Heritability of thyroid peroxidase autoantibody levels in type 1 diabetes: evidence from discordant twin pairs.

AIMS/HYPOTHESIS: The discordance status of (autoimmune) type 1 diabetes within monozygotic twin pairs points to the importance of environmental factors. The aim of this study was to investigate whether the environmental events causing type 1 diabetes influence thyroid autoimmunity. METHODS: Monozygotic and dizygotic twins discordant for type 1 diabetes from the UK and USA were tested for thyroid peroxidase autoantibodies (TPOA) by radioimmunoassay. Using quantitative genetic model fitting of a liability-threshold model we estimated the contribution of genetic (heritability) and environmental factors to TPOA. RESULTS: TPOA positivity was higher in females than in males in both cohorts and was associated with later age at diagnosis in the UK and combined cohorts (p < 0.01). TPOA did not specifically segregate with type 1 diabetes in the twin pairs (p > 0.2 in all groups). The best-fitting models showed heritability (95% CI) estimates for TPOA of 63% (37%, 80%) for the UK and 80% (51%, 92%) for US twins, while the best-fitting meta-analysis model of the two twin cohorts combined included additive genetic and unique environmental factors with a heritability estimate of 69% (50%, 82%). CONCLUSIONS/INTERPRETATION: Risk of thyroid autoimmunity, defined by TPOA, in the context of autoimmune diabetes is, substantially, genetically determined in discordant twin pairs. Environmental factors leading to type 1 diabetes were not the same as those involved with thyroid autoimmunity. It follows that it is as important to investigate for thyroid autoimmunity in relatives of type 1 diabetes patients as it is in the patients themselves.

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 ß-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.

Challenges in developing endpoints for type 1 diabetes intervention studies.

Development of efficient and safe intervention strategies for preserving and/or restoring endogenous insulin production in type 1 diabetes has encountered a wide range of challenges, including lack of standardized trial protocols and of consensus on appropriate efficacy endpoints. For the greatest part, difficulties resided in choosing the most suitable assay(s) and parameter(s) to assess the beta-cell function. It is now an accepted approach to evaluate endogenous insulin secretion by measuring C-peptide levels (with highly sensitive and normalized measurement methods) in response to a physiologic stimulus (liquid mixed-meal) under standardized conditions. Preventive interventions mandate the identification of well-defined, reliable and validated mechanistic or immunological markers of efficacy that would correlate with (and predict) the clinical outcome. This has not been consistently achieved to date. However, it has been generally agreed that for preventive studies performed very early in the disease course (in subjects without signs of autoimmunity against beta-cells) development of two or more islet related autoantibodies could be employed as biomarkers of disease and thereafter, diagnostic criteria of diabetes serve as suitable endpoints.This report summarizes the conclusions of the D-Cure workshop of international experts held in Barcelona in April 2007 and the current recommendations and updates in the field.

Altered monocyte cyclooxygenase response to lipopolysaccharide in type 1 diabetes.

Type 1 diabetes is caused by adaptive immune responses, but innate immunity is important because monocytes infiltrate islets. Activated monocytes express cyclooxygenase (COX)-2, promoting prostaglandin-E(2) (PGE(2)) secretion, whereas COX-1 expression is constitutive. We aimed to define monocyte COX expression in type 1 diabetes basally and after lipopolysaccharide (LPS) stimulation. Isolated CD14(+) monocytes were analyzed for COX mRNA and protein expression from identical twins (discordant for type 1 diabetes) and control subjects. Basal monocyte COX mRNA, protein expression, and PGE(2) secretion were normal in type 1 diabetic subjects. After LPS, twins and control subjects showed a COX mRNA isoform switch with decreased COX-1 mRNA (P < 0.01), increased COX-2 mRNA (P < 0.01), and increased COX-2 protein expression (P < 0.01). Compared with control subjects, both diabetic and nondiabetic twins showed greater LPS-induced downregulation of monocyte COX-1 mRNA (P = 0.02), reduced upregulation of COX-2 mRNA and protein (P < 0.03), and greater inhibition by the COX-2 inhibitor di-isopropylfluorophosphate (DFP) of monocyte PGE(2) (P < 0.007). We demonstrate an alteration in monocyte COX mRNA expression as well as monocyte COX-2 and PGE(2) production after LPS in type 1 diabetic patients and their nondiabetic twins. Because COX-2 response to LPS is proinflammatory, an inherited reduced response would predispose to chronic inflammatory diseases such as type 1 diabetes.

Evidence for independent heritability of the glycation gap (glycosylation gap) fraction of HbA1c in nondiabetic twins.

OBJECTIVE: HbA(1c) (A1C) is substantially determined by genetic factors not shared in common with glucose. Fractions of the variance in A1C, the glycation gap (GG; previously called the glycosylation gap) and the hemoglobin glycosylation index, correlate with diabetes complications. We therefore tested whether GG (measured A1C - A1C predicted from glycated serum proteins [GSPs]) was genetically determined and whether it accounted for the heritability of A1C. RESEARCH DESIGN AND METHODS: We conducted a classic twin study on A1C and GSP collected in 40 and 46 pairs of monozygotic and dizygotic healthy female twins, respectively. The predicted A1C was based on the regression line between A1C and GSP in a separate population spanning the pathophysiologic range. RESULTS: GG was more strongly correlated between monozygotic (r = 0.65) than dizygotic (r = 0.48) twins, adjusted for age and BMI. The best-fitting quantitative genetic model adjusted for age and BMI showed that 69% of population variance in GG is heritable, while the remaining 31% is due to unique environmental influences. In contrast, GSP was similarly correlated between monozygotic (r = 0.55) and dizygotic (r = 0.49) twins, hence not genetically determined. GG was strongly correlated to A1C (r = 0.48), attributable mostly to genetic factors. About one-third of the heritability of A1C is shared with GG; the remainder is specific to A1C. CONCLUSIONS: Heritability of the GG accounts for about one-third of the heritability of A1C. By implication, there are gene(s) that preferentially affect erythrocyte lifespan or glucose and/or nonenzymatic glycation or deglycation in the intracellular, rather than extracellular, compartment.

Clinical review: Type 1 diabetes and latent autoimmune diabetes in adults: one end of the rainbow.

CONTEXT: The aim of this review was to explore the pathogenic and clinical spectrum of type 1 diabetes, which includes a form of adult onset autoimmune diabetes usually referred to as latent autoimmune diabetes in adults (LADA). We looked at this entire range of forms of autoimmune diabetes as a spectrum of genetic and nongenetic environmental influences, diabetes-associated immune responses, and metabolic changes. EVIDENCE ACQUISITION: We assessed epidemiological, genetic, immunological, and clinical data from major articles on autoimmune diabetes, including LADA and type 1 diabetes, published since 1992. EVIDENCE SYNTHESIS: Data analysis of autoimmune diabetes indi-cates that type 1 diabetes and LADA occupy different poles of the same spectrum. CONCLUSION: Evidence is presented that LADA represents one end of a rainbow encompassing type 1 diabetes. The clinical nature and management of autoimmune diabetes poses important therapeutic questions regarding conventional therapy for hyperglycemia as well as therapy aiming to protect residual beta-cell function. Limiting loss of endogenous insulin secretion using immunomodulation could be valuable, not only for LADA but also for type 1 diabetes.

No evidence for genetically determined alteration in insulin secretion or sensitivity predisposing to type 1 diabetes: a study of identical twins.

OBJECTIVE: To determine whether inherited changes in insulin secretion or sensitivity could predispose to type 1 diabetes, we studied identical twins of type 1 diabetic patients. RESEARCH DESIGN AND METHODS: We studied prospectively a consecutive series of 27 identical twins of patients with type 1 diabetes who were initially nondiabetic, as well as 14 control subjects, over a period of 18 years. Of these 27 twins, 15 remain nondiabetic (now estimated at low disease risk) and 12 developed diabetes (pre-diabetic twins). Subjects were tested when not diabetic on at least two occasions with an intravenous glucose tolerance test (IVGTT), and we estimated insulin secretion as first-phase insulin response (FPIR), glucose clearance (K(g)), and insulin sensitivity both by homeostasis model assessment of insulin resistance (HOMA-IR) and relative to insulin response by the basal HOMA-IR-to-FPIR ratio. RESULTS: Twins now at low risk and control subjects had similar fasting blood glucose and insulin levels, FPIR, K(g), HOMA-IR, and HOMA-IR-to-FPIR ratio. In contrast, pre-diabetic twins compared with control twins had higher fasting insulin levels (10.3 +/- 6.0 vs. 4.6 +/- 4.0 mIU/ml), lower FPIR (245 +/- 129 vs. 796 +/- 622 mIU . ml(-1) . 10 min(-1)), lower K(g) (1.5 +/- 0.6 vs. 2.6 +/- 0.8% per min), and higher HOMA-IR-to-FPIR ratio (0.007 +/- 0.005 vs. 0.001 +/- 0.0009) (all P < 0.01). CONCLUSIONS: These observations in low-risk nondiabetic identical twins failed to identify a familial alteration in either insulin secretion or sensitivity predisposing to type 1 diabetes.

Level of an advanced glycated end product is genetically determined: a study of normal twins.

Reducing sugars react with amino groups in proteins, lipids, and nucleic acids to produce advanced glycation end products (AGEs), including N(epsilon)-carboxymethyl lysine (CML), which have been implicated in oxidative stress and vascular damage. The aim of this study was to determine whether genetic factors influence serum CML levels in normal subjects. We performed a classical twin study of CML in healthy nondiabetic female twins, 39 monozygotic and 45 dizygotic pairs, aged 21-74 years. Serum CML levels were estimated by enzyme-linked immunosorbent assay. Twin correlations (r) for serum CML levels were higher in monozygotic (r = 0.71) compared with dizygotic (r = 0.50) twin pairs, suggesting a substantial genetic effect and confirmed by quantitative genetic model fitting. Additive genetic effects (heritability) explained 74% (95% CI 58-84) of population variance in CML. Heritability (%) of fasting glucose (51%) and HbA(1c) (62%) could not explain CML heritability, which was not associated with them. CML levels are, therefore, predominantly genetically determined and independent of genes influencing fasting glucose or HbA(1c). Thus familial, largely genetic factors influence AGE implicating these glycoxidation products in the genetic contribution to macro- and microvascular disease.

Clinical evidence for the safety of GAD65 immunomodulation in adult-onset autoimmune diabetes.

The purpose of this Phase II study was to evaluate if alum-formulated human recombinant GAD65 is safe and does not compromise beta cell function. The study was conducted as a randomized, double blind, placebo-controlled, dose-escalation clinical trial in a total of 47 Latent Autoimmune Diabetes in Adults (LADA) patients who received either placebo or 4, 20, 100, or 500 microg Diamyd subcutaneously at Weeks 1 and 4. Safety evaluations, including neurology, beta cell function tests, diabetes status assessment, hematology, biochemistry, and cellular and humoral immunological markers, were repeatedly assessed over 24 weeks. None of the patients had significant study-related adverse events (AE). Fasting c-peptide levels at 24 weeks were increased compared with placebo (P=.0015) in the 20 microg but not in the other dose groups. In addition, both fasting (P=.0081) and stimulated (P=.0236) c-peptide levels increased from baseline to 24 weeks in the 20 microg dose group. GADA log levels clearly increased (P=.0002) in response to 500 microg Diamyd. The (CD4+)(CD25+)/(CD4+)(CD25-) cell ratio increased (P=.0128) at 24 weeks in the 20 microg group. No sudden increase in HbA1c or plasma glucose or decrease in beta cell function was observed in any of the dose groups. These positive findings for clinical safety further support the clinical development of Diamyd as a therapeutic to prevent autoimmune diabetes.

Humoral and cellular autoimmune responses in stiff person syndrome.

Stiff person syndrome (SPS) is a chronic autoimmune disease associated with humoral and cellular immune responses to glutamic acid decarboxylase (GAD) 65. Another chronic autoimmune disease, type 1 diabetes (T1D), is also associated with autoimmune responses to this antigen, but T1D patients develop SPS only extremely rarely and only a third of SPS patients develop T1D (mostly mild manifestations in adulthood). In a previous study, we described important differences between T1D and SPS in the autoimmune response to GAD 65: (1) T cells of SPS patients recognize epitopes in the middle of GAD 65 (amino residues 81-171 and 313-403), whereas patients with T1D preferentially recognize another middle (161-243) and a C-terminal region (473-555); and (2) GAD antibodies (Abs) were nearly exclusively of the Th1-associated IgG1 type in T1D, whereas SPS patients had both Th1- and Th2-associated IgG4 and IgE GAD Abs. These differences were not simply related to different HLA alleles. Fine epitope mapping revealed further distinct T cell epitopes in both diseases despite similar HLA background. Therefore, a single autoantigen can elicit different immune responses causing distinct chronic autoimmune diseases possibly related to a Th1 or Th2 bias of the disease.

Preventing type 1 diabetes mellitus: hype and hope.

Autoimmune diseases affect 10% or more of the UK population. In organ-specific autoimmune diseases a particular tissue is targeted by the aggressive immune response. Type 1 diabetes is due to destruction of insulin-secreting islet cells. Both genetic and environmental factors cause type 1 diabetes and environmental events can operate very early, even in utero. The early induction of diabetes-associated autoantibodies and the long prediabetic period means that autoimmune diabetes in children can be predicted by detecting these autoantibodies. The purpose of prediction is prevention and initial studies suggest that it is possible to modify the disease process. However, the impact of such therapy on the disease is as yet extremely modest.

Biologic variability in plasma glucose, hemoglobin A1c, and advanced glycation end products associated with diabetes complications.

Plasma glucose plays a key role in the complications of diabetes mellitus. Hemoglobin A1c (HbA1c) and circulating concentrations of advanced glycation end products (AGEs) are central to diabetes clinical care and pathophysiology. However, there is evidence for variation between individuals in the relationship of plasma glucose to both these measures and to specific complications. The glycation gap (GG) and hemoglobin glycation index represent tools for quantitating the variability in the relationship between plasma glucose and HbA1c useful for identification of underlying mechanisms. Recent evidence demonstrates the heritability of HbA1c, the GG, and AGEs, yet not of glycated serum proteins. There has been tremendous effort devoted to identifying the heritable basis of types 1 and 2 diabetes; however, studies on the heritable contributors to these mediators of glucose effect on complications are only beginning. New evidence for normal biologic variation in the distribution of glucose into the red blood cell (RBC) intracellular compartment and RBC lifespan in people with and without diabetes represent candidates for heritable mechanisms and contributors to the rise in HbA1c with age. Taken as a whole, genetic and mechanistic evidence suggests new potential targets for complications prevention and improvement in complications risk estimation. These observations could help tilt the risk-benefit balance in glycemic control toward a more beneficial outcome.

Plasmacytoid dendritic cells are proportionally expanded at diagnosis of type 1 diabetes and enhance islet autoantigen presentation to T-cells through immune complex capture.

OBJECTIVE: Immune-mediated destruction of beta-cells resulting in type 1 diabetes involves activation of proinflammatory, islet autoreactive T-cells, a process under the control of dendritic cells of the innate immune system. We tested the hypothesis that type 1 diabetes development is associated with disturbance of blood dendritic cell subsets that could enhance islet-specific autoimmunity. RESEARCH DESIGN AND METHODS: We examined blood dendritic cells (plasmacytoid and myeloid) in 40 patients with recent-onset diabetes (median duration 28 days) and matched control subjects. We also examined the relative ability of different dendritic cell subsets to process and present soluble or immune complexed islet cell autoantigen (the islet tyrosine phosphatase IA-2) to responder CD4 T-cells. RESULTS: The balance of blood dendritic cells was profoundly disturbed at diabetes diagnosis, with a significantly elevated proportion of plasmacytoid and reduction of myeloid cells compared with control subjects. Dendritic cell subset distribution was normal in long-standing disease and in patients with type 2 diabetes. Both dendritic cell subsets processed and presented soluble IA-2 to CD4 T-cells after short-term culture, but only plasmacytoid dendritic cells enhanced (by as much as 100%) autoantigen presentation in the presence of IA-2(+) autoantibody patient serum. CONCLUSIONS: The plasmacytoid subset of dendritic cells is overrepresented in the blood close to diabetes onset and shows a distinctive ability to capture islet autoantigenic immune complexes and enhance autoantigen-driven CD4 T-cell activation. This suggests a synergistic proinflammatory role for plasmacytoid dendritic cells and islet cell autoantibodies in type 1 diabetes.

Identical twins discordant for type 1 diabetes show a different pattern of in vitro CD56+ cell activation.

BACKGROUND: Recent studies in animal models indicate a role for natural killer (NK) cells in the protection against type 1 diabetes. In humans, a reduction of NK cell numbers has been reported in identical twins discordant for type 1 diabetes, irrespective of whether they have the disease. Here we have tested whether the activation and expansion of human NK cells with lipopolysaccharide (LPS) reveals differences between these twins. METHODS: Proportions of CD56(+) NK cells and T-cells and Va24Vb11(+) NK-T cells from diabetic and non-diabetic twins was assessed before and after activation using flow cytometry. NK receptor usage was monitored by PCR and flow cytometry. RESULTS: The profile of the expressed Killer Cell immunoglobulin-like receptor (KIR) repertoire (using mRNA) in freshly isolated NK cells was identical in pairs of identical twins, despite marked variation among individual twins as well as controls. Basal numbers of CD56(+) and CD94(+) (CD3(-) and CD3(+)) cells and Valpha24(+)Vbeta11(+) NK-T cells were similarly strongly correlated between identical twins (p < 0.006 for all correlations). Following LPS stimulation, the pattern of KIR mRNA expression remained unaltered in twins and the proportion of NK cells and Valpha24(+)Vbeta11(+) NK-T cells remained correlated between pairs of twins. However, there was a significant reduction in the proportion of CD56(+) cells and CD94(+) cells (whether defined as CD3(-) or CD3(+)) responding to LPS in the diabetic compared to the non-diabetic twin (p = 0.031 and 0.025, respectively). CONCLUSION: This reduction in NK cell expansion in response to LPS in patients with type 1 diabetes is consistent with a non-genetically determined alteration in the innate immune response either predisposing to or resulting from the disease.