Hoyong Lim

Follicular regulatory T cells expressing Foxp3 and Bcl-6 suppress germinal center reactions

Foxp3+ regulatory T (Treg) cells suppress different types of immune responses to help maintain homeostasis in the body. How Treg cells regulate humoral immunity, including germinal center reactions, is unclear. Here we identify a subset of Treg cells expressing CXCR5 and Bcl-6 that localize to the germinal centers in mice and humans. The expression of CXCR5 on Treg cells depends on Bcl-6. These CXCR5+Bcl-6+ Treg cells are absent in the thymus but can be generated de novo from CXCR5−Foxp3+ natural Treg precursors. A lack of CXCR5+ Treg cells leads to greater germinal center reactions including germinal center B cells, affinity maturation of antibodies and the differentiation of plasma cells. These results unveil a Bcl-6-CXCR5 axis in Treg cells that drives the development of follicular regulatory T (TFR) cells that function to inhibit the germinal center reactions.

Proatherogenic conditions promote autoimmune T helper 17 cell responses in vivo

Patients with systemic autoimmune diseases show increased incidence of atherosclerosis. However, the contribution of proatherogenic factors to autoimmunity remains unclear. We found that atherogenic mice (herein referred to as LDb mice) exhibited increased serum interleukin-17, which was associated with increased numbers of T helper 17 (Th17) cells in secondary lymphoid organs. The environment within LDb mice was substantially favorable for Th17 cell polarization of autoreactive T cells during homeostatic proliferation, which was considerably inhibited by antibodies directed against oxidized low-density lipoprotein (oxLDL). Moreover, the uptake of oxLDL induced dendritic-cell-mediated Th17 cell polarization by triggering IL-6 production in a process dependent on TLR4, CD36, and MyD88. Furthermore, self-reactive CD4(+) T cells that expanded in the presence of oxLDL induced more profound experimental autoimmune encephalomyelitis. These findings demonstrate that proatherogenic factors promote the polarization and inflammatory function of autoimmune Th17 cells, which could be critical for the pathogenesis of atherosclerosis and other related autoimmune diseases.

Negative Regulation of Pulmonary Th17 Responses by C3a Anaphylatoxin during Allergic Inflammation in Mice

Activation of complement is one of the earliest immune responses to exogenous threats, resulting in various cleavage products including anaphylatoxin C3a. In addition to its contribution to host defense, C3a has been shown to mediate Th2 responses in animal models of asthma. However, the role of C3a on pulmonary Th17 responses during allergic inflammation remains unclear. Here, we show that mice deficient in C3a receptor (C3aR) exhibited (i) higher percentages of endogenous IL-17-producing CD4+ T cells in the lungs, (ii) higher amounts of IL-17 in the bronchoalveolar lavage fluid, and (iii) more neutrophils in the lungs than wild-type mice when challenged with intranasal allergens. Moreover, adoptive transfer experiments showed that the frequencies of antigen-specific IL-17-producing CD4+ T cells were significantly higher in the lungs and bronchial lymph nodes of C3aR-deficient recipients than those of wild-types recipients. Bone-marrow reconstitution study indicated that C3aR-deficiency on hematopoietic cells was required for the increased Th17 responses. Furthermore, C3aR-deficient mice exhibited increased percentages of Foxp3+ regulatory T cells; however, depletion of these cells minimally affected the induction of antigen-specific Th17 cell population in the lungs. Neutralization of IL-17 significantly reduced the number of neutrophils in bronchoalveolar lavage fluid of C3aR-deficient mice. Our findings demonstrate that C3a signals negatively regulate antigen-specific Th17 responses during allergic lung inflammation and the size of Foxp3+ regulatory T cell population in the periphery.

Regulation of autoimmune germinal center reactions in lupus-prone BXD2 mice by follicular helper T cells.

BXD2 mice spontaneously develop autoantibodies and subsequent glomerulonephritis, offering a useful animal model to study autoimmune lupus. Although initial studies showed a critical contribution of IL-17 and Th17 cells in mediating autoimmune B cell responses in BXD2 mice, the role of follicular helper T (Tfh) cells remains incompletely understood. We found that both the frequency of Th17 cells and the levels of IL-17 in circulation in BXD2 mice were comparable to those of wild-type. By contrast, the frequency of PD-1+CXCR5+ Tfh cells was significantly increased in BXD2 mice compared with wild-type mice, while the frequency of PD-1+CXCR5+Foxp3+ follicular regulatory T (Tfr) cells was reduced in the former group. The frequency of Tfh cells rather than that of Th17 cells was positively correlated with the frequency of germinal center B cells as well as the levels of autoantibodies to dsDNA. More importantly, CXCR5+ CD4+ T cells isolated from BXD2 mice induced the production of IgG from naïve B cells in an IL-21-dependent manner, while CCR6+ CD4+ T cells failed to do so. These results together demonstrate that Tfh cells rather than Th17 cells contribute to the autoimmune germinal center reactions in BXD2 mice.

Dynamic control of Th2 cell responses by STAT3 during allergic lung inflammation in mice.

Signal transducer and activator of transcription (STAT) family molecules play essential roles during the differentiation of helper T cells from naïve precursors. Although the role of STAT3 in driving Th17 cell polarization has been well established, its role on Th2 responses to allergens remains incompletely understood. By employing T cell-specific STAT3 deficient mice, we demonstrate that STAT3 in T cells plays diverse role on Th2 cells depending on their locations in an animal model of allergic asthma. In the bronchial lymph nodes, STAT3-deficient T cells produced significantly reduced levels of Th2 cytokines. The frequencies of Th2 cells among CD4(+) T cells in the lung were comparable between STAT3-sufficient and STAT3-deficient T cells. By contrast, STAT3-deficient T cells in the airway exhibited significantly enhanced production of Th2 cell cytokines compared to STAT3-sufficient T cells. Interestingly, a major population of IL-4/5 producers among STAT3-deficient T cells in the airway co-produced IFNγ. The frequency of Th17 cells was significantly diminished whereas that of Th1 cells was increased in all the lung-associated tissues. Our results demonstrate the dynamic and opposing roles of STAT3 during the development of Th2 cells from bronchial lymph nodes to the airway and propose the need of careful consideration on STAT3-targeting approaches for the treatment of lung diseases.

Distinct regulation of Th2 and Th17 responses to allergens by pulmonary antigen presenting cells in vivo

Accumulating evidence demonstrates that both Th2 and Th17 responses are involved in the pathogenesis of allergic airway inflammation in animals as well as in humans. The lung contains diverse types of antigen presenting cells. However, the mechanism by which these antigen presenting cells regulate Th2 versus Th17 responses in the lung remains incompletely understood. Here, we show that intranasal administration of fungal protease allergen induced both Th2 and Th17 responses in the lung with different kinetics. Notably, depletion of CD11c(+) cells or macrophages greatly diminished the numbers of allergen-specific Th2 cells in the lung, the infiltration of eosinophils into the airway and airway hyperreactivity. In sharp contrast, depletion of the same antigen presenting cells significantly increased the numbers of allergen-specific Th17 cells in the lung and the infiltration of neutrophils into the airway. Moreover, although a subpopulation of lung epithelial cells express MHC II, lack of MHC II expression in parenchymal cells did not alter pulmonary Th2 and Th17 responses. Our results demonstrate that antigen presenting cells differentially regulate the generation of pulmonary Th2 and Th17 cells in response to intranasal protease allergens.

Fibrinogen cleavage products and Toll-like receptor 4 promote the generation of programmed cell death 1 ligand 2–positive dendritic cells in allergic asthma

Background: Inhaled protease allergens preferentially trigger TH2‐mediated inflammation in allergic asthma. The role of dendritic cells (DCs) on induction of TH2 cell responses in allergic asthma has been well documented; however, the mechanism by which protease allergens induce TH2‐favorable DCs in the airway remains unclear. Objective: We sought to determine a subset of DCs responsible for TH2 cell responses in allergic asthma and the mechanism by which protease allergens induce the DC subset in the airway. Methods: Mice were challenged intranasally with protease allergens or fibrinogen cleavage products (FCPs) to induce allergic airway inflammation. DCs isolated from mediastinal lymph nodes were analyzed for surface phenotype and T‐cell stimulatory function. Anti‐Thy1.2 and Mas‐TRECK mice were used to deplete innate lymphoid cells and mast cells, respectively. Adoptive cell transfer, bone marrow DC culture, anti–IL‐13, and Toll‐like receptor (TLR) 4–deficient mice were used for further mechanistic studies. Results: Protease allergens induced a remarkable accumulation of TH2‐favorable programmed cell death 1 ligand 2 (PD‐L2)+ DCs in mediastinal lymph nodes, which was significantly abolished in mice depleted of mast cells and, to a lesser extent, innate lymphoid cells. Mechanistically, FCPs generated by protease allergens triggered IL‐13 production from wild‐type mast cells but not from TLR4‐deficient mast cells, which resulted in an increase in the number of PD‐L2+ DCs. Intranasal administration of FCPs induced an increase in numbers of PD‐L2+ DCs in the airway, which was significantly abolished in TLR4‐ and mast cell–deficient mice. Injection of IL‐13 restored the PD‐L2+ DC population in mice lacking mast cells. Conclusion: Our findings unveil the “protease–FCP–TLR4–mast cell–IL‐13” axis as a molecular mechanism for generation of TH2‐favorable PD‐L2+ DCs in allergic asthma and suggest that targeting the PD‐L2+ DC pathway might be effective in suppressing allergic T‐cell responses in the airway. Graphical abstract: Figure. No caption available.

Epstein Barr Virus-Induced 3 (EBI3) Together with IL-12 Negatively Regulates T Helper 17-Mediated Immunity to Listeria monocytogenes Infection

Although the protective functions by T helper 17 (Th17) cytokines against extracellular bacterial and fungal infection have been well documented, their importance against intracellular bacterial infection remains unclear. Here, we investigated the contribution of Th17 responses to host defense against intracellular bacteria Listeria monocytogenes and found that Th17 cell generation was suppressed in this model. Unexpectedly, mice lacking both p35 and EBI3 cleared L. monocytogenes as efficiently as wild-type mice, whereas p35-deficient mice failed to do so. Furthermore, both innate cells and pathogen-specific T cells from double-deficient mice produced significantly higher IL-17 and IL-22 compared to wild-type mice. The bacterial burden in the liver of double-deficient mice treated with anti-IL-17 was significantly increased compared to those receiving a control Ab. Transfer of Th17 cells specific for listeriolysin O as well as administration of IL-17 and IL-22 significantly suppressed bacterial growth in p35-deficient mice, indicating the critical contribution of Th17 responses to host defense against the intracellular pathogen in the absence of IL-12 and proper Th1 responses. Our findings unveil a novel immune evasion mechanism whereby the intracellular bacteria exploit IL-27EBI3 to suppress Th17-mediated protective immunity.

Atherogenic dyslipidemia promotes autoimmune follicular helper T cell responses via IL-27.

The incidence of atherosclerosis is higher among patients with systemic lupus erythematosus (SLE); however, the mechanism by which an atherogenic environment affects autoimmunity remains unclear. We found that reconstitution of atherosclerosis-prone Apoe–/– and Ldlr–/– mice with bone marrow from lupus-prone BXD2 mice resulted in increased autoantibody production and glomerulonephritis. This enhanced disease was associated with an increase in CXCR3+ follicular helper T cells (TFH cells). TFH cells isolated from Apoe–/– mice had higher expression of genes associated with inflammatory responses and SLE and were more potent in inducing production of the immunoglobulin IgG2c. Mechanistically, the atherogenic environment induced the cytokine IL-27 from dendritic cells in a Toll-like receptor 4 (TLR4)-dependent manner, which in turn triggered the differentiation of CXCR3+ TFH cells while inhibiting the differentiation of follicular regulatory T cells. Blockade of IL-27 signals diminished the increased TFH cell responses in atherogenic mice. Thus, atherogenic dyslipidemia augments autoimmune TFH cell responses and subsequent IgG2c production in a TLR4- and IL-27-dependent manner.Dyslipidemia and autoimmune disease are often associated. Chung and colleagues demonstrate a mechanistic pathway by which dyslipidemia leads to the induction of pathogenic autoantibodies.

Concomitant suppression of TH2 and TH17 cell responses in allergic asthma by targeting retinoic acid receptor-related orphan receptor γt

Background: Allergic asthma is a heterogeneous chronic inflammatory disease of the airways with a massive infiltration of eosinophils or neutrophils mediated by allergen‐specific TH2 and TH17 cells, respectively. Therefore successful treatment of allergic asthma will require suppression of both TH2 and TH17 cells. Objective: We sought to investigate the role of the TH17 cell pathway in regulating TH2 cell responses in allergic asthma. Methods: Allergic asthma was induced by intranasal challenge with proteinase allergens in C57BL/6, Il17a−/−Il17f−/−, and retinoic acid receptor–related orphan receptor &ggr;t (ROR&ggr;t)gfp/gfp mice. A pharmacologic ROR&ggr;t inhibitor was used to evaluate its preventive and therapeutic effects in allergic asthma. Characteristics of allergic airway inflammation were analyzed by using flow cytometry, histology, quantitative real‐time PCR, and ELISA. Mixed bone marrow chimeric mice, fate mapping analysis, short hairpin RNA transduction, and in vitro T‐cell differentiation were used for mechanistic studies. Results: Mice deficient in IL‐17A and IL‐17F, as well as ROR&ggr;t, exhibited a significant reduction not only in TH17 cell responses but also in TH2 cell responses in an animal model of allergic asthma. Similarly, mice treated with an ROR&ggr;t inhibitor had significantly diminished TH17 and TH2 cell responses, leading to reduced neutrophil and eosinophil numbers in the airway. ROR&ggr;t‐deficient T cells were intrinsically defective in differentiating into TH2 cells and expressed increased levels of B‐cell lymphoma 6 (Bcl6). Bcl6 knockdown resulted in a remarkable restoration of TH2 cell differentiation in ROR&ggr;t‐deficient T cells. Blockade of ROR&ggr;t also significantly hampered the differentiation of human TH2 and TH17 cells from naive CD4+ T cells. Conclusion: ROR&ggr;t in T cells is required for optimal TH2 cell differentiation by suppressing Bcl6 expression; this finding suggests that targeting ROR&ggr;t might be a promising approach for the treatment of allergic asthma by concomitantly suppressing TH17 and TH2 cell responses in the airway.