Microbiology & Immunology - Research Publications
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ItemDynamics of immunoglobulin sequence diversity in HIV-1 infected individuals(ROYAL SOC, 2015-09-05)Advances in immunoglobulin (Ig) sequencing technology are leading to new perspectives on immune system dynamics. Much research in this nascent field has focused on resolving immune responses to viral infection. However, the dynamics of B-cell diversity in early HIV infection, and in response to anti-retroviral therapy, are still poorly understood. Here, we investigate these dynamics through bulk Ig sequencing of samples collected over 2 years from a group of eight HIV-1 infected patients, five of whom received anti-retroviral therapy during the first half of the study period. We applied previously published methods for visualizing and quantifying B-cell sequence diversity, including the Gini index, and compared their efficacy to alternative measures. While we found significantly greater clonal structure in HIV-infected patients versus healthy controls, within HIV patients, we observed no significant relationships between statistics of B-cell clonal expansion and clinical variables such as viral load and CD4(+) count. Although there are many potential explanations for this, we suggest that important factors include poor sampling resolution and complex B-cell dynamics that are difficult to summarize using simple summary statistics. Importantly, we find a significant association between observed Gini indices and sequencing read depth, and we conclude that more robust analytical methods and a closer integration of experimental and theoretical work is needed to further our understanding of B-cell repertoire diversity during viral infection.
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ItemNo Preview AvailableCD4+ T Cells Specific for C. difficile Toxins are a Marker of Patients with Active Relapsing Disease(Oxford University Press (OUP), 2017-10-01)Background: The bacterial pathogen Clostridium difficile is the leading cause of nosocomial infectious diarrhea. Although C. difficile infection (CDI) can be treated with antibiotics, approximately 25% of patients relapse after treatment. The pathogenicity of CDI requires the activities of its toxins, TcdA and TcdB, but T cell-mediated responses to these toxins remain uncharacterized. Methods: We enrolled two cohorts of patients, one with newly acquired CDI (n = 14) and the other with relapsing CDI (n = 25); and healthy volunteers with no history of CDI (n = 12). We measured peripheral blood CD4+ T cell responses to the toxins using a whole blood flow cytometry assay that identifies antigen-specific CD4+ T cells by co-expression of CD25 and OX40 following 44h incubation with antigen (Fig 1). Results: We found that in patients with recurring CDI, T cell responses to TcdB were significantly higher than in healthy controls (median 1.04% vs. 0.18%; P = 0.003, Fig 2). In contrast, TcdA T cell responses and anti-TcdA/TcdB IgG titres were not different between recurring patients and controls. TcdB, but not TcdA, T cell responses were significantly higher in recurring CDI compared with newly acquired CDI (median 1.04% vs. 0.44%; P = 0.032). In both patient cohorts TcdB-specific CD4+ T cells were functionally heterogeneous, on average: 25% expressed the gut homing marker integrin β7; there was a 1:1 ratio of Tregs to T effectors; and T effectors contained Th1, Th2 and Th17 cells at a 1.5:1:3 ratio. The proportion of Th1 and Th17 cells within TcdB-specific CD4+ T cells was also significantly reduced in recurring, compared with newly acquired, CDI (Fig 3). Analysis of sorted TcdB-specific CD25+OX40+ cells confirmed specificity for TcdB and polarization towards Th17 cells, which are important for intestinal anti-pathogen immunity. Conclusion: This is the first investigation of T cell immunity to C. difficile toxins. Our data show that anti-TcdB CD4+ T cell responses are a more specific marker of disease than IgG titres. Tracking how toxin-specific CD4+ T cell responses change following treatment and/or vaccination not only has the potential to predict relapse, but also to deliver insight into how CD4+ T cell memory develops in response to this prevalent pathogen.
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ItemThe role of FOXP3 in autoimmunity(CURRENT BIOLOGY LTD, 2016-12)FOXP3 controls the development and function of T regulatory cells (Tregs). Autoimmunity is linked to changes in FOXP3 activity that can occur at multiple levels and lead to Treg dysfunction. For example, changes in IL-2 signaling, FOXP3 transcription and/or post-translational modifications can all contribute to loss of self-tolerance. As additional pathways of FOXP3 regulation are elucidated, new therapeutic approaches to increase Treg activity either by cell therapy or pharmacological intervention are being tested. Early success from pioneering studies of Treg-based therapy in transplantation has promoted the undertaking of similar studies in autoimmunity, with emerging evidence for the effectiveness of these approaches, particularly in the context of type 1 diabetes.
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ItemCirculating gluten-specific FOXP3+CD39+ regulatory T cells have impaired suppressive function in patients with celiac disease(MOSBY-ELSEVIER, 2017-12)BACKGROUND: Celiac disease is a chronic immune-mediated inflammatory disorder of the gut triggered by dietary gluten. Although the effector T-cell response in patients with celiac disease has been well characterized, the role of regulatory T (Treg) cells in the loss of tolerance to gluten remains poorly understood. OBJECTIVE: We sought to define whether patients with celiac disease have a dysfunction or lack of gluten-specific forkhead box protein 3 (FOXP3)+ Treg cells. METHODS: Treated patients with celiac disease underwent oral wheat challenge to stimulate recirculation of gluten-specific T cells. Peripheral blood was collected before and after challenge. To comprehensively measure the gluten-specific CD4+ T-cell response, we paired traditional IFN-γ ELISpot with an assay to detect antigen-specific CD4+ T cells that does not rely on tetramers, antigen-stimulated cytokine production, or proliferation but rather on antigen-induced coexpression of CD25 and OX40 (CD134). RESULTS: Numbers of circulating gluten-specific Treg cells and effector T cells both increased significantly after oral wheat challenge, peaking at day 6. Surprisingly, we found that approximately 80% of the ex vivo circulating gluten-specific CD4+ T cells were FOXP3+CD39+ Treg cells, which reside within the pool of memory CD4+CD25+CD127lowCD45RO+ Treg cells. Although we observed normal suppressive function in peripheral polyclonal Treg cells from patients with celiac disease, after a short in vitro expansion, the gluten-specific FOXP3+CD39+ Treg cells exhibited significantly reduced suppressive function compared with polyclonal Treg cells. CONCLUSION: This study provides the first estimation of FOXP3+CD39+ Treg cell frequency within circulating gluten-specific CD4+ T cells after oral gluten challenge of patients with celiac disease. FOXP3+CD39+ Treg cells comprised a major proportion of all circulating gluten-specific CD4+ T cells but had impaired suppressive function, indicating that Treg cell dysfunction might be a key contributor to disease pathogenesis.
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ItemA Novel Whole Blood Assay Detects Flagellin-Specific CD4+ T Cells in Patients with Inflammatory Bowel Disease(W B Saunders Co-Elsevier, 2017-04-01)
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ItemNo Preview AvailableHigh-dimensional Analysis of Human Regulatory T Cells Using Mass Cytometry(American Association of Immunologists, 2016-05-01)
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ItemHow antigen specificity directs regulatory T-cell function: self, foreign and engineered specificity(WILEY, 2016-07)Regulatory T cells (Tregs) are a suppressive subset of T cells that have important roles in maintaining self-tolerance and preventing immunopathology. The T-cell receptor (TCR) and its antigen specificity play a dominant role in the differentiation of cells to a Treg fate, either in the thymus or in the periphery. This review focuses on the effects of the TCR and its antigen specificity on Treg biology. The role of Tregs with specificity for self-antigen has primarily been studied in the context of autoimmune disease, although recent studies have focused on their role in steady-state conditions. The role of Tregs that are specific for pathogens, dietary antigens and allergens is much less studied, although recent data suggest a significant and previously underappreciated role for Tregs during memory responses to a wide range of foreign antigens. The development of TCR- or chimeric antigen receptor (CAR)-transduced T cells means we are now able to engineer Tregs with disease-relevant antigen specificities, paving the way for ensuring specificity with Treg-based therapies. Understanding the role that antigens play in driving the generation and function of Tregs is critical for defining the pathophysiology of many immune-mediated diseases, and developing new therapeutic interventions.
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ItemThe environment of regulatory T cell biology: cytokines, metabolites, and the microbiome(FRONTIERS MEDIA SA, 2015-02-18)Regulatory T cells (Tregs) are suppressive T cells that have an essential role in maintaining the balance between immune activation and tolerance. Their development, either in the thymus, periphery, or experimentally in vitro, and stability and function all depend on the right mix of environmental stimuli. This review focuses on the effects of cytokines, metabolites, and the microbiome on both human and mouse Treg biology. The role of cytokines secreted by innate and adaptive immune cells in directing Treg development and shaping their function is well established. New and emerging data suggest that metabolites, such as retinoic acid, and microbial products, such as short-chain fatty acids, also have a critical role in guiding the functional specialization of Tregs. Overall, the complex interaction between distinct environmental stimuli results in unique, and in some cases tissue-specific, tolerogenic environments. Understanding the conditions that favor Treg induction, accumulation, and function is critical to defining the pathophysiology of many immune-mediated diseases and to developing new therapeutic interventions.