bluuebunny/crossref_metadata_2025_split huggingface dataset:
1461-P: Quantifying Proinsulin Misfolding in the Endoplasmic Reticulum
Authors: ARUNAGIRI ANOOP , ALAM MD MAROOF , HAATAJA LEENA , SAMY PRAVEEN J. , SOLEIMANPOUR SCOTT , SATIN LESLIE S. , ITKIN-ANSARI PAMELA , LIU MING , ARVAN PETER 
Published: May 2022

We recently reported that normal pancreatic islets or beta cell lines contain a subset of proinsulin molecules that have not achieved the three intramolecular disulfide bonds required for native insulin structure. Indeed, we found that non-native proinsulin monomers can form intermolecular disulfide-linked complexes, which are detectable by immunoblotting after nonreducing SDS-PAGE. However, we have recognized difficulties in the quantitative interpretation of these analyses, as we frequently identified conditions in which little or no proinsulin monomer was detected, and non-native disulfide-linked complexes amounted to greater cumulative proinsulin band intensity than the entire population of proinsulin molecules detected by immunoblotting upon reducing SDS-PAGE. Here we have developed several critical modifications of our methodology to rectify these discrepancies to undertake a more accurate determination of the pool of misfolded proinsulin molecules in pancreatic beta cells under conditions of health and disease. Using these modifications, we find that the size of the proinsulin monomer pool has previously been underestimated and the fraction of disulfide-linked complexes overestimated — and this can be readily explained by proinsulin antibody affinity in the immunoblotting protocol. Additionally by immunoblotting, we now resolve native and non-native proinsulin monomers, highlighting the inefficiency of non-native proinsulin transport through the secretory pathway.

Disclosure
A.Arunagiri: None. M.Alam: None. L.Haataja: None. P.J.Samy: None. S.Soleimanpour: None. L.S.Satin: None. P.Itkin-ansari: None. M.Liu: None. P.Arvan: n/a.

Funding
National Institutes of Health (R01-DK48280) , National Institutes of Health (U01-DK127747) , and National Institutes of Health (R24 DK110973)
1466-P: Pancreatic Delta Cells Switched Islet's Ca2+ Oscillation Modes by Inhibiting Endogenous Glucagon from Alpha Cells
Authors: REN HUIXIA , GROUP CHAO TANG , GROUP LIANGYI CHEN 
Published: May 2022

Glucose-stimulated pancreatic islets show different patterns of Ca2+ signaling with fast, slow and mixed fast/slow oscillations. However, it is not clear whether they are characteristic features of individual islets. We report islet spontaneous slowdown from fast to slow/mixed Ca2+ oscillation under high glucose stimulation. The fast oscillation stage typically lasts for several hours and intrinsic to the islet. The modes switch is reversible. Low glucose resting for 30 minutes fully restored the fast oscillation. Microfluidic perfusion delays and static incubation brings forward the mode switch, which suggests the critical role of the paracrine environment. Correspondingly, exogenous glucagon delays and somatostatin brings forward the mode switch. Pharmacologically blocking Sstr2 on α cells triggers endogenous glucagon secretion and switches islets back to fast oscillations. Optoinhibition and relief of δ cells cause islets to repeatably show fast and slow oscillations. Our study highlights the importance of δ-α cell interactions to generate tunable islet oscillation patterns. The design principle of pancreatic islet was revealed, as the minority of δ cells command the majority of α and β cells.

Funding
No
149-LB: Pediatric Exogenous Insulin Antibody Syndrome—Successful Treatment with Mycophenolate Mofetil
Authors: SABA LESLIE , ORANDI AMIR B. , PITTOCK SIOBHAN , CREO ANA 
Published: May 2022

Case Description: A lean, 17-year-old Caucasian male with type 1 diabetes diagnosed at age 9 experienced an abrupt increase in his insulin requirements. He was managed on mixed daily injections. Daily insulin requirements increased from 1.7 units/kg/day to 3.3 units/kg/day over 6 months. His total insulin level was 7 mcIU/mL with a free insulin level of 4.8 mcIU/mL, indicating exogenous insulin antibodies. Switching from insulin aspart to glulisine, due to presumed antibodies against insulin aspart was unsuccessful, and his insulin requirements continued to increase to 4.4 units/kg/day. Immunosuppressive therapy was commenced with mycophenolate mofetil, decreasing his insulin requirements to 1.1 units/kg/day after 7 months of treatment. Repeat testing revealed an improvement in the discrepancy between total and free insulin levels, with a total insulin level of 5.2 mcIU/mL and free insulin level of 4.6 mcIU/mL.
Discussion: Exogenous insulin antibody syndrome should be considered in patients with poorly controlled diabetes and requiring greater than 2 units/kg/day of insulin without obesity. Classically, antibodies against exogenous insulin were found in patients receiving insulin derived from animal sources; however, recombinant human insulin can very rarely trigger an antibody response in select patients. Immunosuppression using mycophenolate mofetil greatly reduced insulin requirements and this case highlights its novel use in children with diabetes who develop insulin antibody syndrome.

Disclosure
L. Saba: None. A. B. Orandi: None. S. Pittock: None. A. Creo: None.
15-LB: Safety and Efficacy of Tetramethylpyrazine Nitrone in Treating Patients with Type 2 Diabetes and Microalbuminuria—A Multicenter, Randomized, Double-Blind, Placebo-Controlled Phase II Clinical Trial
Authors: YANG JINKUI 
Published: May 2022

Background and purpose: Tetramethylpyrazine nitrone (TBN) is a multi-functional agent that has shown kidney protective effects in preclinical studies. TBN has a unique mechanism of action, i.e., scavenging free radicals and activating AMPK/PGC-1α/NRF2 pathway. This phase II clinical trial investigated the safety and efficacy of TBN in patients with early diabetic kidney disease (DKD) .
Methods: This multicenter, randomized, double-blind, placebo-controlled trial enrolled 143 Chinese type 2 diabetic patients (T2DM) with microalbuminuria (urinary albumin-to-creatinine ratio ≥ 30 to &lt;300 mg/g creatinine) treated with or without renin-angiotensin system inhibitor who had eGFR ≥60 ml/min per 1.73 m2. Participants were randomized (1:1:1:1) to receive 300 mg, 600 mg or 1200 mg TBN bid or placebo for 180 days. The primary outcome was change in urinary albumin-to-creatinine ratio (UACR) from baseline to day 180. The safety outcome included any adverse events during 180 days.
Results: TBN was safe and well-tolerated. There were no statistical differences among the incidences of drug-related adverse event among the TBN and the placebo groups. TBN (600 mg, bid) decreased UACR by -46.56% [90% CI,-61.15, -1.74, p&gt;0.99] from baseline to day 180 versus -14.33% [90% CI, -38.26, 19.39] in the placebo group. The proportion of patients with ≥ 30% reduction in UACR was 54.8% [95% CI, -9.10, 38.26, p=0.216] in the TBN (600 mg) group versus 39.4% in the placebo group. The change in eGFR from baseline to day 180 was 0 [95% CI, -6.66, 5.00, p&gt;0.99] in the TBN (600 mg) group versus a decline of -2.12% [95% CI, -11.90, -0.63] in the placebo group.
Conclusions: TBN is a promising new treatment for early DKD and its safety and efficacy will be further investigated in clinical trials.

Disclosure
J. Yang: None.

Funding
Guangzhou Magpie Pharmaceutical Co. Ltd (CTR20220455)
181-LB: Fracture Risk of Antidiabetic Agents in Type 2 Diabetes—A Network Meta-analysis
Authors: HSUAN TSAI WEN , KE KONG SIANG , TSAI MING-CHIEH 
Published: May 2022

Background: Diabetes is associated with greater risk of fracture. Our aim is to evaluate the correlation of different antidiabetic agents with the risk of fracture among subjects with type 2 diabetes.
Methods: The literature search was conducted in PubMed, Embase, and ClinicalTrials.gov on 22 October 2020. Keywords included “type 2 diabetes,” “fracture,” “randomized controlled trial,” “metformin,” “insulin,” “thiazolinedione,” “sodium-glucose co-transporter-2 inhibitor,” “glucagon-like peptide-1 receptor agonists,” “dipeptidyl peptidase 4 inhibitors” and “sulfonylurea.” Two authors (W-H. T and M-C.T) conducted study selection and data extraction independently. The primary main outcome was the fracture events during the followed-up period pooled by random-effected models.
Results: A total of 214686 patients with diabetes in 188 studies were included and 2850 patients had fracture events. Patients taking sodium-glucose co-transporter-2 inhibitor had higher risk of fracture when compared with glucagon-like peptide-1 receptor agonists (risk ratio [RR] 1.43 [95% confidence interval (CI) 1.08–1.91]) and sulfonylurea (RR 1.45 [95% CI 1.07–1.98]) . Patients taking dipeptidyl peptidase 4 inhibitors also had higher risk of fracture when compared with glucagon-like peptide-1 receptor agonists (risk ratio [RR] 1.4 [95% confidence interval (CI) 1.03–1.91]) and sulfonylurea (RR 1.42 [95% CI 1.03–1.95]) . Metformin, glucagon-like peptide-1 receptor agonists and sulfonylurea had protective effect on fracture when compared with thiazolinedione.
Conclusions: Patients with diabetes taking sodium-glucose co-transporter-2 inhibitor or dipeptidyl peptidase 4 inhibitors had higher risk of fracture when compared with glucagon-like peptide-1 receptor agonists and sulfonylurea. Metformin, glucagon-like peptide-1 receptor agonists and sulfonylurea had protective effect on fracture when compared with thiazolinedione.

Disclosure
W. Tsai: None. S. Kong: None. M. Tsai: None.
191-OR: Hepatic Expression of the Branched-Chain Keto Acid Dehydrogenase Kinase Is Sufficient to Cause Glucose Intolerance in Lean Mice
Authors: WALEJKO JACQUELYN , MCGARRAH, III ROBERT W. , WHITE PHILLIP J. 
Published: May 2022

Perturbed branched-chain amino acid homeostasis is a feature of obesity, insulin-resistance and type 2 diabetes. Induction of the branched-chain keto acid dehydrogenase kinase (BDK) in liver is one mechanism that is thought to contribute to elevated BCAA levels in these conditions. Whether higher levels of BDK in liver are sufficient to disturb systemic BCAA homeostasis and promote glucose intolerance in the absence of obesity or insulin resistance has never been directly tested. We treated 12 week-old C57BL6/J mice with adeno-associated virus, serotype 8 (AAV8) -BDK or control AAV8-GFP to promote long term expression of BDK specifically in liver of lean mice. Mice were then maintained on a standard low-fat chow diet for 8 weeks (n= 9 AAV8-GFP and n=AAV8-BDK) . AAV8-BDK significantly raised BDK protein and mRNA expression in liver but not adipose, skeletal muscle, or cardiac tissue compared to AAV8-GFP control mice. The higher expression of BDK in liver was associated with robust inhibitory phosphorylation of the branched-chain keto acid dehydrogenase (BCKDH) on ser293. AAV8-mediated induction of BDK expression in liver in our study was sufficient to promote significant elevations in circulating levels of the direct substrate of BCKDH, the branched chain α-keto acids as well as their cognate BCAA. There was no effect of AAV8-BDK to alter food intake, weight gain or adiposity. However, glucose tolerance measured by a 1g/kg intraperitoneal glucose tolerance test at week 7 was significantly impaired in mice receiving AAV8-BDK compared to AAV8-GFP control. These data demonstrate for the first time that induction of BDK in liver, as occurs with obesity, is sufficient to disturb systemic BCAA and glucose homeostasis in lean mice.

Disclosure
J.Walejko: None. R.W.Mcgarrah: None. P.J.White: None.

Funding
American Diabetes Association (1-16-INI-17)
193-OR: Chronotherapy with the Glucokinase Activator AZD1656 Greatly Affects Its Ability to Improve Metabolism in Obese Zucker Rats
Authors: KROON TOBIAS , HAGSTEDT THERESE , ALEXANDERSSON IDA , FERM ANNETT , PETERSSON MARIE , MAURER STEFANIE , ZARROUKI BADER , WALLENIUS KRISTINA , OAKES NICHOLAS D. , BOUCHER JEREMIE 
Published: May 2022

Circadian rhythms play critical roles in regulating metabolism, including daily feeding/fasting periods and oscillating expressions of certain genes. Glucokinase (Gck) , expressed in the liver and pancreas, is central for whole-body glucose homeostasis by acting as a glucose sensor and regulating metabolism according to prevailing glucose levels. Gck expression has also been shown to follow a circadian rhythm. Gck activators (GKAs) effectively reduce hyperglycemia, however, adverse events have also involved hypoglycemia, hyperlipidemia, and hepatic steatosis. Given the circadian rhythmicity and natural postprandial activation of Gck, we hypothesized that GKA treatment would benefit from being timed specifically during feeding periods. We compared the metabolic effects following differentially timed administrations of the GKA AZD1656 in the obese, insulin resistant Zucker rat. Four weeks of continuous treatment significantly improved glycemic control; however, hepatic steatosis and inflammation manifested. In contrast, a robust reduction in hepatic steatosis and inflammation on top of improved glycemia, was observed when AZD1656 administration was timed to feeding periods only. When AZD1656 was timed to fasting periods, detrimental metabolic effects were observed, including impaired glycemic control, severe liver steatosis and elevated inflammation. Mechanistically, timing AZD1656 administration to feeding periods was found to divert de novo lipogenesis-derived lipids away from hepatic storage towards VLDL synthesis and secretion, without affecting plasma triglyceride levels. In conclusion, intermittent AZD1656 dosing timed to feeding periods promotes glucose disposal when needed the most, while the daily drug holiday allows for maintained metabolic flexibility and avoids hepatic steatosis. Chronotherapeutic approaches should be considered for the development of GKAs and other drugs acting on metabolic targets.

Disclosure
T. Kroon: Employee; AstraZeneca. I. Alexandersson: Employee; AstraZeneca. A. Ferm: Employee; AstraZeneca. M. Petersson: Employee; AstraZeneca. S. Maurer: Employee; AstraZeneca, Merck Sharp & Dohme Corp. B. Zarrouki: Employee; AstraZeneca. K. Wallenius: Employee; AstraZeneca. N.D. Oakes: Employee; AstraZeneca. J. Boucher: Employee; AstraZeneca, Evotec International GmbH.
195-OR: A Novel 13C5 Glutamine Tracer Method (Q Flux) Reveals a Key Role of Succinyl CoA Anaplerosis in Promoting Increased Rates of Hepatic Gluconeogenesis during Hyperglucagonemia
Authors: HUBBARD BRANDON T. , SHULMAN GERALD I. 
Published: May 2022

Hyperglucagonemia is a hallmark of type 2 diabetes mellitus (T2DM) and contributes to increased rates of endogenous glucose production (EGP) , mostly through increased rates of hepatic gluconeogenesis (GNG) , but the anaplerotic pathways and respective substrates that support this process are unclear. To address this question, we developed a novel and extensively validated 13C5 glutamine-based in vivo metabolic flux analysis method (Q Flux) , which allows for quantitation of both the relative and absolute rates of hepatic gluconeogenesis from pyruvate (via pyruvate carboxylase [PC]) , succinyl CoA (via methylmalonyl CoA mutase [MUT]) , glutamine (via glutaminase [GLS2]) , and glycerol. Male Sprague-Dawley rats (∼300 g) were infused for 90 minutes with either low-dose (LD) glucagon (1.25 ng/[kg-min]) or high-dose (HD) glucagon (ng/[kg-min]) , plus insulin (0.5 mU/[kg-min]) , somatostatin (4 μg/[kg-min]) , and 13C5 glutamine (3 μmol/[kg-min]) after a 16-hour fast to deplete hepatic glycogen. HD animals displayed significantly increased plasma glucose concentrations and EGP relative to LD controls. Analyses of liver tissues collected at sacrifice revealed that in LD animals, 70% of mitochondrial GNG was from pyruvate, 15% from succinyl CoA, and 15% from glutamine. In HD animals, 59% of mitochondrial GNG was derived from pyruvate (p=0.vs. LD) , 26% from succinyl CoA (p=0.0vs. LD) , and 15% from glutamine (p=0.99 vs. LD) .
Conclusion: We find an unexpected role for succinyl CoA anaplerosis and methylmalonyl CoA mutase flux in supporting increased rates of mitochondrial GNG during hyperglucagonemia. Furthermore, relative contributions of glutamine to hepatic gluconeogenesis were unchanged, contrary to previous reports. Taken together, these results identify novel targets to reduce hyperglucagonemia-induced increases in rates of gluconeogenesis and hyperglycemia in T2DM.

Disclosure
B.T.Hubbard: None. G.I.Shulman: Advisory Panel; 89bio, Inc., AstraZeneca, Equator Therapeutics, Inc., Janssen Research & Development, LLC, Merck & Co., Inc., Consultant; DiCerna Pharmaceuticals, Inc. , Novo Nordisk, Other Relationship; Generian Pharmaceuticals, iMetabolic Biopharma Corporation, Maze Therapeutics, The Liver Company, Stock/Shareholder; Levels Health, Inc. .
196-OR: (Pro) renin Receptor Signaling in Tyrosine Hydroxylase Neurons in the Hypothalamic Paraventricular Nucleus Is Required for Obesity-Associated Glucose Metabolic Impairment
Authors: SOUZA LUCAS AC. , WORKER CALEB J. , GAYBAN ARIANA JULIA , SCHWARTZ MICHAEL W. , EARLEY YUMEI FENG 
Published: May 2022

We recently reported that neuron-specific (pro) renin receptor (PRR) deletion reduces fasting blood glucose (BG) levels and improves glucose tolerance in high fat diet (HFD) -treated mice. To investigate underlying mechanisms, we focused on tyrosine hydroxylase (TH) neurons in the hypothalamic paraventricular nucleus (THPVN neurons) , a largely GABAergic population that expresses PRR and projects to autonomic centers important for BG regulation. To delete PRR from THPVN neurons, we injected into the PVN of PRR-loxp mice either an adeno-associated virus expressing cre recombinase under the control of the rat TH promoter (AAV2-TH-Cre) or AAV2-TH-eGFP as a control. After placing animals on a HFD for 6 wk, we found that the obesity-induced increase of fasting BG levels was blocked by PRR deletion from THPVN neurons (118.0 ± 3.7 vs. 139.3 ± 6.6 mg/dL, n=6/group, p= 0.04) , accompanied by a trend towards improved i.p. glucose tolerance (AUC: 12990 ± 1136 vs. 9467 ± 1298, n=6, p=0.07) . To further investigate the role of THPVN neurons in glucose homeostasis, we employed a Designer Receptors Exclusively Activated by Designer Drugs (DREADD) approach whereby AAV2-TH-Cre was injected bilaterally into the PVN of either hM3Dq-loxp (excitatory) or hM4Di-loxp (inhibitory) mice, and continuous glucose monitoring was performed in conscious, freely moving mice. Following i.p. injection of clozapine-N-oxide (CNO, 1 mg/Kg) or vehicle, we found that whereas basal BG levels were not altered by THPVN neuron activation, THPVN neuron inhibition significantly increased basal BG levels compared to vehicle in male (ΔBG: 31.6 ± 8.6 mg/dL vs. 11.0 ± 6.7 mg/dL, n=4, p=0.03) but not female mice. These findings establish a role for THPVN neuron PRR signaling in the pathogenesis of obesity-associated glucose metabolic impairment, and show that in male mice, basal THPVN neuron activity is required for normal glucose homeostasis.

Disclosure
L.Ac. souza: None. C.J.Worker: None. A.Gayban: None. M.W.Schwartz: None. Y.Feng earley: None.

Funding
National Institutes of Health (R01HL122770, R35HL155008, RDK83042, RDK124238)
199-LB: Artemisia scoparia Attenuates Glucocorticoid-Induced Lipolysis in Adipocytes
Authors: HARVEY INNOCENCE , STEPHENS JACQUELINE M. 
Published: May 2022

Prescription glucocorticoid use is widespread across developed countries for the treatment of several inflammatory conditions. Elevated glucocorticoids are known to promote lipolysis and often result in metabolic disorders such as insulin resistance and hepatic lipid accumulation. An extract of Artemisia scoparia (SCO) has been shown to reduce lipolysis and promote metabolic health but has not been investigated in the context of glucocorticoid-mediated metabolic disease. We assessed the ability of SCO to modulate glucocorticoid-induced lipolysis in 3T3-L1 adipocytes. Mature adipocytes were pretreated with vehicle (DMSO) , SCO (50 ug/mL) , or rosiglitazone for three days. Cells from each condition were then exposed to either vehicle (ethanol/BSA) , Dexamethasone (synthetic glucocorticoid; 500 nM) , or TNFa (0.75 nM) , in the presence or absence of RU486 (glucocorticoid receptor (GR) antagonist; 250 nM) for 20 hours. Separate experiments were performed to determine the involvement of the GR in SCO’s effects using an siRNA approach. Glycerol and free fatty acids (FFA’s) were measured from the media to evaluate changes in lipolysis. The expression and phosphorylation status of several lipolytic genes and proteins were also assessed via qPCR and western blot, respectively. SCO significantly attenuated dexamethasone-induced glycerol and FFA release, with the combination of SCO and RU486 resulting in the greatest effects. SCO led to significant reductions in the monomeric forms of PDE3B and PDE4D, but greater expression of multimeric complexes were observed. Reduction in GR expression did not interfere with the effects of SCO, suggesting the GR is not required for SCO-mediated attenuation of lipolysis. These observations shed some light on the protective effects of SCO under conditions of elevated glucocorticoids, but additional studies are needed to identify the underlying mechanisms that confer beneficial effects of SCO to combat both diet- and glucocorticoid-induced metabolic dysfunction.

Disclosure
I. Harvey: None. J. M. Stephens: None.

Funding
National Center for Complementary and Integrative Health T32 (AT004094)
207-LB: Aldose Reductase Inhibition by AT-001 Limits Diastolic Dysfunction and Adverse Remodelling in Diabetic Cardiomyopathy
Authors: KARWI QUTUBA G. , GOPAL KESHAV , AMIRHOSSEIN TABATABAEI DAKHILI SEYED , WAGG CORY S. , ZHANG LIYAN , SUN QIUYU , SAED CHRISTINA T. , PANIDARAPU SAI , PERFETTI RICCARDO , RAMASAMY RAVICHANDRAN , USSHER JOHN R. , LOPASCHUK GARY 
Published: May 2022

Diabetic cardiomyopathy (DbCM) increases mortality and morbidity in type 2 diabetes (T2D) subjects. Increased cardiac aldose reductase (AR) activity has been correlated with impaired cardiac function and less effective energy metabolism in DbCM subjects. The aim of the present study was to evaluate the effect AT-001, a potent and selective AR inhibitor, on cardiac function, structure, and energy metabolism in diabetic cardiomyopathic mice that overexpress the human AR (hAR-Tg) . The effects of AT-001 were compared to those of dapagliflozin, a sodium-glucose cotransporter inhibitor that decreases the risk of cardiovascular death. DbCM was induced in human AR overexpressing transgenic (hAR-Tg) mice by subjecting them to a high-fat diet (60% kcal from lard) for 10-wk with a single intraperitoneal streptozotocin injection (75 mg/kg) at 4-wk. Male mice were randomized to receive either vehicle, AT-001 (40 mg/kg/day) ,or dapagliflozin (1 mg/kg/day) for the final 3-wk. AT-001 treatment improved diastolic function in vivo (a decrease in the E/e’ ratio and an increase in the E’/A’) and reduced left ventricular mass (LV) mass in DbCM mice compared to vehicle-treated DbCM mice. Cardioprotection by AT-001 was associated with reduced cardiac AR activity, decreased circulating blood sorbitol levels, and a reduction in cardiac fatty acid oxidation rates in the DbCM mice. Treatment with dapagliflozin did not significantly affect either diastolic function or LV mass in DbCM mice. In addition, dapagliflozin did not have a significant effect on cardiac fatty acid oxidation rates.
In summary, the present study demonstrates that AT-001 attenuates diastolic dysfunction and cardiac hypertrophy in DbCM. AT-001-induced cardioprotection is accompanied by reduced cardiac fatty oxidation rates in DbCM.

Disclosure
Q. G. Karwi: None. R. Ramasamy: Consultant; Applied Therapeutics. J. R. Ussher: None. G. Lopaschuk: None. K. Gopal: None. S. Tabatabaei dakhili: None. C. S. Wagg: None. L. Zhang: None. Q. Sun: None. C. T. Saed: None. S. Panidarapu: None. R. Perfetti: Employee; Applied Therapeutics.