Increase In Glucagon Levels Research Articles

929-P: A Study to Evaluate Clinical and Metabolic Profile of Type 2 Diabetes Patients Receiving SGLT2 Inhibitors

Aim: The aim of the study was to determine the efficacy of SGLT2i and its impact on metabolic profile of type 2 diabetes (T2DM) patients with reference to serum glucagon and blood ketones. Methodology: The study was conducted in T2DM patients with inadequate glycemic control on a background therapy of either metformin and sulphonylurea (Group A) or metformin and DPP4 inhibitor(Group B) Blood samples were taken for estimation of fasting plasma glucose (FPG), post prandial plasma glucose (PPPG), HbA1c, beta-hydroxybutyrate (β-HB) and glucagon levels. Empagliflozin 10mg, was added to the existing anti-diabetic medication. The metabolic parameters were reassessed after 12 weeks. Results: At the end of 12 weeks glucagon levels increased significantly (28.8 ± 28.74 at baseline vs 108 ± 74 pg/ml; p<0.001). A significant increase in β-HB levels was also observed (0.1 ± 0.07 mmol/L at baseline versus 0.12 ± 0.07 mmol/L (p=0.03). FPG, PPPG and HbA1c decreased significantly by 30.33mg%, 53.16mg% and 1.05% respectively(p<0.01) In group A, FPG and PPPG decreased by 34.43 mg/dl and 51.71mg/dl respectively (p<0.001) with a mean HbA1c reduction of 1.09% (p<0.001) Serum glucagon and β-HB levels increased significantly [82.63 pg/ml (p<0.001) and 0.04 mmol/L (p<0.01) respectively]. In group B, there was decrease in FPG (mean difference 23.6mg %; p=0.003) and PPPG (mean difference 55.5mg%; p<0.001) at 12 weeks. HbA1c reduction of 0.99% was observed. (p=0.003) There was a significant increase in glucagon levels (mean difference +73.52 pg/ml; p<0.001). Conclusion: Addition of Empagliflozin to T2DM patients with inadequate glycemic control on either metformin and sulphonylurea or metformin and DPP4 inhibitors significantly improved efficacy. Plasma glucagon and β-HB levels increased with chronic empagliflozin treatment suggesting a change in substrate utilization towards fat. Disclosure S. Kandula: None. M.A. Meshram: None. A.C. Sonwane: None.

Evaluating the effectiveness of a novel somatostatin receptor 2 antagonist, ZT-01, for hypoglycemia prevention in a rodent model of type 2 diabetes.

Background: Elevated levels of somatostatin blunt glucagon counterregulation during hypoglycemia in type 1 diabetes (T1D) and this can be improved using somatostatin receptor 2 (SSTR2) antagonists. Hypoglycemia also occurs in late-stage type 2 diabetes (T2D), particularly when insulin therapy is initiated, but the utility of SSTR2 antagonists in ameliorating hypoglycemia in this disease state is unknown. We examined the efficacy of a single-dose of SSTR2 antagonists in a rodent model of T2D. Methods: High-fat fed (HFF), low dose streptozotocin (STZ, 35mg/kg)-induced T2D and HFF only, nondiabetic (controls-no STZ) rats were treated with the SSTR2 antagonists ZT-01/PRL-2903 or vehicle (n = 9-11/group) 60min before an insulin tolerance test (ITT; 2-12U/kg insulin aspart) or an oral glucose tolerance test (OGTT; 2g/kg glucose via oral gavage) on separate days. Results: This rodent model of T2D is characterized by higher baseline glucose and HbA1c levels relative to HFF controls. T2D rats also had lower c-peptide levels at baseline and a blunted glucagon counterregulatory response to hypoglycemia when subjected to the ITT. SSTR2 antagonists increased the glucagon response and reduced incidence of hypoglycemia, which was more pronounced with ZT-01 than PRL-2903. ZT-01 treatment in the T2D rats increased glucagon levels above the control response within 60min of dosing, and values remained elevated during the ITT (glucagon Cmax: 156 ± 50 vs. 77 ± 46pg/mL, p < 0.01). Hypoglycemia incidence was attenuated with ZT-01 vs. controls (63% vs. 100%) and average time to hypoglycemia onset was also delayed (103.1 ± 24.6 vs. 66.1 ± 23.6min, p < 0.05). ZT-01 administration at the OGTT onset increased the glucagon response without exacerbating hyperglycemia (2877 ± 806 vs. 2982 ± 781), potentially due to the corresponding increase in c-peptide levels (6251 ± 5463 vs. 14008 ± 5495, p = 0.013). Conclusion: Treatment with SSTR2 antagonists increases glucagon responses in a rat model of T2D and results in less hypoglycemia exposure. Future studies are required to determine the best dosing periods for chronic SSTR2 antagonism treatment in T2D.

219-OR: Glucagon Counterregulation in a Hypoglycemic Clamp in Type 1 Diabetes Is Increased by ZT-01, a Novel Somatostatin Receptor 2 Antagonist—A Phase 1b Study

This study evaluated the effect of ZT-01 on glucagon counterregulation during hypoglycemia in adults with type 1 diabetes, in a Phase 1b randomized, three-arm, double-blind crossover trial in 23 hypoglycemia aware adults with T1D. Participants underwent three hypoglycemic clamp procedures with 3 or 20 mg ZT-01 or placebo dosed 1 h prior to stepwise glucose clamp conditions using variable rate IV insulin infusion: euglycemia (5.0 ± 0.5 mmol/L), level 1 (3.5 ± 0.3 mmol/L, L1), and level 2 hypoglycemia (2.6 ± 0.2 mmol/L, L2). With ZT-01, peak glucagon increased by up to 15.1 ± 10.5 and 27.1 ± 17.3 pg/mL, in L1 and L2 hypoglycemia, respectively, compared with no increase in L1 and an 8.4 ± 8.4 pg/mL increase in L2 hypoglycemia with placebo (p&amp;lt;0.0001, figure). A counterregulatory response of increased glucose level coinciding with an increased glucagon level was observed more frequently with ZT-01 than placebo. No clinically significant effects on other counterregulatory hormones or hypoglycemia symptom scores were observed with ZT-01. A transient rise in glucagon and blood glucose was observed with ZT-01 post-dose prior to hypoglycemia. No SAEs were reported and no TEAEs were observed with ZT-01. This study demonstrates proof-of-concept that ZT-01 significantly enhances the defective counterregulatory response in people with T1D during L1 and L2 hypoglycemia. Disclosure A.Abitbol: Advisory Panel; Abbott Diabetes, Lilly Diabetes, Dexcom, Inc., Novo Nordisk Canada Inc., Janssen Pharmaceuticals, Inc., Research Support; Abbott Diabetes, Lilly Diabetes, Zucara Therapeutics, Novo Nordisk Canada Inc., Moderna, Inc., Senseonics, Speaker's Bureau; Boehringer Ingelheim (Canada) Ltd., Lilly Diabetes, Amgen Canada, Dexcom, Inc., Novo Nordisk Canada Inc., HLS Therapeutics Inc., Janssen Pharmaceuticals, Inc., Merck &amp; Co., Inc., Sanofi. S.Peers: Employee; Zucara Therapeutics. E.Simonson: None. M.Midmer: Board Member; Zucara Therapeutics, Employee; Zucara Therapeutics, Stock/Shareholder; Zucara Therapeutics. M.Riddell: Advisory Panel; Zealand Pharma A/S, Zucara Therapeutics, Indigo Diabetes, Consultant; Lilly Diabetes, Eli Lilly and Company, Jaeb Center for Health Research, Speaker's Bureau; Dexcom, Inc., Novo Nordisk, Sanofi, Stock/Shareholder; Supersapiens, Zucara Therapeutics. R.Liggins: Employee; Zucara Therapeutics, Stock/Shareholder; Zucara Therapeutics.

175-LB: Altered Glycogen Cell Number in Placental mTOR Signaling Deficiency Model in Mice

Fetal Growth Restriction (FGR) is a determinant for the development of chronic diseases, such as type II diabetes in adulthood. FGR is caused by the failure of the placenta to provide adequate nutrients and oxygen for fetal development. mTOR regulates cell metabolism and growth, and it is reduced in the FGR placenta. We previously reported that placental mTOR ablation induces insulin resistance and glucose intolerance in adult mice. We hypothesize that mTOR signaling regulates glucose utilization in the placenta, which stores glucose reservoirs as glycogen. Appropriate localization and breakdown of glycogen cells is crucial for normal fetal growth. We leveraged our mouse model lacking placental mTOR signaling to ask whether glycogen cell number was altered. First, we found that critical glycogenesis and glycogenolysis enzymes, as well as the glucagon receptor, were expressed in placental tissue, indicating the capability for glycogen cell synthesis and breakdown, like glycogen-rich tissues such as the liver. Next, we show increased glycogen cell counts in the junctional zone of the mTORKO placenta compared to littermate controls by IHC. However, whole placental glycogen content, measured by ELISA, demonstrates no changes between KO and control. These data suggest that mTOR signaling modulates the localization and migration of glycogen cells during high metabolic needs. Glucagon mRNA is expressed in the placenta, with significant increases in the male KO placenta, indicating the placenta may produce its own glucagon. However, IHC staining indicates increased glucagon levels in female KO placenta in the labyrinth zone. Together, these data suggest that the placenta may have the capability to regulate its own glycogen breakdown by altering glucagon expression. Our results indicate that the FGR placenta induced by mTOR deletion has altered glycogen cell content and mislocalization, which mimics the phenotypes of placenta impacted by gestational diabetes, suggesting a shared mechanistic pathway. Disclosure B. Clifton: None. M. Beetch: None. E. Alejandro: None. Funding National Institutes of Health (R01DK115720)

1-LB: Evaluating the Effectiveness of the Novel Somatostatin Receptor Antagonist (SSTR2a) ZT-01 for Hypoglycemia Prevention in a Rodent Model of Type 2 Diabetes

Background: Few pharmacological strategies exist to prevent hypoglycemia in diabetes, other than reducing the intensity of glucose-lowering therapy. SSTR2a treatment increases glucagon counterregulation and reduces the time spent in hypoglycemia in rodent models of type 1 diabetes (T1D), but the utility of this approach in type 2 diabetes (T2D) is unclear. The purpose of this study was to test the effect of acute SSTR2a treatment in a rodent model of hypoglycemia in T2D. Methods: High fat fed (HFF), low dose streptozotocin (35 mg/kg) T2D rats received SSTR2a treatment (subcutaneous ZT-01 3 mg/kg) or vehicle (N=9-11/group) 60 min before the induction of hypoglycemia using rapid-acting insulin (12 U/kg). Glycemia, glucagon and c-peptide levels were measured intermittently pre- and post-dosing. Results: T2D in the rats was characterized by significantly higher baseline glucose (21.8 ± 4.7 vs 6.5 ± 0.7 mmol/L, p&amp;lt;0.05) and HbA1c (8.6 ± 0.6% vs 5.5 ± 0.5%; p&amp;lt;0.0001) levels relative to HFF controls (n=3). T2D rats also displayed reduced c-peptide levels relative to controls (3.1 ± 1.3 vs 6.1 ± 1.2 ng/ml, p&amp;lt;0.05). During hypoglycemic challenge (T2D rats only), SSTR2a elevated glucagon levels relative to vehicle treatment (glucagon Cmax: 156 ± 50 vs 77 ± 46 pg/ml, p&amp;lt;0.01) and reduced the incidence of hypoglycemia by ~40% (5/8 SSTR2a treated vs 9/9 vehicle treated rats). In those that developed hypoglycemia, the average time to hypoglycemia onset was delayed (103.1± 24.6 vs 66.1 ± 23.6 min, p&amp;lt;0.05) with SSTR2a compared to vehicle. Conclusion: SSTR2a treatment increases glucagon levels in this rodent model of T2D, which appears to reduce the risk for hypoglycemia following bolus insulin dosing. Disclosure N. C. D'souza: None. M. Nejad-mansouri: None. R. Liggins: Employee; Zucara Therapeutics, Stock/Shareholder; Zucara Therapeutics. O. Chan: None. M. C. Riddell: Advisory Panel; Zealand Pharma A/S, Zucara Therapeutics, Indigo Diabetes, Consultant; Lilly Diabetes, Eli Lilly and Company, Jaeb Center for Health Research, Speaker's Bureau; Dexcom, Inc., Novo Nordisk, Sanofi, Stock/Shareholder; Supersapiens, Zucara Therapeutics. P. Zaree bavani: None. J. Aiken: None. E. G. Hoffman: None. S. C. Atherley: None. S. Champsi: None. N. Aleali: None. D. Shakeri: None. M. El-zahed: None. N. Akbarian: None. Funding GlycoNet (CD-85)

Impaired hepatic glucose metabolism and liver-α-cell axis in mice with liver-specific ablation of the Hepatocyte Nuclear Factor 4α (Hnf4a) gene

BackgroundHnf4a gene ablation in mouse liver causes hepatic steatosis, perturbs HDL structure and function and affects many pathways and genes related to glucose metabolism. Our aim here was to investigate the role of liver HNF4A in glucose homeostasis. MethodsSerum and tissue samples were obtained from Alb-Cre;Hnf4afl/fl (H4LivKO) mice and their littermate Hnf4afl/fl controls. Fasting glucose and insulin, glucose tolerance, insulin tolerance and glucagon challenge tests were performed by standard procedures. Binding of HNF4A to DNA was assessed by chromatin immunoprecipitation assays. Gene expression analysis was performed by quantitative reverse transcription PCR. ResultsH4LivKO mice presented lower blood levels of fasting glucose, improved glucose tolerance, increased serum lactate levels and reduced response to glucagon challenge compared to their control littermates. Insulin signaling in the liver was reduced despite the increase in serum insulin levels. H4LivKO mice showed altered expression of genes involved in glycolysis, gluconeogenesis and glycogen metabolism in the liver. The expression of the gene encoding the glucagon receptor (Gcgr) was markedly reduced in H4LivKO liver and chromatin immunoprecipitation assays revealed specific and strong binding of HNF4A to the Gcgr promoter. H4LivKO mice presented increased amino acid concentration in the serum, α-cell hyperplasia and a dramatic increase in glucagon levels suggesting an impairment of the liver-α-cell axis. Glucose administration in the drinking water of H4LivKO mice resulted in an impressive extension of survival. The expression of several genes related to non-alcoholic fatty liver disease progression to more severe liver pathologies, including Mcp1, Gdf15, Igfbp-1 and Hmox1, was increased in H4LivKO mice as early as 6 weeks of age and this increased expression was sustained until the endpoint of the study. ConclusionsOur results reveal a novel role of liver HNF4A in controlling blood glucose levels via regulation of glucagon signaling. In combination with the steatotic phenotype, our results suggest that H4LivKO mice could serve as a valuable model for studying glucose homeostasis in the context of non-alcoholic fatty liver disease.

ISPAD Clinical Practice Consensus Guidelines 2022: Ramadan and other religious fasting by young people with diabetes.

ISPAD Clinical Practice Consensus Guidelines 2022: Ramadan and other religious fasting by young people with diabetes.

Alpha Cell Thioredoxin-interacting Protein Deletion Improves Diabetes-associated Hyperglycemia and Hyperglucagonemia.

Thioredoxin-interacting protein (TXNIP) has emerged as a key factor in pancreatic beta cell biology, and its upregulation by glucose and diabetes contributes to the impairment in functional beta cell mass and glucose homeostasis. In addition, beta cell deletion of TXNIP protects against diabetes in different mouse models. However, while TXNIP is ubiquitously expressed, its role in pancreatic alpha cells has remained elusive. We generated an alpha cell TXNIP knockout (aTKO) mouse and assessed the effects on glucose homeostasis. While no significant changes were observed on regular chow, after a 30-week high-fat diet, aTKO animals showed improvement in glucose tolerance and lower blood glucose levels compared to their control littermates. Moreover, in the context of streptozotocin (STZ)-induced diabetes, aTKO mice showed significantly lower blood glucose levels compared to controls. While serum insulin levels were reduced in both control and aTKO mice, STZ-induced diabetes significantly increased glucagon levels in control mice, but this effect was blunted in aTKO mice. Moreover, glucagon secretion from aTKO islets was >2-fold lower than from control islets, while insulin secretion was unchanged in aTKO islets. At the same time, no change in alpha cell or beta cell numbers or mass was observed, and glucagon and insulin expression and content were comparable in isolated islets from aTKO and control mice. Thus together the current studies suggest that downregulation of alpha cell TXNIP is associated with reduced glucagon secretion and that this may contribute to the glucose-lowering effects observed in diabetic aTKO mice.

Glucose metabolism in amyotrophic lateral sclerosis: it is bitter-sweet.

Glucose metabolism in amyotrophic lateral sclerosis: it is bitter-sweet.

Diabetic Ketoacidosis and Dysregulation of Proglucagon Family of Molecules

ObjectivesAn increasing body of evidence supports a significant contributory role for pancreatic α-cell dysfunction in the pathophysiology of diabetic ketoacidosis (DKA). In normal physiology, high glucose levels stimulate insulin and inhibit glucagon secretion, while low glucose stimulates glucagon but inhibits insulin release. This regulatory mechanism is progressively lost in diabetes mellitus (DM), where diminished insulin levels are accompanied by hyperglucagonemia leading to the further aggravation of hyperglycemia caused by insulin deficiency. Glucagon is produced via the posttranslational modification of proglucagon in a process that also produces other proglucagon molecules including major proglucagon fragment (MPGF) and glucagon like peptide-1 (GLP-1). Here we hypothesized that DKA is associated with α-cell dysfunction leading to elevated levels of proglucagon molecules. MethodsWe measured levels of glucagon, GLP-1 and MPGF as indices of glucagon synthesis/α-cell function in the plasma of patients with DKA and in healthy controls. The inclusion criteria for DKA patients were glucose > 250 mg/dL, pH < 7.2, bicarbonate < 15mEq/L, presence of ketones and patients were part of a larger randomized trial. Samples were obtained at baseline prior to trial interventions. An independent sample t-test was used to compare differences. ResultsWe analyzed samples of 25 healthy controls and 59 individuals with DKA (47 T1DM and 12 T2DM) and found significant increase in glucagon levels in all of the DKA groups (controls 47 pg/mL (36, 67), T1DM 63 (40,135), T2DM 84 (61,105), p = 0.013). The differences were significant in MPGF too (controls 0.5 ng/mL (0.4, 0.8), T1DM 1.1 (0.7, 1.9), T2DM 1.3 (0.9, 2.0), p < 0.001), while differences in GLP-1 between groups did not reach statistical significance. C-peptide levels were significantly diminished in DKA and the levels of glucagon and C-peptide were strongly associated in healthy controls but not in DKA. ConclusionsOur data indicate that proglucagon family molecules may be actively contributing to the pathogenesis of DKA and may represent a potential site for future therapeutic interventions. Funding SourcesThe parental study of “Thiamine as adjunctive therapy for diabetic ketoacidosis” was supported through a grant from the National Institute of Health.

776-P: SGLT2i Increased the Fasting Glucagon Level in Patients with Diabetes: A Meta-analysis

Aims: Although it has long been assumed that increased glucagon level participants in the ketosis during sodium-glucose cotransporter 2 inhibitors (SGLT2i) treatment, the effect of SGLT2i on glucagon in diabetic patients remains controversial. Hence, we conducted this metanalysis to assess the overall effect of SGLT2i treatment on plasma glucagon level. Methods: PubMed/MEDLINE, Embase, and Cochrane databases were searched for studies published before August 2020. Clinical trials in type 1 diabetes (T1DM) and type 2 diabetes (T2DM) patients with the report of glucagon variation before and after SGLT2i treatment was included. Result: In total, 10 trials were included. Overall, SGLT2i treatment was associated with significantly increased fasting plasma glucagon level (WMD, 35.00 pg/ml; 95% CI, 24.48 to 45.52 pg/ml, P Conclusions: The increase in fasting glucagon level may be associated with the reduction of insulin, which may decrease the level of insulin-glucagon ratio and make patients susceptible to ketosis. Disclosure X. Zhu: None. C. Lin: None. L. Li: None. S. Hu: None. X. Cai: None. L. Ji: None. Funding National Natural Science Foundation of China (81970698, 81970708); Beijing Natural Science Foundation (7202216)

Neprilysin Inhibition Increases Glucagon Levels in Humans and Mice With Potential Effects on Amino Acid Metabolism.

ContextInhibitors of the protease neprilysin (NEP) are used for treating heart failure, but are also linked to improvements in metabolism. NEP may cleave proglucagon-derived peptides, including the glucose and amino acid (AA)-regulating hormone glucagon. Studies investigating NEP inhibition on glucagon metabolism are warranted.ObjectiveThis work aims to investigate whether NEP inhibition increases glucagon levels.MethodsPlasma concentrations of glucagon and AAs were measured in eight healthy men during a mixed meal with and without a single dose of the NEP inhibitor/angiotensin II type 1 receptor antagonist, sacubitril/valsartan (194 mg/206 mg). Long-term effects of sacubitril/valsartan (8 weeks) were investigated in individuals with obesity (n = 7). Mass spectrometry was used to investigate NEP-induced glucagon degradation, and the derived glucagon fragments were tested pharmacologically in cells transfected with the glucagon receptor (GCGR). Genetic deletion or pharmacological inhibition of NEP with or without concomitant GCGR antagonism was tested in mice to evaluate effects on AA metabolism.ResultsIn healthy men, a single dose of sacubitril/valsartan significantly increased postprandial concentrations of glucagon by 228%, concomitantly lowering concentrations of AAs including glucagonotropic AAs. Eight-week sacubitril/valsartan treatment increased fasting glucagon concentrations in individuals with obesity. NEP cleaved glucagon into 5 inactive fragments (in vitro). Pharmacological NEP inhibition protected both exogenous and endogenous glucagon in mice after an AA challenge, while NEP-deficient mice showed elevated fasting and AA-stimulated plasma concentrations of glucagon and urea compared to controls.ConclusionNEP cleaves glucagon, and inhibitors of NEP result in hyperglucagonemia and may increase postprandial AA catabolism without affecting glycemia.

SGLT2i increased the plasma fasting glucagon level in patients with diabetes: A meta-analysis

Increased glucagon level was hypothesized to participate in the ketoacidosis associated with sodium-glucose co-transporter 2 inhibitors (SGLT2i) treatment. However, the effect of SGLT2i on glucagon remains controversial. Hence, we conducted this meta-analysis to assess the overall effect of SGLT2i treatment on plasma fasting glucagon level in patients with diabetes. PubMed/MEDLINE, Embase, and Cochrane databases were searched for studies published before August 2020. Clinical trials in patients with type 1 diabetes mellitus and type 2 diabetes mellitus with reports of glucagon changes before and after SGLT2i intervention were included. Eligible trials were analyzed by fixed-effect model, random effect model, and meta-regression analysis accordingly. In total, ten trials were included in this meta-analysis. Compared with the non-SGLT2i treatment group, SGLT2i treatment resulted in increased plasma fasting glucagon levels with significance (WMD, 8.35 pg/ml; 95% CI, 2.17–14.54 pg/ml, P＜0.01) in patients with diabetes mellitus. Besides, when compared with non-SGLT2i control group, the insulin level decreased (WMD, −2.78 μU/ml; 95% CI, −5.11 to −0.46 μU/ml, P = 0.02) and ketone body level increased (WMD, 0.17 mmol/l; 95% CI, 0.09–0.25 mmol/l, P＜0.01) in patients with type 2 diabetes. In conclusion, our result indicated SGLT2i intervention would increase the plasma fasting glucagon level in patients with diabetes mellitus. The increase in plasma fasting glucagon level may be associated with reduced insulin level. The increased glucagon-insulin ratio after the use of SGLT2i may make diabetic patients susceptible to ketosis.

Organ-Specific Metabolic Pathways Distinguish Prediabetes, Type 2 Diabetes and Normal Tissues

Defects in pancreatic islets and the progression of multi-tissue insulin resistance in combination with environmental factors are the main causes of type 2 diabetes (T2D). Mass spectrometry-based proteomics of five key-metabolic tissues on a cohort of 42 multi-organ donors provided deep coverage of the proteomes of pancreatic islets, visceral adipose tissue (VAT), liver, skeletal muscle and serum. Enrichment analysis of gene ontology (GO) terms built a tissue-specific map of the chronological order of altered biological processes across healthy controls (CTRL), pre-diabetes (PD) and T2D subjects. This unique dataset allowed us to explore alterations of entire biological pathways and individual proteins in multiple tissues. We confirmed the significant decrease of the citric acid cycle and the respiratory electron transport in VAT and muscle of T2D and we provided a thorough visual representation of the complete set of downregulated proteins. Importantly, we found widespread novel alterations in relevant biological pathways including the increase in hemostasis in pancreatic islets of PD, the increase in the complement cascade in liver and pancreatic islets of PD and the elevation in cholesterol biosynthesis in liver of T2D. Overall, our findings suggest inflammatory, immune and vascular impairments in pancreatic islets as potentially causal factors of insufficient insulin production and increased glucagon levels in the early stages of T2D. In contrast alterations in lipid metabolism and mitochondrial function in the liver and VAT/muscle, respectively, became evident later in manifest T2D. This first multi-tissue proteomic map indicates the temporal order of tissue-specific metabolic dysregulation in T2D development.

The liver\u2013alpha cell axis associates with liver fat and insulin resistance: a validation study in women with non-steatotic liver fat levels

Aims/hypothesisMany individuals who develop type 2 diabetes also display increased glucagon levels (hyperglucagonaemia), which we have previously found to be associated with the metabolic syndrome. The concept of a liver–alpha cell axis provides a possible link between hyperglucagonaemia and elevated liver fat content, a typical finding in the metabolic syndrome. However, this association has only been studied in individuals with non-alcoholic fatty liver disease. Hence, we searched for a link between the liver and the alpha cells in individuals with non-steatotic levels of liver fat content. We hypothesised that the glucagon–alanine index, an indicator of the functional integrity of the liver–alpha cell axis, would associate with liver fat and insulin resistance in our cohort of women with low levels of liver fat.MethodsWe analysed data from 79 individuals participating in the Prediction, Prevention and Subclassification of Type 2 Diabetes (PPSDiab) study, a prospective observational study of young women at low to high risk for the development of type 2 diabetes. Liver fat content was determined by MRI. Insulin resistance was calculated as HOMA-IR. We conducted Spearman correlation analyses of liver fat content and HOMA-IR with the glucagon–alanine index (the product of fasting plasma levels of glucagon and alanine). The prediction of the glucagon–alanine index by liver fat or HOMA-IR was tested in multivariate linear regression analyses in the whole cohort as well as after stratification for liver fat content ≤0.5% (n = 39) or >0.5% (n = 40).ResultsThe glucagon–alanine index significantly correlated with liver fat and HOMA-IR in the entire cohort (ρ = 0.484, p < 0.001 and ρ = 0.417, p < 0.001, respectively). These associations resulted from significant correlations in participants with a liver fat content >0.5% (liver fat, ρ = 0.550, p < 0.001; HOMA-IR, ρ = 0.429, p = 0.006). In linear regression analyses, the association of the glucagon–alanine index with liver fat remained significant after adjustment for age and HOMA-IR in all participants and in those with liver fat >0.5% (β = 0.246, p = 0.0.23 and β = 0.430, p = 0.007, respectively) but not in participants with liver fat ≤0.5% (β = −0.184, p = 0.286).Conclusions/interpretationWe reproduced the previously reported association of liver fat content and HOMA-IR with the glucagon–alanine index in an independent study cohort of young women with low to high risk for type 2 diabetes. Furthermore, our data indicates an insulin-resistance-independent association of liver fat content with the glucagon–alanine index. In summary, our study supports the concept that even lower levels of liver fat (from 0.5%) are connected to relative hyperglucagonaemia, reflecting an imminent impairment of the liver–alpha cell axis.

154-OR: Neprilysin Inhibition Increases Glucagon Levels with Possible Implications for Hepatic Amino Acid Metabolism

Glucagon (gcg) regulates hepatic glucose and amino acid (AA) metabolism, and increased gcg levels (hyperglucagonemia) contribute to hyperglycemia in diabetes. We hypothesized that the enzyme neprilysin (NEP) contributes to gcg degradation. We measured plasma gcg levels during a mixed meal in nine healthy men after a single dose of the NEP-inhibitor/angiotensin II receptor blocker (194 mg sacubitril/206 mg valsartan, 30 min prior to meal), a DPP-4 inhibitor (sitagliptin, 2x100mg), these combined, or the meal alone. Postprandial gcg levels were 2.7-fold higher in sacubitril/valsartan treated individuals compared to controls (P=0.005) and this was not significantly altered by the addition of sitagliptin (P=0.28). Sacubitril/valsartan also lowered postprandial plasma AA levels (P=0.01), but glucose levels were unaffected (P=0.76). In obese individuals (n=7), eight weeks sacubitril/valsartan treatment increased fasting gcg levels (P=0.02). To test whether NEP degrades gcg and diminishes its signaling, we performed mass-spectrometry and assessed cells transfected with the gcg receptor (gcgr). We found that NEP cleaves gcg and that the gcg fragments produced were unable to activate the gcgr. In non-sedated C57BL/6JRj female mice (n=8) NEP inhibition (sacubitril, 0.7 nmol/g) increased gcg levels (P=0.02) and tended to increase AA disappearance (P=0.076) and urea formation (P=0.08) during an AA challenge. A gcgr antagonist (Novo Nordisk; 25-2648, 100 mg/kg) abolished the increase in urea formation observed with sacubitril alone. Gcg levels were increased 1.9-fold (P&amp;lt;0.002) with sacubitril compared to without, after a single injection of gcg (96 ng/g). In mice with genetic ablation of NEP (n=10), fasting plasma urea (P=0.003) but surprisingly not gcg levels (P=0.57) were increased compared to controls. NEP degrades gcg and thus inhibitors of NEP may result in hyperglucagonemia with potential metabolic perturbations on hepatic AA metabolism. Disclosure S. Kjeldsen: None. S. Zraika: Research Support; Self; Novartis Pharmaceuticals Corporation. S.M. Mongovin: None. L.H. Hansen: None. D. Terzic: None. P.D. Mark: None. P. Plomgaard: None. J.P. Gøtze: None. M. Winther-Soerensen: None. J. Hunt: None. K.D. Galsgaard: None. M.M. Rosenkilde: Board Member; Self; Synklino. Consultant; Self; Antag Therapeutics, Bainan Biotech. G.H. Goossens: None. E.E. Blaak: None. J.J. Holst: Advisory Panel; Self; AstraZeneca, Merck Sharp &amp; Dohme Corp., Novo Nordisk A/S, Zealand Pharma A/S. Other Relationship; Spouse/Partner; Antag Therapeutics. N.J. Wewer Albrechtsen: Research Support; Self; Mercodia, Novo Nordisk A/S, Novo Nordisk Foundation. Speaker’s Bureau; Self; Merck Sharp &amp; Dohme Corp. Funding Novo Nordisk Foundation (NNF19OC0055001)

Epinephrine Responsiveness is Reduced in Livers from Trained Mice

Catecholamines increase liver glucose production at least in part through increases in glycogenolysis and gluconeogenesis. To date, the effects of prior exercise training on the liver’s response to epinephrine, in vivo, have not been fully elucidated. To examine the role of epinephrine signaling on indices of liver glucose production in trained mice, male C57BL/6 mice were subject to either 12 days of voluntary wheel running (TR) or remained sedentary (SED). Epinephrine (0.5 mg/kg body weight), or vehicle were injected intraperitoneally following 12 days of training and tissues harvested 15 minutes post‐injection. The rise in blood glucose following epinephrine injection was significantly blunted in trained compared to sedentary mice and this was paralleled by a blunted reduction in liver glycogen in trained mice. There was a main effect of epinephrine to increase the phosphorylation of protein kinase‐A substrates (pPKAs), which was driven by increases in the sedentary, but not trained, mice. Similarly, epinephrine‐induced increases in the mRNA expression of α‐ and β‐adrenergic receptor 1 and 2 and glucose‐6‐phosphatase were greater in SED compared to TR mice. Exercise and epinephrine decreased serum insulin levels and increased glucagon levels, however, there were no differences within epinephrine treated groups. Changes in phosphorylated AKT, AMPK, and phosphodiesterase 4B protein content, negative regulators of PKA signaling, do not explain the protective effects of exercise training against epinephrine induced increases in blood glucose and liver glycogen breakdown. Taken together our data suggest that prior exercise training reduces the liver’s response to epinephrine. This could be beneficial in the context of training‐induced sparing of glycogen during exercise.Support or Funding InformationThis work was funded, in part, through a National Sciences and Engineering Research Council of Canada (NSERC) grant to D.C.W., who is a Tier II Canada Research Chair in Lipids, Metabolism, and Health. H.A.D was supported by an Ontario Graduate Scholarship. G.L.M. was supported by an Ontario Graduate Scholarship and subsequently an NSERC Canada Graduate Scholarship. L.K.T. was supported by a Dairy Farmers of Ontario Doctoral Research Assistantship and an NSERC Postgraduate Scholarship.

Fasting serum total bile acid levels are associated with insulin sensitivity, islet β-cell function and glucagon levels in response to glucose challenge in patients with type 2 diabetes.

Type 2 diabetes (T2D) is characterized by islet β-cell dysfunction and impaired suppression of glucagon secretion of α-cells in response to oral hyperglycaemia. Bile acid (BA) metabolism plays a dominant role in maintaining glucose homeostasis. So we evaluated the association of fasting serum total bile acids (S-TBAs) with insulin sensitivity, islet β-cell function and glucagon levels in T2D. Total 2,952 T2D patients with fasting S-TBAs in the normal range were recruited and received oral glucose tolerance tests for determination of fasting and postchallenge glucose, C-peptide and glucagon. Fasting and systemic insulin sensitivity were assessed by homeostasis model assessment (HOMA) and Matsuda index using C-peptide, i.e., ISHOMA-cp and ISIM-cp, respectively. Islet β-cell function was assessed by the insulin-secretion-sensitivity-index-2 using C-peptide (ISSI2cp). The area under the glucagon curve (AUCgla) was used to assess postchallenge glucagon. The results showed ISHOMA-cp, ISIM-cp and ISSI2cp decreased, while AUCgla notably increased, across ascending quartiles of S-TBAs but not fasting glucagon. Moreover, S-TBAs were inversely correlated with ISHOMA-cp, ISIM-cp and ISSI2cp (r = -0.21, -0.15 and -0.25, respectively, p < 0.001) and positively correlated with AUCgla (r = 0.32, p < 0.001) but not with fasting glucagon (r = 0.033, p = 0.070). Furthermore, after adjusting for other clinical covariates by multiple linear regression analyses, the S-TBAs were independently associated with ISHOMA-cp (β = -0.04, t = -2.82, p = 0.005), ISIM-cp (β = -0.11, t = -7.05, p < 0.001), ISSI2cp (β = -0.15, t = -10.26, p < 0.001) and AUCgla (β = 0.29, t = 19.08, p < 0.001). Increased fasting S-TBAs are associated with blunted fasting and systemic insulin sensitivity, impaired islet β-cell function and increased glucagon levels in response to glucose challenge in T2D.

Insights Into GLP-1 and GIP Actions Emerging From Vildagliptin Mechanism Studies in Man.

Vildagliptin blocks glucagon like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) inactivation of the meal induced increases in GLP-1 and GIP so that elevated GLP-1 and GIP levels are maintained over 24 h. The primary insulin secretion effect of vildagliptin is to improve the impaired sensitivity of the β-cells to glucose in subjects with impaired fasting glucose (IFG) and impaired glucose tolerance (IGT) and in patients with type 2 diabetes mellitus (T2DM); this effect was seen acutely and maintained over at least 2 years in patients with T2DM. Vildagliptin was also associated with improved β-cell function that is likely secondary to the improved metabolic state. Although there was no evidence of restoration of β-cell mass, the preponderance of the vildagliptin data does indicate that for at least 2 years β-cell function was maintained in vildagliptin treated patients but not in the untreated patients. Vildagliptin suppressed an inappropriate glucagon response to an oral glucose challenge in patients with T2DM, to a mixed meal challenge in patients with T2DM and type 1 diabetes mellitus, and to a mixed meal challenge in subjects with IGT and IFG. The improved glucagon response was maintained for at least 2 years in patients with T2DM and there was no change in the glucagon response in normoglycemic individuals. Vildagliptin lowered glucose levels into the normal range without increasing hypoglycemia. These hypoglycemic benefits appear to be secondary in large part to the improved sensitivity of both the β and α-cell to glucose. In the case of the α-cell, if glucose levels are high, GLP-1 attenuates the glucagon levels and if glucose levels are low, GIP increases glucagon levels. Vildagliptin reduces fatty acid flux from the adipocyte leading to reduced liver fat which in turn leads to increased glucose utilization. The reduced glycosuria and reduced lipo-toxicity associated with vildagliptin therapy does not lead to weight gain presumably due to increased fat mobilization and oxidation during meals and to reduced fat extraction from the gut.

Hypoxia causes reductions in birth weight by altering maternal glucose and lipid metabolism

Hypoxia of residence at high altitude (>2500 m) decreases birth weight. Lower birth weight associates with infant mortality and morbidity and increased susceptibility to later-in-life cardiovascular and metabolic diseases. We sought to determine the effects of hypoxia on maternal glucose and lipid metabolism and their contributions to fetal weight. C57BL6/NCrl mice, housed throughout gestation in normobaric hypoxia (15% oxygen) or normoxia, were studied at mid (E9.5) or late gestation (E17.5). Fetal weight at E17.5 was 7% lower under hypoxia than normoxia. The hypoxic compared with normoxic dams had ~20% less gonadal white adipose tissue at mid and late gestation. The hypoxic dams had better glucose tolerance and insulin sensitivity compared with normoxic dams and failed to develop insulin resistance in late gestation. They also had increased glucagon levels. Glucose uptake to most maternal tissues was ~2-fold greater in the hypoxic than normoxic dams. The alterations in maternal metabolism in hypoxia were associated with upregulation of hypoxia-inducible factor (HIF) target genes that serve, in turn, to increase glycolytic metabolism. We conclude that environmental hypoxia alters maternal metabolism by upregulating the HIF-pathway, and suggest that interventions that antagonize such changes in metabolism in high-altitude pregnancy may be helpful for preserving fetal growth.