bluuebunny/crossref_metadata_2025 huggingface dataset:
1281-P: Understanding the Interaction between Type 2 Diabetes and COVID-via Extracellular Vesicle Proteomic and Phosphoproteomic Signatures
Authors: NUNEZ LOPEZ YURY O. , ILIUK ANYON , CASU ANNA , LUPU DANIEL S. , PRATLEY RICHARD E.
Published: May 2022

Evidence supporting the involvement of EVs in the pathogenesis/severity of SARS-CoV-2 infection is starting to accumulate. However, little is known about their specific associations in the context of COVID-and type 2 diabetes interaction. Our study included 48 plasma samples (N=12/group) obtained from COVID-patients with and without diabetes and from patients with non-COVID-acute respiratory diagnosis (RSP) with and without diabetes. Participants were identified from a set of 494 patients hospitalized at AdventHealth in June-August 2020. Important efforts were made to ensure the homogeneity of the study cohort. Patients with type 1 diabetes, or pregnant, or that went directly into the ICU were excluded, and 4 balanced groups were identified after 10,000 random cohorts were generated and differences in age, gender, race, and ethnicity statistically assessed. EVs were isolated using EVTRAP (Tymora) . Mass spectrometry-based methods were used to detect the global EV proteome and phosphoproteome. Differentially expressed features, enriched pathways, and enriched tissue-specific protein sets were identified. Multidimensional scaling of all EV proteomic and phosphoproteomic data and unsupervised clustering of differentially expressed (absolute fold change ≥ 2, P < 0.05, FDR < 0.05) EV proteins and phosphoproteins successfully distinguished the 4 study groups with close to 100% accuracy. Importantly, we detected enriched pathway networks that suggest the potential therapeutic utility of PKC inhibitors such as bisindolylmaleimide IX, sotrastaurin, and enzastaumn, and inhibitors of ROCK1 such the isoquinoline derivative Fasudil.

In conclusion, we characterized the proteomic landscape of the interaction between type 2 diabetes and COVID-and defined disease-specific EV proteomic signatures that provide insight into the disease pathobiology and druggable targets with potential clinical utility.

Disclosure

Y.O.Nunez lopez: None. A.Iliuk: None. A.Casu: None. D.S.Lupu: None. R.E.Pratley: Other Relationship; Bayer AG, Corcept Therapeutics, Dexcom, Inc., Hanmi Pharm. Co., Ltd., Merck & Co., Inc., Metavention, Novo Nordisk, Pfizer Inc., Poxel SA, Sanofi, Scohia Pharma Inc., Sun Pharmaceutical Industries Ltd.
1287-P: Deep Immune Phenotyping of Type 1 Diabetes by Machine Learning
Authors: VERA-RAMOS JOSÉ ANTONIO , PRIETL BARBARA , HERBSTHOFER LAURIN , PFEIFER VERENA , LÓPEZ-GARCÍA PABLO , STRADNER MARTIN , PIEBER THOMAS
Published: May 2022

Breakdown of self-tolerance is an important common mechanism in autoimmunity. We use machine learning (ML) to identify common patterns and dissimilarities between type 1 diabetes (T1D) , rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE) based on immune phenotyping. PBMCs were isolated from patients with T1D, RA, SLE, and controls. A FACS approach was applied, and a traditional analysis was compared to a ML method implemented on R and based on self-organizing maps (Fig) . Our pipeline includes unsupervised pre-gating, normalization, FlowSOM clustering, and a statistical model (GLMM) , to check for significant differential abundances of cell populations among the autoimmune conditions. After applying our automated workflow to one T cell panel we could identify 14 new cell clusters present in all the samples. The GLMM test revealed a cluster with a significant difference (p=0.035) and a trending one (p=0.059) on the abundance across the different diseases. In particular, CD4pos T cells expressing high IL-7 receptor (CD127) levels and median amounts of CD15s but low CD25, CD161 and FoxP3 are increased in T1D whereas CD4+CD25++CD15s+FoxP3lowCD161lowCD45RA- cells are increased in SLE.

In conclusion, our ML workflow identifies a new subtype of T cells significantly increased in T1D. This unsupervised analysis approach for large datasets enables the discovery of new biomarkers complementing traditional workflows.

Disclosure

J.Vera-ramos: None. B.Prietl: None. L.Herbsthofer: None. V.Pfeifer: None. P.López-garcía: None. M.Stradner: Consultant; AbbVie Inc., AstraZeneca, Speaker's Bureau; Janssen Pharmaceuticals, Inc. T.Pieber: Advisory Panel; Arecor, Novo Nordisk A/S, Research Support; AstraZeneca, European Union, JDRF, Novo Nordisk Foundation, Sanofi, Speaker's Bureau; Novo Nordisk A/S, Roche Diagnostics.

Funding

JDRF, LRA, & NMSS Joint RFA (2-SRA-2021-1043-S-B)
1290-P: A Rare Case of HNF1A-MODY
Authors: VELAYATI SARA , SHANIK MICHAEL , ROMAO ISABELA J.
Published: May 2022

Introduction: Maturity onset diabetes of the young (MODY) is a monogenic nonimmune type of diabetes mellitus. It is rare and sometimes misdiagnosed as type 1 or type 2 diabetes (T2DM) . The age of onset is usually below 25 years. It is more common in the European population. Hepatocyte nuclear factor 1-alpha (HNF1A) is one of the most common mutations and is associated with cardiovascular and renal complications.1 Genetic testing in patients with strong hereditary factors results in definitive diagnosis and subsequent appropriate treatment.2 The aim of this report is to emphasize the importance of distinguishing MODY by considering the family history of diabetes and relative clinical features.

Case presentation: A 58-year-old Filipino woman was diagnosed with T2DM at age 15. Her mother, 5 of 8 siblings and her daughter all had diabetes. They all died in the 2nd or 3rd decades of their life due to diabetes complications. Her grandfather was European. She has renal insufficiency, coronary heart disease and heart failure. Treatment included glimepiride 2 mg daily and sitagliptin 25 mg daily. The physical exam was unremarkable. Body mass index (BMI) was 21. GAD 65 antibody and Islet cell antibody were negative, C peptide: 3.6 ng/mL, blood sugar: 1mg/dL, HbA1c: 7.3%, creatinine: 2.15 mg/dL and GFR: 25 mL/min/1.73m2. A diagnosis of MODY was considered due to strong family history of diabetes, young age at the onset of diabetes and normal BMI. DNA sequencing demonstrated positive HNF1A mutation compatible with HNF1A-MODY. Due to post prandial hyperglycemia, glimepiride was replaced with pre-meal lispro.

Conclusion: This is a rare case of HNF1A-MODY found in a Filipino patient. MODY should be included in the differential diagnosis of diabetic patients with an extensive family history of diabetes, young age of onset of diabetes and negative pancreatic antibodies. Early diagnosis with genetic profiling can guide therapy and hopefully decrease the vascular complications of long-term poor glycemic control.

Disclosure

S.Velayati: None. M.Shanik: Consultant; Bigfoot Biomedical, Inc., Proteomics International, Research Support; Better Therapeutics, Speaker's Bureau; Abbott, Boehringer Ingelheim International GmbH, Novo Nordisk, Xeris Pharmaceuticals, Inc. I.J.Romao: None.
1295-P: Learning from Immune Privileged Tissue Stem Cells to Generate Hypoimmunogenic Islets
Authors: AGUDO JUDITH
Published: May 2022

The ultimate goal to accomplish a definitive cure for Type 1 Diabetes (T1D) is to replace the lost beta-cell mass while ensuring its long-lasting survival. This could be done by transplantation of pancreatic islets or stem cell-derived insulin-producing cells. However, any replacement will irrevocably succumb to the same autoimmune attack that killed the original beta-cells. Moreover, a mismatch between donor cells and recipient patients adds an allo-graft rejection. Thus, it is of the utmost importance to develop strategies to protect transplanted pancreatic islets from immune attack. Unfortunately, many approaches aiming at restoring tolerance by manipulating immune cells have been tested and, although safe, have shown only limited efficacy. Inflammatory processes, infections or major lifestyle changes can easily alter the balance again towards immune activation. We recently discovered the existence of immune privileged stem cells in the skin and muscle, and found that their ability to resist direct attack by T cells was not dependent on a physical barrier. Instead, it is a cell-autonomous process that allows independent cloaking from immune cells. Here we propose to exploit the molecular circuits controlling this striking phenomenon (that already naturally occurs in our bodies) to provide similar protection to islets for transplantation. We identified a molecular pathway that reduces antigen presentation allowing protection from T cells while preserving enough MHC class I to not trigger a Natural Killer (NK) cell response. Since this mechanism of protection is cell intrinsic, changes in overall immune function should not alter the ability of engineered islets to hide from immune rejection allowing for long-lasting survival.

Disclosure

J.Agudo: None.

Funding

American Diabetes Association (1-20-ACE-08)
13-LB: A Prediction Model of Chronic Kidney Disease Progression among Individuals with Type 2 Diabetes in the United States
Authors: LIN YILU , SHAO HUI , ANDERSON AMANDA H. , FONSECA VIVIAN , BATUMAN VECIHI , SHI LIZHENG
Published: May 2022

CKD progression among individuals with T2D is associated with poor outcomes and high costs. The risk factors for CKD progression in T2D have not been fully studied. This study aimed to predict CKD progression among individuals with T2D. Using the ACCORD clinical trial, a time-varying Cox model was developed to predict the risk of CKD progression among patients with CKD. CKD progression was defined as a 50% decline, or 25 mL/min/1.73 m2 decline in eGFR from baseline, doubling of the serum creatinine, or onset of ESKD. A list of candidate variables included demographic characteristics, physical exam, laboratory, medical history, drug use, and healthcare utilization. A stepwise algorithm was used for variable selection. Data was separated into training and validation set. Model performance was evaluated by Brier score (BS) and C statistics. Confidence intervals (CI) were calculated by bootstraping. Decomposition analysis was conducted to assess the predictor contribution. Generalizability was assessed on patient-level data of the HARMONY Outcome clinical trial. A total of 6,982 T2D patients with CKD were used for model development, with a median follow-up of 4 years and 3,346 CKD progression events. The predictors for CKD progression included female sex, age at T2D diagnosis, smoking status, BP, HR, HbA1c, ALT, eGFR, UACR, retinopathy event, hospitalization, interaction of SBP and smoking, and SBP and ALT. The model demonstrated good discrimination (C-statistics 0.745 [95% CI 0.723-0.763]) and calibration (BS 0.0923 [95% CI 0.0873-0.0965]) . The most contributing predictors for CKD progression were eGFR, HbA1c, and SBP. There were 2,954 patients with CKD extracted from HARMONY with 640 CKD progression events and median follow-up of 2 years. The model demonstrated acceptable discrimination (C-statistics: 0.742 [95% CI 0.721-0.7645]) and calibration (BS: 0.0988 [95% CI 0.0952-0.1022]) in the external data. For high-risk patients with both diabetes and CKD, the tool as a dynamic risk prediction of CKD progression may help develop novel strategies to lower the risk of CKD progression.

Disclosure

Y. Lin: None. H. Shao: Board Member; BRAVO4HEALTH, LLC. A. H. Anderson: None. V. Fonseca: Consultant; Abbott, Asahi Kasei Corporation, Bayer AG, Novo Nordisk, Sanofi, Research Support; Fractyl Health, Inc., Jaguar Gene Therapy, Stock/Shareholder; Abbott, Amgen Inc., BRAVO4Health, Mellitus Health. V. Batuman: None. L. Shi: None.
1302-P: Inhibitory Effect of Advanced Glycation End Products on Proinsulin Processing
Authors: BRINGS SEBASTIAN , MIER WALTER , BEIJER BARBRO , SZENDROEDI JULIA M. , NAWROTH PETER P. , FLEMING THOMAS H.
Published: May 2022

Advanced glycation end-products (AGEs) form at an accelerated rate in diabetes and are associated with the development of diabetic complications. So far, the effect of AGEs on proteases was mostly investigated with substrates modified non-specifically with hexoses amongst others in combination with proteases of broad specificity. Here, we investigated the effect of AGEs on the proinsulin processing peptidases preprotein convertase 1 (PC1) and carboxypeptidase E (CPE) via two ways: - Measuring of activity towards peptide substrates. These contained chemically defined AGEs carboxymethyllysine (CML) or methylglyoxyl-hydroimidazolone-1 (MGH1) or unmodified amino acids. - Recombinant human proinsulin was incubated with methylglyoxal for 24 h and was processed by addition of PC1 and CPE in vitro. Formation of insulin was compared to unmodified proinsulin. In parallel formation of AGE-modified proinsulin was quantified.

Artificial peptide substrates were synthesized by solid phase peptide synthesis and purified by HPLC. A protected building block was used for MGH1 incorporation while CML was introduced on the resin. Protease activity towards artificial substrates was measured by fluorescence assay or LC MS/MS. Proinsulin, insulin and AGE-modified proinsulin were quantified by LC MS.

Strong activity of PC1 and CPE was seen towards arginine containing substrates. Activity was completely absent towards modified amino acids MGH1 and CML for both, PC1 and CPE. Incubation of proinsulin with MG resulted in MGH1 modified proinsulin. At low MG (µM) concentrations this was associated with a 25 % reduction of insulin formation while high MG concentrations (95 µM) led to a 75 % reduction in insulin formation.

Here we demonstrate for the first time the effect of substrate modifications with AGEs on physiologically important proteases PC1 and CPE. The latter are not only involved in the processing of proinsulin but a range of endocrinologically and neurophysiologically important peptide hormones.

Disclosure

W. Mier: None. B. Beijer: None. J.M. Szendroedi: Consultant; Boehringer Ingelheim International GmbH. T.H. Fleming: None.

Funding

DFG (SFB 1118)
131-OR: Effect of a Value-Based Medication Benefit on Acute Preventable Diabetes Complications and Related Disparities
Authors: WHARAM J. FRANK , ZHANG FANG , LAKOMA MATTHEW , GARABEDIAN LAURA F. , ARGETSINGER STEPHANIE , ROSS-DEGNAN DENNIS
Published: May 2022

Employers are increasingly purchasing "preventive drug list" (PDL) insurance plans that make cardiometabolic medications $0. PDL designs improve adherence, but effects on health outcomes are unknown. We analyzed 2004-2017 commercial health insurance claims using a large, national data set. The intervention group included 12,853 commercially-insured diabetes patients aged 12-64 years who experienced employer-mandated PDL switches; the control group included 12,989 matched counterparts whose employers offered no PDL. We defined the index date as the day the intervention group received the PDL, and matched controls had a contemporaneous index date. Both groups had a 1-year baseline and 1-year follow-up study period. We measured monthly rates of aggregated acute preventable diabetes complication episodes (e.g, hyperglycemic coma, angina, and pneumonia) presenting to (1) the outpatient setting or (2) the emergency department, observation, or inpatient settings ("high acuity") . We constructed cumulative interrupted time series plots of these two measures and conducted segmented regression analysis and marginal effects estimation. Low-income members were a subgroup of interest. Annual outpatient and high-acuity complication episodes decreased by -13.5% (-15.2%, -11.7%) and -13.9% (-15.3%, -12.5%) , respectively, in the PDL group relative to controls. Low-income PDL members also experienced decreased outpatient (-27.2% [-32.1%, -22.4%]) and high-acuity complication episodes (-4.4% [-6.5%, -2.4%]) . This rigorous, quasi-experimental study is the first to demonstrate that a value-based medication benefit is associated with improved health outcomes and reduced disparities. Employer adoption of PDLs thus represents a relatively expedient approach to improving health and equity, and our results support a strategy of incentivizing such targeted cost sharing reductions for vulnerable diabetes populations.

Disclosure

J.Wharam: None. F.Zhang: Research Support; GlaxoSmithKline plc. M.Lakoma: None. L.F.Garabedian: None. S.Argetsinger: None. D.Ross-degnan: None.

Funding

CDC and NIDDK
1310-P: Effects of Insulin Resistance on Brain Responses to Caloric Sugars and Nonnutritive Sweetener
Authors: KULLMANN STEPHANIE , VEIT RALF , JANN KAY , YUNKER ALEXANDRA G. , LUO SHAN H. , ANGELO BRENDAN C. , ALVES JASMIN M. , MONTEROSSO JOHN R. , PAGE KATHLEEN A.
Published: May 2022

Evidence suggests that insulin resistance (IR) affects feeding behavior by modulating neural responses to post-ingestive signals, including nutrients and taste. We examined associations between IR and activation of the hypothalamus and insula, brain regions involved in homeostasis and taste processing, to ingestion of sugars and the non-nutritive sweetener (NNS) , sucralose. Seventy-four participants (23.4±4 years; 42%male; BMI 27.2±5; HOMA-IR 5.3±5) completed 4 MRI visits with ingestion of drinks containing 75g sucrose, 75g glucose, sucralose, or water in a randomized cross-over design. Arterial spin labeling MRI was performed before, 15min and 30min after drink ingestion to measure cerebral blood flow (CBF) . Primary outcomes were associations between HOMA-IR and hypothalamic and insular CBF responses to glucose vs. sucrose, sucralose vs. water, and sucrose vs. sucralose. Mixed effects models included the following factors: drink contrasts, sex, time point, and BMI and HOMA-IR as continuous variables. Bonferroni-Holm was used to adjust for multiple comparisons. In the hypothalamus, HOMA-IR, independent of BMI, was associated with a greater hypothalamic CBF response to sucralose vs. water (p<0.03) , but there was no association between HOMA-IR and hypothalamic response to sugar. In the insula, there was a drink by HOMA-IR interaction in the insular response to sucralose vs. water (p<0.02) ; persons with IR showed greater increases in CBF after sucralose vs. water. We also found a main effect of HOMA-IR; persons with higher IR had greater insular CBF increases to sugar (p<0.05) , and a drink by HOMA-IR interaction revealed a stronger association between IR and insular response to glucose vs. sucrose (p<0.03) . There was no association between HOMA-IR and brain responses to sucralose vs. sucrose. These findings show that IR is associated with greater activation of brain regions involved in taste and energy homeostasis to ingestion of sugar and NNS, which may promote eating behavior.

Disclosure

S. Kullmann: Other Relationship; Novo Nordisk. R. Veit: None. K. Jann: None. S.H. Luo: None. B.C. Angelo: None. J.M. Alves: None. J.R. Monterosso: None. K.A. Page: None.

Funding

NIDDK R01DK102794
1316-P: Brainstem Neurons Regulating Energy Homeostasis
Authors: NECTOW ALEXANDER R.
Published: May 2022

Energy balance is maintained by an extended circuitry in the central nervous system that carefully balances food intake with energy expenditure. We have recently shown that the brainstem’s dorsal raphe nucleus (DRN) is an important component of this extended circuit, capable of potently regulating both energy intake and expenditure, and ultimately body weight. Despite its important role in energy homeostasis, the DRN is markedly heterogenous, consequently hindering our ability to manipulate and record from its distinct cell subtypes. Towards this end, we have recently performed nuclear tagging of large classes of DRN neurons, enabling us to molecularly profile these cells at both ensemble and single-cell resolutions. Using this approach, we have found that even among a priori defined major cell types within the DRN - for example, those marked by the vesicular glutamate and GABA transporters - there is a marked degree of molecular heterogeneity. Despite this significant molecular heterogeneity, some subclasses of DRN neurons are hierarchically differentiable, leading to the identification of molecularly dissociable ‘clades.’ These clades have unique molecular handles; these handles are now enabling us to record from and perturb the activity of distinct DRN subpopulations, which were previously genetically inaccessible. Together, this work represents an important conceptual and technical advance, enabling us to classify DRN neurons with new resolution, as well as to genetically access them to establish their cell subtype-specific function.

Disclosure

A.R.Nectow: None.

Funding

American Diabetes Association (1-18-ACE-49)
1318-P: Repetitive Transcranial Magnetic Stimulation: A Potential Treatment for Obesity in Patients with Prader-Willi Syndrome
Authors: FERRULLI ANNA , CANNAVARO DANIELE , MACRÌ CONCETTA , MASSARINI STEFANO , LUZI LIVIO
Published: May 2022

Prader-Willi Syndrome (PWS) is the most common syndromic type of human obesity. Multiple endocrine abnormalities due to hypothalamic dysfunction as well as cognitive and behavioral disorders occur in PWS. Hypothalamic dysfunction, dysregulated mesocorticolimbic reward system, and impaired prefrontal cortex (PFC) function are implicated in the pathophysiology of hyperphagia and obesity in PWS. All available therapeutic tools have not fully targeted at counteract the mechanisms underlying obesity in PWS. Previously, we demonstrated the safety and efficacy of repetitive Transcranial Magnetic Stimulation (rTMS) in inducing weight loss in obesity by modulating the brain dopaminergic system and enhancing the inhibitory activity of the PFC on eating behaviour (Ferrulli et al, DOM 2019) . In this case-report, for the first time, we tested the efficacy and safety of rTMS in a 23-year-old patient with PWS, in whom repeated interventions aimed at inducing weight loss have failed. A 5-week high frequency (18 Hz) rTMS treatment, associated with diet, resulted in a 3.5% weight loss (127.4 vs. 123 Kg, BMI: 50.4 vs. 48.6 Kg/m2, WHR: 1.14 vs. 1.12) . Concerning the metabolic parameters, an improvement in lipid profile has been shown (CHO/HDL/LDL/TG: 212/40/142/152 vs. 190/40/124/131 mg/dL) . While no significant changes emerged in the psychometric assessment of food craving, impulsiveness, quality of life, self-esteem, interestingly, a robust improvement in the Mini-Mental State Examination score (corrected for age and schooling) has been found [from 22 (severe impairment) to 27 (moderate impairment) ]. The improvement in the cognitive status assessed through the Montreal Cognitive Assessment was weaker (17/30 to 19/30) . In view of the urgent need of novel effective interventions for obesity in PWS, these findings support a potential role of rTMS in reducing food drive and behaviors impacting hyperphagia and obesity in PWS, with a possible impact also on cognitive functions.

Disclosure

A.Ferrulli: None. D.Cannavaro: None. C.Macrì: None. S.Massarini: None. L.Luzi: Advisory Panel; Eli Lilly and Company, Speaker’s Bureau; Eli Lilly and Company, Novo Nordisk.

Funding

IRCCS Multimedica - Ricerca Corrente
1323-P: Insulin Secretion at Diabetes Remission in Obese Blacks Presenting with Diabetic Ketoacidosis
Authors: STEFANOVSKI DARKO , OLADEJO OMOLADE , UMPIERREZ GUILLERMO E. , VELLANKI PRIYATHAMA
Published: May 2022

In obese Blacks presenting with diabetic ketoacidosis (DKA) at new-onset of diabetes, intensive insulin treatment improves β-cell function so that ˜70% achieve near-normoglycemia remission (HbA1c < 7%, fasting blood glucose [BG]<130 mg/dl while off insulin for ≥1 week) and maintain glycemic control without medications. The 2021 American Diabetes Association (ADA) consensus defined diabetes remission as HbA1c <6.5% or BG 126 mg/dl while off medications for >3 months. It is unknown whether glycemic measures and insulin secretion differ between patients who achieve the new ADA definition of remission (intensive remission [IR]) compared to near-normoglycemia remission (non-intensive remission [NIR]) .We performed 75-g, 240-minute (min) oral glucose tolerance tests (OGTT) in 13 obese (BMI>30 kg/m2) Blacks who presented with DKA (BG>250 mg/dl, HCO3<18 mEq/L, pH<7.3) and achieved remission from insulin. Glucose and C-peptide levels were measured at -15, 0, 10, 20, 30, 60, 90, 120, 180 and 240 min during OGTT. Insulin secretory rates (ISR) , first phase (Φdynamic) , second phase (Φstatic) and total (Φtotal) insulin secretion were calculated using a previously published model. Data was compared between IR, (HbA1c<6.5%, n=7) vs. NIR, (HbA1c≥6.5%, n=6)) There were no differences in age, BMI and mean BG levels during OGTT between groups. Model estimates of Φdynamic were significant (P=0.046) and 5-fold higher in IR (Marginal Mean: 514.00, 95% CI: 134.53-893.47 [109]) vs. NIR (Marginal Mean: 100, 95% CI: 100.45, 95% CI: -41.97-242.87 [109]) . Estimates of Φstatic were significant (P=0.022) and 2-fold higher in the IR (Marginal Mean: 31.84, 95% CI: 19.88-43.80 [min-1]) vs. NIR (Marginal Mean: 14.59, 95% CI: 6.79-22.39 [min-1]) with no significant (P=0.058) group differences between Φtotal.In comparison to NIR, obese Blacks with IR exhibit marked preservation of β-cell function suggesting that HbA1c <6.5% should be targeted for insulin remission after presentation with DKA.

Disclosure

D.Stefanovski: None. O.Oladejo: None. G.E.Umpierrez: Research Support; AstraZeneca, Dexcom, Inc., Novo Nordisk. P.Vellanki: n/a.

Funding

NIH/NIDDK K23 DK11324-01A1
133-LB: Relationship of HbA1c and Glucose by Ethnicity in UK Biobank
Authors: WILLIAMS JOHN A. , KARWATH ANDREAS , ROUND RACHEL A. , STRATTON IRENE M. , GHOSH SANDIP , MOSTAFA SAMIUL , ROBERTS GRAHAM , WEBBER JONATHAN , GKOUTOS GEORGIOS , MANLEY SUSAN E.
Published: May 2022

The UK Biobank recruited 502,481 people aged 40 to 70 years who were invited to participate by post so not necessarily representative of the general population. The 405,363 participants with random glucose, HbA1c and the necessary demographic/clinical data were aged 56.5 (8.09) years, mean SD, with 54% female, and ethnicity 95.0% White W, 1.9 South Asian SA, 1.3% Black B, 0.9% Other O, 0.6% Mixed Background MB, and 0.3% Chinese C. Glucose was measured by hexokinase and HbA1c by BioRad VARIANT II TURBO analysers, reference range 20-42 mmol/mol in a dedicated central laboratory. Cohen's D statistic was used to compare glucose/HbA1c by ethnicity versus the largest group W. Mean glucose ranged from 5.04 to 5.41 mmol/L with HbA1c40.7 mmol/mol for SAversus36.0 for W (D=0.78 medium effect M) , B 39.3 (D=0.51 M) , O 38.4 (D=0.37 small S) , C 37.1 (D=0.17 very small V) & MB 36.6 (D=0.09 V) , all p values <0.001. There were large differences by ethnicity in linear regression slopes for the glucose/HbA1c relationship when compared to W with correlation coefficient comparison Z-statistics for SA 16.74, B 8.69, O 10.94, C 8.96, all p<0.001, except for MB 2.56, p=0.01. Such ethnic differences in HbA1c have been reported previously and questions raised as whether different cut-offs should be used for diagnosis of diabetes. However, more information on the prevalence of microvascular complications and differences in haematological profiles is required for such modifications.

Disclosure

J. A. Williams: None. S. E. Manley: n / a. A. Karwath: None. R. A. Round: None. I. M. Stratton: None. S. Ghosh: Other Relationship; Boehringer Ingelheim International GmbH, Eli Lilly and Company, Novo Nordisk. S. Mostafa: None. G. Roberts: None. J. Webber: None. G. Gkoutos: None.

Funding

Medical Research Council (MR/S003991/1) , MRC HDR UK (HDRUK/CFC/01) , NIHR Birmingham ECMC, NIHR Birmingham SRMRC, Nanocommons H2020-EU (731032) , MAESTRIA (965286)
1339-P: Plasma Heparin Cofactor II Activity Is Inversely Associated with the Development NAFLD in Patients with Diabetes
Authors: HARA TOMOYO , YAMAGAMI HIROKI , MASUDA SHIHO , MITSUI YUKARI , KURAHASHI KIYOE , YOSHIDA SUMIKO , OTODA TOSHIKI , YUASA TOMOYUKI , KURODA AKIO , ENDO ITSURO , MATSUHISA MUNEHIDE , ABE MASAHIRO , AIHARA KEN-ICHI
Published: May 2022

Aims and Introduction: The prevalence and incidence of nonalcoholic fatty liver disease (NAFLD) have been increasing worldwide. Thrombin exerts various pathophysiological functions via activation of protease-activated receptors (PARs) , and activation of PARs promotes the development of NAFLD. Since heparin cofactor II (HCII) specifically inhibits thrombin action by binding to dermatan sulfate proteoglycans in various tissue matrices, we hypothesized that plasma HCII activity is associated with the development of NAFLD.

Materials and Methods: Plasma HCII activity and clinical markers for assessment of hepatic fibrosis including Fibrosis 4 (FIB-4) index, NAFLD fibrosis score (NFS) , and aspartate aminotransferase to platelet ratio index (APRI) were determined in 322 Japanese patients with diabetes mellitus (183 males and 139 females; mean age, 66.0±11.3 years) . The relationships between plasma HCII activities and the clinical markers were statistically evaluated.

Results: The mean plasma HCII activity in all participants was 94.0 ± 17.7%. Multivariate regression analysis with adjustments for confounding factors including age, sex, and serum albumin showed that plasma HCII activity independently contributed to reduction of the FIB-4 index (p < 0.0001) , NFS (p < 0.0001) , and APRI (p < 0.05) .

Conclusions: Plasma HCII activity was independently and inversely associated with the clinical markers of hepatic fibrosis. The results suggest that HCII can serve as a novel therapeutic target for the development of NAFLD in patients with diabetes.

Disclosure

T.Hara: None. I.Endo: None. M.Matsuhisa: Research Support; Nissui , Sysmex Corp., Speaker’s Bureau; Astellas Pharma Inc., Eli Lilly and Company, Merck Sharp & Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Novo Nordisk, Sanofi, Sumitomo Dainippon Pharma Co., Ltd., Takeda Pharmaceutical Company Limited. M.Abe: None. K.Aihara: None. H.Yamagami: None. S.Masuda: None. Y.Mitsui: None. K.Kurahashi: None. S.Yoshida: None. T.Otoda: None. T.Yuasa: None. A.Kuroda: None.
134-LB: Prediabetes Geographic "Hotspots"—Opportunities for Targeted Diabetes Prevention
Authors: SCHMITTDIEL JULIE , YASSIN MAHER , RODRIGUEZ LUIS A. , THOMAS TAINAYAH , MOIN TANNAZ , LEGGETT ALLISON G. , SANCHEZ PERLA , KENRIK DURU O.
Published: May 2022

Over 38% of U.S. adults have prediabetes, or elevated glucose levels linked to type 2 diabetes incidence. The U.S. Preventive Services Task Force recommends giving people who screen positive for prediabetes access to evidence-based prevention strategies such as the Diabetes Prevention Program (DPP) . However, there is little data on prediabetes case clusters (‘hotspots’) to guide in-person program placement and planning. We used ArcGIS software to geospatially map 760,044 adults ages 35-70 with an A1c or FPG screening test in 2019 to census tracts served by Kaiser Permanente Northern California, an integrated health care system with 4.5 million members. A total of 304,727 adults had prediabetes (40%) defined as A1c 5.7-6.4% or FPG 100-125 mg/dL. Seven prediabetes hotspots concentrated in Alameda, Fresno, Madera, Sacramento, San Joaquin, Santa Clara, and Solano counties emerged. Adults living in prediabetes hotspots were less likely to be non-Hispanic White (34% vs. 51%, p<.001) . Hotspots had lower mean Census tract-level median household income ($93,670 vs. $98,216, p<.05) and percent with a college degree or more (34% vs. 43%, p<.001) , but higher percent receiving Medicaid (27% vs. 21%, p<.001) . Prediabetes cases clustered in geographic hotspots with demographics that differed from other areas. Policy makers and health care providers can use this information to target diabetes prevention resources and outreach.

Disclosure

J. Schmittdiel: None. M. Yassin: None. L. A. Rodriguez: None. T. Thomas: None. T. Moin: None. A. G. Leggett: None. P. Sanchez: None. O. Duru: None.

Funding

National Institutes of Health (R18DK122372-01A1)
1342-P: Reduction of Glucokinase and Glucokinase Regulatory Protein Expression by siRNA Is Associated with the Reduction of Lactate and Pyruvate Export from Canine Hepatocytes
Authors: KABIR MORVARID , ADER MARILYN , BERGMAN RICHARD N.
Published: May 2022

Glucokinase (GCK) mediates hepatic glucose uptake during hyperglycemia and is crucial for the regulation of glucose-responsive glycolysis genes. Mutations in GCK are responsible for maturity-onset diabetes of the young, and glucokinase regulatory protein (GKRP) , the regulator of hepatic GCK, is also a diabetes susceptibility locus. Recently in in vivo experiments, we used hepatic lactate export as a surrogate for GCK activity, which was a measure of “glucose effectiveness.” We hypothesized that in vitro GCK and GKRP suppression in hepatocytes promotes a reduction in lactate and pyruvate export from the liver. We used cryopreserved canine hepatocytes and knocked down GCK and GKRP via lipid delivery of siRNA. A control group did not receive siRNA. We measured lactate dehydrogenase D (LDHD) and pyruvate kinase (PK) gene expression and lactate and pyruvate release into the medium 24 and 48 h after siRNA delivery. Selective blocking of GCK and GKRP did not change cell proliferation but reduced the expression of these genes by 90% and 85%, respectively (P<0.0vs. controls; n=4) . GCK and GKRP knockdown reduced LDHD by 71% and 74%, respectively (P<0.001) ; similarly, PK was reduced by 84% and 82%, respectively (P<0.001) . Lactate exported into the medium was reduced by 36% (P<0.001) and 26% (P<0.05) by GCK and GKRP siRNA, respectively, after 48 h of hepatocyte culture. However, pyruvate was reduced by 17% only after 48 h culture of GCK siRNA (P<0.05) . Thus, we showed that direct suppression of GCK and GKRP genes by siRNA in vitro affects glucose-responsive glycolysis genes, consequently reducing lactate export which can be used as a surrogate for reduced glucokinase activity in hepatocytes. The use of antisense oligonucleotides or CRISPR gene-editing methods in animals will be useful methods to study the glucose-lactate relationship in vivo and will evaluate the role of glucose effectiveness in the pathogenesis of type 2 diabetes.

Disclosure

M.Kabir: None. M.Ader: None. R.N.Bergman: Consultant; Fractyl Health, Inc., Novo Nordisk, Research Support; AstraZeneca, Janssen Research & Development, LLC.

Funding

National Institutes of Health R01- DK027619-28
1345-P: Low-Dose Colchicine/Metformin Combination Ameliorates NASH via Regulating AMPK/NRF2/GDF15 Axis in Mice
Authors: HAM SUN AH , CHO HEE-YEON , JEONG CHOLONG , YOUN JU HO
Published: May 2022

Nonalcoholic steatohepatitis (NASH) , a complex metabolic disease by multifactorial pathogenesis is demanded multiple pharmacological modes of action for robust treatment strategies. Colchicine has shown efficacy in various clinical settings which inflammation is a key component, however growing evidences suggest that the therapeutic effect of colchicine is multifaceted by intervening in several different pathways involved in inflammation. Here, we investigated the novel therapeutic effects of low-doses of colchicine (LDC) -metformin (LDC-Met) combination to targeting NASH. To induce NASH disorder, C57BL/6J male mice were fed a CDHFD (choline-deficient high fat diet) for 12 weeks, then were orally administrated by LDC-Met with CDHFD for an additional 12 weeks. In CDHFD induced NASH mice models, LDC-Met markedly decreased the NAS and fibrosis score along with reducing serum level of liver enzymes and total cholesterol increased by CDHFD feeding. Consistent with these data, hepatic gene expression of steatosis, fibrosis, inflammation and apoptosis markers increased by CDHFD was noticeably inhibited by LDC-Met. Interestingly, significant increased circulating GDF15 was found in LDC-Met group, which was accompanied by increased hepatic GDF15 expression. Additionally, LDC-Met significantly augmented hepatic phosphorylation of AMPK and NRF2 in CDHFD-fed mice and NLRP3 inflammasome activity were negatively controlled by LDC-Met. Finally, LDC-Met noticeably reduced CDHFD-induced total body weight gain by approximately 20%, along with epididymal white adipose tissue (eWAT) weight reduction. Taken together, we found that the combination of LDC-Met has the best synergistic effect on NASH resolution, suggesting significant implications for potential drug repurposing and new drug development. Moreover, this novel combination elevates circulating levels of GDF15, which is necessary to obtain beneficial effects on energy balance and body weight.

Disclosure

S.Ham: None. H.Cho: None. C.Jeong: None. J.Youn: None.
1347-P: High Prevalence of NAFLD and Lower Lean Mass in South Asian Youth
Authors: HITT TALIA A. , HUI STEVE C.N. , KAMEL IHAB R. , BARKER PETER B. , ZEMEL BABETTE S. , KELLY ANDREA , MAGGE SHEELA N.
Published: May 2022

Background: South Asians (SA) have high rates of NAFLD. SA adults have NAFLD at lower BMI. We examined differences in ectopic fat between SA and White (W) youth, and its relationship to BMI z-score (BMIZ) , fat depots, and metabolic labs.

Methods: SA and W youth, 12-21y, BMI ≥ 80%ile (or ≥23kg/m2 if ≥18y) underwent MRS to measure liver fat and intramyocellular lipid (IMCL) (soleus) , and DXA to quantify total % body fat (%BF) and lean mass (LM) . Abdominal MRI was used to measure abdominal fat (AF) , visceral fat (VF) , SQ fat, deep SQ, and superficial SQ. HbA1c, fasting glucose (FBG) , and lipids were obtained. Univariate analyses used parametric or non-parametric methods as appropriate. Linear regression adjusted for potential confounding variables. NAFLD defined as >5% liver fat.

Results: Participants (28 SA, 9 W) had similar age, sex, and BMIZ. Liver fat was higher in SA [2.0% (1.1, 7.4) ] compared to W [0.8% (0.7, 1.4) ], but not statistically different. NAFLD was present in 29% of SA youth, but in no W youth. There was no difference in IMCL (p=0.9) or VF between groups (p=0.5) . Adjusting for age, sex, puberty and BMIZ, SA had lower LM [β= -8.1, p=0.003] and higher %BF [β=5.8, p=0.01], AF [β=151.6, p=0.006], total SQ fat [β=65.9, p=0.001], deep SQ fat [β=1.9, p=0.005] and HbA1c [β=3.8, p=0.01] than W. FBG and lipids were similar between groups. In SA, liver fat correlated with HDL-C (p=0.02) , LDL-C (p=0.03) , non-HDL-C (p=0.03) and borderline with FBG (p=0.05) , HbA1c (p=0.06) , but not with BMIZ, LM, %BF, or triglycerides. In SA, VF was correlated with triglycerides (p=0.002) and non-HDL-C (p=0.04) and borderline with LM (p=0.05) and HDL-C (p=0.06) .

Conclusions: These preliminary results of an ongoing study demonstrate that 29% of SA youth had NAFLD compared to no W youth. Liver fat in SA youth was not related to typical metabolic risk factors. Notably SA youth had lower LM, and increased AF, deep SQ fat, and HbA1c compared to W youth, after adjustment. Further study is needed to clarify the mechanisms underlying excess liver fat in SA youth.

Disclosure

T.A.Hitt: None. S.C.N.Hui: None. I.R.Kamel: None. P.B.Barker: None. B.S.Zemel: None. A.Kelly: Research Support; vTv Therapeutics. S.N.Magge: None.

Funding

National Institutes of Health (R01DK115648)
1349-P: Bariatric Surgery Causes an Early Improvement in the Lipidomic Profile and a Decrease in Plasma Ceramides and Diacylglycerols in Patients with Type 2 Diabetes and NAFLD
Authors: KALAVALAPALLI SRILAXMI , BARB DIANA , DILLARD RACHEL E. , GODINEZ EDDISON , FRIEDMAN JEFFREY , FANOUS NADA , WARREN CURTIS O. , BRIL FERNANDO , LOMONACO ROMINA , ZARRINPAR ALI , CUSI KENNETH
Published: May 2022

Bariatric surgery (BarS) is known to improve insulin resistance (IR) , atherogenic dyslipidemia and steatohepatitis in patients (pts) with type 2 diabetes (T2D) . Total ceramides (Cer) and diacylglycerols (DAGs) have been implicated in the development of IR and nonalcoholic fatty liver disease (NAFLD) , but the impact of BarS on their plasma concentration remains unclear. The aim of this study was to assess early changes in the plasma lipidome, before and 3-6 months post BarS. To this end, we recruited 17 pts with NAFLD who underwent BarS (age 47±3, BMI 46±2 kg/m2, 76% with T2D) , and 6 controls without T2D (baseline data only, age 47±6, BMI 31.2±2.1 kg/m2) . Global lipidomic profiling was performed on orbitrap mass spectrometer with UHPLC. Data was analysed using LipidMatch software. Plasma levels of total lipids (TL) (34%; p=0.01) , Cer (20%; p=0.31) , DAGs (50%; p=0.03) and total triglycerides (TG) (79%; p=0.01) were higher in pts with NAFLD compared to controls. Bariatric surgery reduced plasma TL (-26%) , Cer (-31%) , DAGs (-34%) and TG (-43%) levels (all p<0.01) . In pts with vs. without T2D, plasma TL were higher (28%; p=0.01) . There was a reduction in TL (-29%; p<0.01) , Cer (-22%; 0.27) , DAGs (-41%; p=0.02) and TG (-52%; p<0.01) post BarS and a similar pattern was observed across the lipidomic profile (cholesterol esters, phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol) . Reduction in weight after BarS correlated with lower DAGs (r=0.81; p=0.03) and TG (r=0.66; p=0.07) . Reduction in Cer correlated with improved HOMA-IR (r= 0.71; p=0.04) and Adipo-IR (r= 0.63; p=0.09) .

In conclusion, pts with NAFLD and T2D have a significantly worse lipidomic profile compared to pts without T2D. The unfavorable lipidomic profile was normalized after BarS, in particular Cer and DAG levels, known to be linked to IR. Assessment of the lipidomic profile may provide further insights into insulin sensitivity changes after BarS.

Disclosure

S.Kalavalapalli: None. A.Zarrinpar: None. K.Cusi: Consultant; Altimmune, Arrowhead Pharmaceuticals, Inc., AstraZeneca, Boehringer Ingelheim International GmbH, Bristol-Myers Squibb Company, Eli Lilly and Company, Fractyl Health, Inc., Genentech, Inc., Hanmi Pharm. Co., Ltd., Intercept Pharmaceuticals, Inc., Novo Nordisk. D.Barb: None. R.E.Dillard: None. E.Godinez: None. J.Friedman: n/a. N.Fanous: None. C.O.Warren: None. F.Bril: None. R.Lomonaco: None.
1350-P: Elucidation of Terminal Fatty Acid–Derived Acetyl-CoA Preference for Ketogenic Flux
Authors: MCLEOD MARC , RAGAVAN MUKUNDAN , RUSHIN ANNA , MERRITT MATTHEW , GIACALONE ANTHONY G.
Published: May 2022

Insulin resistance is a major contributor to nonalcoholic fatty liver disease (NAFLD) . NAFLD is a progressive disease that leads to impaired hepatic function and liver cirrhosis. Increased ketogenesis has been correlated with a protective phenotype in mice and humans. During our study we identified differences in acetyl-CoA partitioning between the oxidative tricarboxylic acid cycle (TCA) , and ketogenesis. 8 week old C57Bl6 mice were fed a high fat or low fat (60% or 10% fat by calories respectively) diet for 16 weeks before performing liver perfusion in 12-14 hours fasted mice with 1 mM [D15]-octanoate fatty acid tracer. The effluent perfusate was collected and GC-MS analysis was performed to identify the distribution of deuterium label. Enrichments were greater in ketones (˜70%) than the TCA cycle (˜20%) with the unexpected determination that terminal acetyl-CoA units derived from fatty acids were preferentially used for ketogenesis as compared to internal acetyl-CoA units (Figure 1) . These changes were confirmed with fractional enrichment analysis and metabolic modeling of the ketogenic pathway and TCA cycle. This finding elucidates differential handling of acetyl-CoA that could be exploited for therapeutic benefit to NAFLD.

Disclosure

M.Mcleod: None. M.Ragavan: None. A.Rushin: None. M.Merritt: None. A.G.Giacalone: None.
1352-P: Differential Metabolomics and Lipidomics Profiles in Low-Birth-Weight Men Unmasked by Four-Weeks High-Carbohydrate Overfeeding
Authors: VILLUMSEN SOFIE O. , JUSTESEN LOUISE , ELINGAARD-LARSEN LINE OHRT , THUESEN ANNE CATHRINE B. , KIM MIN , LEGIDO-QUIGLEY CRISTINA , DANIELSEN ELSE R. , HANSEN TORBEN , BRØNS CHARLOTTE , VAAG ALLAN A.
Published: May 2022

Background: We hypothesized that low birth weight (LBW) subjects with a known increased risk of type 2 diabetes exhibit increased sensitivity towards the deleterious metabolic effects of 4-weeks high carbohydrate overfeeding (HCOF) compared with normal birth weight (NBW) controls.

Methods: Untargeted serum metabolomics and lipidomics were measured before and after 4-weeks HCOF (+25% energy) in 26 non-obese men with LBW (2789±174 g) and 21 normal birth weight (NBW) men/controls (3804±172 g) matched for BMI and age. Data were analyzed using a combination of advanced statistical and bioinformatical tools.

Results: Among 65 identified metabolites, 8 metabolites were affected differentially by HCOF between the two groups (p<0.05) . In response to HCOF, ingenuity pathway analysis (IPA) revealed increased accumulation and peroxidation of lipids in LBW subjects. In contrast, NBW controls increased albumin synthesis after HCOF. Changes in upstream regulators such as nicotinamide nucleotide transhydrogenase (NNT) , uncoupling protein 1 (UCP1) , and carbonic anhydrase 9 (CA9) were observed in both groups in response to HCOF. NBW controls displayed increased NNT and UCP1 activation, whereas LBW subjects responded with paradoxically decreased NNT and UCP1 activation. CA9 expression was decreased in both groups. Network correlation analysis revealed an upregulation of fatty acids in LBW subjects, contrasting a downregulation among NBW controls. Among 279 lipids identified, the LBW subjects exhibited a significantly higher number of lipids to be increased in response to HCOF (n=26) compared to NBW subjects (n=6) (PFisher=0.0004) , and IPA analysis revealed an activation of the PPARG pathway in NBW as opposed to a paradoxical inhibition in LBW subjects.

Conclusion: The results document severe differential perturbations of lipid metabolism in LBW men exposed to HCOF. Public health initiatives promoting low carbohydrate diets in LBW subjects are warranted.

Disclosure

S.O.Villumsen: None. A.A.Vaag: Stock/Shareholder; AstraZeneca. L.Justesen: None. L.Elingaard-larsen: None. A.B.Thuesen: None. M.Kim: None. C.Legido-quigley: None. E.R.Danielsen: None. T.Hansen: None. C.Brøns: Stock/Shareholder; Novo Nordisk.
1355-P: Carbohydrate Overfeeding Unmasks Severe Dysmetabolic Traits in Low-Birth-Weight Men
Authors: BRØNS CHARLOTTE , THUESEN ANNE CATHRINE B. , JUSTESEN LOUISE , ELINGAARD-LARSEN LINE OHRT , STØRLING JOACHIM , DANIELSEN ELSE R. , HANSEN TORBEN , VAAG ALLAN A.
Published: May 2022

Background: Low birth weight (LBW) individuals are at increased risk of developing Type 2 diabetes (T2D) , especially when exposed to an affluent dietary pattern. Impaired subcutaneous adipose tissue expandability and ectopic fat storage may represent key features explaining the link between LBW and risk of T2D. We hypothesized that a 4-week simple carbohydrate overfeeding challenge (+25% energy) (COF) would adversely influence dysmetabolic traits including hepatic fat content in LBW compared to normal birth weight (NBW) men.

Methods: Twenty-two non-obese LBW and 21 age- and BMI-matched NBW men with a mean age of 37.6 years were included. Body composition, hepatic fat (MR spectroscopy) , glucose and insulin metabolism, hepatic glucose production (HGP; deuterium glucose) , energy metabolism (indirect calorimetry) and selected plasma biomarkers (multiplex ELISA) were measured before and after 4 weeks COF.

Results: Fasting plasma glucose and C-peptide levels, as well as energy expenditure (EE) and fat oxidation (FOX) rates increased significantly more in response to COF among the LBW compared to NBW subjects (all P<0.05) . However, despite significantly increased baseline hepatic fat content, COF resulted in similar body weight and hepatic fat content increments in LBW compared to NBW subjects. HGP and hepatic insulin resistance were unaffected by COF in both groups. Fasting plasma adiponectin levels were significantly reduced in LBW versus NBW subjects before and after COF, whereas plasma leptin and FGF21 levels were increased (both P<0.005) after COF in LBW men only.

Conclusion: LBW compared to NBW subjects respond to COF with differential increments in plasma glucose, C-peptide, leptin and FGF21 levels, as well as increased EE and FOX rates. The results underscore the importance of LBW individuals avoiding simple carbohydrate overfeeding. Further studies are needed to understand the role of increased hepatic fat and insulin resistance as key mechanisms linking LBW with increased risk of T2D.

Disclosure

C.Brøns: Stock/Shareholder; Novo Nordisk. A.B.Thuesen: None. L.Justesen: None. L.Elingaard-larsen: None. J.Størling: None. E.R.Danielsen: None. T.Hansen: None. A.A.Vaag: Stock/Shareholder; AstraZeneca.

Funding

Novo Nordisk Foundation (NNF15OC0016692) , Aase and Ejnar Danielsens Fond, Augustinus Foundation, Simon Spies Fonden, TrygFonden.