Glutamic acid decarboxylase 65 and islet cell antigen 512/IA-2 autoantibodies in relation to human leukocyte antigen class II DR and DQ alleles and haplotypes in type 1 diabetes mellitus.

The frequencies of autoantibodies against glutamic acid decarboxylase 65 (GAD65) and islet cell antigen (ICA) 512/IA-2 (512/IA-2) are functions of the specific human leukocyte antigen (HLA) in type 1 diabetes mellitus (T1D). We investigated the association of HLA class II (DR and DQ) alleles and haplotypes with the presence of GAD and IA-2 autoantibodies in T1D. Autoantibodies were tested in 88 Tunisian T1D patients and 112 age- and gender-matched normoglycemic control subjects by enzyme immunoassay. Among T1D patients, mean anti-GAD antibody titers were higher in the DRB1*030101 allele (P < 0.001), together with the DRB1*030101/DQB1*0201 (P < 0.001) and DRB1*040101/DQB1*0302 (P = 0.002) haplotypes, while lower anti-GAD titers were associated with the DRB1*070101 (P = 0.001) and DRB1*110101 (P < 0.001) alleles and DRB1*070101/DQB1*0201 (P = 0.001) and DRB1*110101/DQB1*030101 (P = 0.001) haplotypes. Mean anti-IA-2 antibody titers were higher in the DRB1*040101 allele (P = 0.007) and DRB1*040101/DQB1*0302 (P = 0.001) haplotypes but were lower in the DRB1*110101 allele (P = 0.010) and the DRB1*110101 (P < 0.001) and DRB1*110101/DQB1*030101 (P = 0.025) haplotypes. Multinomial regression analysis confirmed the positive association of DRB1*030101 and the negative association of DRB1*110101 and DQB1*030101, along with the DRB1*070101/DQB1*0201 and DRB1*110101/DQB1*030101 haplotypes, with anti-GAD levels. In contrast, only the DRB1*040101/DQB1*0302 haplotype was positively associated with altered anti-IA-2 titers. Increased GAD65 and IA-2 antibody positivity is differentially associated with select HLA class II alleles and haplotypes, confirming the heterogeneous nature of T1D.

Identification of specific tumor necrosis factor-α-susceptible and -protective haplotypes associated with the risk of type 1 diabetes.

AIM: We investigated the association of tumor necrosis factor (TNF)α gene polymorphism with type 1 diabetes (T1D). METHODS: TNF-α -1031T/C, -863C/A, -857C/T, -376G/A, -308G/A, -238G/A, and +488G/A single nucleotide polymorphisms (SNPs) were assessed in 198 T1DM patients and 180 age-and gender-matched, normoglycemic control subjects using PCR-restriction fragment length polymorphism (RFLP). RESULTS: Higher frequencies of -863A (p = 8.0 × 10-6), -857T (p = 1.4 × 10-4), and -238A (p = 0.002) alleles were seen in T1D patients than in the control group. Significant differences were noted in the distribution of -863T/C, -857C/T, -376G/A, -308G/A, and -238G/A genotypes between patients and controls. Haploview analysis revealed high linkage disequilibrium (LD) between the -376G/A and -308G/A SNPs, but this was lower between the other polymorphisms. Five-locus TNFα haplotypes were constructed based on the prevalence of individual SNPs and the LD between them. An increased frequency of CTGGG, CCGAG, and ACGGG haplotypes, and a reduced frequency of the CCGGG haplotype was seen in patients. When the Bonferroni correction was applied, differences were significant for the CTGGG (Pc = 1.4 × 10-3), CCGAG (Pc = 0.023), and ACGGG (Pc = 1.2 × 10-3) haplotypes which were greater, and the CCGGG haplotype (Pc = 3.8 × 10-5) which was smaller, among T1D patients, thereby conferring susceptibility to and protection from T1D, respectively. CONCLUSION: These results demonstrate that TNF-α polymorphisms, in particular -863C/A, -857C/T, and -238G/A, are significantly associated with T1D. Additional studies, on other racial groups, are needed to confirm our findings.

Association of single nucleotide polymorphisms in cytotoxic T-lymphocyte antigen 4 and susceptibility to autoimmune type 1 diabetes in Tunisians.

In addition to HLA and insulin genes, the costimulatory molecule CTLA-4 gene is a confirmed type 1 diabetes (T1D) susceptibility gene. Previous studies investigated the association of CTLA-4 genetic variants with the risk of T1D, but with inconclusive findings. Here, we tested the contributions of common CTLA-4 gene variants to T1D susceptibility in Tunisian patients and control subjects. The study subjects comprised 228 T1D patients (47.8% females) and 193 unrelated healthy controls (45.6% females). Genotyping for CTLA-4 CT60A/G (rs3087243), +49A/G (rs231775), and -318C/T (rs5742909) was performed by PCR-restriction fragment length polymorphism (RFLP) analysis. The minor-allele frequencies (MAF) for the three CTLA-4 variants were significantly higher in T1D patients, and significantly higher frequencies of homozygous +49G/G and homozygous CT60G/G genotypes were seen in patients, which was confirmed by univariate regression analysis (taking the homozygous wild type as a reference). Of the eight possible three-locus CTLA-4 haplotypes (+49A/G, -318C/T, and CT60A/G) identified, multivariate regression analysis confirmed the positive association of ACG (odds ratio [OR], 1.93; 95% confidence interval [CI], 1.26 to 2.94), GCG (OR, 2.40; 95% CI, 1.11 to 5.21), and GTA (OR, 4.67; 95% CI, 1.52 to 14.39) haplotypes with T1D, after confounding variables were adjusted for. Our results indicate that CTLA-4 gene variants are associated with increased T1D susceptibility in Tunisian patients, further supporting a central role for altered T-cell costimulation in T1D pathogenesis.

Autoimmune type 1 diabetes genetic susceptibility encoded by human leukocyte antigen DRB1 and DQB1 genes in Tunisia.

Human leukocyte antigen (HLA) class II genes contribute to the genetic susceptibility to type 1 diabetes (T1D), and susceptible alleles and haplotypes were implicated in the pathogenesis of T1D. This study investigated the heterogeneity in HLA class II haplotype distribution among Tunisian patients with T1D. This was a retrospective case control study done in Monastir in central Tunisia. The subjects comprised 88 T1D patients and 112 healthy controls. HLA-DRB1 and -DQB1 genotyping was done by PCR-sequence-specific priming. Significant DRB1 and DQB1 allelic differences were seen between T1D patients and controls; these differences comprised DRB1*030101 and DQB1*0302, which were higher in T1D patients than in control subjects, and DRB1*070101, DRB1*110101, DQB1*030101, and DQB1*060101, which were lower in T1D patients than in control subjects. In addition, the frequencies of DRB1*030101-DQB1*0201 and DRB1*040101-DQB1*0302 were higher in T1D patients than in control subjects, and the frequencies of DRB1*070101-DQB1*0201 and DRB1*110101-DQB1*030101 haplotypes were lower in T1D patients than in control subjects. Multiple logistic regression analysis revealed the positive association of DRB1*030101-DQB1*0201 and DRB1*040101-DQB1*0302 and the negative association of only DRB1*070101-DQB1*0201 haplotypes with T1D. Furthermore, a significantly increased prevalence of DRB1*030101-DQB1*0201 homozygotes was seen for T1D subjects than for control subjects. Our results confirm the association of specific HLA-DR and -DQ alleles and haplotypes with T1D in Tunisians. The identification of similar and unique haplotypes in Tunisians compared to other Caucasians highlights the need for evaluating the contribution of HLA class II to the genetic susceptibility to T1D with regard to haplotype usage and also to ethnic origin and racial background.