Ahmed Delli

Type 1 diabetes patients born to immigrants to Sweden increase their native diabetes risk and differ from Swedish patients in HLA types and islet autoantibodies.

Delli AJ, Lindblad B, Carlsson A, Forsander G, Ivarsson S‐A, Ludvigsson J, Marcus C, Lernmark Å; for the Better Diabetes Diagnosis (BDD) Study Group. Type 1 diabetes patients born to immigrants to Sweden increase their native diabetes risk and differ from Swedish patients in HLA types and islet autoantibodies.

Zinc Transporter 8 Autoantibodies and Their Association With SLC30A8 and HLA-DQ Genes Differ Between Immigrant and Swedish Patients With Newly Diagnosed Type 1 Diabetes in the Better Diabetes Diagnosis Study

We examined whether zinc transporter 8 autoantibodies (ZnT8A; arginine ZnT8-RA, tryptophan ZnT8-WA, and glutamine ZnT8-QA variants) differed between immigrant and Swedish patients due to different polymorphisms of SLC30A8, HLA-DQ, or both. Newly diagnosed autoimmune (≥1 islet autoantibody) type 1 diabetic patients (n = 2,964, <18 years, 55% male) were ascertained in the Better Diabetes Diagnosis study. Two subgroups were identified: Swedes (n = 2,160, 73%) and immigrants (non-Swedes; n = 212, 7%). Non-Swedes had less frequent ZnT8-WA (38%) than Swedes (50%), consistent with a lower frequency in the non-Swedes (37%) of SLC30A8 CT+TT (RW+WW) genotypes than in the Swedes (54%). ZnT8-RA (57 and 58%, respectively) did not differ despite a higher frequency of CC (RR) genotypes in non-Swedes (63%) than Swedes (46%). We tested whether this inconsistency was due to HLA-DQ as 2/X (2/2; 2/y; y is anything but 2 or 8), which was a major genotype in non-Swedes (40%) compared with Swedes (14%). In the non-Swedes only, 2/X (2/2; 2/y) was negatively associated with ZnT8-WA and ZnT8-QA but not ZnT8-RA. Molecular simulation showed nonbinding of the relevant ZnT8-R peptide to DQ2, explaining in part a possible lack of tolerance to ZnT8-R. At diagnosis in non-Swedes, the presence of ZnT8-RA rather than ZnT8-WA was likely due to effects of HLA-DQ2 and the SLC30A8 CC (RR) genotypes.

A novel triple mix radiobinding assay for the three ZnT8 (ZnT8-RWQ) autoantibody variants in children with newly diagnosed diabetes.

BACKGROUND AND AIMS Autoantibodies against the zinc transporter 8 (ZnT8A) are common in type 1 diabetes (T1D). ZnT8A analyses are complicated by the fact that there are three variants of the autoantigen at amino acid position 325 representing ZnT8-R (Arginine), ZnT8-W (Tryptophan) and ZnT8-Q (Glutamin). The aims of the study were: 1) to develop an autoantigen triple mix Radio-Binding Assay (RBA) for ZnT8A; 2) to identify the individual ZnT8-R,-W,-QA reactivity and 3) to validate the triple mix ZnT8A RBA in children with newly diagnosed T1D. METHODS Serum samples were obtained from 2664 (56% males, n=1436) patients in the Swedish nationwide Better Diabetes Diagnosis (BDD) study representing patients with T1D (97%, n=2582), T2D (1.7%, n=46), MODY (1.0%, n=28) and secondary diabetes (0.3%, n=8). cDNA coding for the C-terminal end of each variant was prepared by site-directed mutagenesis and subcloned into a high efficiency in vitro transcription translation vector. The ZnT8 variants were labeled with 35S-methionine and used in a standard RBA separating free from autoantibody-bound autoantigen with Protein A-Sepharose. RESULTS ZnT8-TripleA was detected in 1678 (65%) patients with T1D, 4 (9%) T2D, 3 (11%) MODY and in none (0%) of the patients with secondary diabetes. Among the T1D patients ZnT8-RA was detected in 1351 (52%) patients, ZnT8-WA in 1209 (47%) and ZnT8-QA in 790 (31%) demonstrating that 1661 (64%) had one or several ZnT8A. The ZnT8-TripleA assay showed a false positive rate of 1.9% (n=49). Only 1.2% (n=32) of the T1D patients were false negative for ZnT8-TripleA compared to 0/46 (0%) of the T2D patients. The precision (intra assay CV) and reproducibility (inter assay CV) of the ZnT8-TripleA assay did not differ from the RBA of the individual ZnT8 variants. CONCLUSION We conclude that the ZnT8-TripleA assay had low false positive and false negative rates. The ZnT8-TripleA assay would therefore be highly suitable not only to analyze patient with newly diagnosed diabetes but also for screening the general population since this assay demonstrated high sensitivity and very high specificity.

Triple specificity of ZnT8 autoantibodies in relation to HLA and other islet autoantibodies in childhood and adolescent type 1 diabetes.

To establish the diagnostic sensitivity of and the relationships between autoantibodies to all three Zinc transporter 8 (Zinc transporter 8 autoantibody to either one, two, or all three amino acid variants at position 325, ZnT8A) variants to human leukocyte antigen (HLA)‐DQ and to autoantibodies to glutamic acid decarboxylase (GADA), insulinoma‐associated protein 2 (IA‐2A), and insulin (IAA).

Autoimmunity against INS-IGF2 expressed in human pancreatic islets

Background: Islet INS-IGF2 was examined as a possible autoantigen in type 1 diabetes. Results: INS-IGF2 expression was inversely related to donor HbA1c and glucose-stimulated insulin release. Autoantibodies doubly reactive with INS-IGF2 and insulin were more common in patients with type 1 diabetes than controls. Conclusion: INS-IGF2 is recognized by autoantibodies in type 1 diabetes. Significance: Autoantibodies doubly reactive with both INS-IGF2 and insulin may contribute to type 1 diabetes. Insulin is a major autoantigen in islet autoimmunity and progression to type 1 diabetes. It has been suggested that the insulin B-chain may be critical to insulin autoimmunity in type 1 diabetes. INS-IGF2 consists of the preproinsulin signal peptide, the insulin B-chain, and eight amino acids of the C-peptide in addition to 138 amino acids from the IGF2 gene. We aimed to determine the expression of INS-IGF2 in human pancreatic islets and autoantibodies in newly diagnosed children with type 1 diabetes and controls. INS-IGF2, expressed primarily in beta cells, showed higher levels of expression in islets from normal compared with donors with either type 2 diabetes (p = 0.006) or high HbA1c levels (p < 0.001). INS-IGF2 autoantibody levels were increased in newly diagnosed patients with type 1 diabetes (n = 304) compared with healthy controls (n = 355; p < 0.001). Displacement with cold insulin and INS-IGF2 revealed that more patients than controls had doubly reactive insulin-INS-IGF2 autoantibodies. These data suggest that INS-IGF2, which contains the preproinsulin signal peptide, the B-chain, and eight amino acids of the C-peptide may be an autoantigen in type 1 diabetes. INS-IGF2 and insulin may share autoantibody-binding sites, thus complicating the notion that insulin is the primary autoantigen in type 1 diabetes.

Association between autoantibodies to the Arginine variant of the Zinc transporter 8 (ZnT8) and stimulated C-peptide levels in Danish children and adolescents with newly diagnosed type 1 diabetes

The zinc transporter 8 (ZnT8) was recently identified as a common autoantigen in type 1 diabetes (T1D) and inclusion of ZnT8 autoantibodies (ZnT8Ab) was found to increase the diagnostic specificity of T1D.

Thyroid autoimmunity in relation to islet autoantibodies and HLA-DQ genotype in newly diagnosed type 1 diabetes in children and adolescents.

Aims/hypothesisThe aim of this work was to investigate, in children newly diagnosed with type 1 diabetes: (1) the prevalence of autoantibodies against thyroid peroxidase (TPOAb) and thyroglobulin (TGAb); and (2) the association between TPOAb, TGAb or both, with either islet autoantibodies or HLA-DQ genes.MethodsBlood samples from 2,433 children newly diagnosed with type 1 diabetes were analysed for TPOAb and TGAb in addition to autoantibodies against arginine zinc transporter 8 (ZnT8RA), tryptophan zinc transporter 8 (ZnT8WA), glutamine zinc transporter 8 (ZnT8QA), glutamic acid decarboxylase (GADA), insulin (IAA), insulinoma-associated protein-2 (IA-2A), HLA-DQA-B1 genotypes, thyroid-stimulating hormone (TSH) and free thyroxine (T4).ResultsAt type 1 diabetes diagnosis, 12% of the children had thyroid autoantibodies (60% were girls; p < 0.0001). GADA was positively associated with TPOAb (p < 0.001) and with TGAb (p < 0.001). In addition, ZnT8A was associated with both TPOAb (p = 0.039) and TGAb (p = 0.015). DQB1*05:01 in any genotype was negatively associated with TPOAb (OR 0.55, 95% CI 0.37, 0.83, p value corrected for multiple comparisons (pc) = 0.012) and possibly with TGAb (OR 0.55, 95% CI 0.35, 0.87, pc = 0.07). Thyroid autoimmunity in children newly diagnosed with type 1 diabetes was rarely (0.45%) associated with onset of clinical thyroid disease based on TSH and free T4.Conclusions/interpretationGADA and ZnT8A increased the risk for thyroid autoimmunity at the time of clinical diagnosis of type 1 diabetes, while HLA-DQB1*05:01 reduced the risk. However, the associations between thyroid autoimmunity and HLA-DQ genotype were weak and did not fully explain the co-occurrence of islet and thyroid autoimmunity.

Equal access to health care may diminish the differences in outcome between native and immigrant patients with type 1 diabetes.

Previous studies have found that ethnicity influences glycemic control. We hypothesized that differences between Nordic and non‐Nordic patients are less pronounced for children with type 1 diabetes in high incidence countries in Northern Europe.

Antibodies to influenza virus A/H1N1 hemagglutinin in type 1 diabetes children diagnosed before, during and after the Swedish A(H1N1)pdm09 vaccination campaign 2009-2010.

We determined A/H1N1‐hemagglutinin (HA) antibodies in relation to HLA‐DQ genotypes and islet autoantibodies at clinical diagnosis in 1141 incident 0.7‐to 18‐year‐old type 1 diabetes patients diagnosed April 2009–December 2010. Antibodies to 35S‐methionine‐labelled A/H1N1 hemagglutinin were determined in a radiobinding assay in patients diagnosed before (n = 325), during (n = 355) and after (n = 461) the October 2009–March 2010 Swedish A(H1N1)pdm09 vaccination campaign, along with HLA‐DQ genotypes and autoantibodies against GAD, insulin, IA‐2 and ZnT8 transporter. Before vaccination, 0.6% patients had A/H1N1‐HA antibodies compared with 40% during and 27% after vaccination (P < 0.0001). In children <3 years of age, A/H1N1‐HA antibodies were found only during vaccination. The frequency of A/H1N1‐HA antibodies during vaccination decreased after vaccination among the 3 < 6 (P = 0.006) and 13 < 18 (P = 0.001), but not among the 6 < 13‐year‐olds. HLA‐DQ2/8 positive children <3 years decreased from 54% (15/28) before and 68% (19/28) during, to 30% (9/30) after vaccination (P = 0.014). Regardless of age, DQ2/2; 2/X (n = 177) patients had lower frequency (P = 0.020) and levels (P = 0.042) of A/H1N1‐HA antibodies compared with non‐DQ2/2; 2/X (n = 964) patients. GADA frequency was 50% before, 60% during and 51% after vaccination (P = 0.009). ZnT8QA frequency increased from 30% before to 34% during and 41% after vaccination (P = 0.002). Our findings suggest that young (<3 years) along with DQ2/2; 2/X patients were low responders to Pandemrix®. As the proportion of DQ2/8 patients <3 years of age decreased after vaccination and the frequencies of GADA and ZnT8QA were enhanced, it cannot be excluded that the vaccine affected clinical onset of type 1 diabetes.

Neuropeptide Y is a minor autoantigen in newly diagnosed type 1 diabetes patients.

Autoantibodies (A) against Neuropeptide Y (NPY), was reported in 9% newly diagnosed type 1 diabetes (T1D) patients. A single nucleotide polymorphism (SNP) at rs16139 (T1128C) within the NPY‐gene identified an amino acid substitution from leucine (L) to proline (P) (L7P) associated with both glucose tolerance and type 2 diabetes. We aimed to determine: (i) the influence of autoantibodies to leucine neuropeptide Y (NPY‐LA) and autoantibodies to proline neuropeptide Y (NPY‐PA) on the diagnostic sensitivity of type 1 diabetes (T1D), (ii) the association of NPYA with major islet autoantibodies, and (iii) the association of NPYA with HLA‐DQ genotypes in newly diagnosed T1D patients.