles in the transcriptional response to IFN-␣/␤ and in the induction of antiviral activity, the role of NF-B in IFN signaling and action has not been studied extensively. To identify the functional role of NF-B in IFN action, sensitivity to the antiviral effect of IFN was examined in fibroblasts that either had normal NF-B function or had a functional deletion of the IFN-induced NF-B pathway by germ line disruption of the Rel p50 and p65 proteins. Antiviral assays demonstrated that NF-B knockout (KO) fibroblasts were sensitized to the antiviral action of IFN-␤. To determine the relationship between gene regulation by IFN-␤ and antiviral activity, microarray analysis was performed on RNA samples collected from IFN-treated murine fibroblasts. Microarray analysis identified several classical IFN-stimulated genes (ISGs) involved in the antiviral action of IFN. Quantitative real-time PCR demonstrated that, whereas the IFN-induced expression of some ISGs was enhanced in NF-B KO cells relative to mouse wild-type fibroblasts, the IFN-induced expression of other ISGs was lower in NF-B KO cells. Our results demonstrate the distinctive role of NF-B in the regulation of ISGs and in the induction of antiviral activity. Thus, the IFN-activated NF-B pathway not only counterbalances apoptosis, but also regulates the expression of ISGs and the induction of antiviral activity. EXPERIMENTAL PROCEDURESBiological Reagents and Cell Culture-Recombinant Chinese hamster ovary cell-expressed rat IFN-␤ was obtained from Biogen Idec, Inc. (6). The b...

Variation in responses to pathogens is influenced by exposure history, environment and the host's genetic status. We recently demonstrated that human leukocyte antigen class II allelic differences are a major determinant of the severity of invasive group A streptococcal (GAS) sepsis in humans. While in-depth controlled molecular studies on populations of genetically wellcharacterized humans are not feasible, it is now possible to exploit genetically diverse panels of recombinant inbred BXD mice to define genetic and environmental risk factors. Our goal in this study was to standardize the model and identify genetic and nongenetic covariates influencing invasive infection outcomes. Despite having common ancestors, the various BXD strains (n strains ¼ 33, n individuals ¼ 445) showed marked differences in survival. Mice from all strains developed bacteremia but exhibited considerable differences in disease severity, bacterial dissemination and mortality rates. Bacteremia and survival showed the expected negative correlation. Among nongenetic factors, age -but not sex or weight -was a significant predictor of survival (P ¼ 0.0005). To minimize nongenetic variability, we limited further analyses to mice aged 40-120 days and calculated a corrected relative survival index that reflects the number of days an animal survived post-infection normalized to all significant covariates. Genetic background (strain) was the most significant factor determining susceptibility (Pp0.0001), thus underscoring the strong effect of host genetic variation in determining susceptibility to severe GAS sepsis. This model offers powerful unbiased forward genetics to map specific quantitative trait loci and networks of pathways modulating the severity of GAS sepsis.

Several studies suggest that TNF-alpha contributes to the development of insulin resistance (IR). We compared transcriptional profiles of rat H-411E liver cells exposed to insulin in the absence or presence of TNF-alpha. We identified 33 genes whose expression was altered by insulin, and then reversed by TNF-alpha. Twenty-six of these 33 genes created a single network centered around: insulin, TNF-alpha, p38-MAPK, TGFb1; SMAD and STAT1; and enzymes and cytokines involved in apoptosis (CASP3, GADD45B, IL2, TNF-alpha, etc.). We analyzed our data together with other data of gene expression in adipocytes and found a number of processes common to both, for example, cell death and inflammation; intercellular signaling and metabolism; G-Protein, IL-10 and PTEN signaling. Moreover, the two datasets combined generated a single molecular network that further identified PTEN (a phosphatase) as a unique new link between insulin signaling, IR, and apoptosis reflecting the pathophysiology of "metabolic syndrome".

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