Differentiation-dependent regulation of the Ifng cytokine gene locus in T helper (Th) cells has emerged as an excellent model for functional study of distal elements that control lineage-specific gene expression. We previously identified a cis-regulatory element located 22 kb upstream of the Ifng gene (Conserved Non-coding Sequence -22, or CNS-22) that is a site for recruitment of the transcription factors T-bet, Runx3, NF-κB and STAT4, which act to regulate transcription of the Ifng gene in Th1 cells. Here, we report the generation of mice with a conditional deletion of CNS-22 that has enabled us to define the epigenetic and functional consequences of its absence. Deletion of CNS-22 led to a defect in induction of Ifng by the cytokines IL-12 and IL-18, with a more modest effect on induction via T-cell receptor activation. To better understand how CNS-22 and other Ifng CNSs regulated Ifng transcription in response to these distinct stimuli, we examined activation-dependent changes in epigenetic modifications across the extended Ifng locus in CNS-22-deficient T cells. We demonstrate that in response to both cytokine and TCR driven activation signals, CNS-22 and other Ifng CNSs recruit increased activity of histone acetyl transferases (HATs) that transiently enhance levels of histones H3 and H4 acetylation across the extended Ifng locus. We also demonstrate that activation-responsive increases in histone acetylation levels are directly linked to the ability of Ifng CNSs to acutely enhance Pol II recruitment to the Ifng promoter. Finally, we show that impairment in IL-12+IL-18 dependent induction of Ifng stems from the importance of CNS-22 in coordinating locus-wide levels of histone acetylation in response to these cytokines. These findings identify a role for acute histone acetylation in the enhancer function of distal conserved cis-elements that regulate of Ifng gene expression.
Differentiation of multipotent naïve T cell precursors into functionally mature effector cells that control different types of immune responses is an excellent model to study lineage-specific regulation of gene expression. A number of cis-regulatory elements have been reported to control expression of the gene that encodes the cytokine IFN-γ which is a signature product of effector T cells of the Th1 lineage. However, none of these elements has been analyzed for effects on gene expression and chromatin remodeling through deletional analysis in the native Ifng gene locus. Here we have generated mice in which a key element previously implicated in control of Ifng gene expression (CNS-22) was conditionally deleted from the genome. Th1 cells in which CNS-22 was deleted had activation-specific deficits in Ifng expression and demonstrated defects in epigenetic changes across the Ifng locus. Mapping epigenetic consequences of CNS-22 deletion led to identification of acute hyperacetylation of histones immediately flanking this and other cis-regulatory elements that was associated with Ifng gene transcription, as well as more global defects in histone acetylation. These findings support a mechanism whereby regulatory sites that have acquired baseline histone acetylation marks during lineage specification undergo acute, activation-dependent increases in histone acetyl transferase activity that enhance transcription of inducible genes.
Interleukin-22 (IL-22) is central to host protection against bacterial infections at barrier sites. Both innate lymphoid cells (ILCs) and T cells produce IL-22. However, the specific contributions of CD4+ T cells and their developmental origins are unclear. We found that the enteric pathogen Citrobacter rodentium induced sequential waves of IL-22 producing ILCs and CD4+ T cells that were each critical to host defense during a primary infection. Whereas IL-22 production by ILCs was strictly IL-23–dependent, development of IL-22 producing CD4+ T cells occurred via an IL-6–dependent mechanism that was augmented by, but not dependent on, IL-23, and was dependent on both transcription factors T-bet and AhR. Transfer of CD4+ T cells differentiated with IL-6 in the absence of TGF-β (“Th22” cells) conferred protection of infected IL-22-deficient mice whereas transferred Th17 cells did not. These findings establish Th22 cells as an important component of mucosal anti-microbial host defense.
Sensitization to fungi, such as the mold Aspergillus fumigatus, is increasingly becoming linked with asthma severity. We have previously shown that lung responses generated via the beta-glucan receptor Dectin-1 are required for lung defense during acute, invasive A. fumigatus infection. Unexpectedly, in an allergic model of chronic lung exposure to live A. fumigatus conidia, beta-glucan recognition via Dectin-1 led to the induction of multiple proallergic (Muc5ac, Clca3, CCL17, CCL22 and IL-33) and proinflammatory (IL-1β and CXCL1) mediators that compromised lung function. Attenuated proallergic and proinflammatory responses in the absence of Dectin-1 was not associated with changes in Ido (indoleamine 2,3-dioxygenase), Il12p35/Ebi3 (IL-35), IL-10 or TGF-β levels. Assessment of T helper responses demonstrated that purified lung CD4+ T cells produced IL-4, IL-13, IFN-γ and IL-17A, but not IL-22, in a Dectin-1 dependent manner. In contrast, we observed robust, Dectin-1 dependent IL-22 production by unfractionated lung digest cells. Intriguingly, the absence of IL-22 alone mimicked the attenuated proallergic and proinflammatory responses observed in the absence of Dectin-1, suggesting that Dectin-1 mediated IL-22 production potentiated responses that led to decrements in lung function. To this end, neutralization of IL-22 improved lung function in normal mice. Collectively, these results indicate that the beta-glucan receptor Dectin-1 contributes to lung inflammation and immunopathology associated with persistent fungal exposure via the production of IL-22.
γδ T cells contribute uniquely to host immune defense. However, how they function remains an enigma. Although it is unclear what most γδ T cells recognize, common dogma asserts that they recognize self-antigens. While they are the major initial Interleukin-17 (IL-17) producers in infections, it is unclear what is required to trigger these cells to act. Here, we report that a noted B cell antigen, the algae protein-phycoerythrin (PE) is an antigen for murine and human γδ T cells. PE also stained specific bovine γδ T cells. Employing this specificity, we demonstrated that antigen recognition, but not extensive clonal expansion, was required to activate naïve γδ T cells to make IL-17. In this activated state, γδ T cells gained the ability to respond to cytokine signals that perpetuated the IL-17 production. These results underscore the adaptability of lymphocyte antigen receptors and suggest a previously unrecognized antigen-driven rapid response in protective immunity prior to the maturation of classical adaptive immunity.
Levels of reactive free radicals are elevated in the airway during asthmatic exacerbations, but their roles in the pathophysiology of asthma remain unclear. We have identified subsets of myeloid-derived suppressor-like cells as key sources of nitric oxide and superoxide in the lungs of mice with evolving experimental allergic airway inflammation and established these cells as master regulators of the airway inflammatory response. The profiles of free radicals they produced depended on expression of iNOS, arginase, and NADPH oxidase. These radicals controlled the pro- and anti-inflammatory potential of these cells, and also regulated the reciprocal pattern of their infiltration into the lung. The nitric oxide-producing cells were Ly-6C+Ly-6G− and down-modulated T cell activation, recruited Treg cells, and dramatically down-regulated antigen-induced airway hyperresponsiveness. The superoxide-producing cells were Ly-6C−Ly-6G+ and expressed proinflammatory activities, exacerbating airway hyperresponsiveness in a superoxide-dependent fashion. A smaller population of Ly-6C+Ly-6G+ cells also suppressed T cell responses, but in an iNOS- and arginase-independent fashion. These regulatory myeloid cells represent important targets for asthma therapy.
Development of T helper (Th) 17 cells requires transforming growth factor (TGF)-β and interleukin (IL)-6 and is independent of the Th1 pathway. Although T cells that produce interferon (IFN)-γ are a recognized feature of Th17 cell responses, mice deficient for STAT4 and T-bet—two prototypical Th1 transcription factors—are protected from autoimmunity associated with Th17 pathogenesis. To examine the fate and pathogenic potential of Th17 cells and origin of IFN-γ-producing T cells that emerge during Th17 immunity, we developed IL-17F reporter mice that identify cells committed to expression of IL-17F and IL-17A. Th17 cells required TGF-β for sustained expression of IL-17F and IL-17A. In the absence of TGF-β, both IL-23 and IL-12 acted to suppress IL-17 and enhance IFN-γ production in a STAT4- and T-bet-dependent manner, albeit with distinct efficiencies. These results support a model of late Th17 developmental plasticity with implications for auto-immunity and host defense.
Similar to hematopoietic stem cells, memory lymphocytes self renew, while their clonally expanded effector progeny differentiate to fight infection and tumors. Recently, Muranski et al. report in Immunity (2011) that a subset of Th17 effector cells functions as memory cells and retain stem cell properties.
Regulatory B cells that are functionally defined by their capacity to express IL-10 (B10 cells) downregulate inflammation and autoimmunity. In studies using well-defined IL-10-reporter mice, this rare B10 cell subset was also found to maintain a capacity for plasma cell differentiation. During a transient period of il10 transcription, the blimp1 and irf4 transcription factors were induced in B10 cells while pax5 and bcl6 were downregulated as a significant fraction of B10 cells completed the genetic and phenotypic program leading to antibody-secreting cell differentiation in vitro and in vivo. B10 cell-derived IgM reacted with both self and foreign Ags, whereas B10 cells generated Ag-specific IgG in response to immunizations. Moreover, B10 cells represented a significant source of serum IgM and IgG during adoptive transfer experiments, and produced Ag-specific, polyreactive and autoreactive antibody specificities that were consistent with their expression of a diverse Ag receptor repertoire. Thereby, B10 cells not only limit inflammation and immune responses by the transient production of IL-10, but may also facilitate clearance of their eliciting Ags through an inherent capacity to quickly generate polyreactive and/or Ag-specific antibodies during humoral immune responses.
Human neonates are at significantly greater risk of serious infection than immunocompetent adults. In particular, very low birth weight infants in the neonatal intensive care nursery are at high risk of developing life-threatening bacterial and fungal infections. Recent studies have identified Th17 cells as critical mediators of immunity to bacterial and fungal infections at epithelial barriers. Little is known, however, about the ontogeny of Th17 responses in humans. The frequency of serious bacterial infections in preterm infants and the importance of Th17 cells in providing protection against such infections in animal studies prompted us to study Th17 development in human neonates. NaÔve CD4 T cells from extremely preterm infants, term infants, and adults were assayed for their capacity to develop into Th17 effector cells. Surprisingly, Th17 capacity was inversely related to developmental age. Neonates expressed higher levels of IL-23R, RORγt, and STAT3 prior to activation and showed a significant Th17 bias after activation. In contrast, adult cells expressed more TBX21 with a corresponding Th1 bias. CD161 expression on Th17 precursors was also developmentally regulated. Our results suggest there is significant developmental regulation of CD4 effector lineages with a strong bias toward Th17 development early in life.
Th17 cell; premature infant; CD161; neonatal immunity; T-helper subsets
B cells contribute to the pathogenesis of chronic autoimmune disorders like systemic lupus erythematosus (SLE) via multiple effector functions. However, B cells are also implicated in regulating SLE and other autoimmune syndromes via release of IL-10. B cells secreting IL-10 have been termed “Breg” and have been proposed as a separate subset of cells, a concept that remains controversial. The balance between pro- and anti-inflammatory effects could determine the success of B cell targeted therapies for autoimmune disorders and it is therefore pivotal to understand the significance of B cell-secreted IL-10 in spontaneous autoimmunity. By lineage specific deletion of Il10 from B cells we demonstrate that B cell-derived IL-10 is ineffective in suppressing the spontaneous activation of self-reactive B and T cells during lupus. Correspondingly, severity of organ disease and survival rates in mice harboring Il10 deficient B cells are unaltered. Genetic marking of cells that transcribe Il10 illustrates that the pool of IL-10 competent cells is dominated by CD4 T cells and macrophages. IL-10 competent cells of the B lineage are rare in vivo and among them short-lived plasmablasts have the highest frequency, suggesting an activation rather than lineage-driven phenotype. Putative Breg phenotypic subsets such as CD1dhiCD5+ and CD21hiCD23hi B cells are not enriched in Il10 transcription. These genetic studies demonstrate that in a spontaneous model of murine lupus, IL-10 dependent B cell regulation does not restrain disease and thus the pathogenic effects of B cells are not detectably counterbalanced by their IL-10 dependent regulatory functions.
The interleukin-17 (IL-17) family of cytokines phylogenetically pre-dates the evolution of T cells in jawed vertebrates, suggesting that the ontogeny of the Th17 cell lineage must have arisen to confer an evolutionary advantage to the host over innate sources of IL-17. Using a model of mucosal immunization with the encapsulated bacteria Klebsiella pneumoniae, B cells largely recognized polysaccharide capsular antigens, which limited protection to only the vaccine strain. In contrast, memory Th17 cells proliferated in response to conserved outer membrane proteins and conferred protection against several serotypes of K. pneumoniae, including the recently described multi-drug resistant New Dehli metallolactamase strain. Notably, this heterologous, clade specific protection was antibody-independent, demonstrating the Th17 cell lineage confers a host advantage by providing heterologous mucosal immunity independent of serotype specific antibody.
We have previously reported that mice deficient in the beta-glucan receptor Dectin-1 displayed increased susceptibility to Aspergillus fumigatus lung infection in the presence of lower interleukin 23 (IL-23) and IL-17A production in the lungs and have reported a role for IL-17A in lung defense. As IL-23 is also thought to control the production of IL-22, we examined the role of Dectin-1 in IL-22 production, as well as the role of IL-22 in innate host defense against A. fumigatus. Here, we show that Dectin-1-deficient mice demonstrated significantly reduced levels of IL-22 in the lungs early after A. fumigatus challenge. Culturing cells from enzymatic lung digests ex vivo further demonstrated Dectin-1-dependent IL-22 production. IL-22 production was additionally found to be independent of IL-1β, IL-6, or IL-18 but required IL-23. The addition of recombinant IL-23 augmented IL-22 production in wild-type (WT) lung cells and rescued IL-22 production by lung cells from Dectin-1-deficient mice. In vivo neutralization of IL-22 in the lungs of WT mice resulted in impaired A. fumigatus lung clearance. Moreover, mice deficient in IL-22 also demonstrated a higher lung fungal burden after A. fumigatus challenge in the presence of impaired IL-1α, tumor necrosis factor alpha (TNF-α), CCL3/MIP-1α, and CCL4/MIP-1β production and lower neutrophil recruitment, yet intact IL-17A production. We further show that lung lavage fluid collected from both A. fumigatus-challenged Dectin-1-deficient and IL-22-deficient mice had compromised anti-fungal activity against A. fumigatus in vitro. Although lipocalin 2 production was observed to be Dectin-1 and IL-22 dependent, lipocalin 2-deficient mice did not demonstrate impaired A. fumigatus clearance. Moreover, lung S100a8, S100a9, and Reg3g mRNA expression was not lower in either Dectin-1-deficient or IL-22-deficient mice. Collectively, our results indicate that early innate lung defense against A. fumigatus is mediated by Dectin-1-dependent IL-22 production.
Regulatory T (Treg) cells are plastic, but the in vivo mechanisms by which they are converted into Foxp3+interferon (IFN)-γ+ T cells, and whether these converted cells retain the ability to inhibit colitis, are not clear.
Foxp3+ Treg cells were generated by culture of naïve CD4+ T cells from Foxp3GFP CBir1 T-cell receptor (TCR) transgenic (CBir1-Tg) mice, which are specific for CBir1 flagellin (an immunodominant microbiota antigen), with transforming growth factor (TGF)-β. Foxp3GFP+ CBir1-Tg Treg cells were isolated by fluorescence-activated cell sorting and transferred into TCRβxδ−/− mice. Colitis was induced by transfer of naïve CBir1-Tg CD4+ T cells into immunodeficient mice.
Microbiota antigen-specific Foxp3+ Treg cells were converted, in the intestine, to IFN-γ+ T-helper (Th)1 cells, interleukin (IL)-17+ Th17 cells, and Foxp3+ T cells that coexpress IFN-γ and/or IL-17. Conversion of Treg cells into IFN-γ-producing Th1 cells and Foxp3+IFN-γ+ T cells required innate cell production of IL-12 in the intestine; blocking IL-12 with an antibody inhibited their conversion to Th1 and Foxp3+IFN-γ+ T cells in the intestines of mice that were recipients of Treg cells. Addition of IL-12, but not IL-23, promoted conversion of Treg cells into Th1 and Foxp3+IFN-γ+ T cells, in vitro. Foxp3+IFN-γ+ T cells had regulatory activity, because they suppressed proliferation of naïve T cells, in vitro, and inhibited induction of colitis by microbiota antigen-specific T cells. IFN-γ+ Th1 cells were not converted into Treg cells; Foxp3+IFN-γ+ T cells differentiated into IFN-γ+ but not Foxp3+ T cells.
IL-12 promotes conversion of Treg cells into IFN-γ-expressing cells; Foxp3+IFN-γ+ T cells retain their regulatory functions and develop during the transition of Foxp3+ Treg cells into IFN-γ+ Th1 cells.
IBD; immune regulation; inflammation; Crohn’s disease
We have previously reported that compromised interleukin 17A (IL-17A) production in the lungs increased susceptibility to infection with the invasive fungal pathogen Aspergillus fumigatus. Here we have shown that culturing lung cells from A. fumigatus-challenged mice ex vivo demonstrated Dectin-1-dependent IL-17A production. In this system, neutralization of IL-23 but not IL-6, IL-1β, or IL-18 resulted in attenuated IL-17A production. Il23 mRNA expression was found to be lower in lung cells from A. fumigatus-challenged Dectin-1-deficient mice, whereas bone marrow-derived dendritic cells from Dectin-1-deficient mice failed to produce IL-23 in response to A. fumigatus in vitro. Addition of recombinant IL-23 augmented IL-17A production by wild-type (WT) and Dectin-1-deficient lung cells, although the addition of IL-6 or IL-1β did not augment the effect of IL-23. Intracellular cytokine staining of lung cells revealed lower levels of CD11b+ IL-17A+ and Ly-6G+ IL-17A+ cells in A. fumigatus-challenged Dectin-1-deficient mice. Ly-6G+ neutrophils purified from the lungs of A. fumigatus-challenged Dectin-1-deficient mice displayed lower Il17a mRNA expression but surprisingly had intact Rorc and Rora mRNA expression. We further demonstrated that Ly-6G+ neutrophils required the presence of myeloid cells for IL-17A production. Finally, upon in vitro stimulation with A. fumigatus, thioglycolate-elicited peritoneal neutrophils were positive for intracellular IL-17A expression and produced IL-17A in a Dectin-1- and IL-23-dependent manner. In summary, Dectin-1-dependent IL-17A production in the lungs during invasive fungal infection is mediated in part by CD11b+ Ly-6G+ neutrophils in an IL-23-dependent manner.
Study of the development of distinct CD4+ T-cell subsets from naive precursors continues to provide excellent opportunities for dissection of mechanisms that control lineage-specific gene expression or repression. Whereas it had been thought that the induction of transcription networks that control T-lineage commitment were highly stable, reinforced by epigenetic processes that confer heritability of functional phenotypes by the progeny of mature T cells, recent findings support a more dynamic view of T-lineage commitment. Here, we highlight advances in the mapping and functional characterization of cis elements in the Ifng locus that have provided new insights into the control of the chromatin structure and transcriptional activity of this signature T-helper 1 cell gene. We also examine epigenetic features of the Ifng locus that have evolved to enable its re-programming for expression by other T-cell subsets, particularly T-helper 17 cells, and contrast features of the Ifng locus with those of the Il17a–Il17f locus, which appears less promiscuous.
cytokines; gene transcription; epigenetics; transcription factors; immunoregulation
During many chronic infections the responding CD8 T cells become exhausted as they progressively lose their ability to elaborate key effector functions. Unlike prototypic memory CD8 cells, which rapidly synthesize IFN-γ following activation, severely exhausted T cells fail to produce this effector molecule. Nevertheless, the ontogeny of exhausted CD8 T cells as well as the underlying mechanisms that account for their functional inactivation remains ill-defined. We have utilized cytokine reporter mice, which mark the transcription of IFN-γ mRNA by the expression of Thy1.1, to decipher how activation events during the early stages of a chronic infection dictate the development of exhaustion. We show that virus-specific CD8 T cells clearly respond during the early stages of chronic lymphocytic choriomeningitis virus (LCMV) infection, and that this early T cell response is more pronounced than that initially observed in acutely infected hosts. Thus, exhausted CD8 T cells appear to emerge from populations of potently activated precursors. Unlike acute infections, which result in massive expansion of the responding T cells, there is a rapid attenuation of further expansion during chronic infections. The exhausted T cells that subsequently emerge in chronically infected hosts are incapable of producing the IFN-γ protein. Surprisingly, high levels of the IFN-γ transcript are still present in exhausted cells, demonstrating that ablation of IFN-γ production by exhausted cells is not due to transcriptional silencing. Thus, post-transcription regulatory mechanisms likely disable this effector module.
Distal cis-regulatory elements play essential roles in the T lineage-specific expression of cytokine genes. We have mapped interactions of three transacting factors – NF-κB, STAT4 and T-bet – with cis elements in the Ifng locus. We find that RelA is critical for optimal Ifng expression and is differentially recruited to multiple elements contingent upon T cell receptor (TCR) or interleukin-12 (IL-12) plus IL-18 signaling. RelA recruitment to at least four elements is dependent on T-bet-dependent remodeling of the Ifng locus and co-recruitment of STAT4. STAT4 and NF-κB therefore cooperate at multiple cis elements to enable NF-κB–dependent enhancement of Ifng expression. RelA recruitment to distal elements was similar in Th1 and Tc1 effector cells, although T-bet was dispensable in CD8 effectors. These results support a model of Ifng regulation in which distal cis-regulatory elements differentially recruit key transcription factors in a modular fashion to initiate gene transcription induced by distinct activation signals.
Phenotypic plasticity of T helper 17 (Th17) cells suggests instability of chromatin structure of key genes of this lineage. Here we identify epigenetic modifications across the clustered Il17a and Il17f, and Ifng loci before and after differential IL-12 or TGFβ signaling, which induce divergent fates of Th17 cell precursors. We find that Th17 precursors have substantial remodeling of the Ifng locus but undergo critical additional modifications to enable high-level expression when stimulated by IL-12. Permissive modifications across the Il17a-Il17f locus are amplified by TGFβ signaling in Th17 cells, but are rapidly reversed downstream of IL-12–induced, STAT4– and T-bet–mediated silencing of the Rorc gene. These findings reveal substantial chromatin instability of key transcription factor and cytokine genes of Th17 cells and support a model of Th17 lineage plasticity in which cell-extrinsic factors modulate Th17 cell fates through differential effects on the epigenetic status of Th17 lineage factors.
Crohn’s disease and ulcerative colitis are the two major forms of chronic relapsing inflammatory disorders of the human intestines collectively referred to as inflammatory bowel disease (IBD). Though a complex set of autoinflammatory disorders that can be precipitated by diverse genetic and environmental factors, a feature that appears common to IBD pathogenesis is a dysregulated effector T cell response to the commensal microbiota. Due to the heightened effector T cell activity in IBD, developmental and functional pathways that give rise to these cells are potential targets for therapeutic intervention. In this review, we highlight recent advances in our understanding of effector T cell biology in the context of intestinal immune regulation and speculate on their potential clinical significance.
Th2 lymphocytes deliver essential signals for induction of asthmatic airway inflammation. We previously found that airway antigen challenge induces recruitment of Gr-1+ neutrophils prior to the recruitment of Th2 cells. We examined, therefore, whether Gr-1+ cells contribute to the development of Th2-dependent airway inflammation. Systemic depletion of Gr-1+ cells using the RB6-8C5 monoclonal antibody reduced Th2 cell recruitment following intranasal antigen challenge. The levels of both matrix metalloproteinase (MMP)-9 and the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) mRNA were up-regulated in the lungs of mice 12 h after intranasal antigen challenge. Up-regulation of TIMP-1 was independent of Gr-1+ cells, whereas up-regulation of MMP-9 RNA and total gelatinolytic activity were dramatically reduced in mice depleted of Gr-1+ cells. At 24 h after challenge, total lung collagenolytic activity was also up-regulated, in a Gr-1+ cell-dependent fashion. Systemic inhibition of MMP-8 and MMP-9 reduced the airway recruitment of Th cells, resulting in significantly reduced eosinophilic inflammation. These data suggest that antigen challenge via the airway activates Gr-1+ cells and consequently MMPs to facilitate the recruitment of Th cells in the airway inflammatory response.
asthma; airway inflammation; MMP; TIMP
TGF-β, together with IL-6 and IL-21, promotes Th17 cell development. IL-6 and IL-21 induce activation of STAT3, which is crucial for Th17 cell differentiation, as well as the expression of SOCS3, a major negative feedback regulator of STAT3-activating cytokines that negatively regulates Th17 cells. However, it is still largely unclear how TGF-β regulates Th17 cell development, and which TGF-β signaling pathway is involved in Th17 cell development. In this report, we demonstrate that TGF-β inhibits IL-6- and IL-21-induced SOCS3 expression, thus enhancing as well as prolonging STAT3 activation in naïve CD4+CD25− T cells. TGF-β inhibits IL-6-induced SOCS3 promoter activity in T cells. Also, SOCS3 siRNA knockdown partially compensates for the action of TGF-β on Th17 cell development. In mice with a dominant-negative form of TGF-β receptor II (TGF-β RII DN) and impaired TGF-β signaling, IL-6-induced CD4+ T cell expression of SOCS3 is higher, whereas STAT3 activation is lower compared to wild type B6 CD4+ T cells. Addition of a TGF-β RI kinase inhibitor that blocks Smad-dependent TGF-β signaling greatly, but not completely, abrogates the effect of TGF-β on Th17 cell differentiation. Our data indicate that inhibition of SOCS3 and thus enhancement of STAT3 activation is at least one of the mechanisms of TGF-β promotion of Th17 cell development.
We report that like other T cells cultured in the presence of transforming growth factor (TGF) β, Th17 cells also produce interleukin (IL) 9. Th17 cells generated in vitro with IL-6 and TGF-β as well as purified ex vivo Th17 cells both produced IL-9. To determine if IL-9 has functional consequences in Th17-mediated inflammatory disease, we evaluated the role of IL-9 in the development and progression of experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis. The data show that IL-9 neutralization and IL-9 receptor deficiency attenuates disease, and this correlates with decreases in Th17 cells and IL-6–producing macrophages in the central nervous system, as well as mast cell numbers in the regional lymph nodes. Collectively, these data implicate IL-9 as a Th17-derived cytokine that can contribute to inflammatory disease.
Activator protein 1 (AP-1) transcription factors are dimers of Jun, Fos, MAF and activating transcription factor (ATF) family proteins characterized by basic region and leucine zipper domains1. Many AP-1 proteins contain defined transcriptional activation domains (TADs), but Batf and the closely related Batf3 (refs 2, 3) contain only a basic region and leucine zipper and have been considered inhibitors of AP-1 activity3–8. Here we show that Batf is required for the differentiation of IL-17-producing T helper (TH17) cells9. TH17 cells comprise a CD4+ T cell subset that coordinates inflammatory responses in host defense but is pathogenic in autoimmunity10–13.Batf
−/−mice have normal TH1 and TH2 differentiation, but show a defect in TH17 differentiation, and are resistant to experimental autoimmune encephalomyelitis (EAE).Batf
−/−T cells fail to induce known factors required for TH17 differentiation, such as RORγt11 and the cytokine IL-21 (refs 14–17). Neither addition of IL-21 nor overexpression of RORγt fully restores IL-17 production in Batf−/− T cells. The IL-17 promoter is Batf-responsive, and upon TH17 differentiation, Batf binds conserved intergenic elements in the IL-17A/F locus and to the IL-17, IL-21 and IL-22 (ref 18) promoters. These results demonstrate that the AP-1 protein Batf plays a critical role in TH17 differentiation.
LFA-1 (CD11a/CD18) is a member of the β2-integrin family of adhesion molecules important in leukocyte trafficking and activation. Although LFA-1 is thought to contribute to the development of experimental autoimmune encephalomyelitis (EAE) primarily through its functions on effector T cells, its importance on other leukocyte populations remains unexplored. To address this question, we performed both adoptive transfer EAE experiments involving CD11a-/- mice and trafficking studies using bioluminescent T cells expressing luciferase under the control of a CD2 promoter (T-lux cells). Transfer of encephalitogenic CD11a-/- T cells to wild type mice resulted in a significant reduction in overall EAE severity compared to control transfers. We also observed, using in vivo imaging techniques, that CD11a-/- T-lux cells readily infiltrated lymph nodes and the CNS of wild type recipients with kinetics comparable to CD11a+/+ transfers, although their overall numbers in these organs were reduced. Surprisingly, transfer of encephalitogenic wild type T cells to CD11a-/- mice induced a severe and sometimes fatal EAE disease course, associated with massive T cell infiltration and proliferation in the CNS. These data indicate that LFA-1 expression on leukocytes in recipient mice plays an important immunomodulatory role in EAE. Thus, LFA-1 acts as a key regulatory adhesion molecule during the development of EAE, serving both pro- and anti-inflammatory roles in disease pathogenesis.
adhesion molecules; β2-integrins; T lymphocytes; bioluminescence; neuroimmunology
Recent progress in our understanding of mechanisms by which the immunosuppressive cytokine interleukin-10 (IL-10) participates in an ever-increasing diversity of T-cell lineages to maintain immune homeostasis has broadened the framework for defining regulatory and effector T cells and has blurred the lines between them. In this review, we highlight established and emerging roles for IL-10 produced by distinct CD4+ T-cell lineages that underlie its non-redundant role in curbing immune responses to the intestinal microbiota at steady state and its role to limit T-cell-driven inflammation in responses to pathogens.
T cells; Th1/Th2/Th17 cells; cytokines; lineage commitment