GM-CSF producing autoreactive CD4+ T cells in type 1 diabetes.

The phenotype of autoreactive T cells in type 1 diabetes is described as Th1, Th17 and/or Th21, but is largely uncharacterized. We combined multi-parameter cytokine profiling and proliferation, and identified GM-CSF producing cells as a component of the response to beta cell autoantigens proinsulin and GAD65. Overall cytokine profiles of CD4+ T cell were not altered in type 1 diabetes. In contrast, patients with recent onset type 1 diabetes had increased frequencies of proinsulin-responsive CD4+CD45RA- T cells producing GM-CSF (p=0.002), IFNγ (p=0.004), IL-17A (p=0.008), IL-21 (p=0.011), and IL-22 (p=0.007), and GAD65-responsive CD4+CD45RA- T cells producing IL-21 (p=0.039). CD4+ T cells with a GM-CSF+IFNγ-IL-17A-IL-21-IL-22- phenotype were increased in patients for responses to both proinsulin (p=0.006) and GAD65 (p=0.037). GM-CSF producing T cells are a novel phenotype in the repertoire of T helper cells in type 1 diabetes and consolidate a Th1/Th17 pro-inflammatory pathogenesis in the disease.

Association of Dendritic Cell Signatures With Autoimmune Inflammation Revealed by Single-Cell Profiling.

OBJECTIVE: To identify single-cell transcriptional signatures of dendritic cells (DCs) that are associated with autoimmunity, and determine whether those DC signatures are correlated with the clinical heterogeneity of autoimmune disease. METHODS: Blood-derived DCs were single-cell sorted from the peripheral blood of patients with rheumatoid arthritis, systemic lupus erythematosus, or type 1 diabetes as well as healthy individuals. DCs were analyzed using single-cell gene expression assays, performed immediately after isolation or after in vitro stimulation of the cells. In addition, protein expression was measured using fluorescence-activated cell sorting. RESULTS: CD1c+ conventional DCs and plasmacytoid DCs from healthy individuals exhibited diverse transcriptional signatures, while the DC transcriptional signatures in patients with autoimmune disease were altered. In particular, distinct DC clusters, characterized by up-regulation of TAP1, IRF7, and IFNAR1, were abundant in patients with systemic autoimmune disease, whereas DCs from patients with type 1 diabetes had decreased expression of the regulatory genes PTPN6, TGFB, and TYROBP. The frequency of CD1c+ conventional DCs that expressed a systemic autoimmune profile directly correlated with the extent of disease activity in patients with rheumatoid arthritis (Spearman's r = 0.60, P = 0.03). CONCLUSION: DC transcriptional signatures are altered in patients with autoimmune disease and are associated with the level of disease activity, suggesting that immune cell transcriptional profiling could improve our ability to detect and understand the heterogeneity of these diseases, and could guide treatment choices in patients with a complex autoimmune disease.

A divergent population of autoantigen-responsive CD4+ T cells in infants prior to ß cell autoimmunity.

Autoimmune diabetes is marked by sensitization to ß cell self-antigens in childhood. We longitudinally followed at-risk children from infancy and performed single-cell gene expression in ß cell antigen-responsive CD4+ T cells through pre- and established autoimmune phases. A striking divergence in the gene signature of ß cell antigen-responsive naïve CD4+ T cells from children who developed ß cell autoimmunity was found in infancy, well before the appearance of ß cell antigen-specific memory T cells or autoantibodies. The signature resembled a pre-T helper 1 (TH1)/TH17/T follicular helper cell response with expression of CCR6, IL21, TBX21, TNF, RORC, EGR2, TGFB1, and ICOS, in the absence of FOXP3, IL17, and other cytokines. The cells transitioned to an IFNG-TH1 memory phenotype with the emergence of autoantibodies. We suggest that the divergent naïve T cell response is a consequence of genetic or environmental priming during unfavorable perinatal exposures and that the signature will guide future efforts to detect and prevent ß cell autoimmunity.

Incomplete immune response to coxsackie B viruses associates with early autoimmunity against insulin.

Viral infections are associated with autoimmunity in type 1 diabetes. Here, we asked whether this association could be explained by variations in host immune response to a putative type 1 etiological factor, namely coxsackie B viruses (CVB). Heterogeneous antibody responses were observed against CVB capsid proteins. Heterogeneity was largely defined by different binding to VP1 or VP2. Antibody responses that were anti-VP2 competent but anti-VP1 deficient were unable to neutralize CVB, and were characteristic of children who developed early insulin-targeting autoimmunity, suggesting an impaired ability to clear CVB in early childhood. In contrast, children who developed a GAD-targeting autoimmunity had robust VP1 and VP2 antibody responses to CVB. We further found that 20% of memory CD4(+) T cells responding to the GAD65247-266 peptide share identical T cell receptors to T cells responding to the CVB4 p2C30-51 peptide, thereby providing direct evidence for the potential of molecular mimicry as a mechanism for GAD autoimmunity. Here, we highlight functional immune response differences between children who develop insulin-targeting and GAD-targeting autoimmunity, and suggest that children who lose B cell tolerance to insulin within the first years of life have a paradoxical impaired ability to mount humoral immune responses to coxsackie viruses.

Activation of islet autoreactive naïve T cells in infants is influenced by homeostatic mechanisms and antigen-presenting capacity.

Islet autoimmunity precedes type 1 diabetes onset. We previously found that islet autoimmunity rarely starts before 6 months of age but reaches its highest incidence already at ∼1 year of age. We now examine whether homeostatic expansion and immune competence changes seen in a maturating immune system may account for this marked variation in islet autoimmunity risk in the first year of life. We found naïve proinsulin- and GAD65-responsive T cells in cord blood (CB) of healthy newborns, with highest responses observed in children with type 1 diabetes-susceptible HLA-DRB1/DQB1 genotypes. Homeostatic expansion characteristics with increased IL-7 concentrations and enhanced T-cell responsiveness to IL-7 were observed throughout the first year of life. However, the ability of antigen-presenting cells to activate naïve T cells was compromised at birth, and CB monocytes had low surface expression of CD40 and HLA class II. In contrast, antigen presentation and expression of these molecules had reached competent adult levels by the high incidence age of 8 months. We propose that temporal changes in islet autoimmunity seroconversion in infants are a consequence of the changing balance between homeostatic drive and antigen presentation competence. These findings are relevant for early prevention of type 1 diabetes.