Browsing by Subject "T-Lymphocytes"
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Item BTB-Kelch Proteins and TAk1 Kinase N Immune Function(2006-12-19) Liu, Hong-Hsing; Chen, ZhijianBTB-kelch proteins are putative components of E3 ligases with a BTB domain at the N terminus and several kelch repeats at the C terminus. KLHL6 and mKELCH are two members of this family. Conditional ablation of Klhl6 in B cells resulted in mild developmental phenotypes in bone marrow precursors. The number of peripheral B cells was decreased by half, and responded defectively in germinal center formation after antigen stimulations. mKELCH is a novel protein cloned from hearts. Knocked-in LacZ expressed predominantly at muscles and several photosensitive organs. TAK1 is a member of MAPKKK. Deletion of TAK1 prevented the maturation of CD4+ or CD8+ single positive thymocytes, leading to reduction of T cells in peripheral tissues. Thymocytes lacking TAK1 failed to activate NF-κB and JNK, and were prone to apoptosis upon stimulation. All three mouse models have provided important evidences in elucidating biological functions for each protein in vivo.Item Counter-Regulation of Human Cd4+ T Helper 2 Lymphocyte Development and Stability by Type I Interferon(2013-01-16) Huber, Jonathan Philip; Farrar, J. David, Ph.D.Innate cytokines shape the differentiation of CD4+ T cells from naïve precursors into multiple functional subsets in order carry out effective adaptive immune responses to diverse immunological stimuli. Interleukin-12 controls the development of T helper 1 (Th1) cells, which fight infection by intracellular pathogens such as bacteria and viruses. Helminth parasites and allergens induce the production of IL-4, which drives differentiation to Th2. The IL-12 and IL-4 signaling pathways also counter-regulate each other, though this balance favors Th2 as IL-4 signaling overrides the effects of IL-12. In addition, IL-4 induces expression of the GATA3 transcription factor that stabilizes the Th2 phenotype through auto-regulatory maintenance of its own expression. A role for type I interferon (IFN-a/b), a major antiviral cytokine, has been described more recently in Th1 cells, but the potential role of IFN-a/b in counter-regulating Th2 development is poorly understood. My work reveals that IFN-a/b blocks IL-4-induced Th2 differentiation of human CD4+ T cells by inhibiting expression of GATA3. The loss of GATA3 in developing Th2 cells leads to reduced secretion of the Th2 cytokines, IL-4, IL-5, and IL-13, and reduced expression of chemoattractant receptor expressed in Th2 cells (CRTh2). IFN-a/b also suppresses GATA3 in committed Th2 cells, leading to a loss of the Th2 phenotype. GATA3 transcription utilizes two distinct first exons, exon 1A and 1B, which are controlled by separate promoters, but IL-4 signaling in Th2 cells leads to preferential utilization of the upstream exon 1A transcript. IFN-a specifically blocks expression of the 1A but not 1B transcript, indicating a specific regulation of Th2 cells. Furthermore, IFN-a appears to induce epigenetic silencing of an upstream conserved non-coding sequence I, which may be required for optimal exon 1A transcription in Th2 cells. This work reveals an unexpected role for IFN-a in selectively inhibiting Th2 but not Th1 differentiation, which may be important for ensuring appropriate development of antiviral immunity. In addition, the ability of IFN-a to suppress both developing Th2 cells and previously committed Th2 cells suggests that IFN-a may be useful as a novel therapeutic for atopic diseases.Item The Function of the TCR zeta zeta Module in T Cells(2005-04-29) Pitcher, Lisa Anne; Oers, Nicolai vanThe alpha beta T cell receptor complex (TCR) has the unique ability to discriminate and differentially respond to peptide/MHC ligands encountered on the surface of antigen presenting cells. The ligation of the TCR with peptide/MHC complexes is translated into intracellular signals through a conserved sequence motif, termed ITAM, or immunoreceptor tyrosine-based activation motif, which are present in one or more copies in the cytoplasmic portions of the TCR zeta and CD3 gamma, delta and epsilon chains. A distinctive feature of the TCR complex is that it contains ten ITAMs, in contrast to other antigen receptor complexes which contain two or four ITAMs. The ten TCR ITAMs are distributed as one in each CD3 chain and three in the TCR zeta subunit (TCR alpha beta epsilon delta epsilon gamma zeta zeta). It has been proposed that the TCR is comprised of two autonomous signaling modules, TCR zeta zeta and CD3 gamma epsilon/delta epsilon. Following receptor ligation, TCR zeta is the most heavily tyrosine-phosphorylated subunit of the TCR, developing into two stable intermediates of 21- and 23-kDa (p21 and p23). Based on the number of ITAMs it contributes, TCR zeta zeta was initially presumed to be the predominant signaling module in the TCR complex, with p21 and p23 being linked to virtually all aspects of T cell biology. To clearly define the functions of the TCR zeta zeta module, we generated a series of TCR zeta transgenic mice, with modified zeta molecules, that selectively express p21 alone, no p21 or p23, or no phospho-zeta intermediates. In a wild type or high affinity TCR system (P14), T cell development was completely normal in the TCR zeta transgenic lines. Surprisingly, when peripheral T cells were analyzed for their functionality in response to various stimuli, including peptide/MHC-, T cell mitogen- and superantigenic- stimulation, equivalent dose response curves were observed, regardless of phospho-zeta. Notably, these data also eliminated a possible inhibitory role for the partially phosphorylated p21 intermediate of TCR zeta. To more carefully examine the roles of p21 and p23, the TCR zeta transgenics were mated to a TCR transgenic line (HY) bearing T cells with a low affinity TCR. In this system, important roles for phospho-zeta during T cell selection were revealed. Specifically, TCR zeta ITAMs functioned additively during positive selection events. These selection events appear to be independent of traditional signaling pathways, as the signaling capacity of unselected T cells in the absence of all phospho-zeta was equivalent to T cells with wild-type TCR zeta subunits. These results imply that the CD3 gamma epsilon/delta epsilon module is the predominant signaling module in the TCR complex. Our studies also identified a unique role for p21 during negative selection events. The select expression of p21 in T cells attenuated negative selection of thymocytes, resulting in the generation of a population of potentially autoreactive cells. Based on these data, a revised model of TCR signal transmission is proposed. Within this model, the TCR zeta zeta and CD3 gamma epsilon/delta epsilon modules contribute both redundant and unique functions to T cells. The CD3 gamma epsilon/delta epsilon module is primarily responsible for classical TCR-mediated signaling pathways leading to T cell activation. The TCR zeta zeta and CD3 gamma epsilon/delta epsilon modules contribute redundant functions to thymocyte positive selection. These redundant functions are mediated by the ten TCR ITAMs. Phosphorylated intermediates of the TCR zeta zeta module also contribute to thymocyte positive selection, likely through alternative signaling pathways. In addition, the TCR zeta zeta module functions in a unique manner during thymocyte negative selection, with p21 attenuating negative selection of thymocytes. Furthermore, preliminary evidence suggests novel roles for the TCR zeta zeta module in the maintenance of peripheral T cells and in the adaptive immune response to bacterial pathogens.Item In Vivo Identification of SLE1B: LY108 Mediates Autoantibody Production(2008-05-12) Chan, Alice; Wakeland, Edward K.In the NZM2410 model of murine lupus, Sle1b mediates anti-nuclear autoantibody (ANA) production. Our goal is to determine the causative gene in the Sle1b locus. Seven members of the SLAM/CD2 family are located within the Sle1b interval, and previous work has shown that structural and expression polymorphisms in lymphocytes distinguish two major SLAM/CD2 haplotypes. To further narrow the interval, we utilized a BAC transgenic rescue approach whereby BACs carrying the lupus-resistant B6 alleles were bred to B6.Sle1b mice to identify the region mediating ANA suppression. One BAC carrying Cd84 and Ly108 suppressed autoantibody production. We then generated BAC transgenic mice carrying the lupus-susceptible (129) and lupus-resistant (B6) alleles of Ly108 on the B6 and B6.Sle1b genetic background, respectively. The B6 allele of Ly108 suppresses ANA production on the lupus-susceptible B6.Sle1b background while the 129 allele induces ANA on the lupus-resistant B6 genome. Taken together, these data identify Ly108 as a causative gene in Sle1b. While Ly108 is needed to mediate the breach in tolerance, we have also identified other SLAM family members as genetic modifiers necessary to recapitulate fully penetrant, high titer ANA production as seen in Sle1b. We found that in vitro stimulation of B6.Sle1b CD4 T cells led to altered cytokine production, such as decreased IL4 production. Interestingly, these phenotypes have also been reported in knockouts of SLAM/CD2 family members as well as in the absence of the SLAM family adaptor, SAP. Our data indicates that the presence of the Sle1b haplotype, derived from either NZM2410 or 129, recapitulates these phenotypes, independent of the absence of these molecules. While recent reports have suggested a role for SAP in ANA development, we find that the breach in tolerance in Sle1b mice is SAP-independent. However, SAP is necessary to potentiate the autoantibody production. ANAs is an important biomarker for autoimmune diseases including, Systemic Lupus Erythematosus (SLE), and potentially identifies an autoimmune-prone state. We have identified genes which contribute to the production of ANAs. Elucidating the pathways these genes dysregulate will provide critical insight into our understanding of tolerance and how tolerance can be breached.Item Itk is a Critical Regulator of Spatiotemporal Localization at the Immunological Synapse(2010-05-14) Singleton, Kentner Leroy; Wulfing, ChristophThe activation of T cells by antigen presenting cells (APCs) is an important step in the initiation of the adaptive immune response. Itk, a member of the Tec family of non-receptor protein tyrosine kinases, is important for T cell activation – Itk-/- CD4+ T cells are hyporesponsive, displaying decreased calcium flux, proliferation, and IL2 production compared to wildtype T cells. The mechanism by which Itk mediates this effect is not fully elucidated. Here we show that Itk is a key regulator of spatiotemporal localization of receptors and proximal signaling intermediates at the T cell / APC interface. As part of this organizational regulation, we found that Itk, through the recruitment of SLAT, mediates activation of Cdc42 at the center of the interface, which is critically required for actin polymerization. We show that targeting activated Cdc42 to the center of the interface restores actin polymerization in the Itk-/- T cell while the addition of constitutively active Cdc42 to the entirety of the interface cannot. These results provide beginnings of a mechanistic explanation of how Itk both regulates the actin cytoskeleton and acts to amplify T cell signaling. These results further demonstrate that control of protein localization at the immunological synapse can be the critical determinant in protein function and that the center of the interface is a site of active signaling.Item Molecular Characterization of the Human Alloimmune Response(2009-11-18) Smith, Melody; Collins, RobertWhen T lymphocytes from two individuals are placed in a mixed lymphocytic reaction (MLR) and allowed to stimulate each other a powerful proliferative response is elicited, reflecting the degree of incompatibility at the major histocompatibiltiy loci. Such in vitro proliferations translate in vivo into graft rejection of stem cell and solid organ transplants and to graft versus host disease (GvHD) in recipients of allogeneic stem cell transplants. Defining the clonal T cell responses elicited in an MLR between two individuals is complicated by the multiplicity of T cell HLA- antigen interactions. We used flow cytometry to investigate the MLR between mismatched individuals to better characterize responder T cell proliferation and the subsets involved in the alloresponse. After observing proliferation in the setting of a completely mismatched one-way MLR and ensuring that the design for the MLR was viable, we sought to characterize clonal T cell responses in a simple system. We attempted to elicit T cell responses to autologous antigen-presenting cells (APC) transfected with a single mismatched HLA-A2 molecule. We failed to detect any alloresponding T cells using a flow cytometric system, except when dendritic cells were used. We then used an HLA-A2 negative responder pre-stimulated in an MLR with an HLA-A2 positive individual for re-challenge by the responder?s own APC transfected with an HLA-A2 plasmid. In this way, we sought to identify an expanding HLA-A2 specific T cell population which could be further characterized by T cell receptor cloning. Nevertheless, we failed to detect an alloimmune response in this T cell line stimulated with HLA-A2 transfected APCs three times over a 21- day culture period. These results suggest that either the frequency of alloreactive T cells was below the limit of detection by these methods or that the particular stimulator-responder pairs used in these experiments were non-reactive. Future work will extend the study to more HLA mismatched stimulator-responder pairings to better define the characteristics of alloreactive T cells.Item Origin and Function of CD8 T Cells in MHC Class Ia-Deficient Mice(2005-12-20) Su, Jie; Forman, JamesB6.H-2Kb-/-Db-/- (DKO) mice are devoid of class Ia but express normal levels of class Ib molecules. They have low levels of CD8 T cells in both the thymus as well as peripheral T cell compartments. Although the percentage of splenic CD8aa T cells is increased in these animals, approximately 90% of CD8 T cells are CD8abeta TCRabeta . In contrast to B6 animals, most of the CD8 T cells from these mice have a memory phenotype (CD44hi) including both CD8abeta and CD8aa subsets. In the thymus of DKO animals, there is a decrease in the percentage of single positive CD8 T cells, although most are CD44low, similar to that seen in B6 mice. Our results indicated that the paucity of CD8 T cells in DKO mice might be in part due in reduced thymic export, lower basal proliferation, high apoptosis, and inability to undergo homeostatic expansion. DKO mice have greatly reduced numbers of mature CD8abeta T cells in their periphery. However, these non class Ia selected CD8abeta cells are able to mediate immune responses to a number of pathogens. Approximately 60% of the CD8abeta T cells in the spleen and peripheral lymph nodes of na?DKO mice display a memory (CD44hi) phenotype. To investigate the origins of these non class Ia selected CD8abeta CD44hi cells, we traced the phenotype of recent thymic emigrants (RTEs) and found that most were CD44lo. We also determined if their appearance was thymus dependent and found that only a small percentage of non class Ia selected CD8abeta CD44hi cells develop in a thymus-independent pathway. Functionally, CD8abeta CD44hi cells from DKO mice are able to secrete IFN-gamma in response to interleukin (IL)-12 and IL-18 in the absence of cognate antigen. When challenged with anti-CD3 in vivo, nearly half of these cells produce IFN-gamma within 3 hours. When purified CD8abeta CD44hi cells from Thy1.2.DKO mice were transferred into Thy1.1 DKO recipients and then challenged with Listeria monocytogenes (LM), an antigen specific anti-LM response was observed six days later. Our data suggest that non class Ia selected CD8abeta CD44hi cells in na?animals can respond to antigen and play a role in the innate as well as the early phase of the acquired immune response.Item PIP5-Kinase Dependent PIP2 Generation Impairs T Cell Activation by Rigidifying the Actin Cortex(2010-05-14) Sun, Yi; Wulfing, ChristophAntigen-specific activation of T cells is initiated by the interaction of T cell receptors (TCRs) with the peptide-loaded MHC complex on antigen presenting cells (APCs). A specialized junction, the immunological synapse (IS), is formed at the T cell-APC interface, where cell surface receptors, signaling molecules as well as cytoskeletal elements rearrange into a highly organized structure (Monks et al., 1998; Grakoui et al., 1999; Cemerski et al., 2006; Fooksman et al., 2009). Dynamics actin and remodeling plays a pivotal role in IS formation and T cell activation (Wülfing et al. 1998, 2003; Dustin et al., 2000; Billadeau et al., 2007; Burkhardt et al., 2008; Gomez et al., 2008). It is believed that the activities of many actin regulatory proteins are spatially and temporally regulated by phosphatidylinositol 4,5-bisphosphate (PIP2) (Sechi et al, 2000; Yin et al., 2003; Logan et al., 2006). How actin dynamics is regulated by PIP2 in T cell activation is unclear. Generation of PIP2 at the plasma membrane can be accomplished by the three isoforms of the type I phosphatidylinositol 4-phosphate 5-kinases (PIP5Ks): α, β, and (Ishihara et al., 1996; Loijens and Anderson, 1996; Oude Weernink et al., 2004). It is proposed that different PIP5K isoforms synthesize distinct pools of PIP2 in the plasma membrane and contribute to the versatility of PIP2 function (Kanaho et al., 2007; van den Bout et al., 2009). The roles of these PIP5Ks in T cell activation have not been addressed. The experiments in this thesis are designed to investigate how regulation of PIP2 levels by changing cellular PIP5K levels affects actin cytoskeletal dynamics at the IS and T cell activation. The major findings of this thesis are: PIP5K isoforms are differentially targeted during T cell activation by APCs. PIP5Kα and γ87 are mainly targeted to the IS, whereas β and γ90 isoforms accumulate at the uropod or distal pole region of the T cells. Overexpression of PIP5Ks caused defective actin polymerization at the periphery of the IS and rigid T cell morphology, impaired TCR proximal signaling and decreased IL-2 secretion. Knockdown of PIP5Kγ instead induced sustained peripheral actin accumulation at the IS, enhanced T cell proximal signaling and IL-2 secretion. Furthermore, we found that the activity of the actin-binding ERM proteins was impaired in PIP5K overexpressing T cells. Taken together, we propose that PIP5K dependent PIP2 production inhibits ERM inactivation triggered by TCR stimulation, which leads to increased T cell rigidity, defective actin dynamics and T cell effector function.Item The Role of Chronically Stimulated and Senescent T Cells in Autoimmunity(2006-09-25) Ratts, Robert Bruce; Racke, Michael K.Myelin-reactive T cells have been hypothesized to play a role in the pathogenesis of MS. If this is the case, these T cells would be expected to be repeatedly stimulated over the course of this disease because of the episodic breakdown of myelin membranes. Continuous stimulation of myelin-reactive T cells may also change the phenotype of these T cells. For example it may drive autoreactive T cells into senescence. Senescent T cells would be unable to divide but may still retain effector functions such as the ability to lyse target cells. The existence of senescent autoreactive T cells in MS patients could explain why therapies that limit the proliferation of T cells have had little effect on the course of MS, particularly later in the course of the disease. Human T cells adopt a CD28-CD57+ phenotype in chronic viral infections and this has been hypothesized to result from chronic stimulation, however this phenotype may also be due to direct viral effects on T cells. Here I make use of human MS patients before and after chronic in vivo administration of the antigen glatiramer acetate to test this hypothesis. Before the initiation of glatiramer acetate treatment, glatiramer acetate -specific CD8 T cells were either CD28-CD57- or CD28+CD57-. This response changed to a predominantly CD28-CD57+ response after one year of continuous stimulation. This phenotype was only observed after chronic stimulation and not in a recall response to mumps. These cells were shown to contain perforin, indicating they likely play a cytotoxic role in vivo. Furthermore CD28-CD57+ CD8 T cells displayed a reduced proliferative capacity indicating they may be senescent or pre-senecent. When myelin-reactive T cell responses were examined a CD28-CD57+ CD8 T cell response could be detected in MS patients but not in healthy controls. These T cells contained mRNA consitant with a cytotoxic role and the abilty to home to the cerebrospinal fluid of MS patients. This observation may explain why therapies that limit the proliferation of T cells have had little effect on the course of MS, particularly later in the course of the disease.Item The Role of the PTPH1-Family of Protein Tyrosine Phosphatases In T Cells(2011-11-04) Young, Jennifer Ann; Van Oers, NicolaiThe alpha beta T cell receptor complex activates intracellular signaling cascades by coupling to several families of protein tyrosine kinases and protein tyrosine phosphatases. Following T cell receptor interactions with cognate peptide/major histocompatibility complexes, Src-family protein tyrosine kinases phosphorylate two tyrosine residues in a conserved amino acid motif termed the immunoreceptor tyrosine-based activation motif. The immunoreceptor tyrosine-based activation motifs are present in one or more copies in the cytoplasmic tails of the T cell receptor invariant chains, CD3 gamma, delta, epsilon and zeta. In all T cell receptor signaling events, the immunoreceptor tyrosine-based activation motifs are transiently phosphorylated. We used a large-scale screen to identify protein tyrosine phosphatase candidates dephosphorylating the T cell receptor zeta immunoreceptor tyrosine-based activation motifs. PTPH1 was identified in this screen, but its endogenous expression was difficult to detect in T cells. Based on sequence homology, PTPN4 was also considered as a putative regulator of phospho-zeta. T cell receptor zeta was bound and dephosphorylated by PTPN4. Overexpression of wild-type PTPN4 inhibited T cell receptor-induced AP-1 and NFkappaB activation in T cells. The overexpression of a substrate-trapping derivative of PTPN4 significantly augmented NFkappaB activation. This finding demonstrates a role for PTPN4 in the regulation of the NFkappaB pathway. PTPN4 knock-out mice were generated to assess the role of this PTPase in lymphocytes. This is the first description of a PTPN4-deficient animal model. No major developmental defects were observed in the PTPN4 knock-out mice. However, PTPN4-deficient animals had altered peripheral effector/memory T cell populations. While early T cell activation and T cell receptor-induced signal transduction events were normal, peripheral T cells from PTPN4-null mice secreted elevated levels of IL-4, IL-5, and IL-13 in the absence of PTPN4. This suggests that PTPN4 has a selective role in regulating effector T cell differentiation. Taken together, both PTPH1 and PTPN4 may regulate T cell receptor zeta phosphorylation, but the unique functions of these two protein tyrosine phosphatases indicate non-overlapping substrates.Item Soluble Peptide Treatment Reverses CD8 T Cell-Induced Disease in a Mouse Model of Spontaneous Tissue-Selective Autoimmunity(2011-10-25) Paek, So Yeon; Katz, Stephen I.Autoimmunity is a complex process that involves recognition of self antigens by autoreactive T cells or tissue targeting by autoantibodies produced by B cells. Specific molecular targets have been identified in several autoimmune diseases but remain unknown in many others. Transgenic (Tg) mice have been utilized to express model antigens that can be recognized by T cells or by autoantibodies. To identify mechanisms by which CD8+ autoreactive T cells cause inflammation, we generated a double transgenic (DTg) murine model of autoimmunity by crossing K14-sOVA mice, which express soluble chicken ovalbumin (OVA), with OT-1 mice, whose CD8 T cells express V2/V5 regions of the T cell receptor that are specific for SIINFEKL peptide (OVA 257-264) in association with class I MHC molecules. The K14-SOVA/OT-1 (#5 and #17) DTg mice develop normally, except that they undergo a destructive process that selectively targets the external pinnae in the first 6 days of life. The purpose of this study was to elucidate the mechanism and attempt to obviate the resulting tissue-specific destruction. By light microscopy, the ear bud area displayed an intense inflammatory infiltrate of V2/V5+CD8+ OT-1 cells when characterized by FACS. Administration of the TCR-recognized SIINFEKL peptide i.v. to pregnant F1 mice on days E16 and E18 in utero and i.p. to newborn pups on days 2 and 4 prevented the inflammatory response and resulted in development of normal-looking ears in 100% of pups. Treatment with the SIINFEKL peptide was shown to down-regulate the CD8 coreceptor and activate T-cells to differentiate into memory T-cells. This model can inform us about mechanisms of peripheral tolerance and potential therapies for autoimmune diseases in which specific molecular targets are known.Item Transcriptional Regulation of T Helper Cell Differentiation in Autoimmunity(2006-12-19) Gocke, Anne Elizabeth; Racke, Michael K.As a means to understand the significance of transcriptional regulation of Th cells in MS pathogenesis, we investigated the role of the transcription factors T-bet and PPARalpha in the differentiation and effector function of T helper cells in the EAE model. Based on its ability to induce a pro-inflammatory immune response, T-bet has been found to regulate genes that have been directly linked to pathogenicity in EAE/MS. We examined the use of T-bet specific AS oligonucleotides and siRNA to silence T-bet expression in autoreactive encephalitogenic T cells and demonstrated that this transcription factor plays a critical role in the differentiation of these cells and in the induction of EAE. siRNA and AS T-bet suppressed T-bet expression, IFN-gamma production, and STAT1 expression during antigen-specific T cell differentiation. In vitro suppression of T-bet in myelin specific cells used for adoptive transfer, and in vivo administration of AS and siRNA T-bet inhibited the induction of EAE. T-bet bound the IFN-gamma and STAT1 promoter regions, suggesting the ability of T-bet and STAT1 to regulate one another in a positive feedback loop, but was not found to regulate the IL- 12/STAT4 pathway. Moreover, we explored the role of T-bet in the IL-23/IL-17 pathway. IL-17 producing T cells have recently been implicated in the pathogenesis of EAE, but the potential role of T-bet in the differentiation of Th17 cells is not completely understood. We demonstrated that therapeutic administration of siRNA T-bet significantly improved the clinical course of established EAE. The improved clinical course was associated with a decrease in T-bet protein expression, reduced IFN-gamma production, and a reciprocal increase in GATA3 expression. We also observed a decrease in IL-23R expression both ex-vivo in splenocytes from siRNA T-bet treated mice and when cells were transfected with siRNA T-bet and subsequently activated in vitro. When T-bet was silenced we observed an ensuing decrease in both IL-23R and IL-17 expression in the CNS. This observation was correlated by the finding that T-bet directly regulates transcription of the IL-23R and, in doing so, influences the fate of pathogenic Th17 cells, which depend on optimal IL-23 responsiveness for survival. Taken together, these data demonstrate that suppression of T-bet ameliorates EAE by limiting differentiation of autoreactive Th1 cells, as well as inhibiting pathogenic Th17 cells via regulation of the IL-23R. In contrast to T-bet, PPARalpha has been demonstrated to regulate anti-inflammatory genes and the PPARalpha agonists gemfibrozil and fenofibrate have been associated with protection from EAE. However, the manner in which these agents ameliorate disease is not well understood. We investigated the mechanism by which gemfibrozil induces immune deviation and protects mice from EAE. We demonstrated that treatment with gemfibrozil increases GATA3 and decreases T-bet expression in vitro and directly ex-vivo. The protective effects of gemfibrozil in EAE were shown to be partially dependent on IL-4 and to occur in a receptor dependent manner. Moreover, PPARalpha was demonstrated, for the first time, to regulate the IL-4 and IL-5 genes and bound the IL-4 promoter in the presence the coactivator SRC-1, suggesting transactivation of the IL-4 gene. Finally, therapeutic administration of gemfibrozil and fenofibrate ameliorated clinically established EAE. The data presented herein illustrate that modulation of specific transcription factors can influence Th cell differentiation and the clinical outcome of EAE, and suggest that targeting T-bet and/or PPARalpha by gene silencing and administration of specific agonists could provide a novel therapy for inflammatory immune-mediated diseases such as MS.