Th1 Cell Lines Research Articles

Th1 and Th2 cytokine expression in hyperkeratotic chronic hand eczema and the role of Tofacitinib a oral JAK inhibitor.

Tissue cytokines in chronic hand eczema (CHE) can predict targeted therapy with novel drugs including JAK inhibitors. Our primary objective was to assess the tissue expression of cytokines of Th1 and Th2 cell lines in CHE patients and to study the efficacy of oral tofacitinib. We recruited patients presenting with recalcitrant CHE. Lesional and non-lesional tissue samples were assessed for Th1(IFN-γ, TNF-α) and Th2 related cytokines (IL-4, IL-13, IL-2,) using real time PCR. Tofacitinib 5mg twice daily was initiated with 4 weekly assessment and we also noted relapses post therapy.Of 21 refractory hyperkeratotic CHE patients, cytokine analysis was performed in 11 patients which showed upregulation of IL-4 [n = 5/11, 1.87-fold increase], TNF-α (n = 5/11, 5.13-fold) and IFN-γ (n = 6/11, 1.98-fold) as compared to uninvolved skin. All patients (100%) had used topical corticosteroids (TCS) and 4/21 (19%) had failed methotrexate and 2/21 (9.5%) had failed acitretin. Tofacitinib 5mg twice daily was given in 15/21 patients. The mean time to achieve hand eczema severity index 90 (HECSI 90) was 4 weeks (mean duration of treatment:5.8 months, n = 12). Side effects were observed in 4/12 (33.3%) patients and relapse was noted in 3/12 (25%) patients after a mean duration of 7 months after discontinuation of tofacitinib. Tofacitinib (pan-JAK inhibitor) showed an excellent response in refractory CHE patients with predominant tissue Th1/Th2 cells related cytokine expression.

ENHO, RXRA, and LXRA polymorphisms and dyslipidaemia, related comorbidities and survival in haemodialysis patients

BackgroundThe energy homeostasis-associated gene (ENHO), retinoid X receptor alpha gene (RXRA), and liver X receptor alpha gene (LXRA) are involved in adipogenic/lipogenic regulation. We investigated whether single-nucleotide polymorphisms in these genes (ENHO rs2281997, rs72735260; RXRA rs749759, rs10776909, rs10881578; LXRA rs2279238, rs7120118, rs11039155) are associated with dyslipidaemia, related comorbidities and survival of haemodialysis (HD) patients also tested for T-helper (Th) cell interleukin genes (IL).MethodsThe study was carried out in 873 HD patients. Dyslipidaemia was diagnosed by the recommendations of the Kidney Disease Outcomes Quality Initiative (K/DOQI) guidelines (2003); atherogenic dyslipidaemia was referred to if the TG/HDL cholesterol ratio was equal to or higher than 3.8. Genotyping of ENHO SNPs, LXRA SNPs, and IL12A rs568408 was carried out using HRM analysis. RXRA SNPs, IL12B rs3212227, and IL18 rs360719 were genotyped using PCR-RFLP analysis. The circulating adropin concentration was determined in 126 patients by enzyme-linked immunosorbent assay. Survival probability was analysed using the Kaplan-Meier method in 440 patients followed through 7.5 years.ResultsDyslipidaemia by K/DOQI was diagnosed in 459 patients (91% revealed hyper-LDL- cholesterolaemia), atherogenic dyslipidaemia was diagnosed in 454 patients, and 231 patients were free of dyslipidaemia by both criteria. The variant allele (T) of ENHO rs2281997 was associated with the hyper-LDL cholesterolaemic pattern of dyslipidaemia by K/DOQI. The frequency of atherogenic dyslipidaemia was lower in T-allele bearers than in CC-genotype patients. The rs2281997 T allele was associated with lower cardiovascular mortality in HD patients showing atherogenic dyslipidaemia. ENHO, RXRA, and LXRA showed epistatic interactions in dyslipidaemia. Circulating adropin was lower in atherogenic dyslipidaemia than in non-atherogenic conditions. RXRA rs10776909 was associated with myocardial infarction. Bearers of LXRA rs2279238, rs7120118 or rs11039155 minor alleles showed higher mortality. ENHO SNP positions fell within the same DNase 1 hypersensitivity site expressed in the Th1 cell line. Epistatic interactions occurred between rs2281997 and Th1 IL SNPs (rs360719, rs568408).ConclusionsAtherogenic dyslipidaemia occurs in HD patients in whom ENHO encodes less adropin. ENHO, RXRA, and LXRA SNPs, separately or jointly, are associated with dyslipidaemia, myocardial infarction, and survival in HD patients. Differences in the availability of transcription binding sites may contribute to these associations.

Inhibition of Inositol-Requiring Enzyme 1 α in Established TH2 Cells Decreases the Secretion of Cytokines IL-5 and IL-13, but Not IL-4

Abstract TH2 cells are crucial in immune responses against extracellular pathogens; however, they can play a predominant role in a number of diseases due to inappropriate IL-4, IL-5, and IL-13 cytokine expression. Therefore, understanding how these cells develop and regulate their effector functions is of great importance. In response to an excess of unfolded proteins, serine-threonine kinase Inositol-Requiring Enzyme 1 α or IRE1α activates the unfolded protein response, UPR, which promotes increased protein folding capacity. T cell activation induces ER stress, and inhibition of IRE1α RNase activity in activated primary cells, via treatment of the cells with the commercially available drug 4μ8c, results in the reduction of IL-5, IL-4, and IL-13. Prior to this work, it was unknown if 4μ8c could inhibit TH2 cytokines in established TH2 cells. Treatment of a mouse TH2 cell line and differentiated human TH2 cells with 4μ8c results in inhibition of IL-5 and IL-13, but not IL-4 as measured by ELISA. The reduced cytokine expression is not due to differences in mRNA stability or mRNA levels; it appears to be due to a defect in secretion, as the cells appear to produce cytokines IL-5 and IL-13 as measured by flow cytometry. This suggests that the inhibition of these cytokines is due to post-translational processes. IL-5 and IL-13 produced by TH2 cells are critical for promoting chronic, inflammatory asthma, and 4μ8c blocks their expression in T cells in vitro. Future studies will determine if 4μ8c treatment can ameliorate the effects of these cytokines in a disease model. This work is supported by the NSU Faculty Research Council grant and OK-INBRE NIH P20GM103447.

A murine model of spontaneous self-reactivity to the ileum

Abstract Inflammatory lesions of Crohn’s disease are commonly found in the ileum yet few mouse models of enterocolitis manifest inflammation at this site. In order to study inflammation targeted to the ileum, we developed a transgenic mouse line (HD5-mHEL/Hb) that expressed a model antigen (mHEL/Hb) using a Paneth cell promoter (HD5). The abundance of Paneth cells is unique to the ileum and variations in their antimicrobial-secreting function are associated with Crohn’s disease. HEL-reactive (3A9) transgenic TH cells accumulate in the ileum of lymphopenic HD5-mHEL/Hb mice following their adoptive transfer. To study spontaneous TH cell reactivity to the ileum, we bred HD5-mHEL/Hb lines to mHEL/Hb-reactive transgenic TH cell lines. A subset of progeny with both the mHEL/Hb antigen and the 3A9 transgene (Bigenic3A9) develop a spontaneous wasting phenotype that is associated with colitis. Bigenic3A9 mice show TH cell activation and accumulate increased numbers of IL-17A-producing TH cells in the ileum. To study the necessity of effector cytokines for the spontaneous disease of Bigenic3A9 mice, Bigenic3A9 mice were bred on IFNγ and IL17A knockout lines. IL17A−/− Bigenic3A9 mice do not show an altered predisposition to the wasting phenotype of IL17A+/+ Bigenic3A9 mice. Unexpectedly, IFNγ−/− Bigenic3A9 mice exhibit an increased incidence of wasting disease which is associated with a skew of TH cell populations towards an IL-17A-producing phenotype. These results identify a significant protective role for IFNγ in Bigenic3A9 mice and suggest protection may result from inhibitory effects on the polarization of IL-17A-producing TH cells.

IL-1β enhances inflammatory TH2 differentiation

IL-1β enhances inflammatory TH2 differentiation

Defining characteristics of classical Hodgkin lymphoma microenvironment T-helper cells

AbstractCD4+ T-helper cells (THs) dominate the classical Hodgkin lymphoma (CHL) microenvironment, but their role is poorly understood. Advances in flow cytometry and immunohistochemistry permit more detailed investigation of this aspect of CHL pathophysiology. To address the hypothesis that the TH-infiltrate, rather than being TH2-enriched, senescent and hypofunctional, is TH1 and activation marker-rich, cytokine-secretory and proliferative, we applied comprehensive flow cytometric immunophenotyping and functional assays of cytokine secretion/proliferation to TH cells from 18 CHL-derived single-cell suspensions (SCSs) compared to reactive lymph nodes (RLNs). CHL-derived TH cells express TH1-associated CXCR3/CCR5 and TNFα/IFNγ/interleukin-2 (IL-2) and less TH2-associated CCR3/CCR4, with no IL-4/IL-13. They lack exhaustion-/suppression-associated PD1, CD57 and terminally differentiated effector memory cells, with more central memory cells, activation-associated partners of Hodgkin Reed Sternberg (HRS) cell-expressed CD30/OX40-L/ICOS-L, and other activation markers. TH cell lines established from CHL and RLN-derived SCSs remain cytokine-secretory. We confirmed and extended these studies using tissue microarray immunohistochemistry (TMA-IHC) from a large CHL tissue bank (n = 122) and demonstrate TH1-associated TBET is abundant in CHL, and TH2-associated CMAF/GATA3 and exhaustion-associated PD1 expressed at significantly lower levels. These molecular insights into the CHL-associated TH offer potential diagnostic, prognostic and pharmacologically modifiable therapeutic targets and do not support the established view of a TH2-enriched, senescent/exhausted, hypofunctional, hypoproliferative infiltrate.

Reprogramming in vivo th17 into th17/th2 by Sirp-α dendritic cells in the lungs

Background Dendritic cells (DCs) play a crucial role in the development of the adaptive immune response. Unbalance DC response can cause Th1, Th17 or Th2-mediated diseases. By in vitro manipulation, Th2 and Th17 cell lines can be reprogrammed into Th1. This highlights the notion of the plasticity of different populations of CD4 T helper cells. So far, the conversion of Th17 memory cells into Th2 cells has not been demonstrated in the tissues.

Airway Inflammation and IgE Production Induced by Dust Mite Allergen-Specific Memory/Effector Th2 Cell Line Can Be Effectively Attenuated by IL-35

CD4(+) memory/effector T cells play a central role in orchestrating the rapid and robust immune responses upon re-encounter with specific Ags. However, the immunologic mechanism(s) underlying these responses are still not fully understood. To investigate this, we generated an allergen (major house dust mite allergen, Blo t 5)-specific murine Th2 cell line that secreted IL-4, IL-5, IL-10, and IL-13, but not IL-9 or TNF-α, upon activation by the cognate Ag. These cells also exhibited CD44(high)CD62L(-) and CD127(+) (IL-7Rα(+)) phenotypes, which are characteristics of memory/effector T cells. Experiments involving adoptive transfer of this Th2 cell line in mice, followed by three intranasal challenges with Blo t 5, induced a dexamethasone-sensitive eosinophilic airway inflammation. This was accompanied by elevation of Th2 cytokines and CC- and CXC-motif chemokines, as well as recruitment of lymphocytes and polymorphic mononuclear cells into the lungs. Moreover, Blo t 5-specific IgE was detected 4 d after the last intranasal challenge, whereas elevation of Blo t 5-specific IgG1 was found at week two. Finally, pulmonary delivery of the pVAX-IL-35 DNA construct effectively downregulated Blo t 5-specific allergic airway inflammation, and i.m. injection of pVAX-IL-35 led to long-lasting suppression of circulating Blo t 5-specific and total IgE. This model provides a robust research tool to elucidate the immunopathogenic role of memory/effector Th2 cells in allergic airway inflammation. Our results suggested that IL-35 could be a potential therapeutic target for allergic asthma through its attenuating effects on allergen-specific CD4(+) memory/effector Th2 cell-mediated airway inflammation.

Identification of Low Frequency Multiple Novel Aspergillus Fumigatus Specific MHC Class II Epitopes and Rapid Generation of An Aspergillus-Specific T-Cell Product Based On Activation Dependent Expression of CD154.

Abstract 1170Poster Board I-192Despite new antifungal drugs, invasive aspergillosis (IA) remains a major cause of morbidity and mortality in patients undergoing myeloablative chemotherapy and allogeneic stem cell transplantation. Invasion of Aspergillus sp. in immuncompetent individuals is primarily controlled by neutrophils, phagocytes and pathogen-specific TH1 CD4+ cells. Therefore, adoptive transfer of Aspergillus-specific CD4+ T-cells could protect patients at risk from IA. Favorable candidates for such an approach are GPI-anchored antigens, which have demonstrated induction of protective adaptive immunity in murine studies.To implement Aspergillus-specific CD4+ T-cells into the clinical setting, we aimed (i) to define which GPI-anchored antigen induces TH1 response, (ii) to map several epitopes and (iii) to generate large amounts of Aspergillus-specific TH1 cells within a short period.Several recombinant proteins were either synthesized or were received by J-P Latge. Amongst all tested proteins, only CRF-1 induced repeatedly TH1 response in healthy individuals.To identify Aspergillus-specific MHC class II epitopes, we mapped peripheral blood mononuclear cells (PBMC) of healthy individuals with a custom made peptide library of CRF-1 consisting of 95 overlapping 15mer peptides. The library was divided into a complete pool, subpools of 10 and 95 single peptides. No precursor frequencies were detected in interferon-gamma (IFN-g)-ELISPOT using the complete and sub-pools. After 7 days of stimulation with subpools, 9 peptides were identified producing high amount of IFN-g in at least 3 donors with different MHC class II alleles (n=6). CD4+ T-cells clones of one peptide were then established by limited dilution cloning. Restriction to DRB1*0401 was identified using LCLs and a tetramer was generated. The peptide-specific T-cell clones showed functional activity against ethanol-inactivated fungus or fungal extracts presented by dendritic cells.To generate Aspergillus-specific TH1 cell lines, we stimulated PBMC with the MHC class II DRB1*0401 restricted peptide. After 7 days of in vitro stimulation (IVS) with interleukin (IL)-2 5U/ml added every other day, tetramer staining was between 0.3 and 11% of CD4+ cells (mean 4.2%, n=5). After 14 days of IVS with 1 restimulation of autologous peptide-pulsed monocytes at a responder: stimulator ratio of 5:1 and IL-7 and IL-15 10ng/ml added after day 7, mean percentages of tetramer staining increased to 37% (range 20-57%, n=4) and absolute cell counts were duplicated. The expansion process was further optimized using IFN-g capture assay and CD154+ MicroBead Kit (Miltenyi). In both assays, PBMC were stimulated for 16 hours, separated by magnetic beads and co-cultured with irradiated autologous PBMC for 14 days using IL-2 5U/ml till day 7 and IL-7 and -15 10ng/ml thereafter. We were unable to expand specific CD4+ cells by IFN-g capture assay in 2 of 3 donors. In contrast, we found that Aspergillus-specific CD4+ cells selected by CD154+ showed comparable tetramer specificity and expansion but higher functional activity in intracellular cytokine assay and IFN-g ELISA than CD4+ cell lines generated from the same donor using the restimulation protocol. The CD4+ cell lines generated by CD154+ expression showed proliferative capacity in CFSE when restimulated with peptides and functional activity in IFN-g ELISA against fungal extracts (n=4).To generate Aspergillus-specific TH1 cell lines recognizing multiple MHC class II epitopes we stimulated PBMC with the previously identified 9 peptides of CRF-1 protein. After 14 days of expansion using CD154+ MicroBead Kit, we measured high IFN-g response towards 4 of 9 peptides (n=3). We are generating T-cell clones and will characterize their HLA-restriction.In summary, we have identified an immunodominant Aspergillus fumigatus protein including 9 MHC class II epitopes. One epitope was characterized specific for an abundant MHC class II allele. CD4+ TH1 cells specific for this epitope can be activated by dendritic cells after uptake of whole fungi or fungal extracts. Furthermore, we have established a GMP-applicable protocol for rapid generation (<14d) of CD4+ T-cell lines specific for Aspergillus fumigatus with low precursor frequency using CD154+ separation. These CD4+ T-cell lines demonstrate functional activity against peptides and fungal extracts that could be prophylactically administered to high risk patients. Disclosures:No relevant conflicts of interest to declare.

Cutting Edge: Antigen-Specific TGFβ-Induced Regulatory T Cells Suppress Th17-Mediated Autoimmune Disease

CD4(+) T cells from the TCR transgenic TxA23 mouse recognize a peptide from the H/K-ATPase alpha-chain. When TxA23 CD4(+) thymocytes are differentiated into Th1, Th2, and Th17 lines, all three subpopulations induced autoimmune gastritis (AIG) upon transfer into nu/nu recipients. The induction of AIG by naive T cells or by Th1 or Th2 cell lines could be prevented by the cotransfer of polyclonal Foxp3(+) T regulatory cells (nTreg), whereas Th17-induced AIG was resistant to suppression. We compared the capacity of different types of Treg to suppress Th17-mediated AIG. Cotransfer of either nTreg or polyclonal TGFbeta-induced Treg (iTreg) did not prevent AIG, while cotransfer of TGFbeta-induced Ag-specific TxA23 iTreg completely prevented the development of disease. Ag-specific iTreg were able to suppress Th17-mediated disease when injected 6 days after the Th17 effectors. The implications of these results for the use of Treg for the cellular biotherapy of autoimmune disease are discussed.

Mechanism of regulatory T cell- induced tumor immune tolerance

regulatory T cell（Treg） is a subset of immunosuppressive Th cell lines with ability to suppress anti-tumor immunity. Tregs are recruited into local tumor microenvironment by tumor-released chemokine CCL22. Tregs induce Th2-type immune responses which contribute the host ignorance and tolerance to tumor cells via releasing of immunosuppressive cytokines and immunosuppressive receptors. Accumulating evidences indicate that inhibiting Treg can break immune tolerance, and subsequently inhibit the growth and progress of cancer. These discoveries and exploration of Treg provide us a new therapeutic target and strategy with promising clinic application. Key words: CD4^＋ CD25^＋ regulatory T cell; Tumor; Immune tolerance

Host T Cells Are the Main Producers of IL-17 within the Central Nervous System during Initiation of Experimental Autoimmune Encephalomyelitis Induced by Adoptive Transfer of Th1 Cell Lines

Experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, has long been thought to be mediated by Th1 CD4(+) T cells. Using adoptive transfer techniques, transfer of CNS specific Th1 T cells was sufficient to induce EAE in naive mice. However, recent studies found a vital role for IL-17 in induction of EAE. These studies suggested that a fraction of IL-17-producing T cells that contaminate Th1 polarized cell lines are largely responsible for initiation of EAE. In this study, we tracked the appearance and cytokine production capacity of adoptively transferred cells within the CNS of mice throughout EAE disease. IL-17-producing, adoptively transferred cells were not enriched over the low percentages present in vitro. Thus, there was no selective recruitment and/or preferential proliferation of adoptively transferred IL-17-producing cells during the induction of EAE. Instead a large number of CNS infiltrating host T cells in mice with EAE were capable of producing IL-17 following ex vivo stimulation. The IL-17-producing T cells contained both alphabeta and gammadelta TCR(+) T cells with a CD4(+)CD8(-) or CD4(-)CD8(-) phenotype. These cells concentrated within the CNS within 3 days of adoptive transfer, and appeared to play a role in EAE induction as adoptive transfer of Th1 lines derived from wild-type mice into IL-17-deficient mice induced reduced EAE clinical outcomes. This study demonstrates that an encephalitogenic Th1 cell line induces recruitment of host IL-17-producing T cells to the CNS during the initiation of EAE and that these cells contribute to the incidence and severity of disease.

Combinatorial Signal Transduction Responses Mediated by Interleukin-2 and -4 Receptors in a Helper TH2 Cell Line

The cytokines interleukin (IL)-2 and IL-4 are important regulators of the adaptive immune response, due in part to their effects on clonal expansion and differentiation of T cells. When IL-2 and IL-4 are administered together, both antagonistic and synergistic effects have been reported, but little is known in general concerning the mechanisms underlying such combinatorial effects. We found evidence for both effects in the proliferation responses of the IL-2 and IL-4 responsive T cell line, HT-2; IL-4 delays the onset of cell growth yet ultimately allows a higher cell density to be achieved in static culture. At the level of signal transduction pathways, we found that IL-4 partially inhibits IL-2 receptor-mediated pathways (PI3K/Akt, Ras/Erk, and STAT5a/b) and does not prolong their transient kinetics. This mode of antagonism, but not the effects on cell proliferation, is overcome at higher concentrations of IL-2 that are sufficient to saturate the signaling responses. By comparison, IL-4-stimulated activation of STAT6 is unaffected by IL-2 and shows sustained kinetics, and we speculate that this or another IL-4 receptor-specific pathway is responsible for the effects of IL-4 on IL-2-stimulated proliferation. A possibly related observation is that IL-4 induces a dramatic cell adhesion phenotype.

T Helper Cell-specific Regulation of Inducible Costimulator Expression via Distinct Mechanisms Mediated by T-bet and GATA-3

Inducible costimulator (ICOS) expression is critical for T cell-mediated immunity. We showed previously that T cell receptor and CD28 co-engagement up-regulate ICOS expression in activated T cells via the induction of NFATc2. Here, we examined the regulation of ICOS expression by Th-specific transcription factors T-bet and GATA-3. Overexpression of T-bet or GATA-3 alone could enhance, and NFATc2 could further synergize with either of them to increase, icos transcription. Although T-bet acted on the icos promoter, GATA-3 operated via an icos 3'-un-translated region element. Interestingly, NFATc2 was found to bind promiscuously the icos promoter in developing Th0, Th1, and Th2 cells but became selectively associated with T-bet at the promoter and with GATA-3 at the 3'-untranslated region in fully differentiated Th1 and Th2 cells, respectively. Collectively, our results reveal a temporally evolving circuit in which the non-selectively expressed NFATc2 cooperates with Th-restricted T-bet or GATA-3 to direct transcription of a costimulatory gene via distinct regulatory elements in different Th cells undergoing differentiation.

Myelin Basic Protein-primed T Cells Induce Neurotrophins in Glial Cells via α5β3 Integrin

Increasing the level of neurotrophins within the central nervous system may have therapeutic efficacy in patients with various neurological diseases. Earlier we have demonstrated that myelin basic protein (MBP)-primed T cells induce the expression of various proinflammatory molecules in glial cells via cell-to-cell contact. Here we describe that after Th2 polarization by gemfibrozil or other drugs, MBP-primed T cells induced the expression of neurotrophic molecules such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3), but not proinflammatory molecules in microglia and astroglia via cell-to-cell contact. MBP-primed Th2 cells expressed alpha5 and beta3 integrins and functional blocking antibodies against both alpha5 and beta3 integrins inhibited the ability of MBP-primed Th2 cells to induce glial neurotrophins. On the other hand, glial cells expressed PDGF-Rbeta and neutralization of this glial receptor abrogated the ability of Th2 cells to induce neurotrophins in glia. Activation of glial cAMP response element-binding protein (CREB) by MBP-primed Th2 cell contact and inhibition of contact-mediated expression of neurotrophins by antisense knockdown of glial CREB suggest that MBP-primed Th2 cell-glia contact induces the expression of neurotrophins through glial activation of CREB. Accordingly, blocking of either alpha5beta3 integrins on T cells or PDGF-Rbeta on glial cells impaired the ability of MBP-primed Th2 cells to induce glial activation of CREB. Furthermore, we demonstrate that these MBP-primed Th2 cells entered into the central nervous system and increased the expression of neurotrophins in vivo in the brain. This study illuminates the importance of alpha5beta3 and PDGF-Rbeta in guiding the novel neurotrophic property of neuroantigen-primed T cells via activation of CREB that may be of therapeutic importance in various neurological disorders.

Helicobacter pylori polyclonally activates murine CD4+ T cells in the absence of antigen‐presenting cells

Helicobacter pylori is a Gram-negative bacterium that causes a variety of gastrointestinal diseases, such as duodenal ulcer and gastric carcinoma. The T cell response against H. pylori is thought to contribute to the pathogenesis of these diseases. Here, we show that mouse-adapted H. pylori is able to polyclonally activate murine CD4(+) T lymphocytes, irrespective of their antigen specificity. Murine T helper cell clones as well as short-term cultured, polyclonal Th1 and Th2 cell lines and a human T cell clone, but not naive CD4(+) T cells, could be activated in this manner. The effect was independent of antigen-presenting cells and required direct contact between H. pylori and T cells. Only whole cells of H. pylori, but not lysates or sonicates were able to activate T cells. The activity was lost after long-term culture of H. pylori on agar-plates. Degradation of H. pylori proteins with specific peptidases dramatically reduced the stimulating ability, implicating that the responsible molecule is likely to be a protein. This unexpected polyclonal T cell stimulatory mechanism may contribute to the T cell-mediated pathogenicity characteristic for H. pylori-mediated diseases.

IL-23 and IL-17 in pathogenesis of experimental ocular autoimmunity: requirement for IL–23 may extend beyond its role in sustaining the IL-17 effector response (129.26)

Abstract Experimental autoimmune uveitis (EAU) represents autoimmune uveitis in humans. Here we examined the role of the IL-23/IL-17 pathway in EAU. We immunized IL-23p19, IL-12p35, IL-12p40, IFN-γ and IL-17 KO mice with the uveitogenic Ag IRBP. Alternatively, we treated wild type mice immunized for EAU with Abs to IL-23p19 or to IL-17 throughout the disease course (prevention), or only during the effector phase (reversal). IL-23p19 KO were resistant to EAU, showing reduced Ag-specific DTH, IL-2, IFN-γ, IL-17, IL-1β, IL-6 and IL-5. In contrast, IL-12p35 and IFN-γ KO mice developed exacerbated EAU, DTH and IL-17 responses. Treatment with anti-IL-23p19 protected from EAU and reduced immunological responses, but was unable to reverse disease, indicating that EAU is dependent on early availability of IL-23. In contrast, anti-IL-17 prevented as well as reversed EAU. Interestingly, severe EAU could be induced with a Th1 cell line that does not produce IL-17. In vivo cytokine neutralization revealed requirement for IFN-γ and TNF-α, but not for IL-17, suggesting that under some conditions an antigen-specific Th1 effector is sufficient to induce EAU. In keeping with this, IL-17 KO mice were still susceptible to disease. These data raise the possibility that the nonredundant need for IL-23 in EAU may extend beyond its role in promoting the Th17 effector response and point to IL-23 and IL-17 as new therapeutic targets for uveitis.

An interferon-γ-producing Th1 subset is the major source of IL-17 in experimental autoimmune encephalitis

Th IL-17 (IL-17+/IFN-γ−) cell lines are significantly more encephalitogenic than Th1 (IL-17−/IFN-γ+) cell lines in adoptive transfer EAE models. In actively induced EAE short ex vivo peptide stimulation identifies an IL-17+/IFN-γ+ population of CD4+ CNS-infiltrating MOG 35–55-specific T cells, which outnumber IL-17+/IFN-γ− cells by approximately 3:1 as disease develops. A decrease in numbers of IL-17+/IFN-γ+ cells following in vitro culture is accompanied by an increase in IL-17−/IFN-γ+ cell numbers. Together these ex vivo and in vitro observations imply that the Th1 lineage is more encephalitogenic than is suggested by adoptive transfer of Th1 (IL-17−/IFN-γ+) cell lines which have been terminally differentiated in vitro.

PART I. Peptide ImmunotherapyNatural and Autoantibodies to Human T-Cell Receptor Vβ Segments: Potential Roles in Immunomodulation

Although the manifestation of inflammatory autodestructive disease is the result of major immunological dysfunction, recent evidence indicates that the immune system attempts to compensate by the production of immunomodulatory autoantibodies. Healthy humans have low levels of naturally occurring autoantibodies directed against the first complementarity-determining region (CDR1) and third framework region (FR3) of their own T-cell receptor (TCR) Vbeta segments, but individuals suffering from rheumatoid arthritis (RA) or systemic lupus erythematosus (SLE) can have highly elevated levels of these autoantibodies. We cloned and characterized human anti-TCR monoclonal autoantibodies (mAAbs) from RA and SLE patients. Because of the cross-reactions between distinct CDR1 segments of human TCR Vbeta and corresponding murine homologs, it was possible to show that human mAAbs blocked the capacity of a murine TH1 cell line (DO11.10) to produce IL-2 in response to antigenic stimulation in vitro. These results support the hypothesis that autoantibodies against TCR Vbeta can shut down TH1-mediated inflammatory autodestructive reactions.

T-bet negatively regulates autoimmune myocarditis by suppressing local production of interleukin 17

Experimental autoimmune myocarditis (EAM) appears after infectious heart disease, the most common cause of dilated cardiomyopathy in humans. Here we report that mice lacking T-bet, a T-box transcription factor required for T helper (Th)1 cell differentiation and interferon (IFN)-γ production, develop severe autoimmune heart disease compared to T-bet −/− control mice. Experiments in T-bet −/− IL-4−/− and T-bet −/− IL-4Rα−/− mice, as well as transfer of heart-specific Th1 and Th2 cell lines, showed that autoimmune heart disease develops independently of Th1 or Th2 polarization. Analysis of T-bet −/− IL-12Rβ1−/− and T-bet −/− IL-12p35−/− mice then identified interleukin (IL)-23 as critical for EAM pathogenesis. In addition, T-bet −/− mice showed a marked increase in production of the IL-23–dependent cytokine IL-17 by heart-infiltrating lymphocytes, and in vivo IL-17 depletion markedly reduced EAM severity in T-bet −/− mice. Heart-infiltrating T-bet −/− CD8+ but not CD8− T cells secrete IFN-γ, which inhibits IL-17 production and protects against severe EAM. In contrast, T-bet −/− CD8+ lymphocytes completely lost their capacity to release IFN-γ within the heart. Collectively, these data show that severe IL-17–mediated EAM can develop in the absence of T-bet, and that T-bet can regulate autoimmunity via the control of nonspecific CD8+ T cell bystander functions in the inflamed target organ.