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Abstract Quantifying the contribution of genetics and environmental effects on disease initiation and progression, as well as the shared genetics of different diseases, is vital for the understanding of the disease etiology of multimorbidities. In this study, we leverage nationwide Danish registries to provide a granular atlas of the genetic origin of disease phenotypes for a cohort of all Danes 1978–2018 with partially known pedigree (n = 6.3 million). We estimate the heritability and genetic correlation between thousands of disease phenotypes using a novel approach that can be scaled to nationwide data. Our findings confirm the importance of genetics for a number of known associations and increase the resolution of heritability by adding numerous associations, some of which point to shared biologically origin of different phenotypes. We also establish the heritability of disease trajectories and the importance of sex-specific genetic contributions. Results can be accessed at https://h2.cpr.ku.dk/ .

Abstract Headache disorders are the most common disorders of the nervous system. The lifetime prevalence of headache disorders show that some individuals never experience headache. The etiology of complete freedom from headache is not known. To assess genetic variants associated with complete freedom from headache, we performed a genome-wide association study of individuals who have never experienced a headache. We included 63,992 individuals (2,998 individuals with complete freedom from headache and 60,994 controls) from the Danish Blood Donor Study Genomic Cohort. Participants were included in two rounds, from 2015 to 2018 and in 2020. We discovered a genome-wide significant association, with the lead variant rs7904615[G] in ADARB2 (EAF = 27%, OR = 1.20 [1.13–1.27], p = 3.92 × 10−9). The genomic locus was replicated in a non-overlapping cohort of 13,032 individuals (539 individuals with complete freedom from headache and 12,493 controls) from the Danish Blood Donor Study Genomic Cohort (p

Abstract Essential tremor (ET) is a prevalent neurological disorder with a largely unknown underlying biology. In this genome-wide association study meta-analysis, comprising 16,480 ET cases and 1,936,173 controls from seven datasets, we identify 12 sequence variants at 11 loci. Evaluating mRNA expression, splicing, plasma protein levels, and coding effects, we highlight seven putative causal genes at these loci, including CA3 and CPLX1. CA3 encodes Carbonic Anhydrase III and carbonic anhydrase inhibitors have been shown to decrease tremors. CPLX1, encoding Complexin-1, regulates neurotransmitter release. Through gene-set enrichment analysis, we identify a significant association with specific cell types, including dopaminergic and GABAergic neurons, as well as biological processes like Rho GTPase signaling. Genetic correlation analyses reveals a positive association between ET and Parkinson’s disease, depression, and anxiety-related phenotypes. This research uncovers risk loci, enhancing our knowledge of the complex genetics of this common but poorly understood disorder, and highlights CA3 and CPLX1 as potential therapeutic targets.

Abstract Background The emerging use of biomarkers in research and tailored care introduces a need for information about the association between biomarkers and basic demographics and lifestyle factors revealing expectable concentrations in healthy individuals while considering general demographic differences. Methods A selection of 47 biomarkers, including markers of inflammation and vascular stress, were measured in plasma samples from 9876 Danish Blood Donor Study participants. Using regression models, we examined the association between biomarkers and sex, age, Body Mass Index (BMI), and smoking. Results Here we show that concentrations of inflammation and vascular stress biomarkers generally increase with higher age, BMI, and smoking. Sex-specific effects are observed for multiple biomarkers. Conclusion This study provides comprehensive information on concentrations of 47 plasma biomarkers in healthy individuals. The study emphasizes that knowledge about biomarker concentrations in healthy individuals is critical for improved understanding of disease pathology and for tailored care and decision support tools.

Abstract Social trust is a heritable trait that has been linked with physical health and longevity. In this study, we performed genome-wide association studies of self-reported social trust in n = 33,882 Danish blood donors. We observed genome-wide and local evidence of genetic similarity with other brain-related phenotypes and estimated the single nucleotide polymorphism-based heritability of trust to be 6% (95% confidence interval = (2.1, 9.9)). In our discovery cohort (n = 25,819), we identified one significantly associated locus (lead variant: rs12776883) in an intronic enhancer region of PLPP4, a gene highly expressed in brain, kidneys, and testes. However, we could not replicate the signal in an independent set of donors who were phenotyped a year later (n = 8063). In the subsequent meta-analysis, we found a second significantly associated variant (rs71543507) in an intergenic enhancer region. Overall, our work confirms that social trust is heritable, and provides an initial look into the genetic factors that influence it.

IntroductionDiabetes is associated with dysregulated immune function and impaired cytokine release, while transient acute hyperglycaemia has been shown to enhance inflammatory cytokine release in preclinical studies. Although diabetes and acute hyperglycaemia are common among patients with community-acquired pneumonia (CAP), the impact of chronic, acute, and acute-on-chronic hyperglycaemia on the host response within this population remains poorly understood. This study investigated whether chronic, acute, and acute-on- chronic hyperglycaemia are associated with distinct mediators of inflammatory, endothelial, and angiogenic host response pathways in patients with CAP.MethodsIn a cross-sectional study of 555 patients with CAP, HbA1c, admission plasma (p)-glucose, and the glycaemic gap (admission p-glucose minus HbA1c- derived average p-glucose) were employed as measures of chronic, acute, and acute-on-chronic hyperglycaemia, respectively. Linear regression was used to model the associations between the hyperglycaemia measures and 47 proteins involved in inflammation, endothelial activation, and angiogenesis measured at admission. The models were adjusted for age, sex, CAP severity, pathogen, immunosuppression, comorbidity, and body mass index. Adjustments for multiple testing were performed with a false discovery rate threshold of less than 0.05.ResultsThe analyses showed that HbA1c levels were positively associated with IL-8, IL-15, IL-17A/F, IL-1RA, sFlt-1, and VEGF-C. Admission plasma glucose was also positively associated with these proteins and GM-CSF. The glycaemic gap was positively associated with IL-8, IL-15, IL-17A/F, IL-2, and VEGF-C.ConclusionIn conclusion, chronic, acute, and acute-on-chronic hyperglycaemia were positively associated with similar host response mediators. Furthermore, acute and acute-on-chronic hyperglycaemia had unique associations with the inflammatory pathways involving GM-CSF and IL-2, respectively.

Abstract Genotypes causing pregnancy loss and perinatal mortality are depleted among living individuals and are therefore difficult to find. To explore genetic causes of recessive lethality, we searched for sequence variants with deficit of homozygosity among 1.52 million individuals from six European populations. In this study, we identified 25 genes harboring protein-altering sequence variants with a strong deficit of homozygosity (10% or less of predicted homozygotes). Sequence variants in 12 of the genes cause Mendelian disease under a recessive mode of inheritance, two under a dominant mode, but variants in the remaining 11 have not been reported to cause disease. Sequence variants with a strong deficit of homozygosity are over-represented among genes essential for growth of human cell lines and genes orthologous to mouse genes known to affect viability. The function of these genes gives insight into the genetics of intrauterine lethality. We also identified 1077 genes with homozygous predicted loss-of-function genotypes not previously described, bringing the total set of genes completely knocked out in humans to 4785.

Abstract Objectives Allele counts of sequence variants obtained by whole genome sequencing (WGS) often play a central role in interpreting the results of genetic and genomic research. However, such variant counts are not readily available for individuals in the Danish population. Here, we present a dataset with allele counts for sequence variants (single nucleotide variants (SNVs) and indels) identified from WGS of 8,671 (5,418 females) individuals from the Danish population. The data resource is based on WGS data from three independent research projects aimed at assessing genetic risk factors for cardiovascular, psychiatric, and headache disorders. To enable the sharing of information on sequence variation in Danish individuals, we created summarized statistics on allele counts from anonymized data and made them available through the European Genome-phenome Archive (EGA, https://identifiers.org/ega.dataset:EGAD00001009756 ) and in a dedicated browser, DanMAC5 (available at www.danmac5.dk ). The summary level data and the DanMAC5 browser provide insight into the allelic spectrum of sequence variants segregating in the Danish population, which is important in variant interpretation. Data description Three WGS datasets with an average coverage of 30x were processed independently using the same quality control pipeline. Subsequently, we summarized, filtered, and merged allele counts to create a high-quality summary level dataset of sequence variants.

Abstract We identify biomarkers for disease progression in three type 2 diabetes cohorts encompassing 2,973 individuals across three molecular classes, metabolites, lipids and proteins. Homocitrulline, isoleucine and 2-aminoadipic acid, eight triacylglycerol species, and lowered sphingomyelin 42:2;2 levels are predictive of faster progression towards insulin requirement. Of ~1,300 proteins examined in two cohorts, levels of GDF15/MIC-1, IL-18Ra, CRELD1, NogoR, FAS, and ENPP7 are associated with faster progression, whilst SMAC/DIABLO, SPOCK1 and HEMK2 predict lower progression rates. In an external replication, proteins and lipids are associated with diabetes incidence and prevalence. NogoR/RTN4R injection improved glucose tolerance in high fat-fed male mice but impaired it in male db/db mice. High NogoR levels led to islet cell apoptosis, and IL-18R antagonised inflammatory IL-18 signalling towards nuclear factor kappa-B in vitro. This comprehensive, multi-disciplinary approach thus identifies biomarkers with potential prognostic utility, provides evidence for possible disease mechanisms, and identifies potential therapeutic avenues to slow diabetes progression.

Novel biomarkers are key to addressing the ongoing pandemic of type 2 diabetes mellitus. While new technologies have improved the potential of identifying such biomarkers, at the same time there is an increasing need for informed prioritization to ensure efficient downstream verification. We have built BALDR, an automated pipeline for biomarker comparison and prioritization in the context of diabetes. BALDR includes protein, gene, and disease data from major public repositories, text-mining data, and human and mouse experimental data from the IMI2 RHAPSODY consortium. These data are provided as easy-to-read figures and tables enabling direct comparison of up to 20 biomarker candidates for diabetes through the public website https://baldr.cpr.ku.dk.

Objectives To examine the level of loneliness experienced during the COVID-19 pandemic in Denmark and to identify associated behavioural patterns and demographic factors.Design Cross-sectional cohort study.Setting Includes Danish active and former blood donors.Participants A questionnaire was sent to 124 307 active and former blood donors, of these a total of 50 968 participants completed the study questionnaire (response rate=41%).Primary and secondary outcome measures Subjective experience of loneliness was measured using the 3-item University of California, Los Angeles Loneliness Scale (UCLA-3). Besides the UCLA-3, the respondents answered items on sociodemographic and economic characteristics, items on precautionary measures taken to avoid COVID-19 infection as well as on COVID-19 anxiety.Results The participants indicated their experienced level of loneliness both before and during the pandemic. Comparing the two reports yielded a mean increase in loneliness scores of 14.1% (p

Background: Whether natural selection may have attributed to the observed blood group frequency differences between populations remains debatable. The ABO system has been associated with several diseases and recently also with susceptibility to COVID-19 infection. Associative studies of the RhD system and diseases are sparser. A large disease-wide risk analysis may further elucidate the relationship between the ABO/RhD blood groups and disease incidence. Methods: We performed a systematic log-linear quasi-Poisson regression analysis of the ABO/RhD blood groups across 1,312 phecode diagnoses. Unlike prior studies, we determined the incidence rate ratio for each individual ABO blood group relative to all other ABO blood groups as opposed to using blood group O as the reference. Moreover, we used up to 41 years of nationwide Danish follow-up data, and a disease categorization scheme specifically developed for diagnosis-wide analysis. Further, we determined associations between the ABO/RhD blood groups and the age at the first diagnosis. Estimates were adjusted for multiple testing. Results: The retrospective cohort included 482,914 Danish patients (60.4% females). The incidence rate ratios (IRRs) of 101 phecodes were found statistically significant between the ABO blood groups, while the IRRs of 28 phecodes were found statistically significant for the RhD blood group. The associations included cancers and musculoskeletal-, genitourinary-, endocrinal-, infectious-, cardiovascular-, and gastrointestinal diseases. Conclusions: We found associations of disease-wide susceptibility differences between the blood groups of the ABO and RhD systems, including cancer of the tongue, monocytic leukemia, cervical cancer, osteoarthrosis, asthma, and HIV- and hepatitis B infection. We found marginal evidence of associations between the blood groups and the age at first diagnosis. Funding: Novo Nordisk Foundation and the Innovation Fund Denmark

The underlying genetics of carpal tunnel syndrome is not well understood. Here, the authors perform a GWAS meta-analysis for carpal tunnel syndrome finding variants at 50 loci with connections to the extracellular matrix discovered through various functional analyses.

Little is known about the biology of back pain, a leading cause of disability. Here the authors report 30 new back pain loci, implicating genes involved in cartilage/bone biology, as well as neurological and inflammatory processes.

Florian Uellendahl-Werth et al. conduct cross-tissue transcriptome-wide association studies to explore genetic mechanisms shared across immune-related and psychiatric traits. Their results identify several genes (including PPP3CA) that could mediate the interplay between psychiatric and inflammatory disease.

Abstract Restless Legs Syndrome (RLS) is a neurological sensorimotor disorder negatively impacting sufferers’ quality of sleep and health-related quality of life. The pathophysiology of RLS is poorly understood and research focusing on the link between RLS and inflammation has been limited. Our study aimed to investigate whether chronic inflammation markers C-reactive protein (CRP) and soluble urokinase-type plasminogen activator receptor (suPAR), as well plasma levels of five different cytokine-specific autoantibodies (c-aAb), i.e. modulators of inflammation, associate with RLS in otherwise healthy individuals. CRP, suPAR and c-aAb were measured in plasma samples of participants from the Danish Blood Donor Study in 2010. Returning donors between 2015 and 2018 completed the validated Cambridge-Hopkins RLS-questionnaire for RLS assessment, resulting in datasets with RLS assessment and values for CRP (N = 3564), suPAR (N = 2546) and c-aAb (N = 1478). We performed logistic regression models using the CRP, suPAR or c-aAb as the independent variable and RLS status as the dependent variable, adjusted for appropriate covariates. Our study indicates that a high concentration of CRP is associated with RLS, while an increased probability of experiencing frequent RLS symptoms in those with an elevated plasma suPAR level appears to be mediated through lifestyle factors. We additionally report that a high titer of autoantibodies specific against the cytokine interferon-alpha was associated with RLS. Our results support the existence of links between systemic inflammation and RLS, though further RLS studies on CRP, suPAR and c-aAb in larger cohorts are warranted to confirm our findings and further reveal the hitherto underexplored links between RLS and inflammation.

People with kidney disease are disproportionately affected by atherosclerosis for unclear reasons. Soluble urokinase plasminogen activator receptor (suPAR) is an immune-derived mediator of kidney disease, levels of which are strongly associated with cardiovascular outcomes. We assessed suPAR’s pathogenic involvement in atherosclerosis using epidemiologic, genetic, and experimental approaches. We found serum suPAR levels to be predictive of coronary artery calcification and cardiovascular events in 5,406 participants without known coronary disease. In a genome-wide association meta-analysis including over 25,000 individuals, we identified a missense variant in the plasminogen activator, urokinase receptor (PLAUR) gene (rs4760), confirmed experimentally to lead to higher suPAR levels. Mendelian randomization analysis in the UK Biobank using rs4760 indicated a causal association between genetically predicted suPAR levels and atherosclerotic phenotypes. In an experimental model of atherosclerosis, proprotein convertase subtilisin/kexin–9 (Pcsk9) transfection in mice overexpressing suPAR (suPARTg) led to substantially increased atherosclerotic plaques with necrotic cores and macrophage infiltration compared with those in WT mice, despite similar cholesterol levels. Prior to induction of atherosclerosis, aortas of suPARTg mice excreted higher levels of CCL2 and had higher monocyte counts compared with WT aortas. Aortic and circulating suPARTg monocytes exhibited a proinflammatory profile and enhanced chemotaxis. These findings characterize suPAR as a pathogenic factor for atherosclerosis acting at least partially through modulation of monocyte function.

Dowsett and colleagues used a genome-wide association approach to investigate the genetic influence on soluble urokinase-type plasminogen activator receptor presence in the plasma of humans. Their findings indicate a 60% heritability factor in British twins, and using a wide sample of Northern European genome samples they identify eleven genetic loci associated with an increase or decrease of this chronic inflammation marker.

Abstract Reports of persistent symptoms after hospitalization with COVID-19 have raised concern of a “long COVID” syndrome. This study aimed at determining the prevalence of and risk factors for acute and persistent symptoms in non-hospitalized patients with polymerase chain reaction (PCR) confirmed COVID-19. We conducted a cohort study of non-hospitalized participants identified via the Danish Civil Registration System with a SARS-CoV-2-positive PCR-test and available biobank samples. Participants received a digital questionnaire on demographics and COVID-19-related symptoms. Persistent symptoms: symptoms > 4 weeks (in sensitivity analyses > 12 weeks). We included 445 participants, of whom 34% were asymptomatic. Most common acute symptoms were fatigue, headache, and sneezing, while fatigue and reduced smell and taste were most severe. Persistent symptoms, most commonly fatigue and memory and concentration difficulties, were reported by 36% of 198 symptomatic participants with follow-up > 4 weeks. Risk factors for persistent symptoms included female sex (women 44% vs. men 24%, odds ratio 2.7, 95% CI 1.4–5.1, p = 0.003) and BMI (odds ratio 1.1, 95% CI 1.0–1.2, p = 0.001). In conclusion, among non-hospitalized PCR-confirmed COVID-19 patients one third were asymptomatic while one third of symptomatic participants had persistent symptoms illustrating the heterogeneity of disease presentation. These findings should be considered in health care planning and policy making related to COVID-19.

Bell et al. report 46 new loci associated with biomarkers of iron homeostasis, including ferritin levels, iron binding capacity, and iron saturation, in the Icelandic, Danish and UK populations. The associated loci point to new iron-regulating proteins and important genetic differences between men and women.

Abstract Bell’s palsy is the most common cause of unilateral facial paralysis and is defined as an idiopathic and acute inability to control movements of the facial muscles on the affected side. While the pathogenesis remains unknown, previous studies have implicated post-viral inflammation and resulting compression of the facial nerve. Reported heritability estimates of 4–14% suggest a genetic component in the etiology and an autosomal dominant inheritance has been proposed. Here, we report findings from a meta-analysis of genome-wide association studies uncovering the first unequivocal association with Bell’s palsy (rs9357446-A; P = 6.79 × 10−23, OR = 1.23; N cases = 4714, N controls = 1,011,520). The variant also confers risk of intervertebral disc disorders (P = 2.99 × 10−11, OR = 1.04) suggesting a common pathogenesis in part or a true pleiotropy.

Abstract Background The rising prevalence of type 2 diabetes (T2D) poses a major global challenge. It remains unresolved to what extent transcriptomic signatures of metabolic dysregulation and T2D can be observed in easily accessible tissues such as blood. Additionally, large-scale human studies are required to further our understanding of the putative inflammatory component of insulin resistance and T2D. Here we used transcriptomics data from individuals with (n = 789) and without (n = 2127) T2D from the IMI-DIRECT cohorts to describe the co-expression structure of whole blood that mainly reflects processes and cell types of the immune system, and how it relates to metabolically relevant clinical traits and T2D. Methods Clusters of co-expressed genes were identified in the non-diabetic IMI-DIRECT cohort and evaluated with regard to stability, as well as preservation and rewiring in the cohort of individuals with T2D. We performed functional and immune cell signature enrichment analyses, and a genome-wide association study to describe the genetic regulation of the modules. Phenotypic and trans-omics associations of the transcriptomic modules were investigated across both IMI-DIRECT cohorts. Results We identified 55 whole blood co-expression modules, some of which clustered in larger super-modules. We identified a large number of associations between these transcriptomic modules and measures of insulin action and glucose tolerance. Some of the metabolically linked modules reflect neutrophil-lymphocyte ratio in blood while others are independent of white blood cell estimates, including a module of genes encoding neutrophil granule proteins with antibacterial properties for which the strongest associations with clinical traits and T2D status were observed. Through the integration of genetic and multi-omics data, we provide a holistic view of the regulation and molecular context of whole blood transcriptomic modules. We furthermore identified an overlap between genetic signals for T2D and co-expression modules involved in type II interferon signaling. Conclusions Our results offer a large-scale map of whole blood transcriptomic modules in the context of metabolic disease and point to novel biological candidates for future studies related to T2D.

The Danish health system has been collecting health-related data on the entire Danish population for years. Here the authors present the Danish Disease Trajectory Browser (DTB), which allows users to explore population-wide disease progression patterns from data collected between 1994 and 2018.

Abstract Background Congenital heart disease (CHD) occurs in almost 1% of newborn children and is considered a multifactorial disorder. CHD may segregate in families due to significant contribution of genetic factors in the disease etiology. The aim of the study was to identify pathophysiological mechanisms in families segregating CHD. Methods We used whole exome sequencing to identify rare genetic variants in ninety consenting participants from 32 Danish families with recurrent CHD. We applied a systems biology approach to identify developmental mechanisms influenced by accumulation of rare variants. We used an independent cohort of 714 CHD cases and 4922 controls for replication and performed functional investigations using zebrafish as in vivo model. Results We identified 1785 genes, in which rare alleles were shared between affected individuals within a family. These genes were enriched for known cardiac developmental genes, and 218 of these genes were mutated in more than one family. Our analysis revealed a functional cluster, enriched for proteins with a known participation in calcium signaling. Replication in an independent cohort confirmed increased mutation burden of calcium-signaling genes in CHD patients. Functional investigation of zebrafish orthologues of ITPR1, PLCB2, and ADCY2 verified a role in cardiac development and suggests a combinatorial effect of inactivation of these genes. Conclusions The study identifies abnormal calcium signaling as a novel pathophysiological mechanism in human CHD and confirms the complex genetic architecture underlying CHD.

Abstract Background Allergic rhinitis (AR), allergic conjunctivitis (AC), and asthma composing multiple phenotypes and improved understanding of these phenotypes and their respective risk factors are needed. Objectives The objective of this study was to define the prevalence of AR, AC, and asthma and their association with allergen‐specific immunoglobulin E (sIgE) sensitization in a large cohort of blood donors and identify risk factors. Methods From the nationwide population‐based Danish Blood Donor Study, 52,976 participants completed an electronic questionnaire including AR, AC, asthma, allergic predisposition, and childhood residence. Of these, 25,257 were additionally tested for sIgE to inhalation allergens (Phadiatop). Results The prevalence of sIgE sensitization, AR, AC, and asthma was 30%, 19%, 15%, and 9%, respectively. The youngest birth cohorts had the highest prevalence of sIgE sensitization and symptoms of asthma, AR, and AC, and for asthma, they apparently experienced symptoms at an earlier age. The sIgE sensitization was positively associated with male sex. The sIgE seroprevalence was higher in participants with both AR and AC (ARC) than in participants with either AR or AC. Allergic predisposition and sIgE sensitization increased the risk of the diseases, while farm upbringing was associated with reduced prevalence of ARC, however, only in sIgE sensitized participants. Conclusion Birth year, childhood residence, sIgE sensitization, and allergic predisposition were associated with asthma, AR, and AC prevalence. Individuals with self‐reported ARC represent a primarily sIgE‐positive phenotype, while those with either AR or AC represent more diverse phenotypes.

Background Restless Legs Syndrome (RLS) is a neurological sensorimotor disorder that occurs in the evening and night, thereby impacting quality of sleep in sufferers. The pathophysiology of RLS is poorly understood but inflammation has been proposed as possibly being involved. The neutrophil-to-lymphocyte ratio (NLR) can be used as an inflammation marker but results from small studies have been inconclusive in determining whether NLR is associated with RLS. We aimed to assess whether an association between NLR and RLS exists in a large cohort of healthy individuals. Methods Neutrophils and lymphocytes were measured in blood samples of 13,055 individuals from the Danish Blood Donor Study, all of whom completed the validated Cambridge-Hopkins RLS-questionnaire for RLS assessment. Results In the sample, 661 individuals were determined as current RLS cases (5.1%). A higher proportion of individuals with RLS were females (62.5% vs 47.5%; PConclusion We found no association between RLS and NLR among Danish blood donors after adjusting for sex, age, alcohol consumption, smoking status and BMI. Further studies are needed to determine whether inflammation is a risk factor for RLS.

Diabetes is a diverse and complex disease, with considerable variation in phenotypic manifestation and severity. This variation hampers the study of etiological differences and reduces the statistical power of analyses of associations to genetics, treatment outcomes, and complications. We address these issues through deep, fine-grained phenotypic stratification of a diabetes cohort. Text mining the electronic health records of 14,017 patients, we matched two controlled vocabularies (ICD-10 and a custom vocabulary developed at the clinical center Steno Diabetes Center Copenhagen) to clinical narratives spanning a 19 year period. The two matched vocabularies comprise over 20,000 medical terms describing symptoms, other diagnoses, and lifestyle factors. The cohort is genetically homogeneous (Caucasian diabetes patients from Denmark) so the resulting stratification is not driven by ethnic differences, but rather by inherently dissimilar progression patterns and lifestyle related risk factors. Using unsupervised Markov clustering, we defined 71 clusters of at least 50 individuals within the diabetes spectrum. The clusters display both distinct and shared longitudinal glycemic dysregulation patterns, temporal co-occurrences of comorbidities, and associations to single nucleotide polymorphisms in or near genes relevant for diabetes comorbidities.

Glucagon-like peptide 1 (GLP-1) stimulated insulin secretion has a considerable heritable component as estimated from twin studies, yet few genetic variants influencing this phenotype have been identified. We performed the first genome-wide association study (GWAS) of GLP-1 stimulated insulin secretion in non-diabetic individuals from the Netherlands Twin register (n = 126). This GWAS was enhanced using a tissue-specific protein-protein interaction network approach. We identified a beta-cell protein-protein interaction module that was significantly enriched for low gene scores based on the GWAS P-values and found support at the network level in an independent cohort from Tübingen, Germany (n = 100). Additionally, a polygenic risk score based on SNPs prioritized from the network was associated (P < 0.05) with glucose-stimulated insulin secretion phenotypes in up to 5,318 individuals in MAGIC cohorts. The network contains both known and novel genes in the context of insulin secretion and is enriched for members of the focal adhesion, extracellular-matrix receptor interaction, actin cytoskeleton regulation, Rap1 and PI3K-Akt signaling pathways. Adipose tissue is, like the beta-cell, one of the target tissues of GLP-1 and we thus hypothesized that similar networks might be functional in both tissues. In order to verify peripheral effects of GLP-1 stimulation, we compared the transcriptome profiling of ob/ob mice treated with liraglutide, a clinically used GLP-1 receptor agonist, versus baseline controls. Some of the upstream regulators of differentially expressed genes in the white adipose tissue of ob/ob mice were also detected in the human beta-cell network of genes associated with GLP-1 stimulated insulin secretion. The findings provide biological insight into the mechanisms through which the effects of GLP-1 may be modulated and highlight a potential role of the beta-cell expressed genes RYR2, GDI2, KIAA0232, COL4A1 and COL4A2 in GLP-1 stimulated insulin secretion.

[This corrects the article DOI: 10.1371/journal.pgen.1005378.].

Genome-wide association studies (GWAS) have identified more than 100 genetic variants contributing to BMI, a measure of body size, or waist-to-hip ratio (adjusted for BMI, WHRadjBMI), a measure of body shape. Body size and shape change as people grow older and these changes differ substantially between men and women. To systematically screen for age- and/or sex-specific effects of genetic variants on BMI and WHRadjBMI, we performed meta-analyses of 114 studies (up to 320,485 individuals of European descent) with genome-wide chip and/or Metabochip data by the Genetic Investigation of Anthropometric Traits (GIANT) Consortium. Each study tested the association of up to ~2.8M SNPs with BMI and WHRadjBMI in four strata (men ≤50y, men >50y, women ≤50y, women >50y) and summary statistics were combined in stratum-specific meta-analyses. We then screened for variants that showed age-specific effects (G x AGE), sex-specific effects (G x SEX) or age-specific effects that differed between men and women (G x AGE x SEX). For BMI, we identified 15 loci (11 previously established for main effects, four novel) that showed significant (FDR

BackgroundNumerous gene loci are related to single measures of body weight and shape. We investigated if 55 SNPs previously associated with BMI or waist measures, modify the effects of fat intake on weight loss and waist reduction under energy restriction.Methods and findingsRandomized controlled trial of 771 obese adults. (RegistrationISRCTN25867281.) One SNP was selected for replication in another weight loss intervention study of 934 obese adults. The original trial was a 10-week 600 kcal/d energy-deficient diet with energy percentage from fat (fat%) in range of 20-25 or 40-45. The replication study used an 8-weeks diet of 880 kcal/d and 20 fat%; change in fat% intake was used for estimation of interaction effects. The main outcomes were intervention weight loss and waist reduction. In the trial, mean change in fat% intake was -12/+4 in the low/high-fat groups. In the replication study, it was -23/-12 among those reducing fat% more/less than the median. TFAP2B-rs987237 genotype AA was associated with 1.0 kg (95% CI, 0.4; 1.6) greater weight loss on the low-fat, and GG genotype with 2.6 kg (1.1; 4.1) greater weight loss on the high-fat (interaction p-value; p = 0.00007). The replication study showed a similar (non-significant) interaction pattern. Waist reduction results generally were similar. Study-strengths include (i) the discovery study randomised trial design combined with the replication opportunity (ii) the strict dietary intake control in both studies (iii) the large sample sizes of both studies. Limitations are (i) the low minor allele frequency of the TFAP2B polymorphism, making it hard to investigate non-additive genetic effects (ii) the different interventions preventing identical replication-discovery study designs (iii) some missing data for non-completers and dietary intake. No adverse effects/outcomes or side-effects were observed.ConclusionsUnder energy restriction, TFAP2B may modify the effect of dietary fat intake on weight loss and waist reduction.

Non-alcoholic fatty liver disease (NAFLD) is a common condition, associated with hepatic insulin resistance and the metabolic syndrome including hyperglycaemia and dyslipidemia. We aimed at studying the potential impact of the NAFLD-associated PNPLA3 rs738409 G-allele on NAFLD-related metabolic traits in hyperglycaemic individuals.The rs738409 variant was genotyped in the population-based Inter99 cohort examined by an oral glucose-tolerance test, and a combined study-sample consisting of 192 twins (96 twin pairs) and a sub-set of the Inter99 population (n = 63) examined by a hyperinsulinemic euglycemic clamp (n(total) = 255). In Inter99, we analyzed associations of rs738409 with components of the WHO-defined metabolic syndrome (n = 5,847) and traits related to metabolic disease (n = 5,663). In the combined study sample we elucidated whether the rs738409 G-allele altered hepatic or peripheral insulin sensitivity. Study populations were divided into individuals with normal glucose-tolerance (NGT) and with impaired glucose regulation (IGR).The case-control study showed no associations with components of the metabolic syndrome or the metabolic syndrome. Among 1,357 IGR individuals, the rs738409 G-allele associated with decreased fasting serum triglyceride levels (per allele effect(β) = -9.9% [-14.4%;-4.0% (95% CI)], p = 5.1×10(-5)) and fasting total cholesterol (β = -0.2 mmol/l [-0.3;-0.01 mmol/l(95% CI)], p = 1.5×10(-4)). Meta-analyses showed no impact on hepatic or peripheral insulin resistance in carriers of the rs738409 G-allele.Our findings suggest that the G-allele of PNPLA3 rs738409 associates with reduced fasting levels of cholesterol and triglyceride in individuals with IGR.

Two meta-analyses of genome-wide association studies (GWAS) have suggested that four variants: rs2605100 in lysophospholipase-like 1 (LYPLAL1), rs10146997 in neuroxin 3 (NRXN3), rs545854 in methionine sulfoxide reductase A (MSRA), and rs987237 in transcription factor activating enhancer-binding protein 2 beta (TFAP2B) associate with measures of central obesity. To elucidate potential underlying phenotypes we aimed to investigate whether these variants associated with: 1) quantitative metabolic traits, 2) anthropometric measures (waist circumference (WC), waist-hip ratio, and BMI), or 3) type 2 diabetes, and central and general overweight and obesity.The four variants were genotyped in Danish individuals using KASPar®. Quantitative metabolic traits were examined in a population-based sample (n = 6,038) and WC and BMI were furthermore analyzed in a combined study sample (n = 13,507). Case-control studies of diabetes and adiposity included 15,326 individuals. The major G-allele of LYPLAL1 rs2605100 associated with increased fasting serum triglyceride concentrations (per allele effect (β) = 3%(1;5(95%CI)), p(additive) = 2.7×10(-3)), an association driven by the male gender (p(interaction) = 0.02). The same allele associated with increased fasting serum insulin concentrations (β = 3%(1;5), p(additive) = 2.5×10(-3)) and increased insulin resistance (HOMA-IR) (β = 4%(1;6), p(additive) = 1.5×10(-3)). The minor G-allele of rs10146997 in NRXN3 associated with increased WC among women (β = 0.55cm (0.20;0.89), p(additive) = 1.7×10(-3), p(interaction) = 1.0×10(-3)), but showed no associations with obesity related metabolic traits. The MSRA rs545854 and TFAP2B rs987237 showed nominal associations with central obesity; however, no underlying metabolic phenotypes became obvious, when investigating quantitative metabolic traits. None of the variants influenced the prevalence of type 2 diabetes.We demonstrate that several of the central obesity-associated variants in LYPLAL1, NRXN3, MSRA, and TFAP2B associate with metabolic and anthropometric traits in Danish adults. However, analyses were made without adjusting for multiple testing, and further studies are needed to confirm the putative role of LYPLAL1, NRXN3, MSRA, and TFAP2B in the pathophysiology of obesity.

Candidate genes for non-alcoholic fatty liver disease (NAFLD) identified by a bioinformatics approach were examined for variant associations to quantitative traits of NAFLD-related phenotypes.By integrating public database text mining, trans-organism protein-protein interaction transferal, and information on liver protein expression a protein-protein interaction network was constructed and from this a smaller isolated interactome was identified. Five genes from this interactome were selected for genetic analysis. Twenty-one tag single-nucleotide polymorphisms (SNPs) which captured all common variation in these genes were genotyped in 10,196 Danes, and analyzed for association with NAFLD-related quantitative traits, type 2 diabetes (T2D), central obesity, and WHO-defined metabolic syndrome (MetS).273 genes were included in the protein-protein interaction analysis and EHHADH, ECHS1, HADHA, HADHB, and ACADL were selected for further examination. A total of 10 nominal statistical significant associations (P

BackgroundPolymorphisms in the potassium channel, voltage-gated, KQT-like subfamily, member 1 (KCNQ1) have recently been reported to associate with type 2 diabetes. The primary aim of the present study was to investigate the putative impact of these KCNQ1 polymorphisms (rs2283228, rs2237892, rs2237895, and rs2237897) on estimates of glucose stimulated insulin release.Methodology/principal findingsGenotypes were examined for associations with serum insulin levels following an oral glucose tolerance test (OGTT) in a population-based sample of 6,039 middle-aged and treatment-naïve individuals. Insulin release indices estimated from the OGTT and the interplay between insulin sensitivity and insulin release were investigated using linear regression and Hotelling T2 analyses. Applying an additive genetic model the minor C-allele of rs2237895 was associated with reduced serum insulin levels 30 min (mean+/-SD: (CC) 277+/-160 vs. (AC) 280+/-164 vs. (AA) 299+/-200 pmol/l, p = 0.008) after an oral glucose load, insulinogenic index (29.6+/-17.4 vs. 30.2+/-18.7vs. 32.2+/-22.1, p = 0.007), incremental area under the insulin curve (20,477+/-12,491 vs. 20,503+/-12,386 vs. 21,810+/-14,685, p = 0.02) among the 4,568 individuals who were glucose tolerant. Adjustment for the degree of insulin sensitivity had no effect on the measures of reduced insulin release. The rs2237895 genotype had a similar impact in the total sample of treatment-naïve individuals. No association with measures of insulin release were identified for the less common diabetes risk alleles of rs2237892, rs2237897, or rs2283228.ConclusionThe minor C-allele of rs2237895 of KCNQ1, which has a prevalence of about 42% among Caucasians was associated with reduced measures of insulin release following an oral glucose load suggesting that the increased risk of type 2 diabetes, previously reported for this variant, likely is mediated through an impaired beta cell function.