Transcription Factor Blimp-1 Research Articles

TIGIT Regulates T Cell Inflammation in Airway Inflammatory Diseases.

TIGIT, a co-inhibitory receptor found on T cells and NK cells, transmits inhibitory signals upon binding to its ligand. This interaction suppresses the activation of various signaling pathways, leading to functional exhaustion of cells, ultimately dampening excessive inflammatory responses or facilitating immune evasion in tumors. Dysregulated TIGIT expression has been noted in T cells across different inflammatory conditions, exhibiting varying effects based on T cell subsets. TIGIT predominantly restrains the effector function of pro-inflammatory T cells, upholds the suppressive function of regulatory T cells, and influences Tfh maturation. Mechanistically, the IL27-induced transcription factors c-Maf and Blimp-1 are believed to be key regulators of TIGIT expression in T cells. Notably, TIGIT expression in T cells is implicated in lung diseases, particularly airway inflammatory conditions such as lung cancer, obstructive pulmonary disease, interstitial lung disease, sarcoidosis, and COVID-19. This review emphasizes the significance of TIGIT in the context of T cell immunity and airway inflammatory diseases.

Blimp-1 and c-Maf regulate immune gene networks to protect against distinct pathways of pathobiont-induced colitis

Intestinal immune responses to microbes are controlled by the cytokine IL-10 to avoid immune pathology. Here, we use single-cell RNA sequencing of colon lamina propria leukocytes (LPLs) along with RNA-seq and ATAC-seq of purified CD4+ T cells to show that the transcription factors Blimp-1 (encoded by Prdm1) and c-Maf co-dominantly regulate Il10 while negatively regulating proinflammatory cytokines in effector T cells. Double-deficient Prdm1fl/flMaffl/flCd4Cre mice infected with Helicobacter hepaticus developed severe colitis with an increase in TH1/NK/ILC1 effector genes in LPLs, while Prdm1fl/flCd4Cre and Maffl/flCd4Cre mice exhibited moderate pathology and a less-marked type 1 effector response. LPLs from infected Maffl/flCd4Cre mice had increased type 17 responses with increased Il17a and Il22 expression and an increase in granulocytes and myeloid cell numbers, resulting in increased T cell–myeloid–neutrophil interactions. Genes over-expressed in human inflammatory bowel disease showed differential expression in LPLs from infected mice in the absence of Prdm1 or Maf, revealing potential mechanisms of human disease.

Identification of Intestinal Lamina Propria Plasma Cells by Surface Transmembrane Activator and CAML Interactor Expression.

Plasma cells secrete an abundance of Abs and are a crucial component of our immune system. The intestinal lamina propria harbors the largest population of plasma cells, most of which produce IgA. These Abs can bind to beneficial gut bacteria to reinforce intestinal homeostasis and provide protection against enteric pathogens. Plasma cells downregulate many cell-surface proteins commonly used to identify B cells. In mice, expression of the surface marker CD138 has been widely used to identify plasma cells in lymph nodes, bone marrow, and spleen. Intestinal plasma cells require liberation via extensive tissue processing involving treatment with collagenase. We report that detection of CD138 surface expression is reduced following collagenase treatment. Using a mouse in which yellow fluorescent protein expression is controlled by the plasma cell requisite transcription factor Blimp-1, we show that surface detection of transmembrane activator and CAML interactor captures a significant proportion of Ab-secreting plasma cells in the intestinal lamina propria and gut-draining mesenteric lymph nodes. Additionally, we describe a flow cytometry panel based on the detection of surface markers to identify murine B cell subsets in the intestinal lamina propria and, as a proof of concept, combine it with a cutting-edge fate-tracking system to characterize the fate of germinal center B cells activated in early life. By identifying plasma cells and other key intestinal B subsets in a manner compatible with several downstream applications, including sorting and culturing and invitro manipulations, this efficient and powerful approach can enhance studies of mucosal immunity.

Depletion of follicular B cell-derived antibody secreting cells does not attenuate angiotensin II-induced hypertension or vascular compliance.

Marginal zone and follicular B cells are known to contribute to the development of angiotensin II-induced hypertension in mice, but the effector function(s) mediating this effect (e.g., antigen presentation, antibody secretion and/or cytokine production) are unknown. B cell differentiation into antibody secreting cells (ASCs) requires the transcription factor Blimp-1. Here, we studied mice with a Blimp-1 deficiency in follicular B cells to evaluate whether antibody secretion underlies the pro-hypertensive action of B cells. 10- to 14-week-old male follicular B cell Blimp-1 knockout (FoB-Blimp-1-KO) and floxed control mice were subcutaneously infused with angiotensin II (0.7 mg/kg/d) or vehicle (0.1% acetic acid in saline) for 28 days. BP was measured by tail-cuff plethysmography or radiotelemetry. Pulse wave velocity was measured by ultrasound. Aortic collagen was quantified by Masson's trichrome staining. Cell types and serum antibodies were quantified by flow cytometry and a bead-based multiplex assay, respectively. In control mice, angiotensin II modestly increased serum IgG3 levels and markedly increased BP, cardiac hypertrophy, aortic stiffening and fibrosis. FoB-Blimp-1-KO mice exhibited impaired IgG1, IgG2a and IgG3 production despite having comparable numbers of B cells and ASCs to control mice. Nevertheless, FoB-Blimp-1-KO mice still developed hypertension, cardiac hypertrophy, aortic stiffening and fibrosis following angiotensin II infusion. Inhibition of follicular B cell differentiation into ASCs did not protect against angiotensin II-induced hypertension or vascular compliance. Follicular B cell functions independent of their differentiation into ASCs and ability to produce high-affinity antibodies, or other B cell subtypes, are likely to be involved in angiotensin II-induced hypertension.

Blimp-1 and c-Maf regulate Il10 andnegatively regulate common and unique proinflammatory gene networks in IL-12 plus IL-27-driven T helper-1 cells.

CD4 + Th1 cells producing IFN-γ are required to eradicate intracellular pathogens, however if uncontrolled these cells can cause immunopathology. The cytokine IL-10 is produced by multiple immune cells including Th1 cells during infection and regulates the immune response to minimise collateral host damage. In this study we aimed to elucidate the transcriptional network of genes controlling the expression of Il10 and proinflammatory cytokines, including Ifng in Th1 cells differentiated from mouse naive CD4 + T cells. We applied computational analysis of gene regulation derived from temporal profiling of gene expression clusters obtained from bulk RNA sequencing (RNA-seq) of flow cytometry sorted naïve CD4 + T cells from mouse spleens differentiated in vitro into Th1 effector cells with IL-12 and IL-27 to produce Ifng and Il10, compared to IL-27 alone which express Il10 only , or IL-12 alone which express Ifng and no Il10, or medium control driven-CD4 + T cells which do not express effector cytokines . Data were integrated with analysis of active genomic regions from these T cells using an assay for transposase-accessible chromatin with sequencing (ATAC)-seq, integrated with literature derived-Chromatin-immunoprecipitation (ChIP)-seq data and the RNA-seq data, to elucidate the transcriptional network of genes controlling expression of Il10 and pro-inflammatory effector genes in Th1 cells. The co-dominant role for the transcription factors, Prdm1 (encoding Blimp-1) and Maf (encoding c-Maf) , in cytokine gene regulation in Th1 cells, was confirmed using T cells obtained from mice with T-cell specific deletion of these transcription factors. We show that the transcription factors Blimp-1 and c-Maf each have unique and common effects on cytokine gene regulation and not only co-operate to induce Il10 gene expression in IL-12 plus IL-27 differentiated mouse Th1 cells, but additionally directly negatively regulate key proinflammatory cytokines including Ifng, thus providing mechanisms for reinforcement of regulated Th1 cell responses. These data show that Blimp-1 and c-Maf positively and negatively regulate a network of both unique and common anti-inflammatory and pro-inflammatory genes to reinforce a Th1 response in mice that will eradicate pathogens with minimum immunopathology.

A p38α-BLIMP1 signalling pathway is essential for plasma cell differentiation

Plasma cells (PC) are antibody-secreting cells and terminal effectors in humoral responses. PCs differentiate directly from activated B cells in response to T cell-independent (TI) antigens or from germinal center B (GCB) cells in T cell-dependent (TD) antigen-induced humoral responses, both of which pathways are essentially regulated by the transcription factor BLIMP1. The p38 mitogen-activated protein kinase isoforms have already been implicated in B cell development, but the precise role of p38α in B cell differentiation is still largely unknown. Here we show that PC differentiation and antibody responses are severely impaired in mice with B cell-specific deletion of p38α, while B cell development and the GCB cell response are spared. By utilizing a Blimp1 reporter mouse model, we show that p38α-deficiency results in decreased BLIMP1 expression. p38α-driven BLIMP1 up-regulation is required for both TI and TD PCs differentiation. By combining CRISPR/Cas9 screening and other approaches, we identify TCF3, TCF4 and IRF4 as downstream effectors of p38α to control PC differentiation via Blimp1 transcription. This study thus identifies an important signalling pathway underpinning PC differentiation upstream of BLIMP1, and points to a highly specialized and non-redundant role for p38α among p38 isoforms.

Multi-target regulation of pro-inflammatory cytokine production by transcription factor Blimp-1

ObjectiveCytokine storm syndrome is a fatal condition related to infectious and autoimmune diseases. Here, we aim to investigate the regulatory mechanisms of Blimp-1 on multiple cytokine production.MethodsThe Blimp1 shRNA was transfected into RAW264.7 macrophages, followed by Toll-like receptor (TLR) ligand stimulation. The mRNA and protein levels of cytokines were detected by real-time PCR and flow cytometric bead array. The nuclear translocation of AP-1 and NF-κB p65 was measured by immunofluorescence staining. The transcriptional activity was detected by luciferase reporter assay with 5 × NF-κB reporter or with IL6 promoter reporter.ResultsBlimp-1 significantly inhibited the expression and secretion of IL-1β, IL-6, and IL-18 in macrophages during stimulation with a variety of TLR ligands. The immunofluorescence staining results showed that Blimp-1 strictly controlled the nuclear translocation of NF-κB p65 in LPS-challenged macrophages. Furthermore, Blimp-1 directly inhibited the transcriptional activity of NF-κB and the transcription of IL6 gene.ConclusionBlimp-1 represses the production of multiple pro-inflammatory cytokines by directly binding the genomic region and restricting the nuclear translocation and transcriptional activity of NF-κB. This finding may provide potential therapeutic strategies for the cytokine storm-related diseases.

Astragalus polysaccharide alleviates ulcerative colitis by regulating the balance of Tfh/Treg cells

The immunomodulatory function of natural active ingredients has long been a focus of scientific research, with recent hotspots reporting targeted modulation of the follicular helper T cells (Tfh)/regulatory T cells (Treg) balance as an emerging strategy for the treatment of ulcerative colitis (UC). Here, dextran sodium sulfate induced mice UC and Astragalus polysaccharide (APS, 200 mg/kg/day) was administered simultaneously. In this study, APS effectively alleviated colitis in mice by improving survival rate, disease activity index (DAI), the change rate of body weight, colonic length and weight, and histopathological injury of the colon. Moreover, APS regulated the expression of inflammatory cytokines interleukin (IL)-2, IL-6, IL-12p70, IL-23, Tumour necrosis factor (TNF)-ɑ, and transforming growth factor (TGF)-β1 in colonic tissues of colitis mice. Importantly, APS significantly downregulated Tfh cell and the expression of its related nuclear transcription factors Blimp-1 and Bcl-6, and cytokine IL-21. Meanwhile, APS regulated the differentiation of Tfh subpopulations in colitis mice, with Tfh10 and Tfr significantly upregulated while Tfh1, Tfh17, and Tfh21 significantly downregulated. In addition, APS significantly upregulated Treg cells and the levels of its associated nuclear transcription factor Foxp3, and cytokine IL-10 in colitis mice. In conclusion, APS effectively alleviated UC by reshaping the balance of Tfh/Treg cells.

Mechanism of Tim-3 regulation of CD8+ T cell function

Abstract Tim-3 or transmembrane immunoglobulin and mucin domain-3 is a type I membrane protein expressed by various immune cell types, and has been shown to have a co-stimulatory role in T cells through the PI3K pathway. Tim-3 comprises extracellular, transmembrane and intracellular domains, the latter of which contains five tyrosine residues. We and others have shown that these tyrosines participate in TCR-mediated signaling pathways. Using LCMV infection, we also found that Tim-3 expression influences the formation of short-lived effector and memory precursor CD8+ T cells. We hypothesize that Tim-3 signaling through the tyrosine residues in its cytoplasmic domain modulates CD8+ T cell activation and differentiation. To test this hypothesis, we have used LCMV-specific TCR transgenic (P14) mice crossed to truncated version of Tim-3 and CD8+ conditional deletion of Tim-3. CD8+ T cell activation and memory were analyzed by flow cytometry after infection with LCMV Armstrong to induce an acute infection. The effects of Tim-3 on CD8+ T cells were found to be most prominent at the effector stage of infection and RNA sequencing has shown that loss of Tim-3 signaling promotes acquisition of a memory-like phenotype. Transcription factors T-bet and Blimp-1 were found to be lower in Tim-3 KO CD8+ T cells, while Tcf-1 maintained at higher levels in the absence of Tim-3 signaling. Cytokine production by effector CD8+ T cells was also found to be lower when Tim-3 was absent. Thus, we conclude that Tim-3 signaling via its cytoplasmic tyrosine residues plays a role in CD8+ T cells in an acute infection. These results may lead to a better understanding of CD8+ T cell activation and memory development in different settings. Supported by R01 AI138504-01A1

Co-Expression of the B-Cell Key Transcription Factors Blimp-1 and IRF4 Identifies Plasma Cells in the Pig.

Antibody-secreting plasma cells (PCs) have remained largely uncharacterized for years in the field of porcine immunology. For an in-depth study of porcine PCs, we identified cross-reactive antibodies against three key transcription factors: PR domain zinc finger protein-1 (Blimp-1), interferon regulatory factor 4 (IRF4), and paired box 5 (Pax5). A distinct Blimp-1+IRF4+ cell population was found in cells isolated from blood, spleen, lymph nodes, bone marrow, and lung of healthy pigs. These cells showed a downregulation of Pax5 compared to other B cells. Within Blimp-1+IRF4+ B cells, IgM-, IgG-, and IgA-expressing cells were identified and immunoglobulin-class distribution was clearly different between the anatomical locations, with IgA+ PCs dominating in lung tissue and IgM+ PCs dominating in the spleen. Expression patterns of Ki-67, MHC-II, CD9, and CD28 were investigated in the different organs. A high expression of Ki-67 was observed in blood, suggesting a plasmablast stage. Blimp-1+IRF4+ cells showed an overall lower expression of MHC-II compared to regular B cells, confirming a progressive loss in B-cell differentiation toward the PC stage. CD28 showed slightly elevated expression levels in Blimp-1+IRF4+ cells in most organs, a phenotype that is also described for PCs in mice and humans. This was not seen for CD9. We further developed a FACS-sorting strategy for live porcine PCs for functional assays. CD3-CD16-CD172a– sorted cells with a CD49dhighFSC-Ahigh phenotype contained Blimp-1+IRF4+ cells and were capable of spontaneous IgG production, thus confirming PC identity. These results reveal fundamental phenotypes of porcine PCs and will facilitate the study of this specific B-cell subset in the future.

HIV-1 Vpr drives a tissue residency-like phenotype during selective infection of resting memory Tcells.

SummaryHIV-1 replicates in CD4+ T cells, leading to AIDS. Determining how HIV-1 shapes its niche to create a permissive environment is central to informing efforts to limit pathogenesis, disturb reservoirs, and achieve a cure. A key roadblock in understanding HIV-T cell interactions is the requirement to activate T cells in vitro to make them permissive to infection. This dramatically alters T cell biology and virus-host interactions. Here we show that HIV-1 cell-to-cell spread permits efficient, productive infection of resting memory T cells without prior activation. Strikingly, we find that HIV-1 infection primes resting T cells to gain characteristics of tissue-resident memory T cells (TRM), including upregulating key surface markers and the transcription factor Blimp-1 and inducing a transcriptional program overlapping the core TRM transcriptional signature. This reprogramming is driven by Vpr and requires Vpr packaging into virions and manipulation of STAT5. Thus, HIV-1 reprograms resting T cells, with implications for viral replication and persistence.

The Blimp-1 transcription factor acts in non-neuronal cells to regulate terminal differentiation of the Drosophila eye.

The formation of a functional organ such as the eye requires specification of the correct cell types and their terminal differentiation into cells with the appropriate morphologies and functions. Here, we show that the zinc-finger transcription factor Blimp-1 acts in secondary and tertiary pigment cells in the Drosophila retina to promote the formation of a bi-convex corneal lens with normal refractive power, and in cone cells to enable complete extension of the photoreceptor rhabdomeres. Blimp-1 expression depends on the hormone ecdysone, and loss of ecdysone signaling causes similar differentiation defects. Timely termination of Blimp-1 expression is also important, as its overexpression in the eye has deleterious effects. Our transcriptomic analysis revealed that Blimp-1 regulates the expression of many structural and secreted proteins in the retina. Blimp-1 may function in part by repressing another transcription factor; Slow border cells is highly upregulated in the absence of Blimp-1, and its overexpression reproduces many of the effects of removing Blimp-1. This work provides insight into the transcriptional networks and cellular interactions that produce the structures necessary for visual function.

A Notch/STAT3-driven Blimp-1/c-Maf-dependent molecular switch induces IL-10 expression in human CD4+ T cells and is defective in Crohn\xb4s disease patients

Immunosuppressive Interleukin (IL)−10 production by pro-inflammatory CD4+ T cells is a central self-regulatory function to limit aberrant inflammation. Still, the molecular mediators controlling IL-10 expression in human CD4+ T cells are largely undefined. Here, we identify a Notch/STAT3 signaling-module as a universal molecular switch to induce IL-10 expression across human naïve and major effector CD4+ T cell subsets. IL-10 induction was transient, jointly controlled by the transcription factors Blimp-1/c-Maf and accompanied by upregulation of several co-inhibitory receptors, including LAG-3, CD49b, PD-1, TIM-3 and TIGIT. Consistent with a protective role of IL-10 in inflammatory bowel diseases (IBD), effector CD4+ T cells from Crohn’s disease patients were defective in Notch/STAT3-induced IL-10 production and skewed towards an inflammatory Th1/17 cell phenotype. Collectively, our data identify a Notch/STAT3—Blimp-1/c-Maf axis as a common anti-inflammatory pathway in human CD4+ T cells, which is defective in IBD and thus may represent an attractive therapeutic target.

A System for In Vitro Generation of Mature Murine Plasma Cells Uncovers Differential Blimp-1/Prdm1 Promoter Usage.

Upon encounter with Ag, B cells undergo a sequential process of differentiation to become Ab-secreting plasma cells. Although the key drivers of differentiation have been identified, research has been limited by the lack of in vitro models recapitulating the full process for murine B cells. In this study, we describe methodology using BCR or TLR ligation to obtain plasma cells that are phenotypically mature, have exited cell cycle and express a gene signature concordant with long-lived plasma cells. Dependent on the initial stimuli, the transcriptomes also show variation including the enhanced expression of matrisome components after BCR stimulation, suggestive of unique functional properties for the resultant plasma cells. Moreover, using the new culture conditions we demonstrate that alternative promoter choice regulating the expression of the master transcription factor Blimp-1/Prdm1 can be observed; when the canonical B cell promoter for Prdm1 is deleted, differentiating B cells exhibit flexibility in the choice of promoter, dictated by the initiating stimulus, with preferential maintenance of expression following exposure to TLR ligation. Thus our system provides a readily tractable model for furthering our understanding of plasma cell biology.

Evaluation of MicroRNA-125b-5p and Transcription Factors BLIMP1 and IRF4 Expression in Unsolved Common Variable Immunodeficiency Patients.

Common variable immunodeficiency (CVID) is the most prevalent form of symptomatic primary humoral immunodeficiencies characterized by failure in the final differentiation of B lymphocytes. The majority of CVID cases have no identified genetic defect, and epigenetic alteration could be involved in the pathogenesis of CVID. Hence, we aimed to evaluate the expression of hsa-miR-125b-5p -and, B lymphocyte-induced maturation protein-1(BLIMP-1) and interferon regulatory protein-4 (IRF-4) in a group of CVID patients with no definitive genetic diagnosis in comparison with healthy individuals. Ten CVID patients (all known genes excluded) and 10 age and sex-matched healthy controls participated in the study. B lymphocytes were isolated and expression of miR-125b-5p, IRF4, and BLIMP1 were evaluated by real-time polymerase chain reaction (RT-PCR). Moreover, B cell subsets were analyzed by flow cytometry. The results showed that the relative expression of miR-125b-5p in CVID patients was increased while it was decreased for the BLIMP1 and IRF4 transcription factors compared with the healthy controls. Although a reduction was observed in switched and non-switched memory B cells among all high-miR patients, these subsets were decreased in patients with normal miRexpression (71.0% and 85.0%, respectively). Our results suggest that overexpression of miR-125b-5p affects the terminal differentiation of B cells in a selected group of CVID patients by downregulating the BLIMP-1 gene and more intensively for the IRF-4 gene expressions.

Remodeling of the tumor microenvironment via disrupting Blimp1+ effector Treg activity augments response to anti-PD-1 blockade

BackgroundAccumulation of Foxp3+ regulatory T (Treg) cells in the tumor often represents an important mechanism for cancer immune evasion and a critical barrier to anti-tumor immunity and immunotherapy. Many tumor-infiltrating Treg cells display an activated phenotype and express the transcription factor Blimp1. However, the specific impact of these Blimp1+ Treg cells and their follicular regulatory T (TFR) cell subset on tumor and the underlying mechanisms of action are not yet well-explored.MethodsVarious transplantable tumor models were established in immunocompetent wild-type mice and mice with a Foxp3-specific ablation of Blimp1. Tumor specimens from patients with metastatic melanoma and TCGA datasets were analyzed to support the potential role of Treg and TFR cells in tumor immunity. In vitro culture assays and in vivo adoptive transfer assays were used to understand how Treg, TFR cells and antibody responses influence tumor control. RNA sequencing and NanoString analysis were performed to reveal the transcriptome of tumor-infiltrating Treg cells and tumor cells, respectively. Finally, the therapeutic effects of anti-PD-1 treatment combined with the disruption of Blimp1+ Treg activity were evaluated.ResultsBlimp1+ Treg and TFR cells were enriched in the tumors, and higher tumoral TFR signatures indicated increased risk of melanoma metastasis. Deletion of Blimp1 in Treg cells resulted in impaired suppressive activity and a reprogramming into effector T-cells, which were largely restricted to the tumor-infiltrating Treg population. This destabilization combined with increased anti-tumor effector cellular responses, follicular helper T-cell expansion, enhanced tumoral IgE deposition and activation of macrophages secondary to dysregulated TFR cells, remodeled the tumor microenvironment and delayed tumor growth. The increased tumor immunogenicity with MHC upregulation improved response to anti-PD-1 blockade. Mechanistically, Blimp1 enforced intratumoral Treg cells with a unique transcriptional program dependent on Eomesodermin (Eomes) expression; deletion of Eomes in Blimp1-deficient Treg cells restored tumor growth and attenuated anti-tumor immunity.ConclusionsThese findings revealed Blimp1 as a new critical regulator of tumor-infiltrating Treg cells and a potential target for modulating Treg activity to treat cancer. Our study has also revealed two FCERIA-containing immune signatures as promising diagnostic or prognostic markers for melanoma patients.

Integration of T helper and BCR signals governs enhanced plasma cell differentiation of memory B cells by regulation of CD45 phosphatase activity.

SUMMARYHumoral immunity relies on the efficient differentiation of memory B cells (MBCs) into antibody-secreting cells (ASCs). T helper (Th) signals upregulate B cell receptor (BCR) signaling by potentiating Src family kinases through increasing CD45 phosphatase activity (CD45 PA).In this study, we show that high CD45 PA in MBCs enhances BCR signaling and is essential for their effective ASC differentiation. Mechanistically, Th signals upregulate CD45 PA through intensifying the surface binding of a CD45 ligand, Galectin-1. CD45 PA works as a sensor of T cell help and defines high-affinity germinal center (GC) plasma cell (PC) precursors characterized by IRF4 expression in vivo. Increasing T cell help in vitro results in an incremental CD45 PA increase and enhances ASC differentiation by facilitating effective induction of the transcription factors IRF4 and BLIMP1. This study connects Th signals with BCR signaling through Galectin-1-dependent regulation of CD45 PA and provides a mechanism for efficient ASC differentiation of MBCs.

ITK regulates IL-10 production by CD8+ T cells and lung immunopathology during influenza infection

Abstract Respiratory viral infections are a major cause of human morbidity and mortality. Influenza (flu) infections cause 250,000 deaths and 3–5 million cases of severe illness during the average flu season. Severe flu infections are associated with a combination of strong pro-inflammatory and weak anti-inflammatory immune responses. Production of the immunomodulatory cytokine IL-10 by T cells restricts immunopathology during flu infections, however our knowledge of the signaling pathways regulating IL-10 induction during flu is limited. Using IL-10GFP reporter mouse models, we found that Interleukin-2 inducible T cell kinase (ITK), a critical component in T cell receptor (TCR) signaling, regulates immunopathology and the development of IL-10-producing CD8+ T cells in the airways during influenza A infection. Utilizing alternative coculture, the model antigen ovalbumin (OVA), and transgenic TCR specific for OVA in CD8+ T cells (OTI) adoptive transfer, we determined that ITK regulates flu antigen-specific IL-10+CD8+ T cell differentiation in a CD8+ T cell intrinsic and extrinsic manner. Exogenous IL-2 rescued IL-10 production by Itk−/− CD8+ T cells, in vitro and in vivo, suggesting that ITK may play a role in IL-2-producing T helper cells that promote IL-10 production in CD8+ T cells. The absence of ITK impaired the expression of transcription factors (TFs) IRF4 and Blimp-1, and retroviral transduction enabling the reintroduction of these TFs could partially rescue IL-10 production in Itk−/− CD8+ T cells. Our data suggests that ITK is a critical regulator of IL-10 production by CD8+ T cells and flu-induced pulmonary immunopathology. Modulating ITK signaling may be a strategy for regulating immunopathology due to viral infections.

Dysregulated follicular regulatory T cells and antibody responses exacerbate CNS autoimmunity

Abstract Follicular regulatory T (TFR) cells are essential for the regulation of germinal center (GC) response and humoral self-tolerance. Dysregulated follicular helper T (TFH) cell-GC-antibody (Ab) response secondary to dysfunctional TFR cells is the root of an array of autoimmune disorders. The contribution of TFR cells to the pathogenesis of multiple sclerosis (MS) and murine experimental autoimmune encephalomyelitis (EAE) remains largely unclear. Here we show that mice with a FoxP3-specific ablation of the transcription factor Blimp1 developed severe EAE and failed to recover compared to control mice, reflecting conversion of unstable regulatory T cells (Tregs) into IL-17A/GM-CSF-producing effector T-cells associated with increased TFH-Ab responses, more IgE deposition in the central nervous system (CNS) and inability to regulate CNS CD11b+ myeloid cells. Notably, serum IgE titers were positively correlated with EAE scores, and culture of CNS CD11b+ cells with sera from these EAE mice enhanced their activation, while transfer of Blimp1-deficient TFR cells promoted Ab production, activation of CNS CD11b+ cells and EAE. Moreover, compared to healthy controls, a small group of MS patients had fewer circulating Tregs and TFR cells that expressed reduced levels of Blimp1, associated with elevated serum IgE. These findings suggest that Blimp1 is essential for the maintenance of TFR cells and Ab responses in EAE, and that dysregulated TFR cells and Ab responses promote CNS autoimmunity.

Tregs facilitate obesity and insulin resistance via a Blimp-1/IL-10 axis.

Interleukin-10 (IL-10) is a critical cytokine used by immune cells to suppress inflammation. Paradoxically, immune cell–derived IL-10 can drive insulin resistance in obesity by suppressing adipocyte energy expenditure and thermogenesis. However, the source of IL-10 necessary for the suppression of adipocyte thermogenesis is unknown. We show here that CD4+Foxp3+ regulatory T cells (Tregs) are a substantial source of IL-10 and that Treg-derived IL-10 can suppress adipocyte beiging. Unexpectedly, Treg-specific loss of IL-10 resulted in increased insulin sensitivity and reduced obesity in high-fat diet–fed male mice. Mechanistically, we determined that Treg-specific loss of the transcription factor Blimp-1, a driver of IL-10 expression by Tregs, phenocopied the Treg-specific IL-10–deficient mice. Loss of Blimp-1 expression in Tregs resulted in reduced ST2+KLRG1+, IL-10-secreting Tregs, particularly in the white adipose tissue. Blimp-1–deficient mice were protected from glucose intolerance, insulin resistance, and diet-induced obesity, through increased white adipose tissue browning. Taken together, our data show that Blimp-1–regulated IL-10 secretion by Tregs represses white adipose tissue beiging to maintain adipose tissue homeostasis.