Natural killer (NK) cells are equipped to innately produce the cytokine gamma interferon (IFN-γ) in part because they basally express high levels of the signal transducer and activator of transcription 4 (STAT4). Type 1 interferons (IFNs) have the potential to activate STAT4 and promote IFN-γ expression, but concurrent induction of elevated STAT1 negatively regulates access to the pathway. As a consequence, it has been difficult to detect type 1 IFN stimulation of NK cell IFN-γ during viral infections in the presence of STAT1 and to understand the evolutionary advantage for maintaining the pathway. The studies reported here evaluated NK cell responses following infections with lymphocytic choriomeningitis virus (LCMV) in the compartment handling the earliest events after infection, the peritoneal cavity. The production of type 1 IFNs, both IFN-α and IFN-β, was shown to be early and of short duration, peaking at 30 h after challenge. NK cell IFN-γ expression was detected with overlapping kinetics and required activating signals delivered through type 1 IFN receptors and STAT4. It took place under conditions of high STAT4 levels but preceded elevated STAT1 expression in NK cells. The IFN-γ response reduced viral burdens. Interestingly, increases in STAT1 were delayed in NK cells compared to other peritoneal exudate cell (PEC) populations. Taken together, the studies demonstrate a novel mechanism for stimulating IFN-γ production and elucidate a biological role for type 1 IFN access to STAT4 in NK cells.

OBJECTIVE

Pathways regulating the complex and sometimes paradoxical effects of cytokines are poorly understood. Accumulating evidence indicates that the biological consequences of type 1 interferon (IFN) exposure are shaped by modifying the concentrations of particular STATs to change access to the different signaling molecules. The results of the experiments presented conclusively demonstrate that NK cell IFN-γ can be induced through type 1 IFN and STAT4 at the first site of infection during a period with high STAT4 but prior to induction of elevated STAT1 in the cells. The response mediates a role in viral defense. Thus, a very early pathway to and source of IFN-γ in evolving immune responses to infections are identified by this work. The information obtained helps resolve long-standing controversies and advances the understanding of mechanisms regulating key type 1 IFN functions, in different cells and compartments and at different times of infection, for accessing biologically important functions.

Patients with systemic lupus erythematosus develop accelerated atherosclerosis independent of traditional risk factors. The 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase inhibitors are widely prescribed for hyperlipidemia, but they also exhibit anti-inflammatory actions that appear to be independent of their suppressive actions on plasma cholesterol levels. In this study, we analyzed the effect of the HMG-CoA reductase inhibitor simvastatin on disease manifestations in gld.apoE-/- mice that lack functional Fas ligand and apolipoprotein E and exhibit accelerated atherosclerosis and aggravated lupus-like features. Wild-type, gld, apoE-/-, and gld.apoE-/- mice were maintained on a high cholesterol Western diet and received daily simvastatin (0.125 mg/kg) or saline for 12 wk. Serum cholesterol levels were unaffected by simvastatin treatment, but atherosclerotic lesion area was reduced in both apoE-/- and gld.apoE-/- mice treated with simvastatin. Simvastatin also reduced the lymphadenopathy, renal disease, and proinflammatory cytokine production seen in gld.apoE-/-, but not gld, mice. The immunomodulatory effects in gld.apoE-/- mice were associated with enhanced STAT6 and decreased STAT4 induction in submandibular lymph node cells. Along with reductions in serum TNF-alpha and IFN-gamma levels, there was also an increase in IL-4 and IL-10 transcript levels in lymph nodes. These data indicate that HMG-CoA reductase inhibitors ameliorate atherosclerosis and lupus-like autoimmunity independent of their cholesterol-lowering effects via a shift from a Th1 to a Th2 phenotype in the gld.apoE-/- model. Thus, the anti-inflammatory activities of statins may have utility for the treatment of both autoimmunity and atherosclerosis in patients with systemic lupus erythematosus.

Complete interleukin-12/interleukin-23 receptor beta1 (IL-12Rbeta1) deficiency is the most frequent known genetic etiology of the syndrome of Mendelian susceptibility to mycobacterial disease. The patients described to date lack IL-12Rbeta1 at the surface of their natural killer (NK) and T cells due to IL12RB1 mutations, which either interrupt the open reading frame or disrupt protein folding. We describe a patient with a large in-frame deletion of 12165 nucleotides (nt) in IL12RB1, encompassing exons 8 to 13 and resulting in the surface expression of nonfunctional IL-12Rbeta1. These 6 exons encode the proximal NH2-terminal half of the extracellular domain downstream from the cytokine-binding domain. Five of 6 monoclonal anti-IL-12Rbeta1 antibodies tested recognized the internally truncated chain on the cell surface. However, IL-12 and IL-23 did not bind normally to the patient's IL-12Rbeta1-containing respective heterodimeric receptors. As a result, signal transducer and activator of transcription-4 (STAT4) was not phosphorylated and interferon-gamma (IFN-gamma) production was not induced in the patient's cells upon stimulation with even high doses of IL-12 or IL-23. The functional defect was completely rescued by retrovirus-mediated IL-12Rbeta1 gene transfer. Thus, the detection of IL-12Rbeta1 on the cell surface does not exclude the possibility of complete IL-12Rbeta1 deficiency in patients with mycobacteriosis or salmonellosis. Paradoxically, the largest IL12RB1 mutation detected is associated with the cell surface expression of nonfunctional IL-12Rbeta1, defining a novel genetic form of IL-12Rbeta1 deficiency.

Crohn' s disease (CD) is a chronic intestinal inflammatory disorder characterized by aberrant mucosal Th1 cell activation and production of IL-12, the major Th1-driving factor. The T cell response to IL-12 is dependent on the expression of a specific receptor composed of two subunits, termed IL-12Rbeta1 and IL-12Rbeta2. The content of IL-12Rbeta2, as measured at the mRNA level, is crucial in regulating Th1 differentiation. In this study we therefore investigated IL-12Rbeta2 RNA transcripts in CD. IL-12Rbeta2 expression was increased in active CD as well as Helicobacter pylori (HP)-associated gastritis and Salmonella colitis compared with that in inactive CD, ulcerative colitis, noninflammatory controls, and celiac disease. In contrast, IL-12Rbeta1 transcripts were expressed at comparable levels in all samples. In CD, IL-12Rbeta2 expression strictly correlated with tyrosine phosphorylation of STAT4, a key component of the IL-12-dependent Th1 polarization. This was associated with a pronounced expression of IFN-gamma. Transcripts for IL-12/p40 were detected in CD, HP-positive, and Salmonella colitis patients, but not in celiac disease, indicating that IL-12Rbeta2 up-regulation occurs only in IL-12-associated Th1 gastrointestinal diseases. Finally, we showed that stimulation of lamina propria mononuclear cells with IL-12 enhanced IL-12Rbeta2, suggesting that IL-12 regulates IL-12Rbeta2 expression in human gastrointestinal mucosa. The data show that the signaling pathway used by IL-12 to induce Th1 differentiation is increased at the site of disease in CD, further supporting the view that IL-12/IL-12R signals contribute to the inflammatory response in this condition.

Recent studies have highlighted a role for the alarmin interleukin (IL)-33 in CD4(+) and CD8(+) T cell activation and function, and have also revealed important distinctions. The IL-33 receptor ST2 is constitutively and abundantly expressed on T-helper-2 (Th2) and GATA-3(+) regulatory T cells in a GATA-3- and STAT5-dependent manner. Upon activation, Th1 and cytotoxic T cells express ST2 transiently, driven by T-bet and/or STAT4. We review these findings here, and critically examine evidence indicating that IL-33 enhances the differentiation and functionality of various T cell subsets through positive feedback loops involving lineage-specifying transcription factors. In this context, we discuss how quantitative and qualitative differences in ST2 expression between effector and GATA-3(+) regulatory T cells may contribute to immune homeostasis, and outline important areas of future inquiry.

Two functionally distinct subsets of B cells that produce Th1- and Th2-like patterns of cytokines have recently been identified. Interleukin-12 (IL-12) is a critical immunoregulatory cytokine that promotes Th1 differentiation through activation of signal transducer and activator of transcription 4 (STAT4). IL-12 has been reported to induce interferon gamma (IFN-gamma) production in B cells, but the relevant signaling pathways are poorly documented. Here, in human primary B cells, we found a functional IL-12 receptor (IL-12R) that internalizes following IL-12 binding. IFN-gamma and, to a lesser extent, IL-12 positively regulated the IL-12Rbeta2 subunit but had no effect on IL-12Rbeta1. On examining the effect of IL-12 on STAT4 and T-bet (2 key factors involved in IFN-gamma promoter activation), we found that IL-12 induced the phosphorylation and nuclear translocation of STAT4. IL-12-dependent constitutive STAT4 activation was also observed in the Epstein-Barr virus (EBV)-transformed B-cell line RPMI 8866 that spontaneously produces IL-12. T-bet expression has been shown to be dependent on STAT1. IL-12 had no direct effect on STAT1 activation or T-bet expression in primary B cells. In contrast, IL-12-induced IFN-gamma led to STAT1 activation, strong expression of T-bet, and IFN-gamma expression. IL-12 therefore initiates a cascade of events in B cells, including STAT4 activation, IL-12Rbeta2 up-regulation, IFN-gamma production, and T-bet up-regulation, potentially leading to Th1-like differentiation.

In vitro human monocyte differentiation to macrophages or dendritic cells (DCs) is driven by GM-CSF or GM-CSF and IL-4, respectively. IFN regulatory factors (IRFs), especially IRF1 and IRF8, are known to play essential roles in the development and functions of macrophages and DCs. In the present study, we performed cDNA microarray and Northern blot analyses to characterize changes in gene expression of selected genes during cytokine-stimulated differentiation of human monocytes to macrophages or DCs. The results show that the expression of IRF4 mRNA, but not of other IRFs, was specifically up-regulated during DC differentiation. No differences in IRF4 promoter histone acetylation could be found between macrophages and DCs, suggesting that the gene locus was accessible for transcription in both cell types. Computer analysis of the human IRF4 promoter revealed several putative STAT and NF-kappaB binding sites, as well as an IRF/Ets binding site. These sites were found to be functional in transcription factor-binding and chromatin immunoprecipitation experiments. Interestingly, Stat4 and NF-kappaB p50 and p65 mRNAs were expressed at higher levels in DCs as compared with macrophages, and enhanced binding of these factors to their respective IRF4 promoter elements was found in DCs. IRF4, together with PU.1, was also found to bind to the IRF/Ets response element in the IRF4 promoter, suggesting that IRF4 protein provides a positive feedback signal for its own gene expression in DCs. Our results suggest that IRF4 is likely to play an important role in myeloid DC differentiation and gene regulatory functions.

BACKGROUND

Genetic factors have a substantial role in determining development of rheumatoid arthritis (RA), and are likely to account for 50-60% of disease susceptibility. Genome-wide association studies have identified non-human leucocyte antigen RA susceptibility loci which associate with RA with low-to-moderate risk.

OBJECTIVE

To investigate recently identified RA susceptibility markers using cohorts from six European countries, and perform a meta-analysis including previously published results.

METHODS

3311 DNA samples were collected from patients from six countries (UK, Germany, France, Greece, Sweden and Denmark). Genotype data or DNA samples for 3709 controls were collected from four countries (not Sweden or Denmark). Eighteen single nucleotide polymorphisms (SNPs) were genotyped using Sequenom MassArray technology. Samples with a >95% success rate and only those SNPs with a genotype success rate of >95% were included in the analysis. Scandinavian patient data were pooled and previously published Swedish control data were accessed as a comparison group. Meta-analysis was used to combine results from this study with all previously published data.

RESULTS

After quality control, 3209 patients and 3692 controls were included in the study. Eight markers (ie, rs1160542 (AFF3), rs1678542 (KIF5A), rs2476601 (PTPN22), rs3087243 (CTLA4), rs4810485 (CD40), rs5029937 (6q23), rs10760130 (TRAF1/C5) and rs7574865 (STAT4)) were significantly associated with RA by meta-analysis. All 18 markers were associated with RA when previously published studies were incorporated in the analysis. Data from this study increased the significance for association with RA and nine markers.

CONCLUSIONS

In a large European RA cohort further evidence for the association of 18 markers with RA development has been obtained.

Interleukin-12 (IL-12) was the first member of the IL-12 family of cytokines to be identified and has therefore become its eponym. It is a heterodimeric protein of two subunits (p35, p40) secreted by phagocytic cells in response to pathogens and mainly acts through STAT4 to induce IFN-γ production in T and NK cells. IFN-γ in turn mediates proinflammatory functions and activates T-bet. As IL-12 engages in TH1 development, it is believed to represent an important link between innate and adaptive immunity. Following its identification and the finding of its association to TH1 commitment, great hopes were placed in IL-12 to become a target for therapeutic applications in multiple settings of autoimmunity and cancer. Though, the discovery of the related members of the IL-12 family and several rather disappointing attempts to translate experimental results into clinical practice, have relativized these hopes. Nevertheless, IL-12 remains a cytokine of outstanding importance with lots of unresolved questions. In this review, we will first briefly depict the biochemistry of the cytokine, its receptor and the related signal transduction, before summarizing the regulation of IL-12 production and its biological functions. We will then describe the current knowledge about the implication of IL-12 in different murine disease models as well as in the corresponding human conditions and comment on possible consequences for future clinical applications.

Natural killer (NK) cells are crucial components of the innate immune system, providing the first line of defense against infectious pathogens and tumors. Interleukin (IL)-12 is an interleukin produced primarily by antigen-presenting cells that play an essential role in the interaction between the innate and adaptive arms of immunity acting upon T and NK cells to generate cytotoxic lymphocytes. In the present study, we explored the effect of IL-12 upregulation on the NK receptor NKG2D and on the promotion of NK cell function. IL-12 enhanced the cytotoxicity of NK cells to different solid and hematological tumor cell lines and promoted interferon-gamma secretion by NK cells. The IL-12-induced cytolytic effect was dependent on the interaction of NKG2D with its ligand, MICA, because blockade of either protein attenuated the effect of IL-12 on NK cytolysis. Reverse transcriptase-polymerase chain reaction and fluorescence-activated cell sorting analyses indicated that IL-12 treatment increased NKG2D transcripts and surface expression in NK cells. Also, IL-12 augmented the expression of cytotoxic effector molecules, TRAIL and perforin, and the phosphorylation of STAT1, STAT4, and ERK1/2, which may also contribute to lysis by NK cells. These results are encouraging for the potential use of IL-12 as part of immunotherapy.

BACKGROUND

Recent studies demonstrated an association of STAT4 variants with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), indicating that multiple autoimmune diseases share common susceptibility genes. We therefore investigated the influence of STAT4 variants on the susceptibility and phenotype of inflammatory bowel diseases (IBD) in a large patient and control cohort.

RESULTS

Genomic DNA from 2704 individuals of Caucasian origin including 857 patients with Crohn's disease (CD), 464 patients with ulcerative colitis (UC), and 1383 healthy, unrelated controls was analyzed for seven SNPs in the STAT4 gene (rs11889341, rs7574865, rs7568275, rs8179673, rs10181656, rs7582694, rs10174238). In addition, a detailed genotype-phenotype analysis was performed. Our analysis revealed an association of the STAT4 SNP rs7574865 with overall decreased susceptibility to CD (p = 0.047, OR 0.86 [95% CI 0.74-0.99]). However, compared to CD patients carrying the wild type genotype, the STAT4 SNP rs7574865 was significantly associated with early CD onset (p = 0.021) and colonic CD (p = 0.008; OR = 4.60, 95% CI 1.63-12.96). For two other STAT4 variants, there was a trend towards protection against CD susceptibility (rs7568275, p = 0.058, OR 0.86 [95% CI 0.74-1.00]; rs10174238, p = 0.057, OR 0.86 [95% CI 0.75-1.00]). In contrast, we did not observe any association with UC susceptibility. Evidence for weak gene-gene interaction of STAT4 with the IL23R SNP rs11209026 was lost after Bonferroni correction.

CONCLUSIONS

Our results identified the STAT4 SNP rs7574865 as a disease-modifying gene variant in colonic CD. However, in contrast to SLE and RA, the effect of rs7574865 on CD susceptibility is only weak.

BACKGROUND

There are now over 30 confirmed loci predisposing to rheumatoid arthritis (RA). Studies have been largely undertaken in patients with anticyclic citrullinated peptide (anti-CCP) positive RA, and some genetic associations appear stronger in this subgroup than in anti-CCP negative disease, although few studies have had adequate power to address the question. The authors therefore investigated confirmed RA susceptibility loci in a large cohort of anti-CCP negative RA subjects.

METHODS

RA patients and controls, with serological and genetic data, were available from UK Caucasian patients (n=4068 anti-CCP positive, 2040 anti-CCP negative RA) and 13,009 healthy controls. HLA-DRB1 genotypes and 36 single nucleotide polymorphisms were tested for association between controls and anti-CCP positive or negative RA.

RESULTS

The shared epitope (SE) showed a strong association with anti-CCP positive and negative RA, although the effect size was significantly lower in the latter (effect size ratio=3.18, p<1.0E-96). A non-intronic marker at TNFAIP3, GIN1/C5orf30, STAT4, ANKRD55/IL6ST, BLK and PTPN22 showed association with RA susceptibility, irrespective of the serological status, the latter three markers remaining significantly associated with anti-CCP negative RA, after correction for multiple testing. No significant association with anti-CCP negative RA was detected for other markers (eg, AFF3, CD28, intronic marker at TNFAIP3), though the study power for those markers was over 80%.

CONCLUSIONS

In the largest sample size studied to date, the authors have shown that the strength of association, the effect size and the number of known RA susceptibility loci associated with disease is different in the two disease serotypes, confirming the hypothesis that they might be two genetically different subsets.

In response to interleukin 12 (IL-12) stimulation, a latent cytoplasmic transcription factor, Stat4 (signal transducer and activator of transcription 4), becomes tyrosine-phosphorylated and translocates into the nucleus where it binds to DNA to activate transcription. Cofactors that can directly bind and regulate Stat4 activity have not been described. We report here that PIASx, a member of the protein inhibitor of activated STAT (PIAS) family, is a negative regulator of Stat4. PIASx becomes associated with Stat4 following IL-12 stimulation in vivo. PIASx inhibits IL-12-stimulated and Stat4-dependent gene activation in human T cells. PIASx does not inhibit the DNA binding activity of Stat4. Instead PIASx is present in the Stat4-DNA binding complex. Finally the inhibitory activity of PIASx on Stat4-mediated gene activation is abolished by the histone deacetylase inhibitor trichostatin A. Our results suggest that PIASx may function as a co-repressor of Stat4.

OBJECTIVE

5-Hydroxytryptamine (5-HT) released from enterochromaffin cells influences intestinal homeostasis by altering gut physiology and is implicated in the pathophysiology of various gut disorders. The mechanisms regulating 5-HT production in the gut remain unclear. This study investigated the T helper (Th) 1/Th2-based immunoregulation of enterochromaffin cell function and 5-HT production in a model of enteric infection.

RESULTS

Trichuris muris-infected AKR (susceptible to infection and generates Th1 response), BALB/c (resistant to infection and generates Th2 response), Stat4-deficient (impaired in Th1 response) and Stat6-deficient (impaired in Th2 response) mice were investigated to assess enterochromaffin cells, 5-HT and cytokines. In association with the generation of a Th2 response we observed higher enterochromaffin cell numbers and 5-HT content in the colon of BALB/c mice compared with AKR mice. Numbers of enterochromaffin cells and amount of 5-HT were significantly lower in Stat6-deficient mice after infection compared with Stat4-deficient mice. In addition, enterochromaffin cell numbers and 5-HT content were significantly higher after reconstitution of severe combined immunodeficient mice with in-vitro polarised Th2 cells.

CONCLUSIONS

The study demonstrated that enterochromaffin cell and 5-HT responses to the same infectious agent are influenced by Th1 or Th2 cytokine predominance and suggests that the immunological profile of the inflammatory response is important in the regulation of enterochromaffin cell biology in the gut. In addition to new data on enterochromaffin cell function in enteric infection and inflammation, this study provides important information on the immuno-endocrine axis in the gut, which may ultimately lead to improved strategies against gut disorders.

Interleukin 12 (IL-12)-induced T helper 1 (Th1) development requires Stat4 activation. However, antigen-activated Th1 cells can produce interferon gamma (IFN-gamma) independently of IL-12 and Stat4 activation. Thus, in differentiated Th1 cells, factors regulated by IL-12 and Stat4 may be involved in IFN-gamma production. Using subtractive cloning, we identified ERM, an Ets transcription factor, to be a Th1-specific, IL-12-induced gene. IL-12-induction of ERM occurred in wild-type and Stat1-deficient, but not Stat4-deficient, T cells, suggesting ERM is Stat4-inducible. Retroviral expression of ERM did not restore IFN-gamma production in Stat4-deficient T cells, but augmented IFN-gamma expression in Stat4-heterozygous T cells. Ets factors frequently regulate transcription via cooperative interactions with other transcription factors, and ERM has been reported to cooperate with c-Jun. However, in the absence of other transcription factors, ERM augmented expression of an IFN-gamma reporter by only 2-fold. Thus, determining the requirement for ERM in Th1 development likely will require gene targeting.

C3H mice infected with Leishmania mexicana fail to develop a protective Th1 response, and are unable to cure. In this study, we show that L. mexicana cysteine proteases suppress the antileishmanial immune response. Previous studies demonstrated that deletion of the entire multicopy cysteine protease B (CPB) gene array in L. mexicana is associated with decreased parasite virulence, potentially attributable to factors related to parasite fitness rather than to direct effects on the host immune response. We now show that C3H mice infected with the L. mexicana deletion mutant (Deltacpb) initially develop lesions that grow at rates comparable to those of wild-type L. mexicana-infected mice. However, in contrast to controls, Deltacpb-induced lesions heal with an accompanying Th1 immune response. Lesion resolution was Th1 dependent, as Deltacpb-infected IL-12p40(-/-) and STAT4(-/-) mice developed high parasite burdens and progressive disease. Moreover, when L. major was transfected with a cosmid expressing multiple L. mexicana CPB genes, this parasite induced a significantly lower IFN-gamma response compared with wild-type L. major. These data indicate that cysteine proteases of L. mexicana are critical in suppressing protective immune responses and that inhibition of CPB may prove to be a valuable immunomodulatory strategy for chronic forms of leishmaniasis.

We have reported that hepatitis B X-interacting protein (HBXIP) promotes the proliferation and migration of breast cancer cells. However, the underlying mechanism is poorly understood. In this study, we report that HBXIP works in the event through up-regulating S100A4. We observed that HBXIP expression was positively correlated to that of S100A4 in 87 clinical breast cancer tissue samples. Then, we identified that HBXIP was able to up-regulate S100A4 expression in breast cancer cells. Notably, we observed the HBXIP nuclear localization, implying that HBXIP may be associated with the promoter of S100A4. Chromatin immunoprecipitation assay (ChIP) and electrophoretic mobility shift assay (EMSA) showed that HBXIP was able to bind to the nucleotides +200~+239 region of S100A4 promoter, containing two putative recognition motif of transcription factor STAT4 and GRβ. It suggests that HBXIP is able to activate S100A4 promoter via interacting with STAT4 in breast cancer cells, leading to the up-regulation of S100A4. In addition, we identified another pathway of S100A4 up-regulation mediated by HBXIP. We found that HBXIP activated the PTEN/PI3K/AKT signaling by inducing DNA methylation of PTEN, which subsequently boosted S100A4 expression. In function, we demonstrated that HBXIP enhanced the growth or migration of breast cancer cells through S100A4 in vivo and in vitro. Collectively, we conclude that HBXIP up-regulates S100A4 through activating S100A4 promoter involving STAT4 and inducing PTEN/PI3K/AKT signaling to promote growth and migration of breast cancer cells. Our finding provides new insight into the mechanism of HBXIP in promotion of the development of breast cancer.

Activation of the transcription factor signal transducer and activator of transcription (STAT)-4 is critical for the differentiation of T-helper 1 cells/type-1 cytotoxic T-cells and the production of interferon (IFN)-gamma. Expression of STAT4, phospho-STAT4, IFN-gamma and T-box expressed in T-cells (T-bet) proteins in bronchial biopsies and bronchoalveolar lavage (BAL)-derived lymphocytes, obtained from 12 smokers with mild/moderate chronic obstructive pulmonary disease (COPD) (forced expiratory volume in one second (FEV1) 59 +/- 16% predicted), 14 smokers with normal lung function (FEV1 106 +/- 12% pred) and 12 nonsmoking subjects (FEV1 111 +/- 14% pred), was examined by immunohistochemistry and immunocytochemistry. In bronchial biopsies of COPD patients, the number of submucosal phospho-STAT4+ cells was increased (240 (22-406) versus 125 (0-492) versus 29 (0-511) cells mm(-2)) when compared with both healthy smokers and control nonsmokers, respectively. In smokers, phospho-STAT4+ cells correlated with the degree of airflow obstruction and the number of IFN-gamma+ cells. Similar results were seen in BAL (2.8 (0.2-5.9) versus 1.03 (0.09-1.6) versus 0.69 (0-2.3) lymphocytes x mL(-1) x 10(3)). In all smokers who underwent lavage, phospho-STAT4+ lymphocytes correlated with airflow obstruction and the number of IFNgamma+ lymphocytes. T-bet expression was not altered in bronchial biopsies and BAL-derived lymphocytes between the three groups. In conclusion, this study suggests that stable mild/moderate chronic obstructive pulmonary disease is associated with an active T-helper 1 cell/type-1 cytotoxic T-cell inflammatory process involving activation of signal transducer and activator of transcription 4 and interferon-gamma production.

Interleukin-23 (IL-23) is a heterodimeric cytokine belonging to the IL-6/IL-12 family that plays a key role in several of autoimmune and inflammatory disorders. This family contains the 34 type I cytokine receptor chains and 27 ligands, which share structural and functional similarities, but on the other hand they display distinct roles in shaping Th cells responses. IL-12 family cytokines have not only proinflammatory effects but they also promote inflammatory responses. IL-23 is composed of the p40 subunit in common with IL-12, and with a unique p19 subunit. IL-23 binding to an IL-23 receptor expressed on dendritic cells, macrophages and monocytes triggers the activation of Jak2 and Tyk2, which in turn phosphorylates STAT1, STAT3, STAT4 and STAT5 as well as induce formation of STAT3-STAT4 heterodimers. IL-23 is one of the essential factors required for the survival and/or expansion of Th17 cells, which produce IL-17, IL-17F, IL-6 and TNF-alpha. Th17 cells stimulated by the IL-23 promote osteoclastogenesis through production of IL-17, which induce receptor activator of NF-kappa B ligand on mesenchymal cells. The IL-23-IL-17 axis includes Th17 cells and plays a key role in the development of autoimmune arthritis.

The Signal Transducer and Activator of Transcription (STAT) family of proteins consists of transcription factors that play a complex and essential role in the regulation of physiologic cell processes, such as proliferation, differentiation, apoptosis and angiogenesis, and serves to organize the epigenetic landscape of immune cells. To date, seven STAT genes have been identified in the human genome; STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b and STAT6. They all account for diverse effects in response to extracellular signaling proteins, mainly by altering gene transcription in the effector cells. Members of the STAT family have been implicated in human cancer development, progression, metastasis, survival and resistance to treatment. Particularly STAT3 and STAT5 are of interest in cancer biology. They are currently considered as oncogenes, but their signaling is embedded into a complex and delicate balance between different (counteracting) transcription factors, and thus, in some contexts, they can have a tumor suppressive role. Assessing STAT signaling mutations as well as screening for aberrant STAT pathway activation may have a role to predict sensitivity to immunotherapy and targeted STAT inhibition. In the present comprehensive review of the literature, we discuss in-depth the role of each STAT family member in cancer, assemble cutting-edge information on the use of these molecules as potential biomarkers and targets for treatment, and address why their clinical implementation is controversy.

Respiratory syncytial virus (RSV) infection of BALB/c mice previously immunized with a recombinant vaccinia virus (vacv) expressing the attachment (G) protein of RSV (vacvG) results in pulmonary eosinophilia, which mimics the response of formalin-inactivated RSV-vaccinated children, as well as increased weight loss, clinical illness, and enhanced pause (Penh). We show that RSV infection of eosinophil-deficient mice previously immunized with vacvG results in the development of increased weight loss, clinical illness, and Penh similar to that in wild-type controls. These measures of RSV vaccine-enhanced disease are dependent upon STAT4. Interestingly, neither IL-12 nor IL-23, the two most common STAT4-activating cytokines, proved necessary for the development of disease. We demonstrate that IFN-gamma, which is produced following STAT4 activation, contributes to clinical illness and increased Penh, but not weight loss. Our results have important implications for future RSV vaccine design, suggesting that enhancing a Th1 response may exacerbate disease.

Interleukin 12 (IL-12) is a major inducer of interferon gamma (IFN-gamma) and the principal mediator of T helper 1 (Th1) differentiation. To identify IL-12-regulated genes, which might contribute to Th1 differentiation and IFNG regulation, we employed microarray analysis. Surprisingly, a ubiquitously expressed proprotein convertase (PC), furin, was one of the most consistently IL-12-induced genes in T cells, and among PCs was the only one regulated by this cytokine. Furin was preferentially expressed in differentiated Th1 cells in a Stat4-dependent manner. Expression of furin enhanced IFN-gamma secretion, whereas inhibition of furin interfered with IFN-gamma production. Thus, we conclude that IL-12 induction of furin might represent a new aspect of IFN-gamma regulation and control of Th1 differentiation.

NF-kappaB/Rel transcription factors are linked to innate immune responses and APC activation. Whether and how the induction of NF-kappaB signaling in normal CD4(+) T cells regulates effector function are not well-understood. The liberation of NF-kappaB dimers from inhibitors of kappaB (IkappaBs) constitutes a central checkpoint for physiologic regulation of most forms of NF-kappaB. To investigate the role of NF-kappaB induction in effector T cell responses, we targeted inhibition of the NF-kappaB/Rel pathway specifically to T cells. The Th1 response in vivo is dramatically weakened when T cells defective in their NF-kappaB induction (referred to as IkappaBalpha(DeltaN) transgenic cells) are activated by a normal APC population. Analyses in vivo, and IL-12-supplemented T cell cultures in vitro, reveal that the mechanism underlying this T cell-intrinsic requirement for NF-kappaB involves activation of the IFN-gamma gene in addition to clonal expansion efficiency. The role of NF-kappaB in IFN-gamma gene expression includes a modest decrease in Stat4 activation, T box expressed in T cell levels, and differentiation efficiency along with a more prominent postdifferentiation step. Further, induced expression of Bcl-3, a trans-activating IkappaB-like protein, is decreased in T cells as a consequence of NF-kappaB inhibition. Together, these findings indicate that NF-kappaB induction in T cells regulates efficient clonal expansion, Th1 differentiation, and IFN-gamma production by Th1 lymphocytes at a control point downstream from differentiation.

The distinctive differentiated states of the CD4+ T helper cells are determined by the set of transcription factors and the genes transcribed by the transcription factors. In vitro induction models, the major determinants of the cytokines present during the T-cell receptor (TCR)-mediated activation process. IL-12 and IFN-γ make Naive CD4+ T cells highly express T-bet and STAT4 and differentiate to TH1 cells, while IL-4 make Naive CD4+ T cells highly express STAT6 and GATA3 and differentiated to TH2 cells. Even through T-bet and GATA3 are master regulators for TH1/TH2 cells differentiation. There are many other transcription factors, such as RUNX family proteins, IRF4, Dec2, Gfi1, Hlx, and JunB that can impair TH1/TH2 cells differentiation. In recent years, noncoding RNAs (microRNA and long noncoding RNA) join in the crowd. The leukocytes should migrate to the right place to show their impact. There are some successful strategies, which are revealed to targeting chemokines and their receptors, that have been developed to treat human immune-related diseases.