Insulin-dependent diabetes mellitus in Santiago, Chile: the role of immunogenetic and environmental factors

The role of HLA class II alleles in the genetic susceptibility to develop insulin-dependent diabetes mellitus (IDDM) was examined by means of PCR and oligospecific probes in 63 IDDM children and 74 controls subjects. In diabetic patients we found a significant increase in the alleles frequency DR3, DR4, DQB1*0302 and DQA1*0301 compared to the control group, where the most prevalent alleles were DR2, DR14 (DRB1*1402), DQA1*0101 and DQA1*0201. All the risk genotypes in the diabetic group were similar than in other caucasian groups: DR3/DR4-DQB1*0201/0302-DQA1*0301/0501 and DR4/DR4-DQB1*0302/0302-DQA1*0301/0301. The homozygote character no asp57 conferred an absolute risk (AR) of 3.87 and the marker Arg52 an AR of 5.78/100.000 bab year. The homozygosis for both markers (no Asp57 + Arg52) had an AR of 7.56/100.000 bab year. Regarding environmental factors associated with IDDM, our population under study showed a low prevalence of infectious agents (mainly mumps and rubella, specifically associated with IDDM) and a high prevalence of effective breast-feeding (over 3 months). These factors could be exercising a protector role in the development of IDDM. The factors that appear to be important in the low incidence of IDDM in Santiago de Chile are: the low prevalence of infectious agents related to IDDM, the high percentage of breast-feeding children in the population, the reduced frequency of susceptible molecules as DR3, DQB1*0201 (compared to other caucasian groups) and the presence of protective genotypes related to DR13 and DR14 observed in the non diabetic children.     

Taking the pith out of reality: a reflexive methodology for psychiatric nursing research

HLA-DQ genes are the main inherited factors predisposing to IDDM. This gene region harbors long terminal repeat (DQ LTR) elements of the human endogenous retrovirus HER V-K, which we analyzed for a possible association with disease. We first investigated whether LTR segregate with DQ alleles in families. Members (n = 110) of 29 families with at least one diabetic child, unrelated patients with IDDM (n = 159), and healthy controls (n = 173) were analyzed. Genomic DNA was amplified for DQ LTR3 by a nested primer approach as well as for DQA1 and DQB1 second exons, to assign DQA1 and DQB1 alleles. DQ LTR segregated in 24 families along with DQ alleles. Of the 29 families, 20 index patients were positive for DQ LTR. The DQ LTR was in all patients on the haplotype carrying the DQA1 *0301 and DQB1 *0302 alleles. A majority of patients had DQ LTR (62%) compared with controls (38%) (p < 1.3 x 10(-5)), even after matching for the high-risk alleles DQA1 *0501, DQB1 *0201-DQA1 *0301, and DQB1 *0302 (79% of patients and 48% of controls; p < 0.02). Subtyping for DRB1 *04 alleles in all DQB1 *0302+ individuals showed 56% DRB1 *0401, DQB1 *0302 [LTR' patients vs. 29% controls with the same haplotype (p < 0.002)]. In conclusion, these data demonstrate the segregation of DQ LTR with DQA1, DQB1 alleles on HLA haplotypes. Furthermore their presence on DRB1 *0401-, DQA1 *0301-, and DQB1 *0302-positive haplotypes suggest that they contribute to DQ-related susceptibility for IDDM.     

On the HLA-DQ(alpha 1*0501, beta 1*0201)-associated susceptibility in celiac disease: a possible gene dosage effect of DQB1*0201

The HLA-associated susceptibility to develop celiac disease (CD) seems mainly to be conferred by a particular HLA-DQ heterodimer encoded by the DQA1*0501 and DQB1*0201 genes either in cis or in trans position. To study the possible influence of DRB1 or other DQA1 and DQB1 alleles on the CD susceptibility conferred by these DQ genes, we performed genomic HLA typing of 94 CD patients, selected those who carried at least one copy of the DRB1*0301-DQA1*0501-DQB1*0201 haplotype (N = 89) and compared them to 47 random, healthy Norwegians matched with the patients to carry at least one copy of the above haplotype. We found an excess of DQB1*0201 homozygosity in the patients. This was due to an increased frequency of the DRB1*0301-DQA1*0501-DQB1*0201 and DRB1*0701-DQA1*0201-DQB1*0201 haplotypes present on the other chromosome. We propose that, in individuals carrying the DQA1*0501 and DQB1*0201 alleles, the presence of a second copy of the DQB1*0201 allele increases susceptibility to CD.     

Linkage disequilibrium between the human leukocyte antigen class II region of the major histocompatibility complex and Graves' disease: replication using a population case control and family-based study

Early case control studies found association of the DRB1 allele, DR3, with Graves' disease (GD). Recent reports, claim the DQA1 allele, DQA1*0501, to be the primary susceptibility determinant within the human leukocyte antigen (HLA) class II region. We typed 228 GD patients, 364 controls, and 98 families (parents, GD, and unaffected sibling) at the DRB1, DQB1, and DQA1 loci. The case control study showed an increased frequency in GD, compared to controls, of DRB1*0304 (47% vs. 24%; pc < 1.4 x 10(-5)), DQB1*02 (58% vs. 46%; pc < 0.035), DQB1*0301/4 (42% vs. 28%; pc < 3.5 x 10(-3)) and DQA1*0501 (67%, vs. 39%; pc < 7 x 10(-6)). The DRB1*0304-DQB1*02-DQA1*0501 haplotype was increased in GD (47%) vs. controls (24%; pc < 1.8 x 10(-5); odds ratio = 2.72). No independent association of these alleles was observed. Preferential transmission of DRB1*0304-DQB1*02-DQA1*0501 from parents heterozygous for the haplotype to GD siblings (72%) was seen in the families (chi2 = 11.95; 1 d.f.; P = 0.0005). Lack of preferential transmission to unaffected siblings (53%; chi2 = 0.19; 1 d.f.; P = NS) excluded segregation distortion. These results show that linkage disequilibrium between GD and the HLA class II region is due to the extended haplotype DRB1*0304-DQB1*02-DQA1*0501.     

Contribution of DRB1*04 variants to predisposition to or protection from insulin dependent diabetes mellitus is independent of dq

Certain DQ alpha/beta dimeric molecules have been shown to play a major role in determining susceptibility or resistance to IDDM. Whether or not predisposition associated with DR4 haplotypes is exclusively due to linkage to DQB1*0302 and DQA1*0301 alleles is still a controversial issue. A modifying effect of certain DRB1*04 subtypes on the susceptibility encoded by DQ alleles is possible, since not all DRB1*04-DQB1*0302 haplotypes are associated with the disease. The distribution of DRB1*04 subtypes was analysed in 240 DR4-positive Caucasian IDDM patients and 110 DR4-positive healthy controls using allele-specific hybridization after genomic amplification. Although an important contribution to IDDM predisposition was encoded by the DQB1*0302 allele which was found in the majority of patients (94.2% vs 64.7% in controls, Odd's ratio OR = 8.8, P < 0.0001), differences between DRB1*04 variants persisted after the effect of the DQB1 locus was removed by matching patients and controls for DQB1*0302. Thus, the DRB1*0402 allele conferred a strong IDDM-predisposing effect (OR = 3.1, P < 0.02) which was highly significant in the absence of DR3 on the second haplotype (OR = 5.6, P < 0.0001) but was not visible among DR3/4 heterozygote individuals. Conversely, the DRB1*0404 allele conferred a strong protective effect (OR = 0.26, P < 0.0001) which was dominant even in the presence of the associated high risk DR3 haplotype (OR = 0.21, P < 0.03). These data indicate that DQ molecules are not the sole contributors to the DR4-associated IDDM predisposition, and that peculiar DR4 subtypes play a significant role in susceptibility to or protection from the disease. DRB1*0402 differs from DRB1*0404 by only two acidic residues at positions 70 and 71 within the peptide binding groove, instead of amide and basic amino acids. This might induce changes of peptide binding specificity that correlate with the genetic linkage of IDDM predisposition.     

Hashimoto's thyroiditis and insulin-dependent diabetes mellitus: differences among individuals with and without abnormal thyroid function

Insulin-dependent diabetes mellitus probands from the Familial Autoimmune and Diabetes Study were evaluated for autoimmune thyroid disease (n = 265). The prevalence of Hashimoto's thyroiditis was 26.6%; 42.0% of these individuals were euthyroid, and 58.0% were hypothyroid. There was a female predominance among hypothyroid and euthyroid Hashimoto's cases compared to those with no thyroid disease (75% vs. 72.4% vs. 41.6%; P < 0.001). Insulin-dependent diabetes mellitus patients with hypothyroid Hashimoto's thyroiditis were more likely to report another autoimmune disease compared to euthyroid Hashimoto's patients or individuals with no thyroid disease (30.8% vs. 17.2% vs. 13.9%; P < 0.01). Sex-specific analysis revealed that this difference was significant for men but not for women. Both euthyroid and hypothyroid Hashimoto's cases were more likely to have a family history of the disease (66.7% vs. 69.2% vs. 47.7%; P < 0.05). No differences were observed in the prevalence of DQA1*0501-DQB1*0201 or DQA1*0301-DQB1*0302 across the three groups. Body mass index, lipid levels, glycemic control, and diabetes complications were also similar. However, euthyroid Hashimoto's women were more likely to report spontaneous abortions than those with hypothyroid Hashimoto's thyroiditis or no thyroid disease (23.8% vs. 61.5% vs. 29.1%; P < 0.05). These data suggest that gender-specific risk factors may be primary determinants of Hashimoto's thyroiditis and other autoimmune diseases among women. However, disease-specific determinants may also increase susceptibility to other autoimmune diseases.     

Characteristics of the innervation of the head and neck

The association of HLA-DRB1 and DQB1 genes with IDDM in Koreans was assessed using 115 IDDM patients and 140 nondiabetic controls. DQB1*0201 is the only DQB1 allele positively associated with IDDM while DQB*0602, *0601 and *0301 are negatively associated. Three DRB1 alleles (DRB1*0301, DRB1*0407 and DRB1*0901) are positively associated while four DR allele groups (DRB1*15, DRB1*12, DRB1*10 and DRB1*14) are negatively associated. However, Haplotype analyses indicated that DQB1*0302, DRB1*0405 and DRB1*0401 may confer susceptibility because the DRB1*0405-DQB*0302 and DRB1*0401-DQB1*0302 haplotypes are positively associated with the disease. The lack of association in Koreans with the DQB1*0302 allele, which appears predisposing in studies of non-Orientals, is due to its strong linkage disequilibrium (LD) with the protective DRB1*0403 and *0406 alleles, while the lack of association with DRB1*0405 is because of its strong LD with the protective DQB1*0401 allele. Nine DR/DQ genotypes confer significantly increased risk to IDDM. Seven of the nine genotypes (DR3/4s, DR1/4s, DR4s/13, DR4s/8, DR4s/7, DR9/13 and DR3/9) were also found to be at high risk to IDDM in other populations, while the two others (DR1/9 and DR9/9) are only found in Koreans. Surprisingly, DR4/4 homozygotes are not associated with high risk to IDDM in Koreans. This observation can be explained by the high frequency of protective DR4 subtypes and the protective DQ alleles (0301 and 0401) associated with the susceptible DR4 alleles. Our analyses indicate that the counterbalancing act between susceptible DRB1 and protective DQB1, and vice versa, that has already been observed in Chinese and Japanese, is the major factor responsible for the low incidence of diabetes in Koreans.     

A phase I/II study to evaluate the effect of fractionated hemibody irradiation in the treatment of osseous metastases--RTOG 88-22

Previous studies have indicated that certain alleles of HLA-DR and -DQ genes were strongly associated with susceptibility and resistance to insulin-dependent diabetes mellitus (IDDM), and the role of DQ molecule in IDDM has been suggested. To further clarify the association of DQ alleles with IDDM, we determined the nucleotide sequences of full-length cDNA from 13 DQA1 alleles and 14 DQB1 alleles. The sequencing analysis revealed sequence polymorphisms outside the hypervariable region of DQ genes. We then analyzed the DQA1 and DQB1 polymorphisms along with that of DRB genes in 86 B-lymphoblastoid cell lines (B-LCLs) from various ethnic groups and in healthy unrelated Japanese and Norwegian individuals. The allelic and haplotypic distributions in each population revealed the characteristic haplotypic formation in the HLA class II region. HLA genes in 139 Japanese and 100 Norwegian IDDM patients were analyzed. DQB1*0301 was negatively associated with IDDM in both ethnic groups, irrespective of associated DRB1 and DQA1 alleles. In DQB1*0302 positive populations, which represented a positive association with IDDM in both ethnic groups, DRB1*0401, *0404, *0802 haplotypes increased in the patients, whereas DRB1*0406 haplotype decreased. Considering about the hierarchy in DRB1 alleles with IDDM susceptibility (DRB1*0401>*0404>*0403 in Norwegian and DRB1*0802>*0403>*0406 in Japanese), the genetic predisposition to IDDM is suggested to be defined by the combination of DR-associated susceptibility and DQ-associated susceptibility and by the DQ-associated resistance which is a dominant genetic trait.     

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A 39-year-old woman had relapsing polychondritis and Beh?et's disease, which was described as mouth and genital ulcers with inflamed cartilage syndrome (MAGIC). Serologic human leukocyte antigen analysis showed A24 (9), A31 (19), B56 (22), B62 (15), Cw6, DR4, DR9. Human leukocyte antigen allele analysis revealed DRB1* 0406/0901, DQA1* 0301/0301, DQB1* 0302/0303, DPB1* 0201/0501 through determining the genotype using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Treatment with methotrexate (5 mg/week) and pentoxifylline (300 mg/d) was effective to control oral ulcers, erythema nodosum, and arthritis.     

Identification of naturally processed T cell epitopes from glutamic acid decarboxylase presented in the context of HLA-DR alleles by T lymphocytes of recent onset IDDM patients

Glutamic acid decarboxylase (GAD) has been defined as a major target antigen in insulin-dependent diabetes mellitus (IDDM). To identify the molecular ligands triggering a T cell response to GAD, a panel of human GAD65-specific T lymphocyte lines was generated from peripheral blood of three recent onset IDDM patients. All lines derived from a patient expressing the high-risk-conferring HLA-DR*0301/ *0401 haplotypes recognized a single epitope localized between amino acid positions 270 and 283 of GAD65, a stretch that is located in close proximity to the homology region shared with Coxsackie virus P2-C protein. All lines with this specificity were restricted to the DRA, B1*0401 product of the DR4 haplotype. Analysis of the GAD-specific T cell response in a second patient homozygous for DR4 haplotypes demonstrated that the same DRA, B1*0401 allele selected T cells specific for a different determinant. The T cell response profile in a third patient showed that DR*1501/ *1601-encoding haplotypes could present at least three different epitopes to GAD65-specific T lymphocytes. One of these epitopes was presented by a DR allele associated with the resistance-conferring DRB1*1501 haplotype. GAD-specific T cell lines could not be isolated from HLA class II-matched normal individuals. Our data reveal that (a) the T cell response to GAD65 is quite heterogenous in recent onset IDDM patients; (b) HLA-DR, not DQ, seems to be the principal restriction element used by T cells present at the onset of the disease; and (c) T cells responding to epitopes containing identical sequences to Coxsackie virus P2-C protein were not detected.     

Association of HLA class I and class II alleles with myositis in Japanese patients

OBJECTIVE: To investigate the correlation of HLA class I and class II antigens and alleles with various forms of myositis in Japanese patients. METHODS: Eighty-four Japanese patients with myositis [22 with polymyositis (PM), 46 with dermatomyositis (DM), 16 with myositis overlapping with other collagen vascular diseases] were typed serologically for HLA-A, B, C antigens. HLA-DRB1, DQA1, and DQB1 alleles were determined by polymerase chain reaction dependent DNA typing methods. Fifty-eight Japanese controls were typed serologically while HLA-DRB1, DQA1, and DQB1 allele typing was carried out in 175, 95, and 104 controls, respectively. RESULTS: HLA-B7 was higher in patients than controls [20.2 vs 6.9% in controls: p=0.02, odds ratio (OR)=3.4]. The increase of HLA-B7 was largely dependent on the increase in overlap patients (37.5%; p=0.005, OR=8.1). HLA-A24 and B52 were significantly decreased in PM as compared to DM, while CW3 was significantly increased in PM versus DM. DRB1*08 alleles were significantly increased in patients (36.9 vs 20.5% in controls; p=0.004, OR=2.3), especially in PM and DM. DQA1*0501 and DQB1*0301 were significantly decreased in patients [4.8 vs 13.7% in controls; p=0.04, OR=0.32, and 8.3 vs 20.2% in controls; p=0.02, OR=0.36, respectively]. CONCLUSION: HLA-class I and class II alleles associated with Japanese patients with myositis may be different from those associated with Caucasian patients.     

Differential expression of insulin-dependent diabetes mellitus-associated HLA-DQA1 alleles in vivo

The strong association of HLA-DQ genes with insulin-dependent diabetes mellitus (IDDM) susceptibility is persuasive evidence of their central role in the etiology of this autoimmune disease. Among other possibilities, it has been proposed that an unbalanced expression of IDDM-associated DQA, and/or DQB alleles may lead to alterations in the composition of alpha beta heterodimers and preferential expression of a particular heterodimer on the antigen-presenting cell surface, leading to self-recognition. In this report, we demonstrate the differential expression of DQA1 alleles in vivo, in particular of the two diabetogenic alleles DQA1*0301 and DQA1*0501. Family studies suggest that unequal HLA-DQA1 allele expression in heterozygous individuals is not associated in cis with the HLA-DQA1 gene, but may be affected by trans-acting determinant(s). We also discuss the segregation of this phenotype in IDDM-affected members. Furthermore, we examined historical samples of PBL from an IDDM-affected individual and an HLA-identical unaffected sibling acting in a kidney transplant program as donor and recipient, respectively. This analysis allowed us to establish that unbalanced expression of DQA1*0301 and DQA1*0501 can be induced by microenvironmental conditions. Inducible differential expression of HLA-DQA1 alleles may account for the discordance in the outcome of autoimmune disease in monozygotic twins and HLA-identical siblings.     

Clinical and genetic characteristics of type 2 diabetes with and without GAD antibodies

The aim of the study was 1) to establish the prevalence of GAD antibodies (GADab) in a population-based study of type 2 diabetes in western Finland, 2) to genetically and phenotypically characterize this subgroup, and 3) to provide a definition for latent autoimmune diabetes in adults (LADA). The prevalence of GADab was 9.3% among 1,122 type 2 diabetic patients, 3.6% among 558 impaired glucose tolerance (IGT) subjects, and 4.4% among 383 nondiabetic control subjects. Islet antigen 2 antibodies (IA2ab) or islet cell antibodies were detected in only 0.5% of the GADab- patients. The GADab+ patients had lower fasting C-peptide concentrations (median [interquartile range]: 0.46 [0.45] vs. 0.62 [0.44] nmol/l, P = 0.0002) and lower insulin response to oral glucose compared with GADab- patients. With respect to features of the metabolic syndrome, the GADab+ patients had lower systolic (140 [29.1] vs. 148 [26.0] mmHg, P = 0.009) and diastolic (79.2 [17.6] vs. 81.0 [13.1] mmHg, P = 0.030) blood pressure values, as well as lower triglyceride concentrations (1.40 [1.18] vs. 1.75 [1.25] mmol/l, P = 0.003). GADab+ men had a lower waist-to-hip ratio compared with GADab- patients. Compared with GADab- patients and control subjects, the GADab+ patients had an increased frequency HLA-DQB1*0201/0302 (13 vs. 4%; P = 0.002) and other genotypes containing the *0302 allele (22 vs. 12%; P = 0.010). However, the frequency of these high-risk genotypes was significantly lower in GADab+ type 2 patients than in type 1 diabetes of young or adult onset (0201/0302 or 0302/X: 36 vs. 66 vs. 64%, P < 0.001). The GADab+ type 2 group did not differ from control subjects with respect to genotypes containing the protective DQB1-alleles *0602 or *0603, nor with respect to the type 1 high-risk genotype in the IDDM1 (Hph1 +/+). We conclude that GADab+ patients differ from both GADab- type 2 diabetic patients and type 1 diabetic patients with respect to beta-cell function, features of the metabolic syndrome, and type 1 diabetes susceptibility genes. Further, we propose that LADA be defined as GADab positivity (>5 relative units) in patients older than 35 years at onset of type 2 diabetes.     

Effects of residues of deltamethrin in cattle faeces on the development and survival of three species of dung-breeding insect

We report on the role of HLA-DQA1 and DQB1 alleles in determining susceptibility to insulin-dependent diabetes mellitus (IDDM) in Hong Kong Chinese and investigate whether these alleles affect the age of onset of the disease. We studied 76 unrelated Chinese patients and 250 controls. There was no apparent predisposing effect of non-aspartic acid residues at position 57 of the DQ beta chain (Asp57-) but there was an excess of homozygous genotypes containing arginine at position 52 of the DQ alpha chain (Arg52+). This excess was mainly attributable to the genotype DQA1*0301/DQA1*05011 in early-onset disease. There was a significant excess of heterodimers of DQ alpha and DQ beta carrying Arg52+ and Asp57- in both early-onset and late-onset disease, but the excess in early-onset disease was mainly attributable to a single heterodimer formed by DQA1*05011 and DQB1*0201. Of three DQA1/DQB1 genotypes containing a double dose of Arg52+ and Asp57-, only one had a strong association with both early-onset and late-onset disease. We show that early-onset IDDM and late-onset IDDM in Chinese may be separated on the basis of their associated DQA1 and DQB1 genotypes and we conclude that previously reported associations of IDDM with Arg52+ and Asp57- residues in Chinese are secondary to specific combinations of DQA1 and DQB1 alleles. We also show that DRB1 molecules play a distinct role in determining susceptibility to early-onset IDDM but the greatest effect is exerted by specific DR/DQ genotypic combinations.     

The major histocompatibility complex region marked by HSP70-1 and HSP70-2 variants is associated with clozapine-induced agranulocytosis in two different ethnic groups

Genes of the major histocompatibility complex (MHC) have been associated with susceptibility to drug-induced adverse reactions. We previously found that clozapine-induced agranulocytosis (CA) is associated with the HLA-DRB1*0402, DRB4*0101, DQB1*0302, DQA1*0301 haplotype in Ashkenazi Jewish patients and with the HLA-DRB1*1601, DRB5*02, DQB1*0502, DQA1*0102 haplotype in non-Jewish patients. In the present study, we tested the hypothesis that the variants of the heat-shock protein 70 (HSP-70) encoded by the HSP-70 loci located within the MHC region and known to be involved in apoptosis and regulation of cell proliferation could play an important role in molecular mechanisms of CA. First, we analyzed HSP70-2 polymorphism in risk-associated haplotypes from HLA homozygous cells and normal individuals and confirmed that the HSP70-2 9-kb variant was associated invariably with DR4 (HLA-DRB1*0402, DQB1*0302) and DR2 (HLA-DRB1*01601, DQB1*0502, DQA1*0102 and HLA-DRB1*1501, DQB1*0602) haplotypes, which were the haplotypes found increased in Jewish and non-Jewish patients with CA, respectively. The 9.0-kb variant was also found to be associated with HLA-B44, DRB1*0401 and HLA-B44, DRB1*07 haplotypes. Second, in patients with CA (12 Ashkenazi Jewish and 20 non-Jewish patients), HSP70-1 A and HSP70-2 9.0-kb variants were associated with the MHC haplotypes found by us to be markers of susceptibility to CA. The clozapine-treated control group had an excess number of HSP70-1 C and HSP70-2 8.5-kb variants, consistent with genetic resistance to CA associated with those variants. This finding supports our hypothesis that a dominant gene within the MHC region (marked by HSP70-1 and HSP70-2), but not necessarily HLA, is associated with CA in two different ethnic groups.     

Characterization of self-glutamic acid decarboxylase 65-reactive CD4+ T-cell clones established from Japanese patients with insulin-dependent diabetes mellitus

To investigate autoimmunity to glutamic acid decarboxylase (GAD) 65 in Japanese patients with insulin-dependent diabetes mellitus (IDDM, type I diabetes), we established seven CD4+ T-cell clones, by stimulating peripheral blood mononuclear cells (PBMC) of six IDDM patients, using a mixture of overlapping human GAD65 peptides. No GAD65 autoreactive T-cell clones were evidenced in four healthy controls. Specificities of T-cell clones were as follows: (a) two clones specific to GAD65 p111-131 (residue 111 to 131) + DR53 (DRB4*0103); (b) one clone specific to GAD65 p413-433 + DR1 (DRB1*0101); (c) two clones specific to GAD65 p200-217 + either DR9 (DRB1*0901) or DR8 (DRB1*0802); and (d) two clones specific to GAD65 p368-388 + DP2 (DPA1*01 or 0201-DPB1*0201). Two DR53-restricted and one DR1-restricted T-cell clones, responded to a recombinant human GAD65 protein, and showed cytotoxicity against B lymphoblastoid cell lines pre-pulsed with the peptides. Six T-cell clones exhibited the Th1-like phenotype. Interestingly, two DR53-restricted T-cell clones killed a Fas-deficient B lymphoblastoid cell line, thereby indicating that cytotoxicity was not completely dependent on a Fas-Fas ligand interaction. Thus, the T-cell epitopes were mapped in a limited portion of GAD65 protein, with a tendency to be restricted by disease-associated HLA-DR, but not DQ molecules.     

HLA-DQB1-defined genetic susceptibility, beta cell autoimmunity, and metabolic characteristics in familial and nonfamilial insulin-dependent diabetes mellitus. Childhood Diabetes in Finland (DiMe) Study Group

Familial aggregation of insulin-dependent diabetes mellitus (IDDM) is a common phenomenon, but the reasons behind it are poorly understood. To investigate whether there is heterogeneity between familial and nonfamilial forms of IDDM we compared genetic, immunological, and clinical characteristics of diabetic children with and without an affected first-degree relative in a population-based series of Finnish children with IDDM. The frequencies of HLA-DQB1 genotypes known to be associated with high (DQB1*0302/0201) or moderate (*0302/x) IDDM risk in the Finnish population were increased, while the proportions of DQB1 genotypes associated with low or decreased risk for IDDM were reduced in the 121 familial cases as compared with the 574 nonfamilial cases (32.7 vs. 21.3%, 41.3 vs. 35.9%, 18.3 vs. 31.4%, and 7.7 vs. 11.4%, respectively; P = 0.002). The frequencies and serum concentrations of islet cell antibodies, insulin autoantibodies, and antibodies to the 65-kD isoform of glutamic acid decarboxylase were similar at diagnosis in the familial and nonfamilial cases. The 31 first-affected cases in the multiple case families were younger at diagnosis than the nonfamilial cases (6.9 vs. 8.5 yr; P < 0.05). The 90 second-affected familial cases had less severe metabolic decompensation at diagnosis than either the first-affected familial or nonfamilial cases. In conclusion, familial aggregation of IDDM in Finland is at least partly explained by a higher frequency of IDDM susceptibility genes in families with multiple affected individuals. The lack of differences in autoantibody levels between the familial and nonfamilial cases indicates homogeneity rather than heterogeneity in the pathogenetic process of beta cell destruction.     

Absence of strong HLA-DR/DQ-DP linkage disequilibrium in the British and French Canadian Caucasoid populations

HLA-DR/DQ-DP linkage disequilibrium was investigated in healthy, unrelated British (n = 150) and French Canadian (n = 67) Caucasoid subjects. HLA-DR and DQ typing was performed by Taq I DNA-RFLP analysis, while DPB1 typing was performed by PCR-SSOP. chi 2 and Fisher's exact tests were performed for all 2-locus biallelic comparisons and coefficients of linkage disequilibrium determined. In the British population, only one example of linkage disequilibrium, significant at P = 0.05 (after correction for the number of comparisons made) was seen (DPB1*0101-DRB1*0301[17(1)]). Additional associations, significant at P = 0.05 before correction for the number of comparisons were also seen, including DPB1*0401-DRB1*15, DPB1*1101-DRB1*0701(7(1)), DPB1*1701-DRB1*0701/ 2(7(2)), DPB1*0101-DQA1*0501, DPB1*0401-DQA1*0102, DPB1*0501-DQA1*0102, DPB1*0101-DQB1*0201, DPB1*0401-DQB1*0602/0603 and DPB1*1101-DQB1*0201. With one exception (DPB1*1101-DQB1*0201), none of these associations was seen in the French Canadian group. These results indicate that although more frequent than thought hitherto, HLA class II linkage disequilibrium involving DPB1 alleles is generally weak, and can differ even between different caucasoid populations. This may have implications for HLA and disease studies.     

Islet cell antibodies are less predictive of IDDM among unaffected children in the general population than in sibs of children with diabetes. The Childhood Diabetes in Finland Study Group

OBJECTIVE: To examine the controversial issue of whether islet cell antibodies (ICAs) have a higher predictive value for progression to clinical IDDM in first-degree relatives of patients with diabetes than in the general population. RESEARCH DESIGN AND METHODS: ICAs were analyzed with standard immunofluorescence in two population-based groups: 765 sibs of children with recent-onset diabetes and 1,212 unaffected Finnish children <20 years of age at initial screening. Those positive for ICAs were additionally tested for antibodies to GAD (GADAs) and the protein tyrosine phosphatase-related IA-2 antigen (IA-2As). Subsequently, these subjects were observed for the manifestation of clinical IDDM over the next 7 years. RESULTS: The frequency of both detectable ICAs and ICA levels > or =20 Juvenile Diabetes Foundation units (JDF U) was significantly higher among the sibs than in the general population (7.8 vs. 4.1% and 4.8 vs. 2.0%, respectively; P < 0.001). The prevalence of GADAs (37/60 vs. 3/48; P < 0.001) and IA-2As (31/60 vs. 0/48; P < 0.001) was increased among ICA-positive sibs compared with ICA-positive individuals from the background population. Over the next 7 years, 24 sibs (3.1%) and 3 unrelated children positive for ICAs (0.3%) progressed to clinical diabetes. The positive predictive value of ICAs was thus 6% in the general population and 40% among the sibs (P < 0.001), or 13 and 59%, respectively (P < 0.001), with an antibody cutoff level of 20 JDF U. The positive predictive value was related to the number of positive autoantibodies in sibs, which was 57% in those with three antibodies, 50% in those with two antibodies, and only 6% in those with ICAs alone. CONCLUSIONS: These data show that the frequency of multiple autoantibodies is substantially lower in ICA-positive children representing the general population than in ICA-positive sibs of children with IDDM. As a consequence, the predictive value of ICAs for IDDM is higher in sibs of affected children than in the general population. This finding must be taken into account when planning intervention trials aimed at preventing or delaying the manifestation of clinical diabetes in individuals from the general population who test positive for ICAs.     

MHC class II alleles and haplotypes in patients with pemphigus vulgaris from India

Pemphigus vulgaris (PV) is a blistering disease of the skin and mucous membranes characterized by an autoantibody response against a keratinocyte adhesion molecule, desmoglein 3, causing acantholysis and blister formation. We compared high resolution MHC class II alleles and haplotype frequencies (HLA-DRB, DQA1 and DQB1) in 37 patients with PV to 89 haplotypes of normal relatives from New Delhi and Ahmedabad. We found that PV patients had significantly increased frequencies of DRB1*1404 (P < 0.0001), DQA1*0101 (P = 0.001), and DQB1*0503 (P < 0.0001). These associations were due to the increased frequencies of the haplotype HLA-DRB1*1404, DRB3*0202, DQA1*0101, DQB1*0503 in patients compared to control haplotypes (p < 0.0001). Also, patients from Ahmedabad had a significant increase in HLA-DQB1*0302 (p = 0.03). An identical amino acid sequence (Leu-Leu-Glu-Arg-Arg-Arg-Ala-Glu), in positions 67-74 of the beta domain of DRB alleles is restricted to some DR14 alleles. Therefore, there are three possible explanations for class II allele involvement in autoantibody in PV patients with class II haplotypes marked by HLA-DR14. First, the class II alleles could be markers for an unidentified susceptibility gene in linkage disequilibrium with them. Second, the primary association could be with DQB1*0503 and the association with HLA-DR14 alleles would be the result of linkage disequilibrium. Third, the HLA-DRB1 locus susceptibility could involve a specific amino acid sequence in the third hypervariable region shared by several HLA-DR14 alleles.