To Type 1 Diabetes Patients Research Articles

The Impact of Thyroiditis on the Immune Profile in Young Patients with Uncomplicated Type 1 Diabetes.

Autoimmune thyroid disease (AIT) is the most frequently linked autoimmune condition to type 1 diabetes (T1D). The analysis of immune profiles could provide valuable insights into the study of these diseases. This knowledge could play a crucial role in understanding the relationship between immune profiles and microcirculation structures and functions. The present study aimed to test the hypothesis that cytokine levels in T1D patients without and those with comorbid Hashimoto's disease differ significantly. The total study group (total T1D) consisted of 62 diabetic young patients: 43 T1D and 19 T1D + AIT matched for age, age at onset, and duration of diabetes. The control group consisted of 32 healthy young subjects. The levels of cytokines (including TNF-α, IL-35, IL-4, IL-10, IL-18, IL-12, VEGF, and angiogenin) were quantified throughout this investigation. A comparative assessment of the cytokines profiles between the control group and total T1D revealed a statistically significant elevation in the levels of IL-4, TNF-α, IL-18, VEGF, and angiogenin, accompanied by a notable decline in IL-10. However, IL-35 and IL-12 exhibited comparable levels between the two groups. A comparison of cytokine levels between T1D + AIT and T1D groups revealed that only angiogenin levels were statistically significantly higher in T1D + AIT. The results of our study indicated that the alterations in cytokine levels associated with AIT did not correspond to the observed changes in T1D-related outcomes. The sole notable observation was the elevation of angiogenin expression, an angiogenic factor.

Association analysis between the TLR9 gene polymorphism rs352140 and type 1 diabetes.

To a great extent, genetic factors contribute to the susceptibility to type 1 diabetes (T1D) development, and by triggering immune imbalance, Toll-like receptor (TLR) 9 is involved in the development of T1D. However, there is a lack of evidence supporting a genetic association between polymorphisms in the TLR9 gene and T1D. In total, 1513 individuals, including T1D patients (n=738) and healthy control individuals (n=775), from the Han Chinese population were recruited for an association analysis of the rs352140 polymorphism of the TLR9 gene and T1D. rs352140 was genotyped by MassARRAY. The allele and genotype distributions of rs352140 in the T1D and healthy groups and those in different T1D subgroups were analyzed by the chi-squared test and binary logistic regression model. The chi-square test and Kruskal-Wallis H test were performed to explore the association between genotype and phenotype in T1D patients. The allele and genotype distributions of rs352140 were significantly different in T1D patients and healthy control individuals (p=0.019, p=0.035). Specifically, the T allele and TT genotype of rs352140 conferred a higher risk of T1D (OR=1.194, 95% CI=1.029-1.385, p=0.019, OR=1.535, 95% CI=1.108-2.126, p=0.010). The allele and genotype distributions of rs352140 were not significantly different between childhood-onset and adult-onset T1D and between T1D with a single islet autoantibody and T1D with multiple islet autoantibodies (p=0.603, p=0.743). rs352140 was associated with T1D susceptibility according to the recessive and additive models (p=0.015, p=0.019) but was not associated with T1D susceptibility in the dominant and overdominant models (p=0.117, p=0.928). Moreover, genotype-phenotype association analysis showed that the TT genotype of rs352140 was associated with higher fasting C-peptide levels (p=0.017). In the Han Chinese population, the TLR9 polymorphism rs352140 is associated with T1D and is a risk factor for susceptibility to T1D.

Neutrophil Extracellular Trap Induced Dendritic Cell Activation Leads to Th1 Polarization in Type 1 Diabetes.

Neutrophils releasing neutrophil extracellular traps (NETs) infiltrate the pancreas prior to type 1 diabetes (T1D) onset; however, the precise nature of their contribution to disease remains poorly defined. To examine how NETs affect immune functions in T1D, we investigated NET composition and their effect on dendritic cells (DCs) and T lymphocytes in T1D children. We showed that T1D patient NET composition differs substantially from that of healthy donors and that the presence of T1D-NETs in a mixed peripheral blood mononuclear cell culture caused a strong shift toward IFNγ-producing T lymphocytes, mediated through activation of innate immunity cells in T1D samples. Importantly, in a monocyte-derived DC (moDC) culture, NETs induced cytokine production, phenotypic change and IFNγ-producing T cells only in samples from T1D patients but not in those from healthy donors. RNA-seq analysis revealed that T1D-NETs presence causes TGFβ downregulation and IFNα upregulation and creates pro-T1D signature in healthy moDCs.

161-OR: Autoantibodies Directed to Deamidated Post-translationally Modified IA-2 Epitopes in Type 1 Diabetes

Autoimmune disorders such rheumatoid arthritis (RA) harbor autoantibodies against a variety of post-translationally modified (PTM) proteins. We aimed to investigate whether PTM epitopes of IA-2, a major autoantigen related to type 1 diabetes (T1D), are targeted by autoantibody responses. The extracellular domain of IA-2 (IA-2-ec) was synthesized incorporating deamidated amino acid residues (IA-2ec-PTM) previously found to elicit T cell responses in T1D patients. IA-2ec-PTM was then subjected to immunoprecipitation with sera from T1D patients. Notably, autoantibody reactivity towards IA-2ec-PTM was present in 28% of T1D patients as compared to ~9% of serum reactivity against the native IA-2ec. The area under the ROC curve (ROC AUC) of IA-2ec-PTM autoantibodies revealed that these autoantibodies can adequately distinguish between T1D patient and control groups (ROC AUC: 0.7132; P < 0.0001). Relatives of T1D probands (from the TrialNet Pathway to Prevention Study) carrying both IA-2ec-PTM AAb and the high-risk HLA-DRB1*04-DQB1*03:02 haplotype progressed rapidly to onset of T1D (Stage 3) compared to those relatives who did not carry these immunologic abnormalities (P = 0.003). Understanding the mechanisms underlying PTMs of islet autoantigens, such as IA-2ec-PTM, is critical to develop new research to determine the ability of modified epitopes to induce islet autoimmunity, their potential role as biomarkers of islet cell injury as well as breaking immune tolerance to islet cell antigens through neo-antigen formation. Disclosure M.J. Acevedo-Calado: None. S. Pietropaolo: None. L. Yu: None. A.W. Michels: Stock/Shareholder; Self; IM Therapeutics. M. Pietropaolo: None. Funding National Institutes of Health

Eyekon Is a Novel Diagnostic Tool for Diabetic Neuropathy

Diabetic neuropathy (DN) is a major complication of diabetes, causing more than 50% of all hospital accesses related to type 1 diabetes (T1D). However, diagnostic criteria are not yet standardized, and diagnosis also of symptomatic DN is often underestimated. Given the medical need for an accurate non-invasive diagnostic tool, we developed a test based on tracking eye movement for early detection of DN called Eykon. SRLab -Tobii TX300 Eye tracker®, an eye-tracking device, is coupled with a series of screens on a computer to test the velocity and accuracy of gaze movement in healthy controls (CTRL, n=30), T1D patients without neuropathy (T1D, n=19) and T1D patients with neuropathy (T1D-N, n=14). The screens are divided in 5 classes: Resistance, Wideness, Pursuit, Velocity and OKN. They aim to evaluate both smooth and saccadic movement in different directions. We evaluated a total of 483 parameters and observed that 27 (5.6%) were altered in T1D patients without signs of advanced DN at the electromyography as compared to CTRL, while 29 (6%) were altered in T1D-N patients, for a total of 56 (11.6%) significantly altered parameters (T1D vs. CTRL). Our pilot study demonstrated the feasibility of this non-invasive exam and its ability to early detect DN, despite the limitations of a small number of subjects. Disclosure M. Ben Nasr: None. E. Giuliani: None. F. D'Addio: None. A. Maestroni: None. V. Usuelli: None. S. Bianchi Marzoli: None. P. Fiorina: None.

Mitochondrial DNA Activates NLRP3 Inflammasome and Contributes to Endothelial Dysfunction and Inflammation in Type 1 Diabetic Mice

IntroductionNLRP3 inflammasome is a molecular platform of innate immune system that regulates the inflammatory response through the activation of caspase‐1 and IL‐1β. It can be activated by several ligands, for example, mitochondrial DNA (mDNA). Some studies have shown that circulating mDNA is increased in patients with diabetes and play a role in the development of the disease; however, its role in vascular changes is still unknown. In this sense, we tested the hypothesis that mDNA contributes to vascular inflammatory/oxidative processes associated to type 1 diabetes (T1D) via NLRP3 inflammasome activation.MethodsWild‐type and NLRP3‐deficient (NLRP3−/−) mice were treated with vehicle (Veh) or streptozotocin (40 mg/kg) (T1D) for 5 days. Vascular reactivity was determined in mesenteric arteries. Caspase‐1 and IL‐1β expression were evaluated by western blot. Pancreatic mDNA was extracted from control (cmDNA) and diabetic animals (dmDNA) for endothelial cells stimulation. ROS generation was determined by chemiluminescence. Hydrogen peroxide (H2O2) production and calcium influx were determined by fluorescence. Data are presented as mean ± standard error of the mean.ResultsDiabetes increased vascular caspase‐1 and IL‐1β activation [arbitrary units (a.u.), 1.2±0.1 vs. 0.8±0.1; 4.8±1.1 vs. 0.8±0.5 vs. the Veh, respectively, p <0.05]. However, this activation was attenuated in T1D NLRP3−/−. Mesenteric arteries from T1D exhibited decreased ACh‐induced vasodilatation vs. Veh [(Emax), 46.6±4.0 vs. 91.5±2.8, n=4–5, p<0.05], which was not observed in T1D NLRP3−/−. The NLRP3 inhibitor MCC950 acutely improved endothelium‐dependent vasodilation in T1D [(Emax), 62.5±3.8 vs. 85.6±3.7, n=4–5, p<0.05]. ROS generation [(Relative luminescence unit, RLU), 1.5×107±2.3 vs. 48.9×103±1.1, n=4–5, p<0.05] as well as H2O2 production [(Relative fluorescence unit, RFU), 1082±242.6 vs. 232.0±42.8, n=4–5, p<0.05] were lower in mesenteric bed from T1D NLRP3−/− than T1D. Only incubation with dmDNA reduced ACh‐induced vasodilation [(Emax), 48.4 ± 4.1 vs. 90.7 ± 3.4, n=4–5, p<0.05], which was attenuated by the presence of an antioxidant [(Emax), 48.4 ± 4.1 vs. 71.5 ± 3.1, n=4–5, p<0.05]. Similarly, only LPS‐primed cells incubated with dmDNA had significant NLRP3 activation (caspase‐1 and IL‐1β activation, respectively) [(u.a.), 1.6 ± 0.2 vs. 0.9 ± 0.1; and 1.6 ± 0.2 vs. 1.0 ± 0.1, n=4–5, p<0.05]. The dmDNA induced in a time‐dependent way both calcium influx and ROS generation in endothelial cells, being the last reversed by the presence of an antioxidant. Likewise, T1D patients had increased circulating mDNA vs. healthy volunteers and also higher NLRP3 serum expression, and caspase‐1/IL‐1β activation.ConclusionThe T1D increases mDNA release which in turn stimulates increased calcium influx, induces ROS generation, and triggers vascular NLRP3 inflammasome activation contributing to inflammatory response and endothelial dysfunction in diabetes.Support or Funding InformationFinancial Support: FAPESP and CAPES. Ethics Committee number (026/2015).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Association of genes of different functional classes with type 1 diabetes

The genetic structure of susceptibility to type 1 diabetes (T1D) in the population of Tomsk was studied. We had a group of T1D patients (N = 285) and a population sample (N = 300) and we studied 58 SNPs localized in the 47 genes which products are involved in various metabolic pathways and processes as fibrogenesis, endothelial dysfunction, and inflammation. Genotyping was performed by mass spectrometry using the Sequenom MassARRAY system (United States). We compared the group of T1D patients and the population sample and found an association with the predisposition to disease for seven markers: rs3765124 of the ADAMDEC1 gene, genotype AA (p = 0.004), allele A (p = 0.033); rs1007856 of the ITGB5 gene, genotype TT (p = 0.015), allele T (p = 0.036); rs20579 of the LIG1 gene, genotype CC (p = 0.004), allele C (p = 0.002); rs12980602 of the IFNL2 gene, allele C (p = 0.029); rs4986819 of the PARP4 gene, allele C (p = 0.044); rs1143674 of the ITGA4 gene genotype GG (p = 0.002); rs679620 of the MMP3 gene, genotype AA (p = 0.008). Thus, the products of genes associated with T1D belong to different molecular classes: metalloproteases (ADAMDEC1, MMP3), cytokines (IL28A), cell surface receptors (ITGA4), adhesion molecules (ITGB5), DNA ligases (LIG1), and ribosyltransferase enzymes (PARP4). The ADAMDEC1, ITGA4, and ITGB5 genes belong to two biological processes: cell communication and signal transduction. The LIG1 and PARP4 genes regulate the metabolism of nucleic acids, MMP3 is involved in the regulation of protein metabolism, and the IFNL2 is involved in the immune response.

A multiplex assay combining insulin, GAD, IA-2 and transglutaminase autoantibodies to facilitate screening for pre-type 1 diabetes and celiac disease

At the current time, multiple candidate interventions are being proposed to abrogate or slow progression to type 1 diabetes (T1D) among islet autoantibody (iAb) positive subjects, but mass screening for eligible subjects and the general population remains a laborious and inefficient process. We have recently developed and extensively validated nonradioactive iAb assays using electrochemiluminescense (ECL) detection with an excellent sensitivity and specificity compared to the gold-standard radioassays. Using ECL detection on a platform from MesoScale Discovery (MSD) allows the measurement of four antibodies in a single well using a small blood volume (6μl). In the present study using a MSD QuickPlex 4-Spot plate, we successfully combined three iAb to insulin (IAA), GAD65 (GADA), and IA-2 (IA-2A) with tissue transglutaminase autoantibodies (TGA) in a single well of a 96 well plate. We tested 40 new onset T1D patients, all positive for at least one iAb and a half of them positive for TGA by radioassay, as well as 50 healthy controls. The multiplex assay retained 100% sensitivity and 100% specificity for all four autoantibodies in terms of positivity identified in patients versus normal controls compared to the corresponding standard radioassays and our single ECL assays. The multiplex ECL assay was able to identify more positivity than current radioassays for IAA and TGA. The development of this multiplex assay will facilitate high-throughput screening for T1D and celiac disease risk in the general population.

Genetic Variations of PTPN2 and PTPN22: Role in the Pathogenesis of Type 1 Diabetes and Crohn's Disease.

Genome wide association studies have identified several genes that might be associated with increase susceptibility to Type 1 Diabetes (T1D) and Crohn's disease. Both Crohn's disease and T1D have a profound impact on the lives of patients and it is pivotal to investigate the genetic role in patients acquiring these diseases. Understanding the effect of single nucleotide polymorphisms (SNP's) in key genes in patients suffering from T1D and Crohn's disease is crucial to finding an effective treatment and generating novel therapeutic drugs. This review article is focused on the impact of SNP's in PTPN2 (protein tyrosine phosphatase, non-receptor type 2) and PTPN22 (protein tyrosine phosphatase non-receptor type 22) on the development of Crohn's disease and T1D. The PTPN2 gene mutation in T1D patients play a direct role in the destruction of beta cells while in Crohn's disease patients, it modulates the innate immune responses. The PTPN22 gene mutations also play a role in both diseases by modulating intracellular signaling. Examining the mechanism through which these genes increase the susceptibility to both diseases and gaining a better understanding of their structure and function is of vital importance to understand the etiology and pathogenesis of Type 1 Diabetes and Crohn's disease.

Assessment of CD4+ T cell responses to glutamic acid decarboxylase 65 using DQ8 tetramers reveals a pathogenic role of GAD65 121-140 and GAD65 250-266 in T1D development.

Susceptibility to type 1 diabetes (T1D) is strongly associated with MHC class II molecules, particularly HLA-DQ8 (DQ8: DQA1*03:01/DQB1*03:02). Monitoring T1D-specific T cell responses to DQ8-restricted epitopes may be key to understanding the immunopathology of the disease. In this study, we examined DQ8-restricted T cell responses to glutamic acid decarboxylase 65 (GAD65) using DQ8 tetramers. We demonstrated that GAD65121–140 and GAD65250–266 elicited responses from DQ8+ subjects. Circulating CD4+ T cells specific for these epitopes were detected significantly more often in T1D patients than in healthy individuals after in vitro expansion. T cell clones specific for GAD65121–140 and GAD65250–266 carried a Th1-dominant phenotype, with some of the GAD65121–140-specific T cell clones producing IL-17. GAD65250–266-specific CD4+ T cells could also be detected by direct ex vivo staining. Analysis of unmanipulated peripheral blood mononuclear cells (PBMCs) revealed that GAD65250–266-specific T cells could be found in both healthy and diabetic individuals but the frequencies of specific T cells were higher in subjects with type 1 diabetes. Taken together, our results suggest a proinflammatory role for T cells specific for DQ8-restricted GAD65121–140 and GAD65250–266 epitopes and implicate their possible contribution to the progression of T1D.

P055 : INS-VNTR POLYMORPHISM AND GENE EXPRESSION IN TYPE 1, TYPE 2 AND GESTATIONAL IN BRAZILIAN DIABETES MELLITUS PATIENTS

Besides HLA class II alleles, the variable number of tandem repeat polymorphism located at 5’ of the insulin gene (INS-VNTR) at chromosome 11p15.5 is the second most important susceptibility region to type 1 diabetes (T1D). INS-VNTR alleles (ACAGGGGTGTGGGG) is divided into class I (30–60 repeats), class II (60–120) and class III (120–160). INS-VNTR class I allele transcribe lower thymic INS than class III allele favoring escape of auto-reactive thymocytes from negative selection. Since insulin resistance is a major shared feature among diabetes patients and since there no studies evaluating these markers in the major types of the disease, we characterized the INS-VNTR polymorphisms (348patients/339controls) and the differential mRNA/microRNA (40 patients) expression in T1D, gestational (GDM) and type 2 (T2D) Brazilian diabetic patients. INS-VNTR was performed using −23HphI PCR-RFLP, and gene expression profiles were performed using Agilent microarrays. INS-VNTR I:I genotype was associated with susceptibility, whereas the III:III genotype conferred protection against T1D development, and I:III genotype was associated to T2D. The multivariate analyses results showed that the I:I genotype was associated with age and dyslipidemia only in T1D patients. Considering all types of diabetes, the differential gene expression profiles (mRNA/miRNA) for INS-VNTR I:I genotype were distinct from those observed for the III:III genotype. The miRNA expression profile according to INS-VNTR polymorphism yielded 31 differentially expressed transcripts. Networks of functional miRNA target, presenting possible INS interactions, showed 6 upregulated ( I:I genotype), 10 upregulated (I:III genotype), and 6 upregulated and 8 downregulated for III:III genotype. Conclusion: This is the first study of INS-VNTR polymorphisms evaluating the three major types of diabetes in the same population group. Moreover, the INS-VNTR genotypes influenced gene and miRNAs expression profiles.

P102 : GESTATIONAL DIABETES MELLITUS TRANSCRIPTOME ANALYSIS REVEALS INDUCTION OF GENES RELATED TO THE MAJOR HISTOCOMPATIBILITY COMPLEX

Gestational diabetes mellitus (GDM) is the most common metabolic disorder observed during pregnancy, and has been defined as an abnormal glucose metabolism first diagnosed during gestation. GDM women usually present postpartum diabetes, insulin resistance, metabolic syndrome, hypertension and dyslipidemia. Although few genetic markers have been described for GDM, approximately 10% of GDM patients present high probability of progression to type 1 diabetes (T1D). In a previous meta-analysis study, evaluating the transcript profiles of patients with the major types of diabetes, we reported that transcription profile of T1D patients was closer to GDM than to type 2 diabetes. Additionally, it has already been reported association of some HLA alleles with GDM, suggesting a possible immunological feature for GDM. In this study, we compared the transcription profiling of PBMCs of 18 GDM patients and 10 healthy pregnant women. Total RNA samples were hybridized to Agilent® 4 × 44 K oligo microarrays encompassing the whole human functional genome. Differentially expressed mRNAs were obtained by Rank Products analysis, which classifies transcripts according to their fold change values. The hierarchical clustering of mRNAs and samples were performed using the Cluster program, dendrograms and spatial representations of mRNAs were constructed using the TreeView software. A total of 731 differentially and significantly expressed mRNAs were observed when GDM patients were compared to controls. Molecular functions and biological pathways, as analyzed by the DAVID database, identified 130 biological processes and nine signaling pathways ( P

Is type 1 diabetes "going viral"?

Although genetic predisposition to type 1 diabetes (T1D) is a key risk factor, it does not explain the recent and rapid rise in incidence of the disease, particularly among children under the age of 5 (1). Changes in the environment and/or how individuals respond to these changes are now widely believed to be responsible for the recent increases in T1D. While viruses have been hypothesized to have an etiological role in T1D for more than five decades, definitive proof has not yet materialized. Nonetheless, a number of candidate viruses have been proposed, including enteroviruses, rotaviruses, herpesviruses, cytomegalovirus, and endogenous retroviruses (2). The strongest and most clinically significant associations point to enterovirus infection. A recent meta-analysis of 26 studies that assessed enteroviral infection using both molecular and immunological assays showed that compared with nondiabetic control subjects, the likelihood of finding evidence of enterovirus is 10-fold higher in T1D patients and fourfold higher in individuals with diabetes-related autoimmunity (3). Despite this evidence, important questions remain. These include whether a viral infection is causal in the development of T1D, whether it acts to accelerate the onset of disease in individuals in whom the process of β-cell destruction has already been initiated, or whether it is simply present because individuals with diabetes are more susceptible to infection. Two articles published in this issue of Diabetes help to address these questions. Ferreira et al. (4) and Kallionpaa et al. (5) used peripheral blood samples from two longitudinal birth cohorts—the BABYDIET cohort (6) and the Finnish Type 1 Diabetes Prediction and Prevention (DIPP) study (7), respectively. Both studies monitored development of diabetes-related autoantibodies as well as the onset of T1D in “at-risk” populations, thereby enabling access to samples …

Association of PTPN22+1858C/T polymorphism with Type 1 diabetes in the North Indian population.

A nonsynonymous SNP +1858C/T (rs2476601) in the protein tyrosine phosphatase, nonreceptor type 22(PTPN22) gene leading to Arg 620 Trp substitution is known to be associated with susceptibility to type 1 diabetes (T1D) and several other autoimmune diseases. We studied this polymorphism in 145 T1D patients and 210 healthy controls from North India. The minor allele +1858T was observed to be significantly increased among patients as compared to healthy controls (2.76% vs 0.5%, P = 0.027, OR = 5.93; 95% CI = 1.4-24.8). The association was also observed at the level of heterozygous C/T genotype (5.5% vs 0.95%, P = 0.026, OR = 6.07; 95% CI = 1.43-25.6). The T allele and C/T genotype were predominantly found among patients who were positive for both glutamic acid decarboxylase 65 (GAD65) and insulin antigen 2 (IA2) autoantibodies and showed significantly increased frequencies (10%, P = 0.034, OR = 11.67; 95% CI = 1.58-84.1 and 20%, P = 0.031, OR = 13.0; 95% CI = 1.66-97.5, respectively) as compared to patients negative for these autoantibodies (0.95% and 1.9%, respectively). The results suggest that the PTPN22+1858T allele is positively associated with T1D in the North Indian population.

CD226 rs763361 is associated with the susceptibility to type 1 diabetes and greater frequency of GAD65 autoantibody in a Brazilian cohort.

CD226 rs763361 variant increases susceptibility to type 1 diabetes (T1D) in Caucasians. There is no data about CD226 variants in the very heterogeneous Brazilian population bearing a wide degree of admixture. We investigated its association with T1D susceptibility, clinical phenotypes, and autoimmune manifestations (islet and extrapancreatic autoantibodies). Casuistry. 532 T1D patients and 594 controls in a case-control study. Initially, CD226 coding regions and boundaries were sequenced in a subset of 106 T1D patients and 102 controls. In a second step, two CD226 variants, rs763361 (exon 7) and rs727088 (3′ UTR region), involved with CD226 regulation, were genotyped in the entire cohort. C-peptide and autoantibody levels were determined. No new polymorphic variant was found. The variants rs763361 and rs727088 were in strong linkage disequilibrium. The TT genotype of rs763361 was associated with TID risk (OR = 1.503; 95% CI = 1.135–1.991; P = 0.0044), mainly in females (P = 0.0012), greater frequency of anti-GAD autoantibody (31.9% × 24.5%; OR = 1.57; CI = 1.136–2.194; P = 0.0081), and lower C-peptide levels when compared to those with TC + CC genotypes (0.41 ± 0.30 ng/dL versus 0.70 ± 0.53 ng/dL P = 0.0218). Conclusions. The rs763361 variant of CD226 gene (TT genotype) was associated with susceptibility to T1D and with the degree of aggressiveness of the disease in T1D patients from Brazil. Ancestry had no effect.

Infectious triggers in type 1 diabetes: is there a case for epitope mimicry?

Environmental factors are the main contributors to type 1 diabetes (T1D) pathogenesis, yet they remain unidentified. Enteroviruses are proposed candidate triggers due to temporal correlations between infection and T1D autoimmunity and to detection of viral proteins in diseased islets. However, such correlations are not universal and may be relatively uncommon. Furthermore, evidence of a cause-effect relationship is lacking, as infection of non-obese diabetic mice with Coxsackie enteroviruses can either trigger or blunt disease. The proposed mechanisms are either non-antigen-specific (i.e. β-cell destruction and release of sequestered antigens, islet inflammation) or antigen-specific (i.e. epitope mimicry, by which immune responses to enteroviruses may be diverted against homologous β-cell antigens). The case for the latter mechanisms is even less stringent, as there is little evidence of promiscuous antigen recognition at the single T-cell level. Other infectious agents may thus be implicated. Demonstration of their role will require fulfilling the Koch's postulates, namely isolation of the agent preferentially in T1D patients, including before disease onset; and T1D induction when the agent is inoculated into mice. The same is needed for cross-reactive T cells to support epitope mimicry mechanisms. Generation of alternative (humanized) mouse models that could be challenged with candidate microbes is needed.

Protective effect of interleukin-23A (IL23A) haplotype variants on type 1A diabetes mellitus in a Brazilian population

The Interleukin 23 (IL-23) has a central role in autoimmunity. Allelic variants of p19 subunit of IL-23 (IL23A) and IL-23 receptor (IL23R) genes and increased IL-23 serum concentrations were associated with autoimmune diseases. We therefore searched for variants of IL23A and IL23R that could predispose to Type 1 diabetes (T1D). The coding regions and boundary intron sequences of IL23A were sequenced. Variants of IL23A and of IL23R were also genotyped. Pancreatic and extrapancreatic autoantibodies and IL-23 serum levels were determined. The cohort involved 370 patients with T1D and 351 healthy control subjects. We observed only one coding IL23A variant (rs11171806 G>A) out of the 6 described in databases. As the G alleles of rs11171806 and rs2066808 variants of IL23A gene were in strong linkage disequilibrium (D′=−0.825 for controls, p<2.0×10−6 and D′=−0.902, p<2.0×10−17 for patients), further analyses were performed with the haplotypes. The GG haplotype was more frequent in controls (16.7%) than in T1D patients (9.5%), conferring a protection to T1D (OR=0.53; pc=0.0003). No association was found between IL23A allelic variants with age at diagnosis of diabetes, C-peptide levels or frequency of autoantibodies. IL23R variants (rs10889677 and rs11209026) frequency and IL-23 serum concentrations were similar between groups. The GG haplotype of lL23A variants (rs11171806 and rs2066808) was protective against T1D. IL23R variants (rs11209026 and rs10889677) were not associated with T1D. IL-23 serum concentrations did not differ between groups.

HLA-A*24 Is an Independent Predictor of 5-Year Progression to Diabetes in Autoantibody-Positive First-Degree Relatives of Type 1 Diabetic Patients

We investigated whether HLA-A*24 typing complements screening for HLA-DQ and for antibodies (Abs) against insulin, GAD, IA-2 (IA-2A), and zinc transporter-8 (ZnT8A) for prediction of rapid progression to type 1 diabetes (T1D). Persistently Ab+ siblings/offspring (n = 288; aged 0–39 years) of T1D patients were genotyped for HLA-DQA1-DQB1 and HLA-A*24 and monitored for development of diabetes within 5 years of first Ab+. HLA-A*24 (P = 0.009), HLA-DQ2/DQ8 (P = 0.001), and positivity for IA-2A ± ZnT8A (P < 0.001) were associated with development of T1D in multivariate analysis. The 5-year risk increased with the number of the above three markers present (n = 0: 6%; n = 1: 18%; n = 2: 46%; n = 3: 100%). Positivity for one or more markers identified a subgroup of 171 (59%) containing 88% of rapid progressors. The combined presence of HLA-A*24 and IA-2A+ ± ZnT8A+ defined a subgroup of 18 (6%) with an 82% diabetes risk. Among IA-2A+ ± ZnT8A+ relatives, identification of HLA-A*24 carriers in addition to HLA-DQ2/DQ8 carriers increased screening sensitivity for relatives at high Ab- and HLA-inferred risk (64% progression; P = 0.002). In conclusion, HLA-A*24 independently predicts rapid progression to T1D in Ab+ relatives and complements IA-2A, ZnT8A, and HLA-DQ2/DQ8 for identifying participants in immunointervention trials.

IGF2BP2 and IGF2 genetic effects in diabetes and diabetic nephropathy

ObjectiveThe IGF2BP2 gene is located on chromosome 3q27.2 within a region linked to type 1 diabetes (T1D), type 2 diabetes (T2D) and diabetic nephropathy (DN). Its protein functionally binds to 5’-UTR of the imprinting IGF2 gene. The present study aims to evaluate the IGF2BP2-IGF2 genetic effects in diabetes and DN. Materials and MethodsThree cohorts including T1D with and without DN (n=1139) of European descents from the GoKinD study, Swedish T1D with and without DN (n=303) and Czech control subjects without diabetes, T1D, T2D with and without DN (n=1418) were enrolled in TaqMan genotyping experiments for IGF2BP2 rs4402960 and IGF2 rs10770125. Igf2bp2 gene expression in kidney tissues of db/db and control mice at the ages of 5 and 26weeks was examined with real time RT-PCR and Western blot. ResultsAn association of IGF2BP2 rs4402960 with T2D in the Czech population was replicated. This IGF2BP2 polymorphism (P=0.037, OR=0.69 95% CI 0.49–0.98) was found to be associated with DN in male not in female patients with T1D selected from the GoKinD study. In the analyses of combined the GoKinD, Czech and Swedish populations, the association between IGF2BP2 polymorphism and DN in male patients with T1D was still significant (P=0.030, OR=0.73, 95% CI 0.54–0.97). IGF2 rs10770125 was also associated with DN in male T1D patients of the GoKinD population (P=0.038, OR=0.67 95% CI 0.46–0.98). There might be a genetic interaction between IGF2BP2 and IGF2 (P=0.05). The Igf2bp2 gene expression levels were increased in the kidneys of db/db mice compared to controls at the age of 5weeks but not at 26weeks. ConclusionsThe present study has replicated the association of IGF2BP2 rs4402960 with T2D in the Czech population and provided data suggesting that IGF2BP2 may have genetic interaction with IGF2 with a protective effect against DN in male patients with T1D.

HLA-DQB1* alleles and genetic susceptibility to type 1 diabetes mellitus

To determine human leukocyte antigen (HLA)-DQB1 allele association with susceptibility to type 1 diabetes (T1D) and to clinical and laboratory findings. This study was conducted on 85 unrelated Egyptian children with T1D recruited consecutively from the Pediatric Diabetes Endocrinology outpatients Clinic; Mansoura University Children's Hospital, Egypt. Patient mean follow up period was 2.5 years. Patients were subdivided according to level of HbA1c (optimal/suboptimal control < 8.5% and poor control ≥ 8.5%). The control group consisted of 113 unrelated age- and sex-matched healthy subjects without T1D or other autoimmune diseases. Genomic DNA extraction was done for all subjects using a DNA isolation kit. HLA-Class II-DQB1 allele typing was carried out with a polymerase chain reaction-sequence-specific oligonucleotide probe using a INNO-LiPA HLA-DQB1 update kit. Significant differences were detected between Egyptian patients with T1D and control groups in the frequencies of DQB1*02 [44.4% vs 18.6%, corrected P value (Pc) < 0.001] and DQB1*03 (41.2% vs 24.4%, Pc < 0.001). Significant differences were also observed between control groups and T1D patients in the frequencies of DQB1*05 (14.6% vs 7.2%, P = 0.029) and DQB1*06 (34.1% vs 7.2%, P < 0.001). However, after correction for multiple comparisons, the significance was retained for HLA-DQB1*06 (Pc < 0.001) but lost for HLA-DQB1*05. HLA-DQB1*0201, *0202, *030201 were positively associated with T1D (Pc = 0.014, Pc < 0.001, and Pc < 0.001 respectively), while HLA-DQB1*060101 was negatively associated (Pc < 0.001) with the condition. Although the HLA-DQB1 alleles 030101 and 050101 were significantly higher in controls (P = 0.016, P = 0.025 respectively), both of them lost statistical significance after correction of P value. The frequency of the HLA-DQB1 genotypes 02/02, 02/03, and 03/03 was higher in T1D patients, and the frequency of the genotypes 03/06, 05/06, and 06/06 was higher in controls, these differences being statistically significant before correction. After correction, the genotypes 02/02, 02/03 in T1D, and the genotypes 03/06, 06/06 in controls were still significant (Pc = 0.01, Pc < 0.001, Pc < 0.001, and Pc = 0.04, respectively). Non-significant associations were found between the frequency HLA-DQB1 alleles and genotypes in T1D in relation to the grade of diabetic control, Microalbuminuria, age, gender, age of presentation, weight, height, frequency of diabetic ketoacidosis (P = 0.42), serum cholesterol, and fasting and post-prandial level of C-peptide (P = 0.83, P = 0.9, respectively). The Current work suggests that HLA-DQB1 alleles *030201, *0202, *0201, and genotypes 02/03, 02/02 may be susceptibility risk factors for development of T1D in Egyptian children, while the HLA-DQB1*060101 allele, and 03/06, 06/06 genotypes may be protective factors. HLA-DQB1 alleles and genotypes do not contribute to microalbuminuria or grade of diabetic control.