Minh N. Pham

Pro- and anti-inflammatory cytokines in latent autoimmune diabetes in adults, type 1 and type 2 diabetes patients: Action LADA 4.

Aims/hypothesisSystemic pro- and anti-inflammatory cytokines are associated with both type 1 and type 2 diabetes, while their role in latent autoimmune diabetes in adults (LADA) is unclear. Therefore, we compared cytokine concentrations in patients with LADA, type 1 or type 2 diabetes and healthy individuals to test the hypothesis that differences of cytokine concentrations between all groups are attributable to diabetes type and BMI.MethodsThe pro-inflammatory cytokines IL-6 and TNF-α, and the anti-inflammatory cytokines IL-1 receptor antagonist (IL-1RA) and IL-10 were measured in 90 participants with type 1 diabetes, 61 with LADA, 465 with type 2 diabetes and 41 control participants using multiple regression models adjusted for BMI, sex, age, blood pressure and diabetes duration.ResultsPatients with type 2 diabetes had higher concentrations of systemic IL-1RA, IL-6 and TNF-α cytokines than patients with either LADA or type 1 diabetes (p < 0.0001 for all differences). Cytokine concentrations in controls were lower than those in all diabetes types (p < 0.04). Increased BMI was positively associated with higher systemic cytokine concentrations in all diabetes types (p < 0.0001). Despite the association of cytokines with anthropometric data, differences between diabetes forms persisted also after adjusting analysis for the confounders BMI, age, sex, disease duration and blood pressure (p < 0.04).Conclusions/interpretationAlthough body mass associates positively with pro- and anti-inflammatory cytokine levels, patients with type 2 diabetes have higher cytokine levels independent of the prevailing BMI. LADA and type 1 diabetes could not be distinguished by systemic cytokines.

Increased serum concentrations of adhesion molecules but not of chemokines in patients with Type 2 diabetes compared with patients with Type 1 diabetes and latent autoimmune diabetes in adult age: Action LADA 5

Diabet. Med. 29, 470–478 (2012)

Serum adipokines as biomarkers of beta-cell function in patients with type 1 diabetes: positive association with leptin and resistin and negative association with adiponectin

We investigated the adipokines adiponectin, leptin and resistin as serum biomarkers of beta‐cell function in patients with type 1 diabetes.

Comparison of cryopreservation methods on T-cell responses to islet and control antigens from type 1 diabetic patients and controls

Type 1 diabetes (T1D) is a cell‐mediated autoimmune disease characterized by destruction of the pancreatic islet cells. The use of cryopreserved cells is preferable to the use of freshly isolated cells to monitor clinical trials to decrease assay and laboratory variability.

Immunology of Diabetes Society T‐Cell Workshop: HLA class II tetramer‐directed epitope validation initiative

Islet‐antigen‐specific CD4+ T cells are known to promote auto‐immune destruction in T1D. Measuring T‐cell number and function provides an important biomarker. In response to this need, we evaluated responses to proinsulin and GAD epitopes in a multicentre study.

Antigen-Specific Regulatory T Cells and Low Dose of IL-2 in Treatment of Type 1 Diabetes

Regulatory T cells (Tregs) play an important role in preventing effector T-cell (Teff) targeting of self-antigens that can lead to tissue destruction in autoimmune settings, including type 1 diabetes (T1D). Autoimmunity is caused in part by an imbalance between Teff and Tregs. Early attempts to treat with immunosuppressive agents have led to serious side effects, thus requiring a more targeted approach. Low-dose IL-2 (LD IL-2) can provide immunoregulation with few side effects by preferentially acting on Tregs to drive tolerance. The concept of LD IL-2 as a therapeutic approach is supported by data in mouse models where autoimmunity is cured and further strengthened by success in human clinical studies in hepatitis C virus-induced vasculitis, chronic graft-versus-host disease, and Alopecia areata. Treatment will require identification of a safe therapeutic window, which is a difficult task given that patients are reported to have deficient or defective IL-2 production or signaling and have experienced mild activation of NK cells and eosinophils with LD IL-2 therapy. In T1D, an LD IL-2 clinical trial concluded that Tregs can be safely expanded in humans; however, the study was not designed to address efficacy. Antigen-specific therapies have also aimed at regulation of the autoimmune response but have been filled with disappointment despite an extensive list of diverse islet antigens tested in humans. This approach could be enhanced through the addition of LD IL-2 to the antigenic treatment regimen to improve the frequency and function of antigen-specific Tregs, without global immunosuppression. Here, we will discuss the use of LD IL-2 and islet antigen to enhance antigen-specific Tregs in T1D and focus on what is known about their immunological impact, their safety, and potential efficacy, and need for better methods to identify therapeutic effectiveness.

Cellular interferon-γ and interleukin-13 immune reactivity in type 1, type 2 and latent autoimmune diabetes: Action LADA 6

OBJECTIVE Type 1 diabetes and latent autoimmune diabetes in adults (LADA) are thought to result from immune-mediated β-cell destruction. It remains unclear why LADA is clinically less severe compared to type 1 diabetes. This study aimed to compare the pro-inflammatory (interferon-γ, IFN-γ) and anti-inflammatory (interleukin-13, IL-13) T-cell responses in humans with LADA and type 1 diabetes. RESEARCH DESIGN AND METHODS IFN-γ and IL-13 T-cell responses to a panel of 16 (auto)-antigens were tested using an enzyme linked immune-spot technique and peripheral T-cells from 35 patients with type 1 diabetes, 59 patients with type 2 diabetes, 23 LADA patients, and 42 control subjects. RESULTS LADA and type 1 diabetes patients did not display any statistically significant differences in the frequency of IFN-γ or IL-13 responses to auto-antigenic stimuli, positive control or mitogen. Overall very low T cell reactivity to autoantigens was detected in all groups. IL-13 responses but not IFN-γ responses to recall antigen tetanus toxoid were higher in healthy control subjects compared to patients with type 1 or type 2 diabetes or LADA (P<0.05). Diabetes, independent of type, was associated with weaker response to recall antigen tetanus toxoid. CONCLUSIONS LADA patients are indistinguishable from type 1 diabetes patients for cellular IFN-γ and IL-13 responses upon mitogen and recall antigen stimulation. These results extend previous findings showing that systemic cytokine/chemokine and humoral responses in type 1 diabetes and LADA are similar.

Inverse Relationship Between Organ-Specific Autoantibodies and Systemic Immune Mediators in Type 1 Diabetes and Type 2 Diabetes: Action LADA 11.

OBJECTIVE We related organ-specific autoantibodies, including diabetes-associated autoantibodies (DAAs) and non-DAAs to systemic cytokines/chemokines in type 1 and type 2 diabetes. RESEARCH DESIGN AND METHODS From the European Action LADA (latent autoimmune diabetes in adults) cohort, patients with adult-onset type 1 diabetes (n = 80, of whom 50 had LADA and 30 had classic type 1 diabetes) and type 2 diabetes (n = 626) were analyzed for DAAs (GAD antibody [GADA], IA-2 antigen, islet cell antibody, and zinc transporter T8), non-DAAs (transglutaminase, thyroid peroxide autoantibodies, parietal cell antibodies), and 10 immune mediator concentrations (measured by LUMINEX). RESULTS Type 1 diabetes patients (whether having classic type 1 diabetes or LADA), apart from their clinical phenotype, could not be distinguished by either autoantibodies (both DAAs and non-DAAs) or immune mediators. In type 1 diabetes, most immune mediators (9 of 10) were negatively correlated with DAA titers. Type 2 diabetes patients, who by definition were without DAAs, had fewer non-DAAs (P < 0.0005), but had higher levels of proinflammatory immune mediators, especially compared with patients with type 1 diabetes who had high GADA titers (interleukin [IL]-6 [P < 0.001], soluble E-selectin [P < 0.01], and IL-1 receptor antagonist [P = 0.052], for trend). CONCLUSIONS Patients with type 1 diabetes had more DAAs and non-DAAs than did those with type 2 diabetes, whereas the frequency and nature of these autoantibodies was broadly similar in classic type 1 diabetes and LADA. Systemic immune mediator levels, in the main, were negatively correlated with DAA titers, and, for some, were higher in patients with type 2 diabetes, especially when compared with patients who had high GADA titers. Differences in the clinical classification of diabetes are associated with graded differences in adaptive and innate immune reactivity.

Oral insulin (human, murine, or porcine) does not prevent diabetes in the non-obese diabetic mouse.

Studies have shown oral insulin prevents type 1 diabetes (T1D) in mouse models, however human trials were inconclusive. We tested the ability of different insulins to prevent T1D in non-obese diabetic mice. Mice received oral insulin or PBS twice weekly and disease was monitored. Contrary to previous studies, no insulin tested showed significant ability to prevent T1D, nor did testing of linked suppression in a delayed type hypersensitivity model have reproducible effect. To investigate delivery of antigen within the GI tract, blue dye was fed to mice. Dye traveled 5-8 cm from stomach to small intestine within 10s, suggesting orally administered antigen may not get digested in the stomach in mice. Insulin incubated with jejunum extracts was instantly digested. Thus, in humans large doses of insulin may be required to achieve tolerance as antigen may be more vulnerable to digestion in the stomach even before reaching the small intestine.

Metabolically inactive insulin analogue does not prevent autoimmune diabetes in NOD mice

Aims/hypothesisInsulin is widely considered to be a driver antigen in type 1 diabetes in humans and in mouse models of the disease. Therefore, insulin or insulin analogues are candidates for tolerogenic drugs to prevent disease onset in individuals with risk of diabetes. Previous experiments have shown that autoimmune diabetes can be prevented in NOD mice by repeated doses of insulin administered via an oral, nasal or parenteral route, but clinical trials in humans have not succeeded. The hypoglycaemic activity of insulin is dose-limiting in clinical studies attempting tolerance and disease prevention. Here, we aimed to investigate the therapeutic potential of metabolically inactive insulin analogue (MII) in NOD mice.MethodsThe tolerogenic potential of MII to prevent autoimmune diabetes was studied by administering multiple i.v. or s.c. injections of MII to non-diabetic 7–12-week-old female NOD mice in three geographical colony locations. The incidence of diabetes was assessed from daily or weekly blood glucose measurements. The effect of MII on insulin autoantibody levels was studied using an electrochemiluminescence-based insulin autoantibody assay. The effect on the number of insulin-reactive CD8+ and CD4+ T lymphocytes in peripheral lymphoid tissue was studied with MHC class I and MHC class II tetramers, respectively.ResultsWe found that twice-weekly s.c. administration of MII accelerates rather than prevents diabetes. High-dose i.v. treatment did not prevent disease or affect insulin autoantibody levels, but it increased the amount of insulin-reactive CD4+ T lymphocytes in peripheral lymphoid tissue.Conclusions/interpretationOur data suggest that parenteral MII, even when used in high doses, has little or no therapeutic potential in NOD mice and may exacerbate disease.