M. A. Kelly

Replication of 13 genome-wide association (GWA)-validated risk variants for type 2 diabetes in Pakistani populations

Aims/hypothesisRecent genome-wide association (GWA) studies and subsequent replication studies have greatly increased the number of validated type 2 diabetes susceptibility variants, but most of these have been conducted in European populations. Despite the high prevalence of the disease in South Asians, studies investigating GWA-validated type 2 diabetes risk variants in this ethnic group are limited. We investigated 30 single nucleotide polymorphisms (SNPs), predominantly derived from recent GWA studies, to determine if and to what extent these variants affect type 2 diabetes risk in two Punjabi populations, originating predominantly from the District of Mirpur, Pakistan.MethodsThirty SNPs were genotyped in 1,678 participants with type 2 diabetes and 1,584 normoglycaemic control participants from two populations; one resident in the UK and one indigenous to the District of Mirpur.ResultsSNPs in or near PPARG, TCF7L2, FTO, CDKN2A/2B, HHEX/IDE, IGF2BP2, SLC30A8, KCNQ1, JAZF1, IRS1, KLF14, CHCHD9 and DUSP9 displayed significant (p < 0.05) associations with type 2 diabetes, with similar effect sizes to those seen in European populations. A constructed genetic risk score was associated with type 2 diabetes (p = 5.46 × 10−12), BMI (p = 2.25 × 10−4) and age at onset of diabetes (p = 0.002).Conclusions/interpretationWe have demonstrated that 13 variants confer an increased risk of type 2 diabetes in our Pakistani populations; to our knowledge this is the first time that SNPs in or near KCNQ1, JAZF1, IRS1, KLF14, CHCHD9 and DUSP9 have been significantly associated with the disease in South Asians. Large-scale studies and meta-analyses of South Asian populations are needed to further confirm the effect of these variants in this ethnic group.

An FTO variant is associated with Type 2 diabetes in South Asian populations after accounting for body mass index and waist circumference

Diabet. Med. 28, 673–680 (2011)

Both DQA1 and DQB1 genes are implicated in HLA-associated protection from Type 1 (insulin-dependent) diabetes mellitus in a British Caucasian population

SummaryInherited susceptibility to Type 1 (insulin-dependent) diabetes mellitus is partly determined by HLA genes. It has been suggested that protection from disease may be conferred by HLA-DQB1 genes which encode molecules with aspartate at position 57. We investigated the contributions of HLA-DRB1, DQA1 and DQB1 genes to protection from disease. Restriction fragment length polymorphism and sequence specific oligonucleotide analysis in 156 British Caucasian Type 1 diabetic and 116 control subjects showed protection from disease was associated with DR2, DRw6 and DR7 haplotypes. The most protective DQA1 allele was DQA1*0102 which occurred on both DR2 and DRw6 haplotypes. The DQB1 alleles DQB1*0303, DQB1*0602 and DQB1*0603 were associated with protection, as was DQB1*0604, which encodes an Asp-57 negative DQβmolecule. Heterozygosity for both protective and predisposing HLA markers was reduced in diabetic compared with control subjects. We conclude that both DQA1 and DQB1 genes are implicated in HLA-associated protection from Type 1 diabetes in this British Caucasian population. The overall structure of the DQ heterodimer is critical and DQβ-Asp 57 is of secondary importance in determining protection from disease. The effect of protective HLA types may predominate over that of predisposing markers.

Genetic and immunological characteristics of Type I diabetes mellitus in an Indo-Aryan population

Aims/hypothesis. Our aim was to characterise the genetic and immunological features associated with Type I (insulin-dependent) diabetes mellitus in a cohort of Indo-Aryan children resident in the United Kingdom.¶Methods. Children with Type I diabetes (n = 53), unaffected first-degree relatives (n = 146) and unrelated healthy control children (n = 54) were typed for alleles of the HLA-DRB1, HLA-DQA1 and HLA-DQB1 genes. Islet cell antibodies and antibodies to glutamic acid decarboxylase, protein tyrosine phosphatase-2 (IA-2ic) and insulin were measured in the diabetic and control children.¶Results. The DRB1*03.DQA1*05.DQB1*02 haplotype was positively associated with the disease, occurring in 78 % of diabetic children compared with 22.6 % of healthy children (pc < 2.4 × 10–5). In simplex families, this haplotype was transmitted more frequently to the diabetic children than to their unaffected siblings (p < 1 × 10–4). The DRB1*04.DQA1* 03.DQB1*0302 haplotype was also transmitted preferentially to the diabetic probands (p < 0.025) but was not associated with disease in the case control study. Islet-related autoantibodies were detected in 89.6 % of diabetic patients compared with 11.8 % of control children (p < 1 × 10–6). Although protein tyrosine phosphatase-2 autoantibodies were detected more frequently among DRB1*04-positive diabetic patients compared with patients lacking this allele, the overall frequency of these autoantibodies was lower than observed in Europid diabetic subjects. This could reflect the absence of a disease association with DRB1*04 in the Indo-Aryan cohort.¶Conclusion/interpretation. Type I diabetes in our Indo-Aryan cohort is similar to the disease observed in Anglo-Europeans but has important immunogenetic differences. The low frequency of protein tyrosine phosphatase-2 autoantibodies among the Indo-Aryan diabetic children could have important implications for the design of future strategies for disease prediction in this population. [Diabetologia (2000) 43: 450–456]

Differential expression of HLA-DQ alleles in peripheral blood mononuclear cells: alleles associated with susceptibility to and protection from autoimmune type 1 diabetes

Differential expression of human leucocyte antigen (HLA) class II genes has been postulated to influence the risk of developing autoimmune disease. In this study, we investigated the relationship between the level of mRNA expression of DQA1 and DQB1 alleles in peripheral blood mononuclear cells and the influence of the alleles on susceptibility to type 1 diabetes (T1D). Transcripts from pairs of DQA1 and DQB1 alleles were quantified in 59 DQ‐heterozygous individuals (29 patients with T1D and 30 healthy control subjects). Luciferase reporter gene assays were used to investigate the relative promoter activities of the alleles associated with high and low risk of disease. DQA1*0301 and the DQB1*06 group of alleles (*0601, *0602, *0603 and *0604) were generally overexpressed in comparison to other alleles. In contrast, mRNA for DQB1*0201/*0202 was generally less abundant than other DQB1 transcripts. These data correlated well with the relative promoter activities observed for the diabetes‐associated alleles; the strongest promoters were those derived from DQA1*0301 and DQB1*0602, while a 700‐bp fragment derived from the DQB1*0201 promoter showed the lowest activity of the DQB1 constructs. There was no simple correlation between the level of expression of specific DQ alleles and their influence on the risk of diabetes. The functional relevance of our findings and their implications for the pathogenesis of autoimmunity remain to be determined.

HLA alleles and IDDM in children in Hungary: a comparison with Finland.

It has been postulated that variation in the distribution of human leukocyte antigen (HLA)-encoded susceptibility alleles for insulin-dependent diabetes mellitus (IDDM) is the genetic basis for variation in the incidence of the disease between populations. The aim of this study was to characterize HLA-encoded susceptibility to IDDM in Hungary and to identify whether HLA-DRB1/DQ-encoded susceptibility could account for the five times lower incidence of disease in Hungary compared with Finland. The haplotypes DRB1*03-DQA1*05-DQB1*02 (DRB1*03-DQ2) and DRB1*04-DQA1*0301-DQB1*0302 (DRB1*04-DQ8) were significantly associated with disease in both populations. Three genotypes incorporating either or both of these haplotypes accounted for over 70% of the diabetic population in both races. The combined background frequency and the degree of risk as measured by odds ratios of these HLA-DRB1-DQ genotypes were not significantly different in the two countries. Comparison of the DRB1*0401-DQ8 haplotype between the two races suggested a role for HLA-B alleles in susceptibility. These data indicate that the susceptibility associated with high risk DRB1-DQ genotypes alone is insufficient to account for the fivefold variation in incidence of IDDM between Hungary and Finland. Other genetic and/or environmental influences must be involved.

HLA typing and immunological characterization of young‐onset diabetes mellitus in a Hong Kong Chinese population

SUMMARY

Long‐Term Expression of an HLA‐DQ Molecule in the EBV‐Transformed Bare Lymphocyte Cell Line, BLS‐1, using a Plasmid Vector

The HLA class II molecule, DQ6, confers strong natural protection against the development of type 1 diabetes. The mechanism of disease protection is unknown, but is likely to be related to the function of the molecule in antigen presentation. In order to investigate this function, we have created an in vitro model which expresses DQ6 in isolation by introducing the relevant DQ alleles into an Epstein–Barr virus (EBV)‐transformed, human leucocyte antigen (HLA) class II‐deficient B cell line, bare lymphocyte syndrome (BLS)‐1. A recent report suggested that the expression of transferred genes in human EBV‐transformed B cells might be limited in duration. We present a plasmid‐based transfection method that allows long‐term stable expression of the DQ molecule. The DQA1*0102 and DQB1*0602 alleles were cloned into the pCIneo expression vector and the constructs were introduced into BLS‐1 by electroporation. Stable transfectants were selected using magnetic sorting and cloned by limiting dilution. Two clones were shown to express functionally active DQ6 molecules even after 14 months of continuous culture. These clones will be used in functional studies to investigate the antigen binding and T‐cell activation properties of the DQ6 molecule.

The Bgl II Rflp Associated with Type 1 Diabetes in Dr3-Positive Subjects is Not Due to a Dqa1 Promoter Region Polymorphism

Type 1 (insulin-dependent) diabetes is strongly associated with the HLA genes encoding DR3 and DR4 and their associated DQ alleles. While 70% of all Caucasian diabetic patients carry the DR3-associated allele DQA1*0501, this allele also occurs in up to 40% of the healthy population. A DQA1 Bgl II 7.2 kb RFLP has been shown to identify a disease-associated subset of DR3-positive subjects. We examined the frequency of this RFLP pattern in 43 diabetic and 25 control DR3-positive subjects and found it to be present in 27 (65%) and 5 (20%) respectively (p = 0.0012). The promoter of the DR3-associated DQA1*0501 allele was amplified in four diabetic subjects who were positive, and four control subjects who were negative, for the 7.2 kb band. The promoter was digested with Bgl II to determine whether polymorphism within the promoter created a disease-associated Bgl II restriction site, which might influence disease susceptibility by an effect on gene transcription. No amplified promoter fragment contained a Bgl II restriction site, suggesting that the disease-associated 7.2 kb band does not result from DQA1 promoter region polymorphism but may be due to polymorphism elsewhere on the DR3 haplotype.