Diabetic ketoacidosis (DKA) is the primary cause of death for children with diabetes, especially when complicated by cerebral edema. Central nervous system (CNS) involvement is common, however the mechanism of, and predictors of CNS dysfunction/injury are largely unknown. In this observational pilot study, blood was collected from pediatric DKA patients at three time points (consent, 12 hr and 24 hr after beginning treatment), to test genetic markers, ribonucleic acid expression and plasma biomarkers reflecting inflammation (tumor necrosis factor-alpha [TNF-α], interleukin-6 [IL-6]) and cerebral dysfunction and/or possible injury (S100β, glial fibrillary acidic protein [GFAP]). Thirty patients were enrolled in the study. The average age was 11.3 yr, 73% were new onset diabetes and 53% were female. Forty percent exhibited abnormal mentation (Glasgow Coma Scale <15), consistent with CNS dysfunction. IL-6 and TNF-α were elevated in plasma, suggesting systemic inflammation. GFAP was measurable in 45% of patients and correlated positively with GCS. Only two patients had detectable levels of S100β. In conclusion, children with DKA often present with evidence of acute neurologic dysfunction or injury. We have demonstrated the feasibility of exploring genetic and biochemical markers of potential importance in the pathophysiology of CNS dysfunction and/or possible injury in DKA. We have identified IL-6, TNF-α and GFAP as potentially important markers for further exploration. A larger, follow-up study will help to better understand the extent and type of CNS injury in DKA as well as the mechanism underlying this dysfunction/injury.
Cerebral edema; diabetic ketoacidosis; genetic biomarkers; vanguard; pilot
To identify additional diabetes susceptibility markers in the MHC that could be responsible for the differential diabetogenicity of different HLA-DR3 CEHs.
Research Design and Methods
High resolution SNP genotyping of the MHC was performed in 15 T1D patients and 39 non-diabetic controls, homozygous for DR3-DQ2 and with one copy of the A*30-B*18-MICA*4-F1C30-DRB1*0301-DQB1*0201-DPB1*0202 HLA haplotype. Significantly associated SNPs were replicated in an independent sample of 554 T1D patients and 841 controls without HLA matching. Electrophoretic mobility shift assay was used to demonstrate a functional effect of an associated SNP.
Seven SNPs showed evidence of association in the initial discovery experiment. Upon replication, only rs419434 (upstream HLA-DOA gene) remained significant. A functional variant (rs432375) in complete LD with rs419434 was shown to affect USF-1 binding and could be responsible for the association signal in the region.
We have identified a novel susceptibility locus within the MHC with a modest contribution to T1D (OR= 1.93; CI: 1.52-2-44; p=10−8) that is independent of HLA-DRB1 locus.
Type 1 diabetes; Major Histocompatibility Complex; association study; conserved extended haplotype; HLA-DOA
HLA genotyping was performed in African American type 1 diabetic patients (n = 772) and controls (n = 1,641) in the largest study of African Americans and type 1 diabetes reported to date. Cases were from Children’s Hospital and Research Center Oakland and from existing collections (Type 1 Diabetes Genetics Consortium [T1DGC], Diabetes Control and Complications Trial/Epidemiology of Diabetes Interventions and Complications [DCCT/EDIC], and Genetics of Kidneys in Diabetes [GoKinD]). Controls were from the T1DGC and from newborn bloodspot cards. The diversity of HLA DRB1-DQA1-DQB1 haplotypes and genotypes is far greater than that found in Europeans and European Americans. Association analyses replicated many type 1 diabetes risk effects of European-derived haplotypes but also revealed novel effects for African-derived haplotypes. Notably, the African-specific “DR3” haplotype DRB1*03:02-DQA1*04:01-DQB1*04:02 is protective for type 1 diabetes, in contrast to the common and highly-susceptible DR3 DRB1*03:01-DQA1*05:01-DQB1*02:01. Both DRB1*07:01 and DRB1*13:03 haplotypes are predisposing when they include DQA1*03:01-DQB1*02:01g but are protective with DQA1*02:01-DQB1*02:01g. The heterozygous DR4/DR9 genotype, containing the African-derived “DR9” haplotype DRB1*09:01-DQA1*03:01-DQB1*02:01g, exhibits extremely high risk (odds ratio = 30.88), approaching that for DR3/DR4 in European populations. Disease risk assessment for African Americans differs greatly from risk assessment in European populations. This has profound implications on risk screening programs and underscores the need for high-resolution genotyping of multiple populations for the rational design of screening programs with tests that will fairly represent the population being screened.
This study assessed the ability to distinguish between type 1 diabetes–affected individuals and their unaffected relatives using HLA and single nucleotide polymorphism (SNP) genotypes.
RESEARCH DESIGN AND METHODS
Eight models, ranging from only the high-risk DR3/DR4 genotype to all significantly associated HLA genotypes and two SNPs mapping to the cytotoxic T-cell–associated antigen-4 gene (CTLA4) and insulin (INS) genes, were fitted to high-resolution class I and class II HLA genotyping data for patients from the Type 1 Diabetes Genetics Consortium collection. Pairs of affected individuals and their unaffected siblings were divided into a “discovery” (n = 1,015 pairs) and a “validation” set (n = 318 pairs). The discriminating performance of various combinations of genetic information was estimated using receiver operating characteristic (ROC) curve analysis.
The use of only the presence or absence of the high-risk DR3/DR4 genotype achieved very modest discriminating ability, yielding an area under the curve (AUC) of 0.62 in the discovery set and 0.59 in the validation set. The full model—which included HLA information from the class II loci DPB1, DRB1, and DQB1; selected alleles from HLA class I loci A and B; and SNPs from the CTLA4 and INS genes—increased the AUC to 0.74 in the discovery set and to 0.71 in the validation set. A cost-effective alternative is proposed, using genotype information equivalent to typing four SNPs (DR3, DR4-DQB1*03:02, CTLA-4, and INS), which achieved an AUC of 0.72 in the discovery set and 0.69 in the validation set.
Genotyping data sufficient to tag DR3, DR4-DQB1*03:02, CTLA4, and INS were shown to distinguish between subjects with type 1 diabetes and their unaffected siblings adequately to achieve clinically utility to identify children in multiplex families to be considered for early intervention.
Type 1 diabetes mellitus (T1DM) is a chronic disease characterized by the autoimmune destruction of pancreatic β-cells. This paper describes the case of a 19-year-old male patient who presented with glutamic acid decarboxylase (GAD) antibody positive and diabetic ketoacidosis, which mandated intensive insulin treatment. Once the ketoacidosis was controlled, an oral dose of 100 mg of sitagliptin was administered once a day. Ketoacidosis was managed by insulin and insulin daily requirement began to dwindle after one month, until its complete withdrawal at 8 weeks, when partial remission was reached. The patient has now remained on sitagliptin treatment alone for a year, without requiring insulin. The benefit observed with this medication is possibly associated with its immunological effects. Inhibition of dipeptidyl peptidase 4 in animal models deregulates the Th1 immune response, increases secretion of Th2 cytokines, activates CD4+CD25+FoxP3+ regulatory T-cells, and prevents IL17 production.
The use of insulin-dose-adjusted HbA1c constitutes the best way to define partial remission in T1DM patients.The use of sitagliptin in T1DM patients could help to decrease daily requirement of insulin by delaying β-cell loss and improving endogenous insulin production.The determination of antibodies against insulin, islet cells, and GAD permits differentiation of T1DM patients from those with atypical or ketosis-prone diabetes.
The Type 1 Diabetes Genetics Consortium has collected type 1 diabetic families worldwide for genetic analysis. The major genetic determinants of type 1 diabetes are alleles at the HLA-DRB1 and DQB1 loci, with both susceptible and protective DR-DQ haplotypes present in all human populations. The aim of this study is to estimate the risk conferred by specific DR-DQ haplotypes and genotypes.
RESEARCH DESIGN AND METHODS
Six hundred and seven Caucasian families and 38 Asian families were typed at high resolution for the DRB1, DQA1, and DQB1 loci. The association analysis was performed by comparing the frequency of DR-DQ haplotypes among the chromosomes transmitted to an affected child with the frequency of chromosomes not transmitted to any affected child.
A number of susceptible, neutral, and protective DR-DQ haplotypes have been identified, and a statistically significant hierarchy of type 1 diabetes risk has been established. The most susceptible haplotypes are the DRB1*0301-DQA1*0501-DQB1*0201 (odds ratio [OR] 3.64) and the DRB1*0405-DQA1*0301-DQB1*0302, DRB1*0401-DQA1*0301-DQB*0302, and DRB1*0402-DQA1*0301-DQB1*0302 haplotypes (ORs 11.37, 8.39, and 3.63), followed by the DRB1*0404-DQA1*0301-DQB1*0302 (OR 1.59) and the DRB1*0801-DQB1*0401-DQB1*0402 (OR 1.25) haplotypes. The most protective haplotypes are DRB1*1501-DQA1*0102-DQB1*0602 (OR 0.03), DRB1*1401-DQA1*0101-DQB1*0503 (OR 0.02), and DRB1*0701-DQA1*0201-DQB1*0303 (OR 0.02).
Specific combinations of alleles at the DRB1, DQA1, and DQB1 loci determine the extent of haplotypic risk. The comparison of closely related DR-DQ haplotype pairs with different type 1 diabetes risks allowed identification of specific amino acid positions critical in determining disease susceptibility. These data also indicate that the risk associated with specific HLA haplotypes can be influenced by the genotype context and that the trans-complementing heterodimer encoded by DQA1*0501 and DQB1*0302 confers very high risk.
The primary associations of the HLA class II genes, HLA-DRB1 and HLA-DQB1, and the class I genes, HLA-A and HLA-B, with type 1 diabetes (T1D) are well established. However, the role of polymorphism at the HLA-DRB3, HLA-DRB4, and HLA-DRB5 loci remains unclear. In two separate studies, one of 500 subjects and 500 control subjects and one of 366 DRB1*03:01–positive samples from selected multiplex T1D families, we used Roche 454 sequencing with Conexio Genomics ASSIGN ATF 454 HLA genotyping software analysis to analyze sequence variation at these three HLA-DRB loci. Association analyses were performed on the two HLA-DRB loci haplotypes (DRB1-DRB3, -DRB4, or -DRB5). Three common HLA-DRB3 alleles (*01:01, *02:02, *03:01) were observed. DRB1*03:01 haplotypes carrying DRB3*02:02 conferred a higher T1D risk than did DRB1*03:01 haplotypes carrying DRB3*01:01 in DRB1*03:01/*03:01 homozygotes with two DRB3*01:01 alleles (odds ratio [OR] 3.4 [95% CI 1.46–8.09]), compared with those carrying one or two DRB3*02:02 alleles (OR 25.5 [3.43–189.2]) (P = 0.033). For DRB1*03:01/*04:01 heterozygotes, however, the HLA-DRB3 allele did not significantly modify the T1D risk of the DRB1*03:01 haplotype (OR 7.7 for *02:02; 6.8 for *01:01). These observations were confirmed by sequence analysis of HLA-DRB3 exon 2 in a targeted replication study of 281 informative T1D family members and 86 affected family-based association control (AFBAC) haplotypes. The frequency of DRB3*02:02 was 42.9% in the DRB1*03:01/*03:01 patients and 27.6% in the DRB1*03:01/*04 (P = 0.005) compared with 22.6% in AFBAC DRB1*03:01 chromosomes (P = 0.001). Analysis of T1D-associated alleles at other HLA loci (HLA-A, HLA-B, and HLA-DPB1) on DRB1*03:01 haplotypes suggests that DRB3*02:02 on the DRB1*03:01 haplotype can contribute to T1D risk.
Human leukocyte antigen A (HLA-A) genotypes were determined for samples from 283 multiplex, Caucasian, type 1 diabetes families from the Human Biological Data Interchange (HBDI) using an immobilized probe assay. Distribution of HLA-A alleles transmitted to patients was significantly different from that in affected family-based controls (AFBAC) (p = 0.004). Transmission disequilibrium test (TDT) analysis revealed differential transmission of several HLA-A alleles from parents to affected offspring. HLA class II DRB1 and DQB1 loci were also typed, allowing assignment of HLA-A alleles to haplotypes and calculation of linkage disequilibrium values. Some of the apparent effects of HLA-A alleles on type 1 diabetes susceptibility were attributable to linkage disequilibrium with DR and DQ alleles, although others were not. The differences in frequencies between patients and controls of alleles A*0101, A*2402, and A*3002 could not be explained by linkage disequilibrium alone. Our results suggest an important role for class I antigens in modulating susceptibility to type 1 diabetes.
HLA-A genotypes; type 1 diabetes
Alleles of human leukocyte antigen (HLA) class II genes are well known to affect susceptibility to type 1 diabetes (T1D), but less is known about the contribution of HLA class I alleles to T1D susceptibility. In this study, molecular genotyping was performed at the HLA-B and HLA-C loci for 283 multiplex Caucasian families, previously typed for HLA-A and the class II loci. Allele frequencies were compared between affected siblings and affected family-based controls. Linkage disequilibrium coefficients were calculated for HLA-B–HLA-C haplotypes and for class I–class II haplotypes. After adjustment for linkage disequilibrium, the following alleles remain associated with T1D: B*1801, B*3906, B*4403, C*0303, C*0802, and C*1601. B and C allele associations were tested for certain T1D-associated DRB1-DQB1 haplotypes, with the following results: B*3801 is protective on DRB1*0401-DQB1*0302 haplotypes, both C*0701 and C*0702 are predisposing on DRB1*0404-DQB1*0302 haplotypes, and B*3906 is predisposing on DRB1*0801-DQB1*0402 haplotypes. As with previous results for HLA-A, HLA-B and HLA-C are associated with age at T1D onset (mean 11.6 ± 0.3 years). The protective allele B*4403 was associated with older age at onset (15.1 years; p < 0.04), and the predisposing alleles C*0702 and B*3906 were associated with younger age at onset (9.5 years, p < 0.001; and 7.8 years, p < 0.002, respectively). These data support a role for HLA class I alleles in susceptibility to and age at onset of T1D.
type 1 diabetes; HLA class I; HLA-B; HLA-C; age at onset
The goal of the IDAWG is to facilitate the consistent analysis of HLA and KIR data, and the sharing of those data among the immunogenomic and larger genomic communities. However, the data-management approaches currently applied by immunogenomic researchers are not widely discussed or reported in the literature, and the effect of different approaches on data-analyses is not known.
With ASHI’s support, the IDAWG developed a forty-five question survey on HLA and KIR data-generation, data-management, and data-analysis practices. Survey questions detailed the loci genotyped, typing systems used, nomenclature versions reported, computer operating systems and software used to manage and transmit data, the approaches applied to resolve HLA ambiguity, and the methods used for basic population-level analyses. Respondents were invited to demonstrate their HLA ambiguity resolution approaches in simulated data sets.By May 2012, 156 respondents from 35 nations had completed the survey . These survey respondents represent a broad sampling of the Immunogenomic community; 52% were European, 30% North American, 10% Asian, 4% South American, and 4% from the Pacific.
The project will continue in conjunction with the 17th Workshop, with the aim of developing community data-sharing standards, ambiguity resolution documentation formats, single-task data-Management tools, and, novel data-analysis methods and applications. While additional project details and plans for the 17th IHIW will be forthcoming, we welcome the input and participation in these projects from the histocompatibility and immunogenetics community.
Meta-analysis; Statistics; HLA; Genetics
In the last fifteen years published reports have described KIR gene-content frequency distributions in more than 120 populations worldwide. However, there have been limited studies examining these data in aggregate in order to detect overall patterns of variation at regional and global levels. Here, we present a summary of the collection of KIR gene-content data for 105 worldwide populations collected as part of the 15th and 16th International Histocompatibility and Immunogenetics Workshops, and preliminary results for data analysis.
The data were contributed by thirty-four laboratories during the four-year course of this project, including data for the HGDP-CEPH populations. Additionally, data from the 15th IHIW and data contributed to the allelefrequencies.net (AFND) database were combined with the current workshop dataset
KIR; HLA; workshop; report
The major histocompatibility complex (MHC) region is strongly associated with multiple sclerosis (MS) susceptibility. HLA-DRB1*15:01 has the strongest effect, and several other alleles have been reported at different levels of validation. Using SNP data from genome-wide studies, we imputed and tested classical alleles and amino acid polymorphisms in 8 classical human leukocyte antigen (HLA) genes in 5,091 cases and 9,595 controls. We identified 11 statistically independent effects overall: 6 HLA-DRB1 and one DPB1 alleles in class II, one HLA-A and two B alleles in class I, and one signal in a region spanning from MICB to LST1. This genomic segment does not contain any HLA class I or II genes and provides robust evidence for the involvement of a non-HLA risk allele within the MHC. Interestingly, this region contains the TNF gene, the cognate ligand of the well-validated TNFRSF1A MS susceptibility gene. The classical HLA effects can be explained to some extent by polymorphic amino acid positions in the peptide-binding grooves. This study dissects the independent effects in the MHC, a critical region for MS susceptibility that harbors multiple risk alleles.
Multiple sclerosis (MS) is an inflammatory and neurodegenerative disease with a heritable component. Although it has been known for a long time that the strongest MS risk factor maps to the major histocompatibility complex (MHC) on chromosome 6, there are still many unresolved questions as to the identity and the nature of the risk variants within the MHC. Because the MHC has a complex structure, systematic investigation across this region has been challenging. In this study, we used state-of-the-art imputation methods coupled to statistical regression to query variants in the human leukocyte antigen (HLA) class I and II genes for a role in MS risk. Starting from available SNP genotype data, we replicated the strongest risk factor, the HLA-DRB1*15:01 allele, and were able to identify 11 independent effects in total. Functional studies are now needed to understand their mechanism in MS etiology.
The availability of both HLA data and genotypes for thousands of SNPs across the major histocompatibility complex (MHC) in 1240 complete families of the Type 1 Diabetes Genetics Consortium allowed us to analyze the occurrence and extent of megabase contiguous identity for founder chromosomes from unrelated individuals. We identified 82 HLA-defined haplotype groups, and within these groups, megabase regions of SNP identity were readily apparent. The conserved chromosomes within the 82 haplotype groups comprise approximately one third of the founder chromosomes. It is currently unknown whether such frequent conservation for groups of unrelated individuals is specific to the MHC, or if initial binning by highly polymorphic HLA alleles facilitated detection of a more general phenomenon within the MHC. Such common identity, specifically across the MHC, impacts type 1 diabetes susceptibility and may impact transplantation between unrelated individuals.
Type 1 diabetes; MHC; HLA; Extended haplotypes; SNP; 8.1; DR8
Type 1 diabetes (T1D) is a T cell-mediated disease. It is strongly associated with susceptibility haplotypes within the major histocompatibility complex, but this association accounts for an estimated 50% of susceptibility. Other studies have identified as many as 50 additional susceptibility loci, but the effect of most is very modest (odds ratio (OR) <1.5). What accounts for the “missing heritability” is unknown and is often attributed to environmental factors. Here we review new data on the cognate ligand of MHC molecules, the T cell receptor (TCR). In rats, we found that one allele of a TCR variable gene, Vβ13A, is strongly associated with T1D (OR >5) and that deletion of Vβ13+ T cells prevents diabetes. A role for the TCR is also suspected in NOD mice, but TCR regions have not been associated with human T1D. To investigate this disparity, we tested the hypothesis in silico that previous studies of human T1D genetics were underpowered to detect MHC-contingent TCR susceptibility. We show that stratifying by MHC markedly increases statistical power to detect potential TCR susceptibility alleles. We suggest that the TCR regions are viable candidates for T1D susceptibility genes, could account for “missing heritability,” and could be targets for prevention.
Type 1 diabetes (T1D) is one of the most widely studied complex genetic disorders, and the genes in HLA are reported to account for approximately 40% to 50% of the familial aggregation of T1D. The major genetic determinants of this disease are polymorphisms of class II HLA genes encoding DQ and DR. The DR-DQ haplotypes conferring the highest risk are DRB1*03:01-DQA1*05:01-DQB1*02:01 (abbreviated “DR3”) and DRB1*04:01/02/04/05/08-DQA1*03:01-DQB1*03:02/04 (or DQB1*02; abbreviated “DR4”). The risk is much higher for the heterozygote formed by these two haplotypes (OR = 16.59; 95% CI, 13.7–20.1) than for either of the homozygotes (DR3/DR3, OR = 6.32; 95% CI, 5.12–7.80; DR4/DR4, OR = 5.68; 95% CI, 3.91). In addition, some haplotypes confer strong protection from disease, such as DRB1*15:01-DQA1*01:02-DQB1*06:02 (abbreviated “DR2”; OR = 0.03; 95% CI, 0.01–0.07). After adjusting for the genetic correlation with DR and DQ, significant associations can be seen for HLA class II DPB1 alleles, in particular, DPB1*04:02, DPB1*03:01, and DPB1*02:02. Outside of the class II region, the strongest susceptibility is conferred by allele B*39:06 (OR =10.31; 95% CI, 4.21–25.1) and other HLA-B alleles. In addition, several loci in the class III region are reported to be associated with T1D, as are some loci telomeric to class I. Not surprisingly, current approaches for the prediction of T1D in screening studies take advantage of genotyping HLA-DR and HLA-DQ loci, which is then combined with family history and screening for autoantibodies directed against islet-cell antigens. Inclusion of additional moderate HLA risk haplotypes may help identify the majority of children with T1D before the onset of the disease.
Type 1 diabetes; Genetic risk; HLA class II; HLA class I; HLA class III; Risk prediction
To test the hypothesis that closely-related HLA haplotypes containing the DRB1*07:01 gene (“DR7” haplotypes) derived from European and African populations differ in their genetic susceptibility for type 1 diabetes (T1D) depending on the DQ-α molecule present.
Research Design and Methods
A combined total of ninety-eight African American T1D patients from the Type 1 Diabetes Genetics Consortium and from Children’s Hospital and Research Center Oakland were genotyped for the HLA class II loci DRB1, DQA1, and DQB1. DNA samples extracted from newborn blood spot cards from African Americans born in California (n=947) were used as a population-based control group.
Among African American cases, the European-derived DRB1*07:01-DQA1*02:01-DQB1*02:01g haplotype was protective for T1D risk (odds ratio (OR)=0.34; 95% CI 0.14 - 0.78; p<0.011), but the African-derived DRB1*07:01-DQA1*03:01-DQB1*02:01g haplotype increased T1D risk (OR=3.96; 95% CI 1.94 - 8.08; p<5.5E-05).
The effect of DRB1*07:01-DQB1*02:01g on T1D susceptibility depends on the DQA1 allele. DRB1*07:01-DQA1*02:01-DQB1*02:01g is protective for T1D however, the presence of DQA1*03:01 on the DRB1*07:01-DQB1*02:01g haplotype not only renders the DR7 haplotype not protective, it creates a haplotype with significant T1D risk. These data underscore the importance of assessing genetic effects within ethnic context.
type 1 diabetes; genetic risk; HLA class II; HLA DR7; African American
The major histocompatibility complex (MHC) class II transactivator gene (CIITA) encodes an important transcription factor required for HLA class II MHC-restricted antigen presentation. MHC genes, including the HLA class II DRB1*03:01 allele, are strongly associated with systemic lupus erythematosus (SLE). Recently the rs4774 CIITA missense variant (+1632G/C) was reported to be associated with susceptibility to multiple sclerosis. In the current study, we investigated CIITA, DRB1*03:01 and risk of SLE using a multi-stage analysis. In stage 1, 9 CIITA variants were tested in 658 cases and 1,363 controls (N = 2,021). In stage 2, rs4774 was tested in 684 cases and 2,938 controls (N = 3,622). We also performed a meta-analysis of the pooled 1,342 cases and 4,301 controls (N = 5,643). In stage 1, rs4774*C was associated with SLE (odds ratio [OR] = 1.24, 95% confidence interval [95% CI] = 1.07–1.44, P = 4.2 × 10−3). Similar results were observed in stage 2 (OR = 1.16, 95% CI = 1.02–1.33, P = 8.5×10−3) and the meta-analysis of the combined dataset (OR = 1.20, 95% CI = 1.09–1.33, Pmeta = 2.5×10−4). In all three analyses, the strongest evidence for association between rs4774*C and SLE was present in individuals who carried at least one copy of DRB1*03:01 (Pmeta= 1.9×10−3). Results support a role for CIITA in SLE, which appears to be stronger in the presence of DRB1*03:01.
systemic lupus erythematosus; autoimmunity; major histocompatibility complex; HLA; CIITA; MHC2TA
To test the hypothesis that clinical observations made at patient presentation can distinguish type 2 diabetes (T2D) from type 1 diabetes (T1D) in pediatric patients aged 2 to 18.
Subjects and Methods
Medical records of 227 African American and 112 Hispanic American pediatric patients diagnosed as T1D or T2D were examined to compare parameters in the two diseases. Age at presentation, BMI z-score, and gender were the variables used in logistic regression analysis to create models for T2D prediction.
The regression-based model created from African American data had a sensitivity of 92% and a specificity of 89%; testing of a replication cohort showed 91% sensitivity and 93% specificity. A model based on the Hispanic American data showed 92% sensitivity and 90% specificity. Similarities between African American and Hispanic American patients include: (1) age at onset for both T1D and T2D decreased from the 1980s to the 2000s; (2) risk of T2D increased markedly with obesity. Racial/ethnic-specific observations included: (1) in African American patients, the proportion of females was significantly higher than that of males for T2D compared to T1D (p<0.0001); (2) in Hispanic Americans, the level of glycated hemoglobin (HbA1c) was significantly higher in T1D than in T2D (p<0.002) at presentation; (3) the strongest contributor to T2D risk was female gender in African Americans, while the strongest contributor to T2D risk was BMI z-score in Hispanic Americans.
Distinction of T2D from T1D at patient presentation was possible with good sensitivity and specificity using only three easily-assessed variables: age, gender, and BMI z-score. In African American pediatric diabetes patients, gender was the strongest predictor of T2D, while in Hispanic patients, BMI z-score was the strongest predictor. This suggests that race/ethnic specific models may be useful to optimize distinction of T1D from T2D at presentation.
Genetic susceptibility to type 1 diabetes (T1D) has been a subject of intensive study for nearly four decades. This article will present the history of these studies, beginning with observations of the Human Leukocyte Antigen (HLA) association in the 1970s, through the advent of DNA-based genotyping methodologies, through recent large, international collaborations and genome-wide association studies. More than 40 genetic loci have been associated with T1D in multiple studies; however, the HLA region, with its multiple genes and extreme polymorphism at those loci, remains by far the greatest contributor to the genetic susceptibility to T1D. Even after decades of study, the complete story has yet to unfold, and exact mechanisms by which HLA and other associated loci confer T1D susceptibility remain elusive.
More than 40 genetic loci have been associated with type 1 diabetes (T1D). Human leukocyte antigen (HLA) is, by far, the strongest predictor of T1D risk.
We report here genotyping data and type 1 diabetes association analyses for HLA class I loci (A, B, and C) on 1,753 multiplex pedigrees from the Type 1 Diabetes Genetics Consortium (T1DGC), a large international collaborative study.
RESEARCH DESIGN AND METHODS
Complete eight-locus HLA genotyping data were generated. Expected patient class I (HLA-A, -B, and -C) allele frequencies were calculated, based on linkage disequilibrium (LD) patterns with observed HLA class II DRB1-DQA1-DQB1 haplotype frequencies. Expected frequencies were compared to observed allele frequencies in patients.
Significant type 1 diabetes associations were observed at all class I HLA loci. After accounting for LD with HLA class II, the most significantly type 1 diabetes–associated alleles were B*5701 (odds ratio 0.19; P = 4 × 10−11) and B*3906 (10.31; P = 4 × 10−10). Other significantly type 1 diabetes–associated alleles included A*2402, A*0201, B*1801, and C*0501 (predisposing) and A*1101, A*3201, A*6601, B*0702, B*4403, B*3502, C*1601, and C*0401 (protective). Some alleles, notably B*3906, appear to modulate the risk of all DRB1-DQA1-DQB1 haplotypes on which they reside, suggesting a class I effect that is independent of class II. Other class I type 1 diabetes associations appear to be specific to individual class II haplotypes. Some apparent associations (e.g., C*1601) could be attributed to strong LD to another class I susceptibility locus (B*4403).
These data indicate that HLA class I alleles, in addition to and independently from HLA class II alleles, are associated with type 1 diabetes.
To determine the relative risk associated with DPA1 and DPB1 alleles and haplotypes in type 1 diabetes.
RESEARCH DESIGN AND METHODS
The frequency of DPA1 and DPB1 alleles and haplotypes in type 1 diabetic patients was compared to the family based control frequency in 1,771 families directly and conditional on HLA (B)-DRB1-DQA1-DQB1 linkage disequilibrium. A relative predispositional analysis (RPA) was performed in the presence or absence of the primary HLA DR-DQ associations and the contribution of DP haplotype to individual DR-DQ haplotype risks examined.
Eight DPA1 and thirty-eight DPB1 alleles forming seventy-four DPA1-DPB1 haplotypes were observed; nineteen DPB1 alleles were associated with multiple DPA1 alleles. Following both analyses, type 1 diabetes susceptibility was significantly associated with DPB1*0301 (DPA1*0103-DPB1*0301) and protection with DPB1*0402 (DPA1*0103-DPB1*0402) and DPA1*0103-DPB1*0101 but not DPA1*0201-DPB1*0101. In addition, DPB1*0202 (DPA1*0103-DPB1*0202) and DPB1*0201 (DPA1*0103-DPB1*0201) were significantly associated with susceptibility in the presence of the high risk and protective DR-DQ haplotypes. Three associations (DPB1*0301, *0402, and *0202) remained statistically significant when only the extended HLA-A1-B8-DR3 haplotype was considered, suggesting that DPB1 alone may delineate the risk associated with this otherwise conserved haplotype.
HLA DP allelic and haplotypic diversity contributes significantly to the risk for type 1 diabetes; DPB1*0301 (DPA1*0103-DPB1*0301) is associated with susceptibility and DPB1*0402 (DPA1*0103-DPB1*0402) and DPA1*0103-DPB1*0101 with protection. Additional evidence is presented for the susceptibility association of DPB1*0202 (DPA1*0103-DPB1*0202) and for a contributory role of individual amino acids and DPA1 or a gene in linkage disequilibrium in DR3-DPB1*0101 positive haplotypes.
One source of deoxyribonucleic acid (DNA) for genetic studies is the utilization of dried blood spots stored on paper cards (Guthrie cards) collected shortly after birth. These cards represent an important source of material for epidemiologic and population-based genetic studies. Extraction of DNA from these cards can lead to variable amounts of recovered DNA. We report here results of our efforts to maximize yield from this valuable, but nonrenewable, resource.
Commercial methods of DNA extraction from blood cards were used, and protocol modifications were introduced that enhanced DNA yield.
Use of a commercial solvent prior to DNA extraction steps gave greater yields than extraction without the solvent. Modification of the elution step by use of prewarmed extraction buffer and a soaking step at an elevated temperature increased yield by 6- to 10-fold.
The modified DNA extraction method yielded as much as 660 ng of DNA from a single 5-mm-diameter punch of a blood spot card. The DNA performed well in downstream, polymerase chain reaction-based applications.
blood spot; DNA sample preparation; Guthrie card; HLA genotyping
A deletion polymorphism in glutathione S-transferase Mu-1 (GSTM1-null) has previously been implicated to play a role in rheumatoid arthritis (RA) risk and progression, although no prior investigations have examined its associations with anticitrullinated protein antibody (ACPA) positivity. The purpose of this study was to examine the associations of GSTM1-null with ACPA positivity in RA and to assess for evidence of interaction between GSTM1 and HLA-DRB1 shared epitope (SE).
Associations of GSTM1-null with ACPA positivity were examined separately in two RA cohorts, the Veterans Affairs Rheumatoid Arthritis (VARA) registry (n = 703) and the Study of New-Onset RA (SONORA; n = 610). Interactions were examined by calculating an attributable proportion (AP) due to interaction.
A majority of patients in the VARA registry (76%) and SONORA (69%) were positive for ACPA with a similar frequency of GSTM1-null (53% and 52%, respectively) and HLA-DRB1 SE positivity (76% and 71%, respectively). The parameter of patients who had ever smoked was more common in the VARA registry (80%) than in SONORA (65%). GSTM1-null was significantly associated with ACPA positivity in the VARA registry (odds ratio (OR), 1.45; 95% confidence interval (CI), 1.02 to 2.05), but not in SONORA (OR, 1.00; 95% CI, 0.71 to 1.42). There were significant additive interactions between GSTM1 and HLA-DRB1 SE in the VARA registry (AP, 0.49; 95% CI, 0.21 to 0.77; P < 0.001) in ACPA positivity, an interaction replicated in SONORA (AP, 0.38; 95% CI, 0.00 to 0.76; P = 0.050).
This study is the first to show that the GSTM1-null genotype, a common genetic variant, exerts significant additive interaction with HLA-DRB1 SE on the risk of ACPA positivity in RA. Since GSTM1 has known antioxidant functions, these data suggest that oxidative stress may be important in the development of RA-specific autoimmunity in genetically susceptible individuals.
Genetic susceptibility to systemic lupus erythematosus (SLE) is well established, with the HLA class II DRB1 and DQB1 loci demonstrating the strongest association. However, HLA may also influence SLE through novel biologic mechanisms in addition to genetic transmission of risk alleles. Evidence for increased maternal–offspring HLA class II compatibility in SLE and differences in maternal versus paternal transmission rates (parent-of-origin effects) and nontransmission rates (noninherited maternal antigen [NIMA] effects) in other autoimmune diseases have been reported. Thus, we investigated maternal–offspring HLA compatibility, parent-of-origin effects, and NIMA effects at DRB1 in SLE.
The cohort comprised 707 SLE families and 188 independent healthy maternal–offspring pairs (total of 2,497 individuals). Family-based association tests were conducted to compare transmitted versus nontransmitted alleles (transmission disequilibrium test) and both maternally versus paternally transmitted (parent-of-origin) and nontransmitted alleles (using the chi-square test of heterogeneity). Analyses were stratified according to the sex of the offspring. Maternally affected offspring DRB1 compatibility in SLE families was compared with paternally affected offspring compatibility and with independent control maternal–offspring pairs (using Fisher’s test) and was restricted to male and nulligravid female offspring with SLE.
As expected, DRB1 was associated with SLE (P < 1 × 10−4). However, mothers of children with SLE had similar transmission and nontransmission frequencies for DRB1 alleles when compared with fathers, including those for the known SLE risk alleles HLA–DRB1*0301, *1501, and *0801. No association between maternal–offspring compatibility and SLE was observed.
Maternal–offspring HLA compatibility, parent-of-origin effects, and NIMA effects at DRB1 are unlikely to play a role in SLE.