Aito Ueno

Increased prevalence of circulating novel IL-17 secreting Foxp3 expressing CD4+ T cells and defective suppressive function of circulating Foxp3+ regulatory cells support plasticity between Th17 and regulatory T cells in inflammatory bowel disease patients.

Background:IL-17 and Foxp3 double-expressing (DE) CD4+ T lymphocytes are novel crossover immune cell population, but the presence and role of these cells in human intestinal inflammation is unclear. The aim of this study was to investigate the circulating IL-17 and Foxp3 DE CD4+ T lymphocytes in patients with inflammatory bowel disease (IBD). Methods:The entire cohort consisted of 79 subjects: 31 patients with Crohns disease, 28 patients with ulcerative colitis, and 20 healthy control subjects (HC). IBD patients with evidence of active disease at endoscopy were entered into the study. Peripheral blood mononuclear cells were used for ex vivo and in vitro studies to assess the characteristics and generation of these novel cells and the function of circulating Foxp3+ CD4+ regulatory T lymphocytes (Treg) in patients with IBD compared with HC. Results:Patients with IBD had significantly higher prevalence of IL-17 and Foxp3 DE CD4+ T lymphocytes compared with age- and gender-matched HC. These cells expressed ROR&ggr;t. The ability of Treg cells to suppress autologous T-cell proliferation was reduced by approximately 60% in patients with IBD compared with HC. Increased generation of these DE cells was demonstrated by the modulation of cytokine environment of CD4+ lymphocytes in vitro in patients with Crohns disease. Conclusions:Prevalence of circulating IL-17 and Foxp3 DE CD4+ T cells is increased in patients with IBD. Coexpression of ROR&ggr;t and Foxp3 in these cells implies conversion from Treg cells to Th17 cells. This is associated with a decreased suppressive function of Foxp3+ CD4+ T lymphocytes in patients with IBD.

Transient Upregulation of Indoleamine 2,3-Dioxygenase in Dendritic Cells by Human Chorionic Gonadotropin Downregulates Autoimmune Diabetes

OBJECTIVE—Pregnancy induces a state of immunological tolerance that aims at suppressing immune responses against the fetus and has been linked to temporal remission of preexisting autoimmune disorders. To understand the mechanisms of this reversible immune regulation, we investigated the role of a key pregnancy hormone, human chorionic gonadotropin (hCG), in immune tolerance against autoimmune type 1 diabetes in nonobese diabetic (NOD) mice. RESEARCH DESIGN AND METHODS—We injected hCG into cytokine gene–deficient NOD mice and evaluated the effects of hCG administration on T-cells and dendritic cells (DCs). RESULTS—We show that administration of hCG to NOD mice inhibits both the activation of diabetogenic CD4+ and CD8+ T-cells, in vitro and in vivo, and the progression of type 1 diabetes by upregulating the expression of indoleamine 2,3-dioxygenase (IDO) in DCs. IDO upregulation is transient and declined shortly after hCG withdrawal. DC depletion restores the diabetetogenic activity of splenic T-cells from hCG-treated mice, and inhibition of IDO activity by 1-methyl-tryptophan abrogates the hCG-induced T-cell suppression and resistance to type 1 diabetes. CONCLUSIONS—We propose that hCG-induced upregulation of IDO in DCs plays a major role in pregnancy-associated resistance to autoimmunity.

Control of NKT Cell Differentiation by Tissue-Specific Microenvironments

CD1d-restricted Vα14 NKT cells play an important role in both Th1- and Th2-type immune responses. To determine whether NKT cells develop two functionally distinct subsets that provoke different types of responses, we examined the phenotypes and cellular functions of NK1.1+ and DX5+ T cells. We found that both NK1.1+ and DX5+ T cells are CD1d-restricted Vα14 T cells with identical Ag specificities, phenotypes, tissue locations, and functions. Similar to the NK1.1 marker, the DX5 marker (CD49b) is expressed on mature NKT cells in both NK1.1 allele-positive and allele-negative strains. However, when NK1.1+ and DX5+ NKT cells isolated from different tissues were compared, we found that thymic and splenic NKT cells differed not only in their cytokine profiles, but also in their phenotype and requirements for costimulatory signals. Thymic NKT cells displayed the phenotype of activated T cells and could be fully activated by TCR ligation. In contrast, splenic NKT cells displayed the phenotype of memory T cells and required a costimulatory signal for activation. Furthermore, the function and phenotype of thymic and splenic NKT cells were modulated by APCs from various tissues that expressed different levels of costimulatory molecules. Modulation of NKT cell function and differentiation may be mediated by synergic effects of costimulatory molecules on the surface of APCs. The results of the present study suggest that the costimulatory signals of tissue-specific APCs are key factors for NKT cell differentiation, and these signals cannot be replaced by anti-CD28 or anti-CD40 ligand Abs.

Th17 plasticity and its changes associated with inflammatory bowel disease

CD4 T helper (Th) cell differentiation into distinct T cell subsets is critical to the normal function of the immune system. Until recently, the paradigm held that naïve T cells differentiated into distinct subsets under the guidance of environmental cues (e.g., cytokines) and that once polarized, these cells were committed to a particular functional state. However, the existence of transdifferentiated T cell populations, which express signature transcription factors and cytokines associated with more than one Th subset, challenges the immutability of T helper subsets and suggests that plasticity is a feature of multifaceted immune responses. How this process impacts immune dysregulation in diseases such as inflammatory bowel diseases (IBD) and the machinery that underlies this process is far from fully understood. Interleukin (IL)-17 secreting helper T (Th17) cells have been heavily implicated in tissue-specific immune pathology including murine models of IBD, human Crohns disease and ulcerative colitis. Plasticity within this subset is suggested by the existence of IL-17 secreting cells, which, can also secrete interferon-γ, the signature cytokine for Th1 cells or, can co-express the anti-inflammatory transcription factor forkhead box p3, a signature transcription factor of regulatory T cells. In this review we mainly discuss evidence for Th17 plasticity, mechanisms, which govern it, and highlight the potential to therapeutically target this process in human IBD.

Ligand-Dependent Induction of Noninflammatory Dendritic Cells by Anergic Invariant NKT Cells Minimizes Autoimmune Inflammation

Stimulated by an agonistic ligand, α-galactosylceramide (αGalCer), invariant NKT (iNKT) cells are capable of both eliciting antitumor responses and suppressing autoimmunity, while they become anergic after an initial phase of activation. It is unknown how iNKT cells act as either activators or regulators in different settings of cellular immunity. We examined effects of αGalCer administration on autoimmune inflammation and characterized phenotypes and functional status of iNKT cells and dendritic cells in αGalCer-treated NOD mice. Although iNKT cells became and remained anergic after the initial exposure to their ligand, anergic iNKT cells induce noninflammatory DCs in response to αGalCer restimulation, whereas activated iNKT cells induce immunogenic maturation of DCs in a small time window after the priming. Induction of noninflammatory DCs results in the activation and expansion of islet-specific T cells with diminished proinflammatory cytokine production. The noninflammatory DCs function at inflammation sites in an Ag-specific fashion, and the persistence of noninflammatory DCs critically inhibits autoimmune pathogenesis in NOD mice. Anergic differentiation is a regulatory event that enables iNKT cells to transform from promoters to suppressors, down-regulating the ongoing inflammatory responses, similar to other regulatory T cells, through a ligand-dependent mechanism.

Opposing Effects of Smoking in Ulcerative Colitis and Crohn's Disease May Be Explained by Differential Effects on Dendritic Cells

Background:The mechanisms underlying the differential effects of cigarette smoking in patients with Crohns disease (CD) and ulcerative colitis (UC) remain unknown. Smoking has been demonstrated to be protective in UC, whereas in CD it has been shown to be associated with a more severe course, more frequent relapses, and postoperative recurrence. Dendritic cells (DC) play a critical role in T-cell activation and differentiation. Thus, we examined the effects of in vitro exposure to cigarette smoke extract (CSE) on phenotype/function of DC obtained from patients with UC and CD. Methods:Sixty-eight subjects were recruited including 30 patients with CD, 19 patients with UC, and 19 healthy controls. Peripheral blood monocytes were differentiated to DC in presence of IL-4 and granulocyte-macrophage colony-stimulating factor. The influence of CSE on Mo-DC subsets, cytokine expression, and ability to drive T cell proliferation and polarization were examined. Results:CSE affected DC phenotypes including increases in class-2 major histocompatibility complex and costimulatory molecules and decreases in CXCL10 and CCL3 levels in UC compared with CD samples. Furthermore, CSE also altered DC function resulting in increasing T cell proliferation and Th1 polarization in CD, whereas it increased Foxp3+ T cells and decreased the Th1 subset in UC samples. Conclusions:CSE modulates DC phenotype and function in patients with UC leading to increased prevalence of Foxp3+ CD4+ T cells, whereas in patients with CD it skews toward Th1 subsets. Differential DC responses to CSE between CD and UC may contribute to the differential effects associated with cigarette smoking status.

Profiles of Lamina Propria T Helper Cell Subsets Discriminate Between Ulcerative Colitis and Crohn's Disease.

Background:Distinction between 2 forms of inflammatory bowel disease (IBD), ulcerative colitis (UC) and Crohns disease (CD), can be challenging. Aberrant mucosal immunity suggests that CD is a T helper type 1 cell (Th1)-driven disease, whereas UC as Th2-driven response. However, whether this paradigm truly distinguishes CD from UC is controversial. We aimed to clarify the discriminating potential of lamina propria Th subsets in patients with IBD. Methods:Biopsies from 79 patients with IBD and 20 healthy controls were collected for Th subsets analysis (Th1:interferon &ggr; [IFN-&ggr;], T-bet; Th2:interleukin 13 [IL-13], Gata3; Th17:IL-17, ROR&ggr;t; Treg:FoxP3). The receiver-operating characteristic curves were constructed to assess the discriminating ability by calculating the area under the receiver-operating characteristic curve. The equation with the highest area under the receiver-operating characteristic curve was applied to newly diagnosed patients to evaluate discriminating ability. Results:Patients with CD showed increased IFN-&ggr;+ or T-bet+ cells and decreased IL-13+ or Gata3+ cells compared with UC. A discriminant equation composed of 4 markers (IFN-&ggr;+, T-bet+, IL-13+, and Gata3+) yielded the highest area under the receiver-operating characteristic curve. In 36 established CD or UC, the sensitivity, specificity, positive and negative predictive probabilities were 92.6%, 55.6%, 86.2%, and 71.4% and in 14 newly diagnosed patients were 100.0%, 42.9%, 63.6%, and 100.0%. Furthermore, Gata3+ cells were increased in tumor necrosis factor inhibitor therapy nonresponders compared with responders in CD. IFN-&ggr;+ cells were directly and inversely proportional to disease activity in patients with CD and UC, respectively. Conclusions:The Th1/Th2 paradigm can distinguish CD from UC and may be further associated with response to tumor necrosis factor inhibitor in CD and disease activity in patients with IBD.

Immunogenicity of Influenza Vaccine for Patients with Inflammatory Bowel Disease on Maintenance Infliximab Therapy: A Randomized Trial.

Background:In patients with inflammatory bowel disease (IBD) on infliximab, data are limited on immune response to influenza vaccine and the impact of vaccine timing. The study aims were to evaluate immune responses to the influenza vaccine in IBD patients on infliximab and the impact of vaccine timing on immune responses. Methods:In this randomized study, 137 subjects with IBD on maintenance infliximab therapy were allocated to receive the 2012/2013 inactivated influenza vaccine at the time of infliximab infusion (n = 69) or midway between infusions (n = 68). Serum was collected before and after vaccination for hemagglutination inhibition titers. Serologic protection was defined by postvaccine titer of ≥1:40. Results:Comparing subjects vaccinated at the time of infliximab with those vaccinated midway, serologic protection was achieved in 67% versus 66% to H1N1 (P = 0.8), in 43% versus 49% to H3N2 (P = 0.5), and in 69% versus 79% to influenza B (P = 0.2). Although solicited adverse events were common (60%), no subject experienced a serious adverse event requiring additional medical attention. Only 6% of subjects had a clinically significant increase in disease activity score, not impacted by vaccine timing. Conclusions:Serologic protection to influenza vaccine is achieved in only approximately 45% to 80% of IBD patients on maintenance infliximab therapy varying by antigen. Yet, importantly, vaccine timing relative to infliximab infusion does not affect the achievement of serologic protection, and the influenza vaccine is well tolerated. Therefore, influenza vaccination at any point during infliximab scheduling is recommended for patients with IBD and opportunities to broaden the availability and convenience of influenza vaccine to optimize coverage should be explored.

Efficient activation of Vα14 invariant NKT cells by foreign lipid antigen is associated with concurrent dendritic cell-specific self recognition

A burst release of cytokines by Vα14 invariant NKT (iNKT) cells upon their TCR engagement critically regulates innate and adaptive immune responses. However, it remains unclear in vivo why iNKT cells respond efficiently to microbial or intracellular lipid Ags that are at low levels or that possess suboptimal antigenicity. We found that dendritic cells (DCs) potentiated iNKT cells to respond to a minimal amount of ligand α-galactosylceramide (αGalCer) through CD1d-dependent autoreactive responses that require endosomal processing and CD1d trafficking. The ability of potentiation of NKT cells was DC specific and did not depend on costimulatory signals and IL-12 production by DCs. However, DCs that failed to synthesize a major endogenous lipid Ag isoglobotrihexosylceramide were unable to potentiate NKT cells for efficient activation. Further analysis showed that differences in the level and pattern of endogenous lipid Ag presentation differentiate DCs and B cells for effective potentiation and subsequent activation of iNKT cells in the presence of an exogenous Ag. Thus, CD1d-dependent potentiation by DCs may be crucial for iNKT cell-mediated immunity against infectious agents.

Enhanced Early Expansion and Maturation of Semi-Invariant NK T Cells Inhibited Autoimmune Pathogenesis in Congenic Nonobese Diabetic Mice

Semi-invariant NK T cell (iNKT) deficiency has long been associated with the pathogenesis of type 1 diabetes (T1D), but the linkage between this the deficiency and T1D susceptibility gene(s) remains unclear. We analyzed NOD mice subcongenic for resistant alleles of Idd9 locus in search for protective mechanisms against T1D, and found that iNKT cell development was significantly enhanced with a more advanced mature phenotype and function in mice containing Idd9.1 sublocus of B10 origin. The enhanced iNKT cell development and function suppressed effector function of diabetogenic T cells. Elimination of iNKT cells by CD1d deficiency almost abolished T1D protection in these mice. Interestingly, although the iNKT cells were responsible for a Th2 orientated cytokine profile that is often regarded as a mechanism of T1D prevention, our data suggests that the Th2 bias played little if any role for the protection. In addition, dendritic cells from the congenic NOD mice showed increased abilities to engage and potentiate iNKT cells, suggesting that a mechanism mediated by dendritic cells or other APCs may be critical for the enhanced development and maturation of iNKT cells. The products of T1D susceptibility gene(s) in Idd9.1 locus may be a key factor for this mechanism.