Impaired Beta-cell Function Research Articles

Reduced expression of stearoyl-CoA desaturase-1, but not free fatty acid receptor 2 or 4 in subcutaneous adipose tissue of patients with newly diagnosed type 2 diabetes mellitus

BackgroundIn subcutaneous adipose tissue (SAT), higher stearoyl-CoA desaturase-1 (SCD1) expression has been related to improved insulin sensitivity in thiazolidinedione-treated type 2 diabetes mellitus patients. In animal models, deficiency of the free fatty acid receptor (FFAR) 2 associated with higher and FFAR4-deficiency with lower insulin sensitivity. We hypothesized that increased FFAR2 expression and reductions in FFAR4 and SCD1 expression in SAT of type 2 diabetes mellitus patients associate positively with insulin resistance and impaired beta cell function.MethodsTwenty-five type 2 diabetes mellitus patients and 25 glucose-tolerant humans (CON) matched for sex, age, and BMI underwent mixed-meal tests to assess insulin sensitivity (OGIS) and beta cell function (ΔAUC(C-peptide)0–180 min/ΔAUC(glucose)0–180 min) in a cross-sectional study. Gene and protein expression of SCD1 and FFAR2/4 were quantified in SAT biopsies.ResultsInsulin sensitivity was 14% and beta cell function 71% (both p < 0.001) lower in type 2 diabetes mellitus patients. In type 2 diabetes mellitus, SCD1 mRNA was fivefold (p < 0.001) and protein expression twofold (p < 0.01) lower. While FFAR2/4 mRNA and protein expression did not differ between groups, FFAR2 protein levels correlated negatively with beta cell function only in CON (r = −0.74, p < 0.01). However, neither SCD1 nor FFAR2/4 protein expression correlated with insulin sensitivity in both groups.ConclusionsType 2 diabetes patients have lower SCD1, which does not associate with insulin resistance. Only in non-diabetic humans, FFAR2 associated with impaired beta cell function.

Abstract 235: Type 2 diabetes and glycemic traits in relation to colorectal cancer risk: A Mendelian randomization study

Abstract Background: Although type 2 diabetes (T2D) has been linked to an increased risk of colorectal cancer (CRC), a causal relationship remains to be established. Also, the influence of hyperglycemia and impaired beta-cell function on CRC remains to be determined. Methods: We performed a Mendelian randomization (MR) study using common single nucleotide polymorphisms (SNPs) identified from prior genome-wide association studies for T2D (78 SNPs), fasting glucose (36 SNPs), glycosylated hemoglobin (HbA1c, 11 SNPs), and homeostatic model assessment for beta-cell function (HOMA-B, 13 SNPs) (all SNPs reached genome-wide significance at 5 x10-8). We first assessed the association of each SNP with CRC risk using logistic regression in 26,357 CRC cases and 20,505 control participants in the Colon Cancer Family Registry, Colorectal Transdisciplinary Study and the Genetics and Epidemiology of Colorectal Cancer Consortium. The MR estimates for the association of each risk factor with CRC were calculated using the inverse-variance weighted method that combines summary data on the association of the SNPs with each trait derived from previous genome-wide association studies. Given the potential sex difference in the relationship of metabolic factors with CRC, all analyses were performed in men and women separately. Results: We did not find any statistically significant association between each of the genetically predicted risk factors and CRC incidence for sex-stratified or combined analyses. The odds ratios (ORs) of CRC associated with genetic risk of T2D were 1.02 (95% confidence interval [CI], 0.95-1.08, P=0.62) in women and 0.96 (95% CI, 0.90-1.03, P=0.25) in men. For fasting glucose, the ORs of CRC associated with one mmol/L increment was 0.91 (95% CI, 0.68-1.20, P=0.49) in women and 1.20 (95% CI, 0.90-1.59, P=0.22) in men, respectively. For HbA1c, the ORs of CRC associated with one percent increment was 0.83 (95% CI, 0.57-1.22, P=0.35) in women and 0.99 (95% CI, 0.67-1.45, P=0.95) in men. For HOMA-B, the ORs of CRC associated with a unit increase in log-transformed measurement was 0.92 (95% CI, 0.53-1.60, P=0.76) in women and 0.77 (95% CI, 0.44-1.35, P=0.36) in men. Conclusion: Our findings do not support a causal association of T2D, impaired beta-cell function, and hyperglycemia with CRC risk. However, the null findings may be due to weak instrument bias since these common SNPs account for only a limited proportion of the inter-individual variation in these glycemic traits. Citation Format: Mingyang Song, Yiwen Lu, Marc Gunter, Neil Murphy, Barbara L. Banbury, Wenjie Ma, Jennifer Prescott, Graham Casey, Stephen B. Gruber, Edward L. Giovannucci, Ulrike Peters, Andrew T. Chan. Type 2 diabetes and glycemic traits in relation to colorectal cancer risk: A Mendelian randomization study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 235.

Pregnancy Glycemia Reflects Life Course Glycemia of the Mother

Background: Current practice of diagnosis and treatment of GDM is largely restricted to late pregnancy, ignoring peri-conceptional window which is crucial for fetal programming of diabetes. A few longitudinal studies reported that metabolic risk factors of GDM are present before pregnancy but there are no reports on a ‘lifecourse’ trajectory of pregnancy glucose insulin metabolism. Pune Maternal Nutrition Study (PMNS) offers a unique opportunity to investigate the association between gestational glycaemia and pre-pregnancy lifecourse glycaemia because of serial measurements from childhood to pregnancy. Methods: We studied 328 female offspring of PMNS born in 1993-96 for glucose insulin measurements at 6, 12, and 18 year of age. Girls who became pregnant underwent a 75 gm OGTT at 28 weeks gestation. We studied the association between gestational fasting plasma glucose (FPG) with their earlier measurements using multiple linear regression analysis. Results: Up to October 2017, 110 women had an OGTT at 28 weeks gestation and have delivered. Thirteen (11.8%) were diagnosed GDM (IADPSG criteria 2011) In lifecourse analysis, FPG tracked from 6 years of age to pregnancy through 12 and 18 years. FPG at 28 weeks was directly associated with her own FPG at 18, 12 and 6 years of age (β= ∼0.3, p&amp;lt;0.001), and it was inversely associated with disposition index at 18 and 12 years (p&amp;lt;0.05). Conclusions: The world’s first description of lifecourse evolution of pregnancy glycaemia suggests that it tracks from early childhood and is associated with impaired beta cell function. The analysis provides an important clue that GDM women have higher pre and peri-conceptional glycaemia which is a risk factor for fetal programming of diabetes. Current clinical approach to GDM ignores this important window. This could be a potential explanation of the failure of current practice of GDM treatment in late pregnancy to reduce long term risks in the offspring. Disclosure N.S. Memane: None. D. Bhat: None. D.A. Raut: None. S.J. Bondarde: None. R. Ladkat: None. P.C. Yajnik: None. C. Fall: None. C.S. Yajnik: None.

Mild Physiologic Hyperglycemia Impairs Beta-Cell Function and Induces Insulin Resistance in Healthy Normal Glucose Tolerant Subjects

The aim of the present study was to evaluate effect of a physiologic increase (+45 mg/dl) in plasma glucose concentration on insulin secretion in healthy NGT individuals. 20 NGT subjects: 12 without family history of T2DM (FH-) (9M/3F, age = 50± 4 years, BMI = 27 ± 1 kg/m2) and 8 with FH (FH+) (4M/4F, Age = 48±2, BMI = 26±1) received an OGTT and 2-step hyperglycemic (+125 and +300 mg/dl) clamp (duration of each step = 80 minutes) followed by IV arginine (5g) bolus. The acute insulin response (AIR0-10min), 2nd phase (SP) insulin secretion (IR) (SPIS10-80 and SIS90-160 minutes during each hyperglycemic step and following arginine (AIRArg) were assessed. Insulin sensitivity was assessed as the glucose infusion rate/steady state plasma insulin (M/I) during the hyperglycemic clamp. FPG concentration increased from 97±4 to 140±4 mg/dl for 48 hours by a variable IV glucose infusion. First phase insulin secretion (0-10 min) and 2nd phase insulin secretion (10-80 and 90-60 min) increased by 59% and 78%, respectively, following chronic glucose infusion while insulin sensitivity (M/I) during both hyperglycemic clamp steps declined (15± 3.5 to 12±2.8 and 20.8 ± 4.7 to 8.1 ± 2.0, both p&amp;lt;0.02). Consequently, the insulin secretion/insulin resistance (disposition) index declined by 85± 18% (1st clamp step) and 81±45% (2nd clamp step) (both p&amp;lt;0.05) following chronic glucose infusion. Similar changes were observed when indices of insulin secretion were measured using C-peptide. There was no difference in the effect of chronic glucose infusion on insulin secretion and insulin sensitivity between subjects with and without FH of T2DM. These results demonstrate that: (i) sustained physiologic hyperglycemia impairs insulin sensitivity (glucotoxicity); (ii) increases absolute insulin secretion; (iii) impairs beta cell function as measured with the disposition index (glucotoxicity). Disclosure A. Merovci: None. E.R. Maldonado Corchado: None. D. Tripathy: None. R.A. DeFronzo: Speaker's Bureau; Self; AstraZeneca, Novo Nordisk Inc.. Advisory Panel; Self; AstraZeneca, Novo Nordisk Inc., Janssen Pharmaceuticals, Inc., Boehringer Ingelheim Pharmaceuticals, Inc., Elcelyx Therapeutics, Inc., Intarcia Therapeutics, Inc.. Research Support; Self; AstraZeneca, Boehringer Ingelheim Pharmaceuticals, Inc., Janssen Pharmaceuticals, Inc., Takeda Pharmaceuticals U.S.A., Inc..

Pharmacological Neprilysin Inhibition Improves Glucose Homeostasis in a Mouse Model of Type 2 Diabetes

Neprilysin (NEP) is a ubiquitous peptidase whose plasma activity in humans and rodents increases in conditions associated with type 2 diabetes (T2D). In high fat-fed mice, increased plasma NEP activity correlates with insulin resistance and impaired beta-cell function, and we’ve shown genetic ablation of NEP ameliorates these deleterious effects, thereby decreasing glucose levels. Thus, we sought to determine whether clinically relevant, pharmacological inhibition of NEP also confers beneficial effects on glycemic status in a mouse model of T2D. Five-week old C57BL/6 mice were fed high fat diet for 3 weeks, followed by injection of low-dose beta-cell toxin streptozotocin (30 mg/kg i.p. once daily for 3 consecutive days; STZ) to induce diabetes, or vehicle (VEH) as a control. Mice were continued on high fat diet alone (CON) or supplemented with the NEP inhibitor sacubitril (48 mg/kg/day; SAC) for 8 weeks. Body weight did not differ among the 3 groups of mice at baseline (Table), and increased to a similar extent in all groups after 8 weeks of treatment. Further, both fasting and fed glucose levels were higher in STZ-CON vs VEH-CON mice after 8 weeks, but significantly lower in STZ-SAC vs STZ-CON mice. Together, these data show that pharmacological NEP inhibition improves both fasting and fed glucose levels in a mouse model of T2D, and thus may be a useful treatment strategy in human T2D.VEH-CON (n=3)STZ-CON (n=5)STZ-SAC (n=7)Body weight (g)Baseline19.9±0.220.9±0.418.6±0.5Post-Treatment35.1±3.532.1±1.331.0±1.9Plasma glucose levels post-treatment (mg/dl)fasting145±5*314±38184±21*fed292±6*459±41317±29**p&amp;lt;0.vs STZ-CON Disclosure J.H. Parilla: None. S. Mongovin: None. B. Barrow: None. N. Esser: None. S. Zraika: None.

Reduced Lower-Limb Muscle Strength in Type 2 Diabetes Patients with Arthritis Is Associated with Impaired Beta-Cell Function

Insulin levels might affect the integrity of musculoskeletal structures. We hypothesized that in type 2 diabetes patients (T2D) with arthritis musculoskeletal impairment is associated with insulin secretion. This analysis of the German Diabetes Study comprised T2D with (T2D+A (n[m/f]=12[4/8], age 63±9 years, BMI 34.8±7.2 kg/m2) or without arthritis (T2D-A: 22[12/10], 59±13 years, 32.8±6.5 kg/m2) and healthy controls (CON: 18[10/8], 42±16 years, 25.3±4.1 kg/m2). Group comparison analyses were adjusted for age, sex and BMI. T2D duration was &amp;lt;5 years. Beta-cell function was assessed from C-peptide levels during glucagon stimulation test (GST) and intravenous glucose tolerance tests (IVGTT), insulin sensitivity (IS) from hyperinsulinemic-euglycemic clamps. Isometric knee extension force (KEF) was assessed with a dynamometer and range of motion (ROM) was measured using a goniometer. IS was comparable in T2D+A and T2D-A (3.5±0.7 vs. 5.1±2.4 mg*kg-¹*min-¹), but lower than in CON (11.5±3.3 mg*kg-¹*min-¹, p&amp;lt;0.05). Hemoglobin A1c (HbA1c) was similar in T2D+A and T2D-A (53±6 vs. 51±11 mmol/mol), but higher than in CON (33±3 mmol/mol, p&amp;lt;0.05). In T2D+A, C-peptide secretion (IVGTT) was 43% and 63% lower (p&amp;lt;0.05) than in T2D-A and in CON, respectively. C-peptide secretion (IVGTT) correlated inversely with HbA1c levels in T2D+A (r=-0.81, p&amp;lt;0.05) and T2D-A (r=-0.60, p&amp;lt;0.05). In T2D+A, KEF was 60% lower than in CON (19±11 kg vs. 47±12 kg, p&amp;lt;0.05) and correlated negatively with C-peptide secretion (GST; r=-0.99, p&amp;lt;0.05). ROM was 8% and 19% lower (p&amp;lt;0.05) in T2D+A than in T2D-A and CON, respectively. Within T2D+A, ROM was negatively correlated with insulin secretion (IVGTT; r=-0.64, p&amp;lt;0.05). In conclusion, musculoskeletal impairment is associated with decreased beta-cell function in T2D patients suffering from arthritis, possibly mediated by chronic hyperglycemia. Disclosure O.P. Zaharia: None. D. Pesta: None. P. Bobrov: None. Y. Kupriyanova: None. K. Bódis: None. V. Burkart: None. J. Hwang: None. K. Müssig: None. M. Roden: Speaker's Bureau; Self; Boehringer Ingelheim GmbH. Research Support; Self; Boehringer Ingelheim GmbH. Consultant; Self; Poxel SA. Research Support; Self; Danone Nutricia Early Life Nutrition, GlaxoSmithKline plc., Nutricia Advanced Medical Nutrition, Sanofi. J. Szendroedi: None.

Elevated Insulin Clearance Contributes to the Reduced Peripheral Insulin Concentration Observed in Obese Young Asian with Type 2 Diabetes

Introduction: The importance of insulin clearance (CLins) on insulin dynamics and the role exerted in controlling glucose in type 2 subjects has recently been appreciated. Moreover, evidence in mice showed that CLins is reduced by incretin hormones at physiological levels. Aim: To evaluate in 29 young (19±1 years) multi-ethnic Asian type 2 subjects (T2D) the contribution of CLins to insulin dynamics and if an association exists with GLP-1. Methods: T2D (15F/14M; BMI=30.2±0.9 kg/m2; HbA1c=8.8±0.4% [73±4.4 mmol/mol]; mean duration of disease 2.5 years) had fasting glucose Gb=192±14 mg/mL and insulin Ib=19±3 µU/mL. They were compared to 17 (8F/9M) age-matched nondiabetic overweight (BMI=27.8±1.0, p=0.1) control subjects (CT, Gb=82±1, p&amp;lt;0.0001; Ib=12±2, p=0.09). All underwent a 75g 2h OGTT with measurements of glucose, insulin, C-peptide and GLP-1. Insulin secretion (ISR) was assessed by deconvolution from peripheral C-peptide; CLins) by ISR/AUCinsulin; beta cell function (BCF) by AUCC-peptide/AUCglucose; insulin sensitivity by OGIS model. GLP-1 effect on BCF (GLPE) was empirically calculated as BCF/AUCGLP-1. Results: T2D were insulin resistant (OGIS=277±16 mL/min m2 vs. 424±14) and had low BCF (82±11 pmolC-peptide/mmolglucose vs. 123±10), both p&amp;lt;0.0001. Lower total AUCinsulin was observed in T2D (4.7±0.7 U/L 2h vs. 11.0±1.4, p&amp;lt;0.0001), with higher CLins (2.06±0.19 L/min) than in CT (1.46±0.10, p=0.023). CLins directly correlated with OGIS in CT (R=0.55, p=0.02), but not in T2D (p=0.98). Despite no difference was observed in AUCGLP-1 (1.6±0.1 nmol/L 2h vs. 1.7±0.1 in CT, p=0.63), which did not correlate with CLins (p&amp;gt;0.2), GLPE was lower in T2D (6.2±0.9 vs. 23.4±2.3, p&amp;lt;0.0001). Conclusion: Reduced peripheral insulin availability in obese young Asian T2D is due not only to a markedly impaired beta cell function and reduced GLP-1 effect, but also to an increased insulin clearance, which however shows no association to GLP-1 response. Disclosure G. Pacini: None. A. Tura: None. C. Hor: None. S. Lim: None. F. Tan: None. C. Tong: None. J.Y.H. Hong: None. F. Md Zain: None. J.J. Holst: Advisory Panel; Self; Novo Nordisk A/S. Board Member; Self; Zealand Pharma A/S. Speaker's Bureau; Self; Merck Sharp &amp; Dohme Corp., AstraZeneca. Research Support; Self; Danish Diabetes Academy, Novo Nordisk Foundation. Other Relationship; Self; Antag Therapeutics. Other Relationship; Spouse/Partner; Antag Therapeutics. W. Mohamud: None. T. Yeow: None.

Beta cell stress in a 4-year follow-up of patients with type 2 diabetes: A longitudinal analysis of the BetaDecline Study.

Type 2 diabetes mellitus (T2DM) is associated with a progressive deterioration in beta cell function and loss of glycaemic control. Clinical predictors of beta cell failure are needed to guide appropriate therapy. A prospective evaluation of a large set of potential predictors of beta cell stress, measured as change in the proinsulin/insulin (PI/I) ratio, was conducted in a cohort of 235 outpatients with T2DM on stable treatment with oral hypoglycaemic agents or diet followed up for ~4years (median value 3.9years; interquartile range 3.8-4.1years). Overall, metabolic control deteriorated over time, with a significant increase in glycated haemoglobin (HbA1c; P<.0001), proinsulin (P<.0001), and PI/I ratio (P=.001), without significant changes in the homeostatic model assessment of insulin resistance. Multivariate regression analysis showed that for each 1% (10.9mmol/mol) increase from baseline in HbA1c, the risk of beta cell stress increased by 3.8 times; for each 1% (10.9mmol/mol) incremental increase in HbA1c during the study, risk of beta cell stress increased by 2.25 times that at baseline. By contrast, baseline anthropometric and clinical variables, lipid profile, inflammatory markers (PCR, IL-6), non-esterified fatty acids, and current therapies did not independently influence PI/I ratio variation during follow-up. In this cohort of patients with T2DM, beta cell function progressively deteriorated despite current therapies. Among a large set of clinical and biochemical predictors, only baseline HbA1c levels and their deterioration overtime were associated with higher beta cell stress over time.

Exercise training decreases pancreatic fat content and improves beta cell function regardless of baseline glucose tolerance: a randomised controlled trial

Aims/hypothesisPancreatic fat accumulation may contribute to the development of beta cell dysfunction. Exercise training improves whole-body insulin sensitivity, but its effects on pancreatic fat content and beta cell dysfunction are unclear. The aim of this parallel-group randomised controlled trial was to evaluate the effects of exercise training on pancreatic fat and beta cell function in healthy and prediabetic or type 2 diabetic participants and to test whether the responses were similar regardless of baseline glucose tolerance.MethodsUsing newspaper announcements, a total of 97 sedentary 40–55-year-old individuals were assessed for eligibility. Prediabetes (impaired fasting glucose and/or impaired glucose tolerance) and type 2 diabetes were defined by ADA criteria. Of the screened candidates, 28 healthy men and 26 prediabetic or type 2 diabetic men and women met the inclusion criteria and were randomised into 2-week-long sprint interval or moderate-intensity continuous training programmes in a 1:1 allocation ratio using random permuted blocks. The primary outcome was pancreatic fat, which was measured by magnetic resonance spectroscopy. As secondary outcomes, beta cell function was studied using variables derived from OGTT, and whole-body insulin sensitivity and pancreatic fatty acid and glucose uptake were measured using positron emission tomography. The measurements were carried out at the Turku PET Centre, Finland. The analyses were based on an intention-to-treat principle. Given the nature of the intervention, blinding was not applicable.ResultsAt baseline, the group of prediabetic or type 2 diabetic men had a higher pancreatic fat content and impaired beta cell function compared with the healthy men, while glucose and fatty acid uptake into the pancreas was similar. Exercise training decreased pancreatic fat similarly in healthy (from 4.4% [3.0%, 6.1%] to 3.6% [2.4%, 5.2%] [mean, 95% CI]) and prediabetic or type 2 diabetic men (from 8.7% [6.0%, 11.9%] to 6.7% [4.4%, 9.6%]; p = 0.036 for time effect) without any changes in pancreatic substrate uptake (p ≥ 0.31 for time effect in both insulin-stimulated glucose and fasting state fatty acid uptake). In prediabetic or type 2 diabetic men and women, both exercise modes similarly improved variables describing beta cell function.Conclusions/interpretationTwo weeks of exercise training improves beta cell function in prediabetic or type 2 diabetic individuals and decreases pancreatic fat regardless of baseline glucose tolerance. This study shows that short-term training efficiently reduces ectopic fat within the pancreas, and exercise training may therefore reduce the risk of type 2 diabetes.Trial registrationClinicalTrials.gov NCT01344928FundingThis study was funded by the Emil Aaltonen Foundation, the European Foundation for the Study of Diabetes, the Finnish Diabetes Foundation, the Orion Research Foundation, the Academy of Finland (grants 251399, 256470, 281440, and 283319), the Ministry of Education of the State of Finland, the Paavo Nurmi Foundation, the Novo Nordisk Foundation, the Finnish Cultural Foundation, the Hospital District of Southwest Finland, the Turku University Foundation, and the Finnish Medical Foundation.

Postprandial endotoxemia may influence the development of type 2 diabetes mellitus: From the CORDIOPREV study

Insulin resistance (IR) and impaired beta-cell function are key determinants of type 2 diabetes mellitus (T2DM). Intestinal absorption of bacterial components activates the toll-like receptors inducing inflammation, and this in turn IR. We evaluated the role of endotoxemia in promoting inflammation-induced insulin resistance (IR) in the development of T2DM, and its usefulness as predictive biomarker. We included in this study 462 patients from the CORDIOPREV study without T2DM at baseline. Of these, 107 patients developed T2DM according to the American Diabetes Association (ADA) diagnosis criteria after a median follow-up of 60 months (Incident-DIAB group), whereas 355 patients did not developed it during this period of time (Non-DIAB group). We observed a postprandial increase in lipopolysaccharides (LPS) levels in the Incident-DIAB at baseline (P<0.001), whereas LPS levels were not modified in the Non-DIAB. Disease-free survival curves based on the LPS postprandial fold change improved T2DM Risk Assessment as compared with the previously described FINDRISC score (hazard ratio of 2.076, 95% CI 1.149-3.750 vs. 1.384, 95% CI 0.740-2.589). Moreover, disease-free survival curves combining the LPS postprandial fold change and FINDRISC score together showed a hazard ratio of 3.835 (95% CI 1.323-11.114), linked to high values of both parameters. Our results suggest that a high postprandial endotoxemia precedes the development of T2DM. Our results also showed the potential use of LPS plasma levels as a biomarker predictor of T2DM development. CLINICAL TRIALS.GOV. NCT00924937.

Both conditional ablation and overexpression of E2 SUMO-conjugating enzyme (UBC9) in mouse pancreatic beta cells result in impaired beta cell function.

Post-translational attachment of a small ubiquitin-like modifier (SUMO) to the lysine (K) residue(s) of target proteins (SUMOylation) is an evolutionary conserved regulatory mechanism. This modification has previously been demonstrated to be implicated in the control of a remarkably versatile regulatory mechanism of cellular processes. However, the exact regulatory role and biological actions of the E2 SUMO-conjugating enzyme (UBC9)-mediated SUMOylation function in pancreatic beta cells has remained elusive. Inducible beta cell-specific Ubc9 (also known as Ube2i) knockout (KO; Ubc9Δbeta) and transgenic (Ubc9Tg) mice were employed to address the impact of SUMOylation on beta cell viability and functionality. Ubc9 deficiency or overexpression was induced at 8weeks of age using tamoxifen. To study the mechanism involved, we closely examined the regulation of the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2) through SUMOylation in beta cells. Upon induction of Ubc9 deficiency, Ubc9Δbeta islets exhibited a 3.5-fold higher accumulation of reactive oxygen species (ROS) than Ubc9f/f control islets. Islets from Ubc9Δbeta mice also had decreased insulin content and loss of beta cell mass after tamoxifen treatment. Specifically, at day 45 after Ubc9 deletion only 40% of beta cell mass remained in Ubc9Δbeta mice, while 90% of beta cell mass was lost by day 75. Diabetes onset was noted in some Ubc9Δbeta mice 8weeks after induction of Ubc9 deficiency and all mice developed diabetes by 10weeks following tamoxifen treatment. In contrast, Ubc9Tg beta cells displayed an increased antioxidant ability but impaired insulin secretion. Unlike Ubc9Δbeta mice, which spontaneously developed diabetes, Ubc9Tg mice preserved normal non-fasting blood glucose levels without developing diabetes. It was noted that SUMOylation of NRF2 promoted its nuclear expression along with enhanced transcriptional activity, thereby preventing ROS accumulation in beta cells. SUMOylation function is required to protect against oxidative stress in beta cells; this mechanism is, at least in part, carried out by the regulation of NRF2 activity to enhance ROS detoxification. Homeostatic SUMOylation is also likely to be essential for maintaining beta cell functionality.

Increased lipid availability for three days reduces whole body glucose uptake, impairs muscle mitochondrial function and initiates opposing effects on PGC-1α promoter methylation in healthy subjects.

AimsFFA and FFA metabolites cause insulin resistance and impair beta cell function. The goal of our research was to examine whether elevation of plasma FFA impairs mitochondrial function and alters PGC-1α promoter methylation.MethodsIn this uncontrolled, change from baseline study design, insulin sensitivity and glucose-stimulated insulin secretion were measured in 9 normal glucose tolerant subjects before and after 3 day lipid infusion to elevate plasma FFA concentration. Vastus lateralis muscle biopsies were obtained and mitochondrial function, PGC-1α expression, and PGC-1α promoter methylation were quantitated.ResultsIncreased plasma FFA (440±93 μmol/Lto 997±242 μM, p<0.001) decreased insulin-stimulated total glucose disposal (TGD) by 25% (p = 0.008), impaired suppression of endogenous glucose production (p = 0.01), and reduced mitochondrial ATP synthesis with complex 1 (34%, p<0.05) and complex 2 (30%, p<0.05) substrates. Lipid infusion had no effect on muscle PGC-1α RNA expression, total methylation or non-CpG methylation, but methylation of the alternative PGC-1α promoter decreased (1.30±0.30 to 0.84±0.15% methylated residues/patient•strand, p = 0.055). Within PGC-1α promoter there was demethylation of CpT residues (0.72±0.16 vs. 0.28±0.10 methylated residues/patient•strand) (p = 0.002), which was inversely correlated with PGC-1α mRNA expression (r = -0.94, p<0.0001) and ATP synthesis with complex 1 (r = -0.80, p<0.01) and complex 2 (r = -0.69, p<0.05) substrates. Lipid infusion increased DNMT-3B (methyltransferase associated with PGC-1α promoter non-CpG methylation) mRNA expression (0.87 ± 0.09 to 1.62 ± 0.22 arbitrary units, p = 0.005), which correlated inversely with CpT demethylation (r = 0.67, p<0.05).Conclusion/InterpretationPhysiologic plasma FFA elevation in NGT individuals has opposing effects on PGC-1α non-CpG residue methylation (CpT demethylation and increased DNMT-3B expression), which is correlated with changes in PGC-1α expression and skeletal muscle mitochondrial function.

Circulating microRNA (miRNA) Expression Profiling in Plasma of Patients with Gestational Diabetes Mellitus Reveals Upregulation of miRNA miR-330-3p.

Gestational diabetes mellitus (GDM) is characterized by insulin resistance accompanied by low/absent beta-cell compensatory adaptation to the increased insulin demand. Although the molecular mechanisms and factors acting on beta-cell compensatory response during pregnancy have been partially elucidated and reported, those inducing an impaired beta-cell compensation and function, thus evolving in GDM, have yet to be fully addressed. MicroRNAs (miRNAs) are a class of small endogenous non-coding RNAs, which negatively modulate gene expression through their sequence-specific binding to 3′UTR of mRNA target. They have been described as potent modulators of cell survival and proliferation and, furthermore, as orchestrating molecules of beta-cell compensatory response and function in diabetes. Moreover, it has been reported that miRNAs can be actively secreted by cells and found in many biological fluids (e.g., serum/plasma), thus representing both optimal candidate disease biomarkers and mediators of tissues crosstalk(s). Here, we analyzed the expression profiles of circulating miRNAs in plasma samples obtained from n = 21 GDM patients and from n = 10 non-diabetic control pregnant women (24–33 weeks of gestation) using TaqMan array microfluidics cards followed by RT-real-time PCR single assay validation. The results highlighted the upregulation of miR-330-3p in plasma of GDM vs non-diabetics. Furthermore, the analysis of miR-330-3p expression levels revealed a bimodally distributed GDM patients group characterized by high or low circulating miR-330 expression and identified as GDM-miR-330high and GDM-miR-330low. Interestingly, GDM-miR-330high subgroup retained lower levels of insulinemia, inversely correlated to miR-330-3p expression levels, and a significant higher rate of primary cesarean sections. Finally, miR-330-3p target genes analysis revealed major modulators of beta-cell proliferation and of insulin secretion, such as the experimentally validated genes E2F1 and CDC42 as well as AGT2R2, a gene involved in the differentiation of mature beta-cells. In conclusion, we demonstrated that plasma miR-330-3p could be of help in identifying GDM patients with potential worse gestational diabetes outcome; in GDM, miR-330-3p may directly be transferred from plasma to beta-cells thus modulating key target genes involved in proliferation, differentiation, and insulin secretion.

Early deficits in insulin secretion, beta cell mass and islet blood perfusion precede onset of autoimmune type 1 diabetes in BioBreeding rats

Aims/hypothesisGenetic studies show coupling of genes affecting beta cell function to type 1 diabetes, but hitherto no studies on whether beta cell dysfunction could precede insulitis and clinical onset of type 1 diabetes are available.MethodsWe used 40-day-old BioBreeding (BB) DRLyp/Lyp rats (a model of spontaneous autoimmune type 1 diabetes) and diabetes-resistant DRLyp/+ and DR+/+ littermates (controls) to investigate beta cell function in vivo, and insulin and glucagon secretion in vitro. Beta cell mass was assessed by optical projection tomography (OPT) and morphometry. Additionally, measurements of intra-islet blood flow were performed using microsphere injections. We also assessed immune cell infiltration, cytokine expression in islets (by immunohistochemistry and qPCR), as well as islet Glut2 expression and ATP/ADP ratio to determine effects on glucose uptake and metabolism in beta cells.ResultsDRLyp/Lyp rats were normoglycaemic and without traces of immune cell infiltrates. However, IVGTTs revealed a significant decrease in the acute insulin response to glucose compared with control rats (1685.3 ± 121.3 vs 633.3 ± 148.7; p < 0.0001). In agreement, insulin secretion was severely perturbed in isolated islets, and both first- and second-phase insulin release were lowered compared with control rats, while glucagon secretion was similar in both groups. Interestingly, after 5–7 days of culture of islets from DRLyp/Lyp rats in normal media, glucose-stimulated insulin secretion (GSIS) was improved; although, a significant decrease in GSIS was still evident compared with islets from control rats at this time (7393.9 ± 1593.7 vs 4416.8 ± 1230.5 pg islet−1 h−1; p < 0.0001). Compared with controls, OPT of whole pancreas from DRLyp/Lyp rats revealed significant reductions in medium (4.1 × 109 ± 9.5 × 107 vs 3.8 × 109 ± 5.8 × 107 μm3; p = 0.044) and small sized islets (1.6 × 109 ± 5.1 × 107 vs 1.4 × 109 ± 4.5 × 107 μm3; p = 0.035). Finally, we found lower intra-islet blood perfusion in vivo (113.1 ± 16.8 vs 76.9 ± 11.8 μl min−1 [g pancreas]−1; p = 0.023) and alterations in the beta cell ATP/ADP ratio in DRLyp/Lyp rats vs control rats.Conclusions/interpretationThe present study identifies a deterioration of beta cell function and mass, and intra-islet blood flow that precedes insulitis and diabetes development in animals prone to autoimmune type 1 diabetes. These underlying changes in islet function may be previously unrecognised factors of importance in type 1 diabetes development.

IKKβ inhibition prevents fat-induced beta cell dysfunction in vitro and in vivo in rodents.

We have previously shown that oxidative stress plays a causal role in beta cell dysfunction induced by fat. Here, we address whether the proinflammatory kinase inhibitor of (nuclear factor) κB kinase β (IKKβ), which is activated by oxidative stress, is also implicated. Fat (oleate or olive oil) was infused intravenously in Wistar rats for 48h with or without the IKKβ inhibitor salicylate. Thereafter, beta cell function was evaluated in vivo using hyperglycaemic clamps or ex vivo in islets isolated from fat-treated rats. We also exposed rat islets to oleate in culture, with or without salicylate and 4(2'-aminoethyl)amino-1,8-dimethylimidazo(1,2-a)quinoxaline; BMS-345541 (BMS, another inhibitor of IKKβ) and evaluated beta cell function in vitro. Furthermore, oleate was infused in mice treated with BMS and in beta cell-specific Ikkb-null mice. 48h infusion of fat impaired beta-cell function in vivo, assessed using the disposition index (DI), in rats (saline: 1.41±0.13; oleate: 0.95±0.11; olive oil [OLO]: 0.87±0.15; p<0.01 for both fats vs saline) and in mice (saline: 2.51±0.39; oleate: 1.20±0.19; p<0.01 vs saline) and ex vivo (i.e., insulin secretion, units are pmol insulin islet-1h-1) in rat islets (saline: 1.51±0.13; oleate: 1.03±0.10; OLO: 0.91±0.13; p<0.001 for both fats vs saline) and the dysfunction was prevented by co-infusion of salicylate in rats (oleate+salicylate: 1.30±0.09; OLO+salicylate: 1.33±0.23) or BMS in mice (oleate+BMS: 2.25±0.42) in vivo and by salicylate in rat islets ex vivo (oleate+salicylate: 1.74±0.31; OLO+salicylate: 1.54±0.29). In cultured islets, 48h exposure to oleate impaired beta-cell function ([in pmol insulin islet-1h-1] control: 0.66±0.12; oleate: 0.23±0.03; p<0.01 vs saline), an effect prevented by both inhibitors (oleate+salicylate: 0.98±0.08; oleate+BMS: 0.50±0.02). Genetic inhibition of IKKβ also prevented fat-induced beta-cell dysfunction ex vivo ([in pmol insulin islet-1h-1] control saline: 0.16±0.02; control oleate: 0.10±0.02; knockout oleate: 0.17±0.04; p<0.05 control saline vs. control oleate) and in vivo (DI: control saline: 3.86±0.40; control oleate: 1.95±0.29; knockout oleate: 2.96±0.24; p<0.01 control saline vs control oleate). Our results demonstrate a causal role for IKKβ in fat-induced beta cell dysfunction in vitro, ex vivo and in vivo.

Involvement of receptor-interacting protein 140 in palmitate-stimulated macrophage infiltration of pancreatic beta cells.

Receptor-interacting protein 140 (RIP140) in macrophages stimulates the nuclear factor-κB subunit RelA to activate tumor necrosis factor (TNF)-α and interleukin (IL)-6 transcription. However, under lipotoxic conditions, the involvement of RIP140 in the infiltration of beta cells by macrophages remains unknown. In the present study, murine RAW264.7 macrophages were transfected with a RIP140 overexpression plasmid or siRNA prior to macrophage activation with 500 µM palmitate. Palmitate-free conditioned media was then collected and added to murine insulinoma MIN6 cells. Significant decreases were observed in cell viability (P<0.01), glucose-stimulated insulin secretion (P<0.01) and levels of peroxisome proliferator-activated receptor-γ coactivator-1α (P<0.05), phosphoenolpyruvate carboxykinase and proliferating cell nuclear antigen mRNA (P<0.01) in MIN6 cells. In addition, conditioned media from palmitate-treated and RIP140-upregulated macrophages significantly increased the levels of uncoupling protein-2 (P<0.01), inducible nitric oxide synthase 1 (P<0.01) and pancreatic and duodenal homeobox 1 (P<0.05) mRNA and levels of activated Jun N-terminal kinase (JNK) (P<0.01) and extracellular signal-regulated kinase (ERK) 1/2 (P<0.01). In turn, the conditioned media was found to be significantly enriched in TNF-α and IL-6 (both P<0.05). These results were the opposite of those obtained from MIN6 cells treated with conditioned media from palmitate-treated and RIP140-knockdown macrophages. MIN6 cells were transfected with RIP140 overexpression plasmid or siRNA prior to treatment with 500 µM palmitate and supernatant was collected for use in macrophage chemotaxis assays. In the palmitate-activated and RIP140-overexpressing MIN6 cells, TNF-α and IL-6 secretion increased significantly (both P<0.05) and macrophage chemotaxis towards MIN6 cells was enhanced. By contrast, downregulating RIP140 in MIN6 cells had the opposite effect. These data suggest that RIP140 in macrophages mediates the transcription of inflammatory cytokines when concentration of palmitate is high. Macrophage RIP140 may also impair beta cell function by activating the JNK and ERK1/2 signaling pathways and promoting specific gene transcription. Furthermore, expression of RIP140 in pancreatic beta cells may stimulate macrophage chemotaxis, thus triggering local low-grade inflammation.

GLP-1 signalling compensates for impaired insulin signalling in regulating beta cell proliferation in βIRKO mice.

We aimed to investigate potential interactions between insulin and glucagon-like peptide (GLP)-1 signalling pathways in the regulation of beta cell-cycle dynamics in vivo, in the context of the therapeutic potential of GLP-1 to modulate impaired beta cell function. Beta cell-specific insulin receptor knockout (βIRKO) mice, which exhibit beta cell dysfunction and an age-dependent decrease in beta cell mass, were treated with the dipeptidyl peptidase-4 inhibitor vildagliptin. Following this, glucose homeostasis and beta cell proliferation were evaluated and underlying molecular mechanisms were investigated. The sustained elevation in circulating GLP-1 levels, caused by treatment of the knockout mice with vildagliptin for 6weeks, significantly improved glucose tolerance secondary to enhanced insulin secretion and proliferation of beta cells. Treating βIRKO beta cell lines with the GLP-1 analogue, exendin-4, promoted Akt phosphorylation and protein expression of cyclins A, D1 and E two- to threefold, in addition to cyclin D2. Pancreases from the vildagliptin-treated βIRKO mice exhibited increased cyclin D1 expression, while cyclin D2 expression was impaired. Activation of GLP-1 signalling compensates for impaired growth factor (insulin) signalling and enhances expression of cyclins to promote beta cell proliferation. Together, these data indicate the potential of GLP-1-related therapies to enhance beta cell proliferation and promote beneficial outcomes in models with dysfunctional beta cells.

Antibodies against glutamic acid decarboxylase and indices of insulin resistance and insulin secretion in nondiabetic adults: a cross-sectional study.

BackgroundAutoimmunity against insulin-producing beta cells from pancreatic islets is a common phenomenon in type 1 diabetes and latent autoimmune diabetes in adults. Some reports have also related beta-cell autoimmunity to insulin resistance (IR) in type 2 diabetes. However, the extent to which autoimmunity against components of beta cells is present and relates to IR and insulin secretion in nondiabetic adults is uncertain.AimTo explore the association between antibodies against glutamic acid decarboxylase (GADA), a major antigen from beta cells, and indices of whole-body IR and beta-cell capacity/insulin secretion in adults who do not have diabetes.MethodsWe studied 81 adults of both sexes aged 30–70, without known diabetes or any autoimmune disease. Participants underwent an oral glucose tolerance test (OGTT) with determination of plasma glucose and insulin at 0, 30, 60, 90, and 120 minutes. From these results we calculated indices of insulin resistance (homeostasis model assessment of insulin resistance [HOMA-IR] and incremental area under the insulin curve [iAUCins]) and insulin secretion (corrected insulin response at 30 minutes and HOMA beta-cell%). GADAs were measured in fasting plasma using immunoenzymatic methods.ResultsWe found an overall prevalence of GADA positivity of 21.3%, without differences by sex and no correlation with age. GADA titers did not change monotonically across quartiles of any of the IR or insulin secretion indices studies. GADA did not correlate linearly with fasting IR expressed as HOMA-IR (Spearman’s r=−0.18, p=0.10) or postabsorptive IR expressed as iAUCins (r=−0.15, p=0.18), but did show a trend toward a negative correlation with insulin secretory capacity expressed by the HOMA-beta cell% index (r=−0.20, p=0.07). Hemoglobin A1c, body mass index, and waist circumference were not associated with GADA titers.ConclusionGADA positivity is frequent and likely related to impaired beta-cell function among adults without known diabetes.

Glucose-induced β cell production of IL-1β contributes to glucotoxicity in human pancreatic islets.

In type 2 diabetes, chronic hyperglycemia is suggested to be detrimental to pancreatic beta cells, causing impaired insulin secretion. IL-1beta is a proinflammatory cytokine acting during the autoimmune process of type 1 diabetes. IL-1beta inhibits beta cell function and promotes Fas-triggered apoptosis in part by activating the transcription factor NF-kappaB. Recently, we have shown that increased glucose concentrations also induce Fas expression and beta cell apoptosis in human islets. The aim of the present study was to test the hypothesis that IL-1beta may mediate the deleterious effects of high glucose on human beta cells. In vitro exposure of islets from nondiabetic organ donors to high glucose levels resulted in increased production and release of IL-1beta, followed by NF-kappaB activation, Fas upregulation, DNA fragmentation, and impaired beta cell function. The IL-1 receptor antagonist protected cultured human islets from these deleterious effects. beta cells themselves were identified as the islet cellular source of glucose-induced IL-1beta. In vivo, IL-1beta-producing beta cells were observed in pancreatic sections of type 2 diabetic patients but not in nondiabetic control subjects. Similarly, IL-1beta was induced in beta cells of the gerbil Psammomys obesus during development of diabetes. Treatment of the animals with phlorizin normalized plasma glucose and prevented beta cell expression of IL-1beta. These findings implicate an inflammatory process in the pathogenesis of glucotoxicity in type 2 diabetes and identify the IL-1beta/NF-kappaB pathway as a target to preserve beta cell mass and function in this condition.

Endogenous beta-cell CART regulates insulin secretion and transcription of beta-cell genes

Impaired beta-cell function is key to the development of type 2 diabetes. Cocaine- and amphetamine-regulated transcript (CART) is an islet peptide with insulinotropic and glucagonostatic properties. Here we studied the role of endogenous CART in beta-cell function. CART silencing in INS-1 (832/13) beta-cells reduced insulin secretion and production, ATP levels and beta-cell exocytosis. This was substantiated by reduced expression of several exocytosis genes, as well as reduced expression of genes important for insulin secretion and processing. In addition, CART silencing reduced the expression of a network of transcription factors essential for beta-cell function. Moreover, in RNAseq data from human islet donors, CARTPT expression levels correlated with insulin, exocytosis genes and key beta-cell transcription factors. Thus, endogenous beta-cell CART regulates insulin expression and secretion in INS-1 (832/13) cells, via actions on the exocytotic machinery and a network of beta-cell transcription factors. We conclude that CART is important for maintaining the beta-cell phenotype.