Functional islet-specific Treg can be generated from CD4+CD25- T cells of healthy and type 1 diabetic subjects.
Journal - European journal of immunology (Germany )
CD4(+)CD25(+)FOXP3(+) Treg cells require TCR engagement for suppressive function, thus ensuring that suppression occurs only in the presence of specific antigens; however, to date no studies have addressed the function of self-antigen-specific Treg in humans. These studies were designed to determine whether peripheral generation and function of islet antigen-specific adaptive Treg are defective in human subjects with type 1 diabetes (T1D). Islet antigen-specific adaptive Treg were induced in vitro by activation of CD4(+)FOXP3(-) T cells with glutamic acid decarboxylase and islet-specific glucose-6-phosphate catalytic subunit-related protein peptides in the context of T1D-associated HLA-DRbeta alleles. Antigen-specific Treg were characterized using flow cytometry for FOXP3 and class II tetramer and assessed for the ability to inhibit proliferation. These adaptive Treg were then compared with influenza-specific Treg from the same study population. The function of tetramer(+) cells that expressed FOXP3 was similar for both influenza and islet antigens generated from control and T1D subjects. In fact, the potency of suppression correlated with FOXP3 expression, not antigen specificity. Thus, these data suggest that development of functional adaptive Treg can occur in response to islet antigens and activation of islet-specific Treg may potentially be used as a targeted immunotherapy in T1D.
|ISSN : ||1521-4141|
|Mesh Heading : ||Antigens, CD4 Diabetes Mellitus, Type 1 Glucose-6-Phosphatase Glutamate Decarboxylase Hemagglutinin Glycoproteins, Influenza Virus Humans Interleukin-2 Receptor alpha Subunit Islets of Langerhans Peptide Fragments T-Lymphocytes, Regulatory immunology metabolism immunology metabolism immunology metabolism immunology immunology immunology metabolism|
|Mesh Heading Relevant : ||immunology immunology immunology|
Defects in IL-2R signaling contribute to diminished maintenance of FOXP3 expression in CD4+CD25+ regulatory T cells of T1D subjects.
Journal - Diabetes
Objective: In humans, multiple genes in the IL-2/IL-2R pathway are associated with T1D. However, no link between IL-2 responsiveness and CD4(+)CD25(+)FOXP3(+) regulatory T cells (Treg) has been demonstrated in T1D subjects despite the role of these IL-2-dependent cells in controlling autoimmunity. Here, we address whether altered IL-2 responsiveness impacts persistence of FOXP3 expression in Treg of T1D subjects. Research design and methods: Persistence of Treg was assessed by culturing sorted CD4(+)CD25(hi) nTreg with IL-2 and measuring FOXP3 expression over-time by flow cytometry for control and T1D populations. The effects of IL-2 on FOXP3 induction were assessed 48 hours following activation of CD4(+)CD25(-) T cells with anti-CD3 antibody. Cytokine receptor expression and signaling upon exposure to IL-2, IL-7 and IL-15 was determined by flow cytometry and western blot analysis. Results: Maintenance of FOXP3 expression in CD4(+)CD25(+) Treg of T1D was diminished in the presence of IL-2, but not IL-7. Impaired responsiveness was not linked to altered expression of the IL-2R complex. Instead, IL-2R signaling was reduced in Treg and total CD4(+) T cells of T1D. In some individuals, decreased STAT5 phosphorylation correlated with significantly higher expression of protein tyrosine phosphatase N2 (PTPN2), a negative regulator of IL-2R signaling. Conclusions: Aberrant IL-2R signaling in CD4(+) T cells of T1D subjects contributes to decreased persistence of FOXP3 expression that may impact establishment of tolerance. These findings suggest novel targets for treatment of T1D within the IL-2R pathway and suggest that an altered IL-2R signaling signature may be a biomarker for T1D.
Combination of rapamycin and IL-2 increases de novo induction of human CD4(+)CD25(+)FOXP3(+) T cells.
Journal - Journal of autoimmunity (England )
The immune system protects itself from autoreactivity by maintaining a balance between effector and Treg responses. Peripheral induction of Treg is one mechanism by which this balance may be maintained. Thus, it is important to understand factors that influence de novo generation of CD4(+)CD25(+)FOXP3(+) Treg. Here, we focus on the effects of cytokines and the cell cycle inhibitor rapamycin. The cytokines IL-2 and IL-7, but not IL-4, increased initial activation induced FOXP3 expression, increased proliferation and sustained expression of FOXP3(+) cells throughout the culture. Addition of rapamycin to cultures containing IL-2 further increased the frequency and absolute number of functional CD4(+)CD25(+)FOXP3(+) Treg. This increase was not due to selective proliferation of FOXP3 cells, but was instead, the result of an increase in the frequency of FOXP3(+) cells induced in G0 through delayed activation while the addition of IL-2 promoted survival and proliferation of the FOXP3(+) population. Thus, combination of rapamycin and IL-2 may provide improved treatment options in transplantation and autoimmunity by promoting induction, survival, and expansion of functional iTreg from CD4(+)CD25(-) cells.
|ISSN : ||0896-8411|
|Mesh Heading : ||Cell Proliferation Cell Survival Forkhead Transcription Factors Humans Immunosuppressive Agents Interleukin-2 Lymphocyte Activation Sirolimus T-Lymphocytes, Regulatory drug effects drug effects metabolism immunology immunology metabolism|
|Mesh Heading Relevant : ||drug effects pharmacology pharmacology drug effects pharmacology drug effects|
Standardized analysis for the quantification of Vbeta CDR3 T-cell receptor diversity.
Journal - Journal of immunological methods (Netherlands )
Assessment of the diversity of the T-cell receptor (TCR) repertoire is often determined by measuring the frequency and distribution of individually rearranged TCRs in a population of T cells. Spectratyping is a common method used to measure TCR repertoire diversity, which examines genetic variation in the third complementarity-determining region (CDR3) region of the TCR Vbeta chain using RT-PCR length-distribution analysis. A variety of methods are currently used to analyze spectratype data including subjective visual measures, qualitative counting measures, and semi-quantitative measures that compare the original data to a standard, control data set. Two major limitations exist for most of these approaches: data files become very wieldy and difficult to manage, and current analytic methods generate data which are difficult to compare between laboratories and across different platforms. Here, we introduce a highly efficient method of analysis that is based upon a normal theoretical Gaussian distribution observed in cord blood and recent thymic emigrants. Using this analysis method, we demonstrate that PBMC obtained from patients with various diseases have skewed TCR repertoire profiles. Upon in vitro activation with anti-CD3 and anti-CD28 coated beads (Xcyte Dynabeads) TCR diversity was restored. Moreover, changes in the TCR repertoire were dynamic in vivo. We demonstrate that use of this streamlined method of analysis in concert with a flexible software package makes quantitative assessment of TCR repertoire diversity straightforward and reproducible, enabling reliable comparisons of diversity values between laboratories and over-time to further collaborative efforts. Analysis of TCR repertoire by such an approach may be valuable in the clinical setting, both for prognostic potential and measuring clinical responses to therapy.
|ISSN : ||0022-1759|
|Mesh Heading : ||Complementarity Determining Regions Humans Immune System Diseases Leukocytes, Mononuclear Sensitivity and Specificity T-Lymphocytes blood immunology metabolism metabolism|
|Mesh Heading Relevant : ||Receptor-CD3 Complex, Antigen, T-Cell Software immunology|