Multicentre prospective validation of a urinary peptidome-based classifier for the diagnosis of type 2 diabetic nephropathy

Background Diabetic nephropathy (DN) is one of the major late complications of diabetes. Treatment aimed at slowing down the progression of DN is available but methods for early and definitive detection of DN progression are currently lacking. The ‘Proteomic prediction and Renin angiotensin aldosterone system Inhibition prevention Of early diabetic nephRopathy In TYpe 2 diabetic patients with normoalbuminuria trial' (PRIORITY) aims to evaluate the early detection of DN in patients with type 2 diabetes (T2D) using a urinary proteome-based classifier (CKD273). Methods In this ancillary study of the recently initiated PRIORITY trial we aimed to validate for the first time the CKD273 classifier in a multicentre (9 different institutions providing samples from 165 T2D patients) prospective setting. In addition we also investigated the influence of sample containers, age and gender on the CKD273 classifier. Results We observed a high consistency of the CKD273 classification scores across the different centres with areas under the curves ranging from 0.95 to 1.00. The classifier was independent of age (range tested 16-89 years) and gender. Furthermore, the use of different urine storage containers did not affect the classification scores. Analysis of the distribution of the individual peptides of the classifier over the nine different centres showed that fragments of blood-derived and extracellular matrix proteins were the most consistently found. Conclusion We provide for the first time validation of this urinary proteome-based classifier in a multicentre prospective setting and show the suitability of the CKD273 classifier to be used in the PRIORITY tria

Plasma proteome analysis of patients with type 1 diabetes with diabetic nephropathy

As part of a clinical proteomics program focused on diabetes and its complications we are looking for new and better protein biomarkers for diabetic nephropathy. The search for new and better biomarkers for diabetic nephropathy has, with a few exceptions, previously focused on either hypothesis-driven studies or urinary based investigations. To date only two studies have investigated the proteome of blood in search for new biomarkers, and these studies were conducted in sera from patients with type 2 diabetes. This is the first reported in depth proteomic study where plasma from type 1 diabetic patients was investigated with the goal of finding improved candidate biomarkers to predict diabetic nephropathy. In order to reach lower concentration proteins in plasma a pre-fractionation step, either hexapeptide bead-based libraries or anion exchange chromatography, was performed prior to surface enhanced laser desorption/ionization time-of-flight mass spectrometry analysis

New genetic loci link adipose and insulin biology to body fat distribution.

Body fat distribution is a heritable trait and a well-established predictor of adverse metabolic outcomes, independent of overall adiposity. To increase our understanding of the genetic basis of body fat distribution and its molecular links to cardiometabolic traits, here we conduct genome-wide association meta-analyses of traits related to waist and hip circumferences in up to 224,459 individuals. We identify 49 loci (33 new) associated with waist-to-hip ratio adjusted for body mass index (BMI), and an additional 19 loci newly associated with related waist and hip circumference measures (P < 5 × 10(-8)). In total, 20 of the 49 waist-to-hip ratio adjusted for BMI loci show significant sexual dimorphism, 19 of which display a stronger effect in women. The identified loci were enriched for genes expressed in adipose tissue and for putative regulatory elements in adipocytes. Pathway analyses implicated adipogenesis, angiogenesis, transcriptional regulation and insulin resistance as processes affecting fat distribution, providing insight into potential pathophysiological mechanisms

A Genome-Wide Association Study of Diabetic Kidney Disease in Subjects With Type 2 Diabetes

Identification of sequence variants robustly associated with predisposition to diabetic kidney disease (DKD) has the potential to provide insights into the pathophysiological mechanisms responsible. We conducted a genome-wide association study (GWAS) of DKD in type 2 diabetes (T2D) using eight complementary dichotomous and quantitative DKD phenotypes: the principal dichotomous analysis involved 5,717 T2D subjects, 3,345 with DKD. Promising association signals were evaluated in up to 26,827 subjects with T2D (12,710 with DKD). A combined (T1D+T2D) GWAS was performed using complementary data available for subjects with T1D, which, with replication samples, involved up to 40,340 diabetic subjects (and 18,582 DKD cases). Analysis of specific DKD phenotypes identified a novel signal near GABRR1 (rs9942471, p=4.5×10-8) associated with 'microalbuminuria' in European T2D cases. However, no replication of this signal was observed in Asian subjects with T2D, or in the equivalent T1D analysis. There was only limited support, in this substantially enlarged analysis, for association at previously-reported DKD signals, except for those at UMOD and PRKAG2, both associated with 'EGFR'. We conclude that, despite challenges in addressing phenotypic heterogeneity, access to increased sample sizes will continue to provide more robust inference regarding risk-variant discovery for DKD.</p

New susceptibility loci associated with kidney disease in type 1 diabetes

WOS:000309817900008Diabetic kidney disease, or diabetic nephropathy (DN), is a major complication of diabetes and the leading cause of end-stage renal disease (ESRD) that requires dialysis treatment or kidney transplantation. In addition to the decrease in the quality of life, DN accounts for a large proportion of the excess mortality associated with type 1 diabetes (T1D). Whereas the degree of glycemia plays a pivotal role in DN, a subset of individuals with poorly controlled T1D do not develop DN. Furthermore, strong familial aggregation supports genetic susceptibility to DN. However, the genes and the molecular mechanisms behind the disease remain poorly understood, and current therapeutic strategies rarely result in reversal of DN. In the GEnetics of Nephropathy: an International Effort (GENIE) consortium, we have undertaken a meta-analysis of genome-wide association studies (GWAS) of T1D DN comprising ∼2.4 million single nucleotide polymorphisms (SNPs) imputed in 6,691 individuals. After additional genotyping of 41 top ranked SNPs representing 24 independent signals in 5,873 individuals, combined meta-analysis revealed association of two SNPs with ESRD: rs7583877 in the AFF3 gene (P = 1.2×10(-8)) and an intergenic SNP on chromosome 15q26 between the genes RGMA and MCTP2, rs12437854 (P = 2.0×10(-9)). Functional data suggest that AFF3 influences renal tubule fibrosis via the transforming growth factor-beta (TGF-β1) pathway. The strongest association with DN as a primary phenotype was seen for an intronic SNP in the ERBB4 gene (rs7588550, P = 2.1×10(-7)), a gene with type 2 diabetes DN differential expression and in the same intron as a variant with cis-eQTL expression of ERBB4. All these detected associations represent new signals in the pathogenesis of DN.Peer reviewe

Exome sequencing-driven discovery of coding polymorphisms associated with common metabolic phenotypes

This article is published with open access at Springerlink.comThis project was funded by the Lundbeck Foundation and produced by The Lundbeck Foundation Centre for Applied Medical Genomics in Personalised Disease Prediction, Prevention and Care (LuCamp, www.lucamp.org). The Novo Nordisk Foundation Center for Basic Metabolic Research is an independent Research Center at the University of Copenhagen partially funded by an unrestricted donation from the Novo Nordisk Foundation (www.metabol.ku.dk). Further funding came from the Danish Council for Independent Research (Medical Sciences). The Inter99 was initiated by Torben Jørgensen (PI), Knut BorchJohnsen (co-PI), Hans Ibsen and Troels F. Thomsen. The steering committee comprises the former two and Charlotta Pisinger. The study was financially supported by research grants from the Danish Research Council, the Danish Centre for Health Technology Assessment, Novo Nordisk Inc., Research Foundation of Copenhagen County, Ministry of Internal Affairs and Health, the Danish Heart Foundation, the Danish Pharmaceutical Association, the Augustinus Foundation, the Ib Henriksen Foundation, the Becket Foundation, and the Danish Diabetes Association. The Health2006 was initiated by Allan Linneberg (PI) and Torben Jørgensen (co-PI). The study was financially supported by grants from the Velux Foundation, The Danish Medical Research Council, Danish Agency for Science, Technology and Innovation, The Aase and Ejner Danielsens Foundation, ALK-Abelló, (Hørsholm, Denmark) and Research Centre for Prevention and Health (the Capital Region of Denmark). In Finland this work has been supported by the following grants to M. Laakso: Academy of Finland, the Finnish Diabetes Research Foundation, the Finnish Cardiovascular Research Foundation, and EVO grant from the Kuopio University Hospital (5263). In the UK the Collection of the UK type 2 diabetes cases was supported by Diabetes UK, BDA Research and the UK Medical Research Council (Biomedical Collections Strategic Grant G0000649). The UK Type 2 Diabetes Genetics Consortium collection was supported by the Wellcome Trust (Biomedical Collections Grant GR072960). We acknowledge use of DNA from The UK Blood Services collection of Common Controls (UKBS-CC collection), funded by the Wellcome Trust grant 076113/C/04/Z and by NIHR programme grant to NHSBT (RP-PG-0310-1002). The collection was established as part of the Wellcome Trust Case Control Consortium (WTCCC). For the 1958 Birth Cohort, venous blood collection was funded by the Medical Research Council grant G0000934 (awarded under the Health of the Public initiative), peripheral blood lymphocyte preparation by Juvenile Diabetes Research Foundation/Wellcome Trust and the cell-line production, DNA extraction and processing by the Wellcome Trust grant 06854/Z/02/Z. The genotyping was supported by the Wellcome Trust (083270) and EU (ENGAGE: HEALTH-F4-2007- 201413). A. P. Morris is a Wellcome Trust Senior Fellow (081682/Z/06/Z) and M. McCarthy receives funding from the Oxford NIHR Biomedical Research Centre. We acknowledge the contribution of M. Sampson. In the Netherlands the work in this study was financially supported by the Dutch Diabetes Research Foundation grant 2006.00.060 and Biobanking and Biomolecular Research Infrastructure the Netherlands (BBMRI-NL). The D.E.S.I.R. cohort was supported by co-operative contracts between Inserm, CNAMTS, Novartis, Lilly and sanofi-aventis, by Inserm (Réseaux en Santé Publique, Interactions entre les determinants de la santé, Cohortes Santé TGIR 2008), by the Association Diabète Risque Vasculaire, the Fédération Française de Cardiologie, La Fondation de France, ALFEDIAM, ONIVINS, Société Francophone du Diabète; Ardix Medical, Bayer Diagnostics, Becton Dickinson, Cardionics, Lilly, Merck Santé, Novo Nordisk, Pierre Fabre, Roche, sanofi-aventis, Topcon. Work in Sweden was supported by grants from the Swedish Research Foundation (Dnr-349-2006-6589, 2009-1039, 521-2010-3490) and Knut & Alice Wallenberg Foundation. Work in Finland was supported by grants from the Sigrid Juselius Foundation, Folkhälsan Research Foundation and the Finnish Medical Society. The Gene-Lifestyle interactions And Complex traits Involved in Elevated Disease Risk (GLACIER) study is nested within the Northern Swedish Health and Disease Study cohort and the Västerbotten Intervention Programme (VIP). The research programme was approved by the Ethical Review Board in Umeå, Sweden. We are indebted to the study participants who dedicated their time and samples to these studies. We also thank the VIP and Umeå Medical Biobank staff for biomedical data collection and preparation. We specifically thank Å. Ågren (Umeå Medical Biobank) for data organisation, and K. Enqvist and T. Johansson (Västerbottens County Council) for expert technical assistance with DNA preparation. The GLACIER Study was funded by project grants from Novo Nordisk (P. W. Franks [PWF]), the Swedish Heart-Lung Foundation (PWF), the Swedish Diabetes Association (to PWF), Påhlssons Foundation (PWF), the Swedish Research Council (PWF), Umeå University Career Development Award (PWF) and The Heart Foundation of Northern Sweden (PWF)

The endothelial nitric oxide synthase gene and risk of diabetic nephropathy and development of cardiovascular disease in type 1 diabetes

Nitric oxide (NO) is important in the maintenance of vascular tone and regulation of blood pressure. NO may also play a role in the development of both nephropathy and cardiovascular disease (CVD) in patients with diabetes. The susceptibility to nephropathy and CVD depends to some extent on genetic factors, therefore polymorphisms in the gene coding for endothelial NO-synthase, NOS3, can affect the risk of developing these diseases. Type 1 diabetes patients attending the Steno Diabetes Center, Denmark, between 1993 and 2001 were enrolled in this study. A total of 458 cases with diabetic nephropathy (albumin excretion &gt;300 mg/24h) and 319 controls with persistent normoalbuminuria (&lt;30 mg/24h), despite &gt; or =20 years of diabetes duration at follow-up were identified. Patients were followed until death or end of the study. Associations between seven NOS3-gene polymorphisms and nephropathy, progression of nephropathy and CVD were studied. There was significant association between the rs743507 TT-genotype and diabetic nephropathy. When including age at diabetes onset, diabetes duration at follow-up, baseline Hb(A1c), sex and ever smoking in the analysis the OR was 1.43 (95% CI=1.03-2.00), P=0.035. In analyses of CVD development using Cox-regression the rs1799983 GG-genotype was a significant protective factor in normoalbuminuric patients, HR=0.32 (0.12-0.82), P=0.018, but not in patients with macroalbuminuria (covariates were; age at follow-up, baseline Hb(A1c), baseline systolic blood pressure, baseline cholesterol, sex and ever smoking). Our conclusion is that the NOS3-gene may be involved in the development of diabetic nephropathy in patients with type 1 diabetes and can be predictive of CVD during follow-up.</p