Serological evaluation of possible exposure to Ljungan virus and related parechovirus in autoimmune (type 1) diabetes in children.

Exposure to Ljungan virus (LV) is implicated in the risk of autoimmune (type 1) diabetes but possible contribution by other parechoviruses is not ruled out. The aim was to compare children diagnosed with type 1 diabetes in 2005-2011 (n = 69) with healthy controls (n = 294), all from the Jämtland County in Sweden, using an exploratory suspension multiplex immunoassay for IgM and IgG against 26 peptides of LV, human parechoviruses (HPeV), Aichi virus and poliovirus in relation to a radiobinding assay (RBA) for antibodies against LV and InfluenzaA/H1N1pdm09. Islet autoantibodies and HLA-DQ genotypes were also determined. 1) All five LV-peptide antibodies correlated to each other (P < 0.001) in the suspension multiplex IgM- and IgG-antibody assay; 2) The LV-VP1_31-60-IgG correlated with insulin autoantibodies alone (P = 0.007) and in combination with HLA-DQ8 overall (P = 0.022) as well as with HLA-DQ 8/8 and 8/X subjects (P = 0.013); 3) RBA detected LV antibodies correlated with young age at diagnosis (P < 0.001) and with insulin autoantibodies (P < 0.001) especially in young HLA-DQ8 subjects (P = 0.004); 4) LV-peptide-VP1_31-60-IgG correlated to RBA LV antibodies (P = 0.009); 5) HPeV3-peptide-IgM and -IgG showed inter-peptide correlations (P < 0.001) but only HPeV3-VP1_1-30-IgG (P < 0.001) and VP1_95-124-IgG (P = 0.009) were related to RBA LV antibodies without relation to insulin autoantibody positivity (P = 0.072 and P = 0.486, respectively). Both exploratory suspension multiplex IgG to LV-peptide VP1_31-60 and RBA detected LV antibodies correlated with insulin autoantibodies and HLA-DQ8 suggesting possible role in type 1 diabetes. It remains to be determined if cross-reactivity or concomitant exposure to LV and HPeV3 contributes to the seroprevalence. J. Med. Virol. © 2015 Wiley Periodicals, Inc

Islet autoantibodies and residual beta cell function in type 1 diabetes children followed for 3-6 years

Aims: To test if islet autoantibodies at diagnosis of type 1 diabetes (T1DM) and after 3-6 years with T1D predict residual beta-cell function (RBF) after 3-6 years with T1D. Methods: T1D children (n = 260, median age at diagnosis 9.4, range 0.9-14.7 years) were tested for GAD65, IA-2, ZnT8R, ZnT8W and ZnT8Q autoantibodies (A) at diagnosis, and 3-6 years after diagnosis when also fasting and stimulated RBF were determined. Results: For every 1-year increase in age at diagnosis of TID, the odds of detectable C-peptide increased 1.21 (1.09, 1.34) times for fasting C-peptide and 1.28 (1.15, 1.42) times for stimulated C-peptide. Based on a linear model for subjects with no change in IA-2A levels, the odds of detectable C-peptide were 35% higher than for subjects whose IA-2A levels decreased by half (OR = 1.35 (1.09, 1.67), p = 0.006); similarly for ZnT8WA (OR = 1.39 (1.09, 1.77), p = 0.008) and ZnT8QA (OR = 1.55 (1.06, 2.26) p = 0.024). Such relationship was not detected for GADA or ZnT8RA. All OR adjusted for confounders. Conclusions: Age at diagnosis with T1D was the major predictor of detectable C-peptide 3-6 years post-diagnosis. Decreases in IA-2A, and possibly ZnT8A, levels between diagnosis and post-diagnosis were associated with a reduction in RBF post-diagnosis. (C) 2012 Elsevier Ireland Ltd. All rights reserved

Relationship Between Ljungan Virus Antibodies, HLA-DQ8, and Insulin Autoantibodies in Newly Diagnosed Type 1 Diabetes Children

Environmental factors, including viral infections, may explain an increasing and fluctuating incidence of childhood type 1 diabetes (T1D). Ljungan virus (LV) isolated from bank voles have been implicated, but it is unclear whether LV contributes to islet autoimmunity, progression to clinical onset, or both, of T1D. The aim was to test whether LV antibodies (LVAb) were related to HLA-DQ and islet autoantibodies in newly diagnosed T1D patients (n = 676) and controls (n = 309). Patients, 0-18 years of age, diagnosed with T1D in 1996-2005 were analyzed for LVAb, HLA-DQ genotypes, and all seven known islet autoantibodies (GADA, IA-2A, IAA, ICA, ZnT8RA, ZnT8WA, and ZnT8QA). LVAb at 75th percentile, defined as cut off, was 90 (range 6-3936) U/mL and 4th quartile LVAb were found in 25% (170/676) of which 64% were < 10 (n = 108, p < 0.0001), and 27% were < 5 (n = 45; p < 0.0001) years old. The 4th quartile LVAb in children < 10 years of age correlated to HLA DQ2/8, 8/8, and 8/X (p < 0.0001). Furthermore, in the group with 4th quartile LVAb, 55% were IAA positive (p = 0.01) and correlation was found between 4th quartile LVAb and IAA in children < 10 years of age (p = 0.035). It is concluded that 1) LVAb were common among the young T1D patients and LVAb levels were higher in the younger age groups; 2) 4th quartile LVAb correlated with IAA; and 3) there was a correlation between 4th quartile LVAb and HLA-DQ8, particularly in the young patients. The presence of LVAb supports the notion that prior exposure to LV may be associated with T1D

Autoimmunity against INS-IGF2 expressed in human pancreatic islets.

Insulin is a major autoantigen in islet autoimmunity and progression to type 1 diabetes. It has been suggested that the insulin B-chain may be critical to insulin autoimmunity in type 1 diabetes. INS-IGF2 consists of the preproinsulin signal peptide, the insulin B-chain and eight amino acids of the C-peptide in addition to 138 amino acids from the IGF2 gene. We aimed to determine 1) expression of INS-IGF2 in human pancreatic islets and 2) autoantibodies in newly diagnosed type 1 diabetes children and controls. INS-IGF2, expressed primarily in beta cells, showed higher levels of expression in islets from normal compared to donors with either type 2 diabetes (p=0.006) or high HbA1c levels (p<0.001). INS-IGF2 autoantibody levels were increased in newly diagnosed type 1 diabetes patients (n=304) compared to healthy controls (n=355; p<0.001). Displacement with cold insulin and INS-IGF2 revealed that more patients than controls had doubly reactive insulin-INS-IGF2 autoantibodies. These data suggest that INS-IGF2, which contains the preproinsulin signal peptide, the B-chain and eight amino acids of the C-peptide may be an autoantigen in type 1 diabetes. INS-IGF2 and insulin may share autoantibody binding sites, thus complicating the notion that insulin is the primary autoantigen in type 1 diabetes

Loss-of-function mutations in SLC30A8 protect against type 2 diabetes

Loss-of-function mutations protective against human disease provide in vivo validation of therapeutic targets1,2,3, yet none are described for type 2 diabetes (T2D). Through sequencing or genotyping ~150,000 individuals across five ethnicities, we identified 12 rare protein-truncating variants in SLC30A8, which encodes an islet zinc transporter (ZnT8)4 and harbors a common variant (p.Trp325Arg) associated with T2D risk, glucose, and proinsulin levels5–7. Collectively, protein-truncating variant carriers had 65% reduced T2D risk (p=1.7×10−6), and non-diabetic Icelandic carriers of a frameshift variant (p.Lys34SerfsX50) demonstrated reduced glucose levels (−0.17 s.d., p=4.6×10−4). The two most common protein-truncating variants (p.Arg138X and p.Lys34SerfsX50) individually associate with T2D protection and encode unstable ZnT8 proteins. Previous functional study of SLC30A8 suggested reduced zinc transport increases T2D risk8,9, yet phenotypic heterogeneity was observed in rodent Slc30a8 knockouts10–15. Contrastingly, loss-of-function mutations in humans provide strong evidence that SLC30A8 haploinsufficiency protects against T2D, proposing ZnT8 inhibition as a therapeutic strategy in T2D prevention

Loss-of-function mutations in SLC30A8 protect against type 2 diabetes.

Neðst á síðunni er hægt að nálgast greinina í heild sinni með því að smella á hlekkinn View/OpenLoss-of-function mutations protective against human disease provide in vivo validation of therapeutic targets, but none have yet been described for type 2 diabetes (T2D). Through sequencing or genotyping of ~150,000 individuals across 5 ancestry groups, we identified 12 rare protein-truncating variants in SLC30A8, which encodes an islet zinc transporter (ZnT8) and harbors a common variant (p.Trp325Arg) associated with T2D risk and glucose and proinsulin levels. Collectively, carriers of protein-truncating variants had 65% reduced T2D risk (P = 1.7 × 10(-6)), and non-diabetic Icelandic carriers of a frameshift variant (p.Lys34Serfs*50) demonstrated reduced glucose levels (-0.17 s.d., P = 4.6 × 10(-4)). The two most common protein-truncating variants (p.Arg138* and p.Lys34Serfs*50) individually associate with T2D protection and encode unstable ZnT8 proteins. Previous functional study of SLC30A8 suggested that reduced zinc transport increases T2D risk, and phenotypic heterogeneity was observed in mouse Slc30a8 knockouts. In contrast, loss-of-function mutations in humans provide strong evidence that SLC30A8 haploinsufficiency protects against T2D, suggesting ZnT8 inhibition as a therapeutic strategy in T2D prevention.US National Institutes of Health (NIH) Training 5-T32-GM007748-33 Doris Duke Charitable Foundation 2006087 Fulbright Diabetes UK Fellowship BDA 11/0004348 Broad Institute from Pfizer, Inc. NIH U01 DK085501 U01 DK085524 U01 DK085545 U01 DK085584 Swedish Research Council Dnr 521-2010-3490 Dnr 349-2006-237 European Research Council (ERC) GENETARGET T2D GA269045 ENGAGE 2007-201413 CEED3 2008-223211 Sigrid Juselius Foundation Folkh lsan Research Foundation ERC AdG 293574 Research Council of Norway 197064/V50 KG Jebsen Foundation University of Bergen Western Norway Health Authority Lundbeck Foundation Novo Nordisk Foundation Wellcome Trust WT098017 WT064890 WT090532 WT090367 WT098381 Uppsala University Swedish Research Council and the Swedish Heart- Lung Foundation Academy of Finland 124243 102318 123885 139635 Finnish Heart Foundation Finnish Diabetes Foundation, Tekes 1510/31/06 Commission of the European Community HEALTH-F2-2007-201681 Ministry of Education and Culture of Finland European Commission Framework Programme 6 Integrated Project LSHM-CT-2004-005272 City of Kuopio and Social Insurance Institution of Finland Finnish Foundation for Cardiovascular Disease NIH/NIDDK U01-DK085545 National Heart, Lung, and Blood Institute (NHLBI) National Institute on Minority Health and Health Disparities N01 HC-95170 N01 HC-95171 N01 HC-95172 European Union Seventh Framework Programme, DIAPREPP Swedish Child Diabetes Foundation (Barndiabetesfonden) 5U01DK085526 DK088389 U54HG003067 R01DK072193 R01DK062370 Z01HG000024info:eu-repo/grantAgreement/EC/FP7/20201

Studies of the role of complement factor H in hemolytic uremic syndrome

Factor H is the main fluid phase regulator of the alternative pathway of complement. Factor H acts as a co-factor for factor I-mediated C3b degradation, inhibits the formation of the C3bBb convertase and accelerates its decay. By discriminating between host and foreign cells, factor H inhibits complement-mediated injury to host cells. Factor H mutations have been associated with atypical hemolytic uremic syndrome (aHUS) a condition characterized by non-immune hemolytic anemia, thrombocytopenia, and acute renal failure. The mechanism by which factor H mutations lead to aHUS is unclear. The purpose of these studies was therefore to examine the interaction of normal and mutated factor H with platelets and endothelial cells as well as the phenotypic expression of factor H in aHUS patients. Factor H bound to washed human platelets in a dose-dependent manner mainly via the C terminal of the protein and the heparin-binding sites. On platelets, factor H bound via the GPIIb/IIIa receptor as well as thrombospondin. Mutated factor H exhibited less binding, a finding that was verified using mutated factor H purified from the serum of a patient with aHUS. The same patient was found to have activation of the alternative complement pathway on platelets demonstrated by the presence of C3 and C5b-9 on the cell surface. Using the patient's serum we showed that the mutated factor H had reduced ability to protect normal platelets from complement activation. The phenotypic expression of factor H mutations was studied in two other patients in whom we demonstrated that the protein either accumulated in cells or exhibited reduced binding to host cells. Both these mechanisms could result in complement activation on host cells (endothelium and platelets), which would in turn promote endothelial cell injury as well as platelet activation and consumption in thrombi

Osteoprotegerin autoantibodies do not predict low bone mineral density in middle-aged women

Purpose Autoantibodies against osteoprotegerin (OPG) have been associated with osteoporosis. The aim was to develop an immunoassay for OPG autoantibodies and test their diagnostic usefulness of identifying women general population with low bone mineral density. Methods Included were 698 women at mean age 55.1 years (range 50.4–60.6) randomly selected from the general population. Measurement of wrist bone mineral density (g/cm2) was performed of the non-dominant wrist by dual-energy X-ray absorptiometry (DXA). A T-score < − 2.5 was defined as having a low bone mineral density. Measurements of OPG autoantibodies were carried by radiobinding assays. Cut-off levels for a positive value were determined from the deviation from normality in the distribution of 398 healthy blood donors representing the 99.7th percentile. Results Forty-five of the 698 (6.6%) women were IgG-OPG positive compared with 2 of 398 (0.5%) controls (p < 0.0001) and 35 of the 698 (5.0%) women had a T-score < − 2.5. There was no difference in bone mineral density between IgG-OPG positive (median 0.439 (range 0.315–0.547) g/cm2) women and IgG-OPG negative (median 0.435 (range 0.176–0.652) g/cm2) women (p = 0.3956). Furthermore, there was neither a correlation between IgG-OPG levels and bone mineral density (rs = 0.1896; p = 0.2068) nor T-score (rs = 0.1889; p = 0.2086). Diagnostic sensitivity and specificity of IgG-OPG for low bone mineral density were 5.7% and 92.9%, and positive and negative predictive values were 7.4% and 90.8%, respectively. Conclusion Elevated OPG autoantibody levels do not predict low bone mineral density in middle-aged women selected from the general population

Improved efficacy by using the pTnT-rhtTG plasmid for the detection of celiac disease specific tissue transglutaminase autoantibodies in radioligand binding assays.

Background. Tissue transglutaminase (tTG) autoantibodies are serological markers for celiac disease. The aim was to study the efficacy of the pTnT-rhtTG plasmid and subsequent diagnostic accuracy of tTG autoantibodies for childhood celiac disease using radioligand binding assays. Methods. Coupled in vitro transcription and translation of tTG were performed by pTnT-rhtTG as well as by the pGEMt Easy-rhtTG vectors using the TNT SP6 Coupled Reticulocyte Lysate System in the presence of [(35)S] methionine. Sera from 190 celiac disease children and 74 controls were measured for tTG autoantibodies in two separate radioligand binding assays using anti-human IgA agarose and protein A sepharose beads for the detection of IgA-tTG and IgG-tTG, respectively. Results. Median incorporation of [(35)S] methionine into the pTnT-rhtTG was 26% compared to 16% for the pGEMt Easy-rhtTG plasmid (p = 0.0016). Using pTnT-rhtTG (as compared to pGEMt Easy-rhtTG), sensitivities were IgA-tTG = 96.3% (95.7%) and IgG-tTG = 95.8% (97.3%) and specificities were IgA-tTG = 91.9% (90.5%) and IgG-tTG = 94.6% (98.4%). According to receiver operator characteristics for the pTnT (pGEMt Easy) assays, area under the curves were IgA-tTG = 98.4% (98.4%) and IgG-tTG = 97.7% (97.2%), respectively. Conclusion. The pTnT-rhtTG plasmid increased the efficacy of tTG antigen usage without reducing the diagnostic accuracy of IgA-tTG and IgG-tTG for childhood celiac disease. The pTnT-rhtTG plasmid is therefore recommended over the pGEMt Easy-rhtTG for the assessment of IgA-tTG and IgG-tTG using radioligand binding assays

Neuropeptide Y autoantibodies in patients with long-term type 1 and type 2 diabetes and neuropathy.

Aims: The neurotransmitter Neuropeptide Y (NPY) was previously reported as a minor autoantigen in newly diagnosed type 1 diabetes (T1D) patients. The single nucleotide polymorphism at rs16139 (T1128C, L7P) in the NPY gene was associated with an increased risk for the development of type 2 diabetes (T2D). We aimed to develop a radiobinding assay for NPY-L (Leucine) and NPY-P (Proline) autoantibodies (A) to study the levels and the association with other islet autoantibodies and neuropathy. Methods: Autoantibodies against NPY-L, NPY-P, ZnT8, GAD65 and IA-2 were studied in T1D (n=48) and T2D (n=26) patients with duration up to 42 and 31years. A subgroup of T1D (n=32) patients re-examined, 5-8years after first visit, was tested for peripheral (Z-score) and autonomic neuropathy (E/I ratio). Results: NPY-LA and NPY-PA were detected in 23% and 19% in T1D (p<0.001), and 12% and 23% in T2D patients (p<0.001) compared to 2.5% controls (n=398). The levels of NPYA declined during follow-up in the T1D patients (p<0.001). The neuropathy was not related to the NPYA or the other islet autoantibodies. Conclusions: Regardless of the absence of an association between NPYA and neuropathy, NPY may contribute to the pathogenesis of T1D and T2D as a minor autoantigen