The choice of adjuvant determines the cytokine profile of T cells in proteoglycan‐induced arthritis but does not influence disease severity

Rheumatoid arthritis (RA) is a debilitating autoimmune disease characterized by chronic inflammation of the synovial joints. Collagen‐induced arthritis (CIA) and proteoglycan‐induced arthritis (PGIA) are mouse models of inflammatory arthritis; CIA is a T helper type 17 (Th17) ‐dependent disease that is induced with antigen in complete Freund's adjuvant, whereas PGIA is Th1‐mediated and is induced using antigen in dimethyldioctadecyl‐ammonium bromide (DDA) as an adjuvant. To investigate whether the type of adjuvant determines the cytokine profile of the pathogenic T cells, we have compared the effect of CFA and DDA on T‐cell responses in a single arthritis model. No differences in incidence or disease severity between aggrecan‐T‐cell receptor transgenic mice immunized with aggrecan in either CFA or DDA were observed. Immunization with CFA resulted in a higher proportion of Th17 cells, whereas DDA induced more Th1 cells. However, the levels of interleukin‐17 (IL‐17) produced by T cells isolated from CFA‐immunized mice after antigen‐specific stimulation were not significantly different from those found in DDA‐immunized mice, indicating that the increased proportion of Th17 cells did not result in significantly higher ex vivo IL‐17 levels. Hence, the choice of adjuvant can affect the overall proportions of Th1 and Th17 cells, without necessarily affecting the level of cytokine production or disease incidence and severity.     

Prediction and characterization of helper T‐cell epitopes from pneumococcal surface adhesin A

Pneumococcal surface adhesin A (PsaA) is a multifunctional lipoprotein known to bind nasopharyngeal epithelial cells, and is significantly involved in bacterial adherence and virulence. Identification of PsaA peptides that optimally bind human leucocyte antigen (HLA) and elicit a potent immune response would be of great importance to vaccine development. However, this is hindered by the multitude of HLA polymorphisms in humans. To identify the conserved immunodominant epitopes, we used an experimental dataset of 28 PsaA synthetic peptides and in silico methods to predict specific peptide‐binding to HLA and murine MHC class II molecules. We also characterized spleen and cervical lymph node (CLN) ‐derived T helper (Th) lymphocyte cytokine responses to these peptides after Streptococcus pneumoniae strain EF3030 challenge in mice. Individual, yet overlapping, peptides 15 amino acids in length revealed residues of PsaA that consistently caused the highest interferon‐γ, interleukin‐2 (IL‐2), IL‐5 and IL‐17 responses and proliferation as well as moderate IL‐10 and IL‐4 responses by ex vivo re‐stimulated splenic and CLN CD4⁺ T cells isolated from S. pneumoniae strain EF3030‐challenged F₁ (B6 ×  BALB/c) mice. In silico analysis revealed that peptides from PsaA may interact with a broad range of HLA‐DP, ‐DQ and ‐DR alleles, due in part to regions lacking β‐turns and asparagine endopeptidase sites. These data suggest that Th cell peptides (7, 19, 20, 22, 23 and 24) screened for secondary structures and MHC class II peptide‐binding affinities can elicit T helper cytokine and proliferative responses to PsaA peptides.     

Therapeutic effects of a fermented soy product on peanut hypersensitivity is associated with modulation of T‐helper type 1 and T‐helper type 2 responses

Background ImmuBalance^? is a koji fungus (Aspergillus oryzae) and lactic acid fermented soybean product. This unique production process is believed to create a food supplement that helps to induce or maintain normal immune response. Objective To assess possible therapeutic effects of ImmuBalance^? on peanut (PN) hypersensitivity using a murine model of peanut allergy (PNA). Methods PN allergic C3H/HeJ mice were fed standard mouse chow containing 0.5% or 1.0% ImmuBalance (ImmuBalance 2X), radiation‐inactivated 1.0% ImmuBalance (I‐ImmuBalance 2X), or regular diet chow (sham) for 4 weeks, beginning 10 weeks after the initial PN sensitization, and then challenged with PN. Anaphylactic symptom scores, plasma histamine, serum PN specific‐IgE levels and splenocyte cytokine profiles were determined. Results While 100% of sham‐treated PNA mice developed anaphylactic reactions with a median score of 3.3 following PN challenge, only 50% of ImmuBalance, 30% of ImmuBalance 2X and 40% of I‐ImmuBalance 2X‐treated mice developed allergic reactions with median scores of 1.0, 0.4 and 0.5 respectively, which were significantly less than that in the sham‐treated mice (P<0.05). Plasma histamine and PN specific‐IgE levels were also significantly less in all treated mice than in sham‐treated mice (P<0.05). Furthermore, IL‐4, IL‐5 and IL‐13 production by PN‐stimulated splenocytes in vitro from ImmuBalance fed mice were markedly reduced compared with sham‐treated mice, whereas IFN‐γ production was moderately increased. TGF‐β and TNF‐α production were similar. Conclusions ImmuBalance protects against PN‐induced anaphylaxis when administered as a food supplement in this model. Protection was associated with down‐regulation of Th2 responses. This supplement may provide a potential novel therapy for PNA.     

Compartmentalization of dendritic cell and T‐cell interactions in the lymph node: Anatomy of T‐cell fate decisions

Upon receiving cognate and co‐stimulatory priming signals from antigen (Ag)‐presenting dendritic cells (DCs) in secondary lymphoid tissues, naïve CD4⁺ T cells differentiate into distinct effector and memory populations. These alternate cell fate decisions, which ultimately control the T‐cell functional attributes, are dictated by programming signals provided by Ag‐bearing DCs and by other cells that are present in the microenvironment in which T‐cell priming occurs. We know that DCs can be subdivided into multiple populations and that the various DC subsets exhibit differential capacities to initiate development of the different CD4⁺ T‐helper populations. What is less well understood is why different subanatomic regions of secondary lymphoid tissues are colonized by distinct populations of Ag‐presenting DCs and how the location of these DCs influences the type of T‐cell response that will be generated. Here we review how chemokine receptors and their ligands, which position allergen and nematode‐activated DCs within different microdomains of secondary lymphoid tissues, contribute to the establishment of IL‐4 committed follicular helper T and type 2 helper cell responses.     

T‐bet regulates differentiation of forkhead box protein 3+ regulatory T cells in programmed cell death‐1‐deficient mice

Programmed cell death‐1 (PD‐1) plays an important role in peripheral T cell tolerance, but whether or not it affects the differentiation of helper T cell subsets remains elusive. Here we describe the importance of PD‐1 in the control of T helper type 1 (Th1) cell activation and development of forkhead box protein 3 (FoxP3⁺) regulatory T cells (T_regs). PD‐1‐deficient T cell‐specific T‐bet transgenic (P/T) mice showed growth retardation, and the majority died within 10 weeks. P/T mice showed T‐bet over‐expression, increased interferon (IFN)‐γ production by CD4⁺ T cells and significantly low FoxP3⁺ T_reg cell percentage. P/T mice developed systemic inflammation, which was probably induced by augmented Th1 response and low FoxP3⁺ T_reg count. The study identified a unique, previously undescribed role for PD‐1 in Th1 and T_reg differentiation, with potential implication in the development of Th1 cell‐targeted therapy.     

The CRTH2 agonist Pyl A prevents lipopolysaccharide‐induced fetal death but induces preterm labour

We have previously demonstrated that the anti‐inflammatory prostaglandin 15‐deoxy‐Δ 12,14‐prostaglandin J₂ (15dPGJ₂) delays inflammation‐induced preterm labour in the mouse and improves pup survival through the inhibition of nuclear factor‐κB (NF‐κB) by a mechanism yet to be elucidated. 15dPGJ₂ is an agonist of the second prostaglandin D₂ receptor, chemoattractant receptor homologous to the T helper 2 cell (CRTH2). In human T helper cells CRTH2 agonists induce the production of the anti‐inflammatory interleukins IL‐10 and IL‐4. We hypothesized that CRTH2 is involved in the protective effect of 15dPGJ₂ in inflammation‐induced preterm labour in the murine model. We therefore studied the effects of a specific small molecule CRTH2 agonist on preterm labour and pup survival. An intrauterine injection of lipopolysaccharide (LPS) was administered to CD1 mice at embryonic day 16, ± CRTH2 agonist/vehicle controls. Mice were killed at 4.5 hr to assess fetal wellbeing and to harvest myometrium and pup brain for analysis of NF‐κB, and T helper type 1/2 interleukins. To examine the effects of the CRTH2 agonist on LPS‐induced preterm labour, mice were allowed to labour spontaneously. Direct effects of the CRTH2 agonist on uterine contractility were examined ex vivo on contracting myometrial strips. The CRTH2 agonist increased fetal survival from 20 to 100% in LPS‐treated mice, and inhibited circular muscle contractility ex vivo. However, it augmented LPS‐induced labour and significantly increased myometrial NF‐κB, IL‐1β, KC‐GRO, interferon‐γ and tumour necrosis factor‐α. This suggests that the action of 15dPGJ₂ is not via CRTH2 and therefore small molecule CRTH2 agonists are not likely to be beneficial for the prevention of inflammation‐induced preterm labour.     

Marked enhancement of antigen‐specific T‐cell responses by IL‐7‐fused nonlytic,but not lytic,Fc as a genetic adjuvant

IL‐7 plays a crucial role in the homeostatic proliferation, differentiation and survival of T cells, as well as in the survival and proliferation of precursor B cells. Here, we demonstrated that utilizing nonlytic Fc‐fused IL‐7 (IL‐7‐Fc^m) as a genetic adjuvant significantly enhanced not only CD4⁺ but also CD8⁺ T‐cell responses by E7 DNA immunization, in addition to improving protection against TC‐1‐induced tumors in comparison to IL‐7 alone. Similar results were obtained in OT‐1 adoptive transfer experiments with OVA DNA injection, suggesting independence from antigenic nature and experimental conditions. In particular, the increased frequency of CD8⁺ T cells was mainly due to enhanced T‐cell proliferation in T‐cell priming, and not to decreased cellular apoptosis. Interestingly, the enhanced adjuvant effect was not seen in the co‐delivery of lytic Fc‐fused IL‐7 (IL‐7‐Fc) which increases T‐cell apoptosis as well as T‐cell proliferation, suggesting that the T‐cell proliferative effect may be neutralized by T‐cell apoptosis. Thus, our findings suggest that nonlytic Fc, in contrast to lytic Fc, fusion to cytokines may provide an insight in designing a potent genetic adjuvant for inducing CD4⁺ and CD8⁺ T‐cell responses.     

IL‐23‐driven encephalo‐tropism and Th17 polarization during CNS‐inflammation in vivo

IL‐23 but not IL‐12 is essential for the development of autoimmune tissue inflammation in mice. Conversely, IL‐12 and IL‐23 impact on the polarization of Th1 and Th17 cells, respectively. While both polarized T helper populations can mediate autoimmune inflammation, their redundancy in the pathogenesis of EAE indicates that IL‐23 exerts its crucial influence on the disease independent of its T helper polarizing capacity. To study the impact of IL‐23 and IL‐12 on the behavior of encephalitogenic T cells in vivo, we generated BM‐chimeric mice in which we can trace individual populations of IL‐23 or IL‐12 responsive T helper cells during EAE. We observed that T cells, which lack IL‐12Rβ1 (no IL‐12 and IL‐23 signaling), fail to invade the CNS and do not acquire a Th17 phenotype. In contrast, loss of IL‐12 signaling prevents Th1 polarization but does not prevent T‐cell entry into the CNS. The loss of IL‐12R engagement does not appear to alter T‐cell expansion but leads to their accumulation in secondary lymphoid organs. We found that IL‐23 licenses T cells to invade the target tissue and to exert their effector function, whereas IL‐12 is critical for Th1 differentiation, but does not influence the pathogenic capacity of auto‐reactive T helper cells in vivo.     

Regulation of CD4 T‐cell receptor diversity by vaccine adjuvants

New vaccines based on soluble recombinant antigens (Ags) require adjuvants to elicit long‐lasting protective humoral and cellular immunity. Despite the importance of CD4 T helper cells for the generation of long‐lived memory B and CD8 T cells, the impact of adjuvants on CD4 T‐cell responses is still poorly understood. Adjuvants are known to promote dendritic cell (DC) maturation and migration to secondary lymphoid organs where they present foreign peptides bound to class II major histocompatibility complex molecules (pMHCII) to naïve CD4 T cells. Random and imprecise rearrangements of genetic elements during thymic development ensure that a vast amount of T‐cell receptors (TCRs) are present in the naïve CD4 T‐cell repertoire. Ag‐specific CD4 T cells are selected from this vast pre‐immune repertoire based on the affinity of their TCR for pMHCII. Here, we review the evidence demonstrating a link between the adjuvant and the specificity and clonotypic diversity of the CD4 T‐cell response, and consider the potential mechanisms at play.     

Differential T‐cell receptor signals for T helper cell programming

Upon encounter with their cognate antigen, naive CD4 T cells become activated and are induced to differentiate into several possible T helper (Th) cell subsets. This differentiation depends on a number of factors including antigen‐presenting cells, cytokines and co‐stimulatory molecules. The strength of the T‐cell receptor (TCR) signal, related to the affinity of TCR for antigen and antigen dose, has emerged as a dominant factor in determining Th cell fate. Recent studies have revealed that TCR signals of high or low strength do not simply induce quantitatively different signals in the T cells, but rather qualitatively distinct pathways can be induced based on TCR signal strength. This review examines the recent literature in this area and highlights important new developments in our understanding of Th cell differentiation and TCR signal strength.     

Pollen‐derived low‐molecular weight factors inhibit 6‐sulfo LacNAc+ dendritic cells' capacity to induce T‐helper type 1 responses

Background Evidence is accumulating that the pollen exsudate contains an array of non‐allergenic, pro‐inflammatory and immunomodulatory substances acting on the innate and adaptive immune system. In this context, pollen‐associated E₁‐phytoprostanes (PPE₁) were shown to licence human monocyte‐derived dendritic cells for T‐helper type 2 (Th2) polarization of naïve T cells. Objective This study aims at analysing the impact of pollen‐associated lipid mediators on cytokine secretion and maturation of 6‐sulfo LacNAc⁺ dendritic cells (slanDCs), the most abundant native dendritic cell (DC) in human peripheral blood, and further dissecting the biologically active substance(s) within aqueous pollen extracts. Results Aqueous birch pollen extracts dose‐dependently inhibited the lipopolysaccharide (LPS)‐induced IL‐12 p70 production, while the levels of IL‐6 remained unaffected. PPE₁ inhibited secretion of both IL‐12 p70 and IL‐6. Aqueous pollen extracts, but not PPE₁ or F₁‐phytoprostanes significantly reduced the LPS‐induced surface expression of the maturation markers CD80, CD83, CD40 and CCR‐7, an effect that was independent of proteins and that was still present in a 3 kDa cut‐off fraction of the pollen extract. These effects were observed irrespective of the atopy status of the donors. Finally, slanDCs exposed to aqueous pollen extracts were impaired in eliciting an IFN‐γ response in naïve CD4⁺ T cells. Conclusion Our data show that slanDCs, a subset of human blood DCs with constitutively high potency to induce Th1 responses, are susceptible to the Th2 polarizing effect of low molecular weight, non‐protein factors derived from pollen. Cite this as: S. Gilles, D. Jacoby, C. Blume, M. J. Mueller, T. Jakob, H. Behrendt, K. Schaekel and C. Traidl‐Hoffmann, Clinical & Experimental Allergy, 2010 (40) 269–278.     

Interleukin-9 as a T helper type 17 cytokine

The production of interleukin‐9 (IL‐9) by CD4 T cells has gathered renewed interest as the result of the observation that its expression is broader than originally thought. This includes the production of IL‐9 by a recently characterized subset of CD4 helper T (Th) cells that are termed Th9 as well as production by additional T‐cell subsets including Th17 cells. There is an incomplete understanding as to which IL‐9‐producing T‐cell subsets develop under physiological conditions. We describe the conditions used to generate IL‐9 in Th17 cells in vitro. We also summarize conditions where both IL‐9 and IL‐17 are found in vivo and propose that Th17 cells producing IL‐9 may co‐exist and interact with Th9 cells during conditions of autoimmunity, allergy and infection.     

Atopic asthma: differential activation phenotypes among memory T helper cells

Background T cells have been implicated in the pathogenesis of atopic asthma. We have previously shown that memory T helper cells (CD4⁺CD45RO⁺) are preferentially activated relative to naïve T helper cells (CD4⁺CD45RA⁺) after bronchial allergen challenge. However, specific T helper subpopulations that are activated in atopy and/or asthma remain undefined. Objective To determine the T helper subpopulations and activation phenotypes relevant to acute and stable asthma that may be common with or distinct from atopy. Methods Two groups of atopic asthmatics (ten acute and nine stable asthmatics) and two non‐asthmatic groups (14 non‐asthmatic atopics and eight normal non‐atopic controls) were analysed. Ten acute asthmatics were assessed in the emergency room during an acute episode (FEV₁ 43.6% ± 18.4). Nine stable asthmatics were assessed during a symptom‐free period (FEV₁ 85% ± 6). Using multiple colour flow cytometry we analysed T cell subpopulations and the expression of IL‐2‐receptor (IL‐2R) and MHC‐class II antigens (MHC II) on naïve and memory T helper cells in the peripheral blood of asthmatic and non‐asthmatic groups. Results Atopic asthmatics (acute and stable) had an increased percentage of memory T helper cells expressing IL‐2R compared with normal non‐atopics (mean SD 16.1 ± 6%, 12.4 ± 2% and 7.7 ± 1.8%, P < 0.05) but not compared with non‐asthmatic atopics (10 ± 3.5%). Naïve T helper cells had low expression of IL‐2R and MHC II in all four groups. MHC II antigen expression was increased in memory T helper cells of asthmatics (acute and stable) compared with normal non‐atopics (13.9 ± 7.5, 10.6 ± 5 and 4.9 ± 2.5, P < 0.05) but not compared with non‐asthmatic atopics (7.92 4). A novel finding was that IL‐2R and the MHC II molecules were mainly expressed in non‐overlapping populations and coexpression was found predominantly on memory T helper cells. Asthmatics (acute and stable) had higher proportion of double positive memory T helper cells (IL‐2R⁺MHC II⁺) compared with both non‐asthmatic groups (P < 0.05). Conclusions We demonstrate a differential expression of IL‐2R⁺ and MCH II⁺ on CD45RO⁺ T helper cells that would suggest that there are three subsets of activated memory T helper cells in asthmatics. Two non‐overlapping IL‐2R⁺ or MHC II⁺ CD45RO⁺ T helper cells and a third subpopulation of activated cells that coexpress IL‐2R and MHC II (double positives). This latter subpopulation is significantly higher in asthmatics (acute or stable) compared with both non‐asthmatic groups, suggesting a specific T helper activation phenotype distinct to atopic asthmatics as compared with atopic non‐asthmatics.     

Differential regulation and impact of fucosyltransferase VII and core 2 β1,6‐N‐acetyl‐glycosaminyltransferase for generation of E‐selectin and P‐selectin ligands in murine CD4+ T cells

Ligands for E‐selectin and P‐selectin (E‐lig and P‐lig) are induced on CD4⁺ T cells upon differentiation into effector T cells. Glycosyltransferases, especially α 1,3‐fucosyltransferase VII (FucT‐VII) and core 2 β1,6‐N‐acetyl‐glycosaminyltransferase I (C2GlcNAcT‐I), are critical for their synthesis. We here analysed the signals that control the expression of E‐lig, P‐lig and mRNA coding for FucT‐VII and C2GlcNAcT‐I. In line with previous reports, we found that P‐lig expression correlates with the regulation of C2GlcNAcT‐I, whereas E‐lig expression can occur at low levels of C2GlcNAcT‐I mRNA but requires high FucT‐VII mRNA expression. Interestingly, the two enzymes are regulated by different signals. Activation‐induced C2GlcNAcT‐I up‐regulation under permissive (T helper type 1) conditions was strongly reduced by cyclosporin A (CsA), suggesting the involvement of T‐cell receptor‐dependent, calcineurin/NFAT‐dependent signals in combination with interleukin‐12 (IL‐12) ‐mediated signals in the regulation of C2GlcNAcT‐I. In contrast, expression of FucT‐VII mRNA was not significantly inhibited by CsA. Interleukin‐4 inhibited the expression of FucT‐VII but IL‐2 and IL‐7 were found to support induction of FucT‐VII and E‐lig. E‐selectin, P‐selectin and their ligands initially appeared to have rather overlapping functions. These findings however, unravel striking differences in the regulation of E‐lig and P‐lig expression, dictated by the dominance of FucT‐VII and C2GlcNAcT‐I, respectively, and their dependency on signals from either promiscuous or homeostatic cytokines (FucT‐VII) or a strong T‐cell receptor signal in combination with inflammatory cytokines in case of C2GlcNAcT‐I.