bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of Deep Transcranial Magnetic Stimulation (dTMS) on Anti-inflammatory Gut Bacterial Species in Obesity
Authors: FERRULLI ANNA , TOSCANO MARCO , ADAMO MICHELA , TERRUZZI ILEANA , DRAGO LORENZO , LUZI LIVIO
Published: June 2018

Growing evidence highlights the crucial role of gut microbiota in affecting different aspects of obesity. Potential mechanisms for microbial control over eating behavior include microbiota influence on reward and satiety pathways, neurotransmitters release, and hypothalamic-pituitary-adrenal (HPA) axis modulation. A methodology proved to be effective in modulating orexigenic/anorexigenic pathways and food-reward system in obesity is the dTMS. Considering its assets, we hypothesized a potential role of dTMS in inducing weight loss in obesity also via microbiota composition modulation. Twenty-two obese subjects (5 M, 17 F; 44.6±2.5 years; BMI 37.5±1.1 kg/m2) were allocated into 3 groups receiving 15 sessions (3 per week for 5 weeks) of high frequency (18 Hz, HF), low frequency (1 Hz, LF) dTMS, or sham stimulation. Fecal samples were collected at baseline and after 5 weeks of treatment. Total bacterial DNA was extracted from fecal samples using the QIAamp DNA Stool Mini Kit (Qiagen, Italy) and analyzed by a metagenomics approach (Ion Personal Genome Machine). After 5 weeks, a weight loss in HF (-3.4±1.0%; p<0.05) was found, associated with a decrease in norepinephrine (-61.5±15.2%; p<0.01). Furthermore, an increase of Faecalibacterium (+153.3% vs. baseline, p<0.05) and Alistipes (+153.4% vs. baseline, p<0.05) genera, and a significant decrease of Lactobacillus (-77.1% vs. baseline, p<0.05) were found in HF. A significant inverse correlation between Phascolarctobacterium variation and BMI reduction (R:-0.728; p<0.05) was found in HF; norepinephrine change significantly correlated with Eubacterium (R:0.857; p<0.05), Parasutterella (R:0.786; p<0.05), and Streptococcus (R:-0.937; p<0.01) variations.

In conclusion, the weight loss, induced by HF dTMS, improves sympathetic activity and gut microbiota composition in obese subjects, mainly promoting bacterial species with anti-inflammatory properties, that are typical of healthy subjects intestinal flora.

Disclosure

A. Ferrulli: None. M. Toscano: None. M. Adamo: None. I. Terruzzi: None. L. Drago: None. L. Luzi: Speaker's Bureau; Self; A. Menarini Diagnostics, AstraZeneca, Eli Lilly and Company. Research Support; Self; Gelesis. Consultant; Self; McKinsey & Company. Speaker's Bureau; Self; Menarini Group, Merck Sharp & Dohme Corp.. Research Support; Self; Novartis AG. Speaker's Bureau; Self; Novo Nordisk A/S. Research Support; Self; Sunstar Foundation. Speaker's Bureau; Self; Smith & Nephew.
Beneficial Effects of RYGB on ß-Cell Function and Hepatic and Peripheral Insulin Sensitivity Are Maintained Seven Years after Surgery in Both Diabetic and Nondiabetic Subjects
Authors: CAMASTRA STEFANIA , ASTIARRAGA BRENNO D. , BARBIERI CHIARA , GAGGINI MELANIA , PALUMBO MARIA , BERTA ROSSANA , CIOCIARO DEMETRIO , ANSELMINO MARCO , MARI ANDREA , FERRANNINI ELE , GASTALDELLI AMALIA
Published: June 2018

Gastric bypass surgery (RYGB) leads to improvement or remission of type 2 diabetes (T2D). Long-term studies indicate that the remission rate is higher 2 than 10 years post-surgery; whether the positive effects of surgery on ß-cell function (ß-GS) and glucose metabolism are maintained long term after surgery has not been determined. We studied 21 patients (12 T2D and 9 nondiabetic (ND)) before (B), 1 (1y) and 7 years (7ys) after RYGB with the use of a mixed meal test (MTT) in combination with glucose tracers to measure postmeal glucose fluxes and a euglycemic insulin clamp for insulin sensitivity (M). ß-GS (insulin secretion and dose response curve) was assessed by mathematical modelling. Weight loss 1y after RYGB was 30% and 35% of initial weight for ND and T2D respectively, and 24% and 30% at 7ys. T2D was resolved 1y post surgery in T2D and this outcome was maintained at 7ys (HbA1c 58±4 vs. 35±1 vs. 41±2 mmol/mol; B, 1y and 7ys). M value improved at 1y in proportion to the weight lost (5.0±0.8 to 7.2±0.7 in ND, and 4.3±0.8 to 7.5±0.7 mg. kgffm-1. min-1 in T2D, p<0.01 for both), and was maintained at 7ys (7.0±0.8 in ND and 6.9±0.9 mg.kgffm-1. min-1 in T2D, p=ns vs. 1y). Plasma glucose profiles were similar in ND and T2D at 1y and 7ys. After RYGB, most of the oral glucose appeared in the circulation in the first 90 min after meal ingestion in both groups and the dynamic of the oral glucose Ra was unchanged at 7ys. Suppression of endogenous glucose production (EGP) was improved in both groups during the first 90 min at 7ys compared to 1y; in T2D, mean EGP was significantly decreased. In T2D, the improvement in ß-GS seen at 1y (34±6 to 67±11 pmol.min-1.m-2.mM-1, p=0.001) was maintained at 7ys (80±21) at a similar level as in ND (138±18 vs. 87± 9 vs. 82±11, B, 1- and 7ys).

Conclusion: In both ND and T2D, RYGB induces marked improvements in glucose tolerance, insulin sensitivity (both muscle and hepatic) and ß-cell function that are maintained 7 years after surgery.

Disclosure

S. Camastra: None. B.D. Astiarraga: None. C. Barbieri: None. M. Gaggini: None. M. Palumbo: None. R. Berta: None. D. Ciociaro: None. M. Anselmino: None. A. Mari: None. E. Ferrannini: None. A. Gastaldelli: Advisory Panel; Self; Gilead Sciences, Inc.. Other Relationship; Self; European Foundation for the Study of Diabetes. Consultant; Self; INVENTIVA SA.
Impact of the Intragastric Balloon Associated with Diet on the Treatment of Diabetic Patients with Overweight or Grade I Obesity and Its influence on the Production of Enterohormones
Authors: REIMAO SILVIA M. , DA SILVA MARIA ELIZABETH R. , NUNES GABRIEL C. , MESTIERI LUIZ HENRIQUE M. , SANTOS ROSA F. , DE MOURA EDUARDO G.
Published: June 2018

Introduction: Obesity is one of the most important public health problems facing the world today. Intragastric balloon (BIB) is a short term, reversible, minimally invasive method for weight loss. Furthermore, it helps changing eating habits and behavior. It promotes gastric distension, resulting in decreased intake of food, delays gastric emptying and enhances the feeling of satiety. The objective of this study was to evaluate the effects of BIB in patients with type 2 diabetes mellitus (T2DM) and overweight or with grade I obesity, by analyzing the weight loss, body composition, glucose and lipid metabolism, production of enterohormones and quality of life.

Methods: Clinical trial including 40 patients with T2DM and BMI 27-34.9 kg/m2, which underwent placement of BIB, kept for 6 months, and follow-up for another 6 months after withdrawn. A standardized meal test diet was carried out at 0 and 6 months. Glucose, insulin, triglycerides, GLP-1 active and total PYY were measured at times 0, 30, 60, 120 and 180 minutes.

Results: The total weight decreased by 15.6 ± 7.23 kg after 6 months of use of BIB, which corresponds to loss of 17.1% of the total weight. After 6 months of follow-up, mean weight loss was maintained at 13.89 kg (15.2%). There was a significant reduction in body fat mass and area of the glucose curve, insulin and triglycerides (p ranged from <0.001 to 0.003). Of the 24 patients (60%) patients with hypertension at baseline, only 3 (7.5%) remained hypertensive after using the BIB. Serum levels of active GLP-1 and total PYY decreased with BIB. The quality of life improved in all domains analyzed (p < 0.001 to 0.041): physical functioning, physical function, pain, general health, vitality, social functioning, emotional aspects and mental health.

Conclusion: BIB is effective for weight loss and control of DM2 with quality of life improvement. There is no positive correlation with the production of enterohormones.

Disclosure

S.M. Reimao: None. M.R. da Silva: None. G.C. Nunes: None. L.M. Mestieri: None. R.F. Santos: None. E.G. de Moura: None.
Melatonin Improves Neovascularization after Hind-Limb Ischemia Surgery in Streptozocin-Induced Diabetic Mice by Enhancing Endothelial Progenitor Cells Functions
Authors: KUO CHIN-SUNG , HUANG PO-HSUN , LIN SHING-JONG
Published: June 2018

Objective: Diabetic patients show poor endothelial progenitor cell (EPC) functions and are prone to ischemic micro-vascular or macro-vascular complications. Melatonin has been shown to have protective effects in ischemic injury, possibly in part by modulating the functioning of EBCs. We investigated whether pretreatment with melatonin can restore the impaired functions of EPCs in diabetes.

Methods: Human EPC were isolated and cultured in a high-glucose medium for functional testing. Cell proliferation, nitric oxide (NO) production, and apoptosis assay were examined. A streptozocin (STZ)-induced diabetic mouse model was established to evaluate the actions of chronic hyperglycemia on ischemia-induced blood flow recovery. The circulating EPC number in the peripheral blood was determined by flow cytometry (Sca-1+/Flk-1+).

Results: Incubation with a high-glucose medium (25 mM) significantly suppressed EPC proliferation, reduced NO production, and lessened phosphorylations of Akt and eNOS. Moreover, EPC treated with high-glucose medium increased reactive oxygen species production, promoted cellular apoptosis and senescence, and also inhibited EPC tube formation. Treatment of melatonin could recover these harmful effects. Four weeks after hindlimb ischemia surgery, the STZ-induced diabetic mice had significantly reduced tissue reperfusion, EPC mobilization, and impaired neovascularization in the ischemic hindlimbs compared with the nondiabetic mice. In STZ-induced diabetic mice, the melatonin group showed significantly increased ischemic/non-ischemic limb blood perfusion ratios, higher capillary densities, and improved EPC mobilization.

Conclusion: Chronic hyperglycemia impaired blood flow recovery and EPC mobilization in response to tissue ischemia, and pretreatment of melatonin could reduce these effects.

Disclosure

C. Kuo: None. P. Huang: None. S. Lin: None.
Dietary Fat Quality and Genetic Risk of Type 2 Diabetes
Authors: MERINO JORDI , DASHTI HASSAN S. , GUASCH MARTA , ELLERVIK CHRISTINA , SMITH CAREN , KILPELAINEN TUOMAS O. , CHASMAN DANIEL , FLOREZ JOSE C.
Published: June 2018

Background: Type 2 diabetes (T2D) is a complex disease driven by lifestyle and genetic factors. The extent to which dietary fat quality may modify T2D genetic burden on the incidence of T2D is unknown.

Methods: We used Cox proportional-hazards models to calculate adjusted hazard ratios (HRs) for T2D among 103,2participants of European descent from 15 prospective cohort studies. T2D genetic risk profile was characterized by a 68-variant genetic risk score (GRS) weighted by published effect sizes. Diet was recorded using validated cohort-specific dietary assessment tools.

Findings: During a median follow-up of 12 years, 20,451 participants developed T2D. The relative risk of T2D per increment of 10 risk alleles in the GRS was 1.68 (95% confidence interval [CI] 1.62-1.74). Increasing monounsaturated fat intake in place of carbohydrates was associated with a higher risk of T2D (HR per 5% of energy 1.08, 95% CI 1.02-1.15), while increasing polyunsaturated or total ω-3 fat intake in place of carbohydrates was associated with a lower risk of T2D (HR per 5% of energy 0.92, 95% CI 0.85-1.00; and HR per increment of 1g/d 0.95, 95% CI, 0.92-0.99, respectively). We did not observe evidence of significant interactions between dietary fat subtypes and GRS on the risk of T2D.

Interpretation: In the present long-term prospective study including 103,2participants, our results support that genetic risk profile and monounsaturated fat intake were each associated with a higher risk of T2D, whereas polyunsaturated fat intake was associated with a lower risk of T2D. Findings from this study suggest that dietary fat recommendations do not need to be tailored to individual T2D genetic risk profile for the primary prevention of T2D, and that dietary fat subtypes associate with the risk of T2D across the spectrum of T2D genetic risk.

Disclosure

J. Merino: None. H.S. Dashti: None. M. Guasch: None. C. Ellervik: None. C. Smith: None. T.O. Kilpelainen: None. D. Chasman: None. J.C. Florez: Consultant; Self; Intarcia Therapeutics, Inc.. Consultant; Spouse/Partner; Santen.
Wnt Pathway Inhibitor DKK1—A Potential Novel Biomarker for Ectopic Skeletal Muscle Adiposity
Authors: ALI HIRA , ZMUDA JOSEPH M. , CVEJKUS RYAN , KERSHAW ERIN E. , KUIPERS ALLISON L. , BUNKER CLAREANN H. , MILJKOVIC IVA
Published: June 2018

Background: Emerging evidence indicates that ectopic skeletal muscle adiposity (i.e., myosteatosis) is greater in African than in European ancestry individuals, and may be a novel risk factor contributing to increased diabetes risk. However, the biological mechanisms underlying myosteatosis are largely unknown. In vitro studies suggest that a Wnt pathway inhibitor, Dickkopf related protein 1 (DKK1), could be a novel biomarker with a role in adiposity regulation.

Methods: We measured fasting serum DKK1 in a pilot study of 159 elderly Afro-Caribbean men (mean age 63.4 years, mean BMI 27.8 kg/m2), a random subset of a population-based study comprising ∼3000 African Ancestry men with fasting morning blood collections for biomarker analyses. DKK1 was measured in duplicate in previously unthawed serum with ELISA.Also assessed were anthropometrics, DXA measured adiposity, and lower leg CT measured myosteatosis (intermuscular adiposity (mm2)) and skeletal muscle density (which reflects the intra-muscular fat content ((mg/cm3) such that greater intra-muscular adiposity reflects lower muscle density).

Results: In Spearman correlation analysis DKK1 was positively associated with BMI (R=0.19, p=0.01), waist circumference (R=0.15, p=0.05), DXA total body fat (R=0.22, p=0.004), and DXA trunk fat (R=0.19, p=0.01), independent of age. In addition, DKK1 was associated positively with intermuscular fat (R=0.19, p=0.01), and inversely with muscle density (R=-0.29, p=<0.001), independent of age and BMI. No significant association was found between DKK1 and fasting serum glucose and insulin levels, and HOMA-IR.

Conclusion: Our findings suggest for the first time that lower serum levels of DKK1 may be associated with greater overall adiposity, and ectopic skeletal muscle adiposity. Further studies are needed to unravel the potential role of DKK1 in the regulation of body fat among African ancestry and other population groups with a high risk of developing obesity and related metabolic disorders.

Disclosure

H. Ali: None. J.M. Zmuda: None. R. Cvejkus: None. E.E. Kershaw: Consultant; Self; Regeneron Pharmaceuticals, Inc.. A.L. Kuipers: None. C.H. Bunker: None. I. Miljkovic: None.
IER3IP1 Regulates Beta-Cell Survival
Authors: SUN JUAN , POLONSKY KENNETH , REN DECHENG
Published: June 2018

Mutations in the gene for Immediate Early Response 3 Interacting Protein 1 (IER3IP1) cause permanent neonatal diabetes mellitus in human. The mechanisms involved have not been determined and the role of IER3IP1 in β-cell survival has not been characterized. In order to determine if there is a molecular link between IER3IP1 deficiency and β-cell survival, we knocked down IER3IP1 gene expression in mouse MIN6 insulinoma cells. IER3IP1 suppression induced apoptotic cell death which was associated with an increase in expression of the BH3-only molecule Bim. Knockdown of Bim reduced apoptotic β-cell death induced by IER3IP1 suppression. Overexpression of the anti-apoptotic molecule Bcl-xL prevents β-cell death induced by IER3IP1 suppression. Moreover, IER3IP1 also regulates activation of the unfolded protein response (UPR). IER3IP1 suppression impairs the IRE1α and PERK arms of UPR. These results suggest that IER3IP1 suppression induces β-cell death by upregulating Bim and decreasing UPR activation. The studies in vivo also show the beta cell specific knockout IER3IP1 mice develop diabetes related to beta cell death and insulin secretion defect.

Disclosure

J. Sun: None. K. Polonsky: None. D. Ren: None.
Targeted Deletion of ATF4 in ß Cells Leads to the Vulnerability and Dedifferentiation during ER Stress
Authors: KITAKAZE KEISUKE , KURAHASHI KIYOE , MIYAKE MASATO , HAMADA YOSHIMASA , OYADOMARI MIHO , OYADOMARI SEIICHI
Published: June 2018

ATF4 is the main transcriptional regulator of the Unfolded Protein Response (UPR) that promote restoration of normal ER function, although the contribution of ATF4 on β-cells remains poorly understood. Therefore, we developed mature β-cell specific ATF4 knockout mice (βATF4KO), were crossed with mice expressing Cre recombinase under the control of insulin promoter. ATF4-deleted β-cells were comparable to WT under normal condition but were susceptible to ER stress. To examine the role of ATF4 in vivo, we bred the Akita insulin folding mutation onto βATF4KO background. Akita/βATF4KO were dramatically compromised as shown by overt hyperglycemia in their early age and remarkable short life span due to ketoacidosis. Interestingly, we found that increased number of cells staining for glucagon, somatostatin or pancreatic polypeptide in the islets of Akita/βATF4KO, suggesting the increased β-cell dedifferentiation. We further employed the gain-of-function study using β-cell specific transgenic mice expressing Fv2E-PERK, which can activate PERK/ATF4 signaling upon addition of the artificial ligand. The induced activation of PERK/ATF4 signaling resulted in reduced insulin secretion without any sign of β-cell apoptosis. Microarray analysis revealed that several key β-cell specific transcription genes such as Pdx1 and MafA were inhibited by the activation of PERK/ATF4 signaling. We demonstrated that ATF4-deleted β-cells are more vulnerable to dedifferentiation during ER stress and excess ATF4 has inhibitory effect in β-cell function. Overall, our study has uncovered that the importance of ATF4 in governing β-cell adaptation and resilience during ER stress.

Disclosure

K. Kitakaze: None. K. Kurahashi: None. M. Miyake: None. Y. Hamada: None. M. Oyadomari: None. S. Oyadomari: None.
Targeting Cellular Calcium Homeostasis to Prevent Beta-Cell Death in Type 1 Diabetes
Authors: CLARK AMY L. , ABREU DAMIEN , BROWN CRIS M. , URANO FUMIHIKO
Published: June 2018

Cytokines are important mediators of beta cell death leading to type 1 diabetes (T1DM). The mechanism of cytokine-mediated beta cell death has been previously shown to involve alterations in beta cell calcium homeostasis and increases in endoplasmic reticulum (ER) stress. Our group has previously established that pharmacologic modulation of cellular calcium levels with dantrolene and sitagliptin can significantly decrease cytokine and ER stress-mediated activation of the calpain and thioredoxin interacting protein pro-apoptotic pathways and suppress cell death in vitro using INS-1E rat insulinoma cells.

The current study was designed to determine if dantrolene and sitagliptin can prevent beta cell death and diabetes in the low dose streptozotocin (STZ) model of T1DM. Male C57Bl/6 mice were treated with 5 doses of STZ (50 mg/kg) daily by 10 days of intraperitoneal (IP) injections of placebo, dantrolene, or sitagliptin. Blood sugar was monitored via tail vein blood using a glucometer. Blood glucose and insulin levels in response to 2g/kg IP dextrose injection were obtained on day 10. Unmethylated insulin DNA was also measured from plasma drawn on day 0, 8 and 15 to monitor beta cell death.

Sitagliptin treated mice were resistant to low dose STZ induced diabetes as indicated by lower random blood glucose levels and normalization of blood glucose and insulin levels in response to IP dextrose infusion compared to placebo treated animals. In addition, sitagliptin treated animals had significantly lower levels of circulating unmethylated insulin DNA suggesting lower incidence of beta cell death as compared to placebo treated animals.

In summary, in the low dose STZ model of diabetes, sitagliptin lowered random blood sugar, normalized blood glucose and insulin response to IP glucose tolerance testing and significantly decreased STZ mediated beta cell death. This study supports further study on the effect of sitagliptin and DPP-4 inhibition on beta cell function and survival in T1DM.

Disclosure

A.L. Clark: None. D. Abreu: None. C.M. Brown: None. F. Urano: Research Support; Self; Eli Lilly and Company. Stock/Shareholder; Self; CytRx.
Role of NumbL:Mig6 Interactions in Beta-Cell Death during Glucolipotoxicity
Authors: BASAVARAJAPPA HALESHA D. , IRIMIA-DOMINGUEZ JOSE M. , FUEGER PATRICK T.
Published: June 2018

As the development of type 2 diabetes (T2D) requires a loss of functional beta cell mass, preserving functional beta cell mass is essential for preventing T2D. We have focused our efforts on EGF receptor (EGFR) signaling because it controls cellular proliferation, survival, and repair mechanisms—all features relevant to maintaining beta cell mass—and is dampened during glucolipotoxic conditions (GLT) similar to those in T2D. We have established that Mig6, an endogenous feedback inhibitor of EGFR, is induced in both rodent islets cultured in GLT and human T2D islets. In addition, GLT impairs EGFR signaling in rodent and human islets. Further, Mig6 overexpression promotes apoptosis in both rodent beta cell lines and islets. Because Mig6 is an adapter protein and likely functions as a molecular scaffold, here we aimed to define the molecular mechanisms of Mig6 in GLT by identifying the interacting partners of Mig6. We performed mass spectrometry analysis using low-level, flag-tagged Mig6 expression with immunoprecipitation from Ins1-derived 832/13 cells cultured in normal conditions and GLT. We identified that Mig6 interacted with Numb and NumbL, regulators of Notch signaling, in a context dependent manner. Specifically, Mig6 interacted with Numb and NumbL in normal glucose conditions whereas these interactions were disrupted by GLT. However, roles for Numb and NumbL in beta cell function or death are largely unknown. We observed that Numb and NumbL are expressed in both rodent and human islets, and whereas their expression is not altered in GLT, their localization and aforementioned interactions were. To examine their impact in beta cell death, we used siRNA-mediated suppression of Numb and NumbL in 832/13 cells during control and GLT conditions. Whereas Numb suppression had no effect, NumbL suppression reduced apoptosis in GLT. These results suggest that NumbL plays an important role in beta cell survival. Hence, the differential interactions of Mig6 and NumbL may determine beta cell fate during stress.

Disclosure

H.D. Basavarajappa: None. J.M. Irimia-Dominguez: None. P.T. Fueger: None.
Use of Live Murine Pancreatic Slices for the Study of ß-Cell Regeneration
Authors: QADIR FAHD , WEITZ JONATHAN , ALVAREZ-CUBELA SILVIA , KLEIN DAGMAR , LANZONI GIACOMO , GARCIA-SANTANA CARLOS A. , MONTALVO ABELARDO , RICORDI CAMILLO , INVERARDI LUCA , CAICEDO ALEJANDRO , PASTORI RICARDO , DOMÍNGUEZ-BENDALA JUAN
Published: June 2018

Introduction: Bone morphogenetic protein 7 (BMP-7), a protein with BMP-activating and TGF-β inactivating properties, induces the proliferation of a unique population of progenitor-like cells characterized by the expression of PDX1 and the BMP receptor activin-like kinase 3 (ALK3/BMPR1A). These cells can be sorted using ALK3 and the purinergic receptor P2Y1 (P2RY1) as a novel surrogate surface marker for PDX1. Sorted P2RY1+/ALK3bright+ cells form BMP-7 expandable colonies. Upon BMP-7 withdrawal, they differentiate into cells of all three lineages (endocrine, acinar and ductal) of the adult pancreas. PDX1+/ALK3+ cells absent from islets are represented in the major pancreatic ducts and pancreatic duct glands. Preliminary evidence indicates that these cells are also present in the murine pancreas. We hypothesized that islet β-cell regeneration could be studied in real time using live murine pancreatic slices transduced with a loxP-dsRed-loxP-EGFP reporter + insulin promoter-Cre adenovirus. If new β-cells arise from preexisting insulin-expressing-cells they would remain green. However, if neogenesis occurs from non-β-cells, there is a window of time during which the degrading red and the new green protein co-localize in the cell. Allowing us to discriminate between neogenesis and β-cell proliferation.

Methods: Low melting point (LMP) agarose-embedded, vibratome-sectioned live pancreatic slices were generated from CD-1 mice. Transduction with a loxP-dsRed-loxP-EGFP reporter + insulin promoter-Cre adenovirus was done in slices cultured atop optically clear membranes. Slices were treated for 4 days with an ALK3 agonist (THR-123) or saline at the same time of the co-transduction. Bright field and fluorescence images of them were taken every 24h for 7 days.

Conclusions: This is the first report on the use of live pancreatic slices as a novel tool for the study of pancreatic regeneration. Our preliminary results suggest that there are quantifiable β-cell neogenic events in response to BMP-7-like agents.

Disclosure

F. Qadir: None. J. Weitz: None. S. Alvarez-Cubela: None. D. Klein: None. G. Lanzoni: None. C.A. Garcia-Santana: Employee; Self; Ophysio Incorporated. A. Montalvo: None. C. Ricordi: None. L. Inverardi: None. A. Caicedo: None. R. Pastori: None. J. Domínguez-Bendala: Stock/Shareholder; Self; Ophysio.
E-Cadherin Downregulation by Cathepsin L Plays a Role in the Proliferation of Insulin-Producing Cells following Inhibition of Serpin B13
Authors: LO CHI-WEN , SHEU TZONG-JEN , CZYZYK JAN
Published: June 2018

Experimental targeting of the exocrine pancreatic ductal tree stimulates beta-cell rejuvenation thereby underscoring the importance of functional crosstalk between the exocrine and endocrine pancreas. We recently found that antibody directed against serpin B13, a protease inhibitor expressed in the ductal epithelium of the exocrine pancreas, increased the catalytic activity of it’s protease target, cathepsin L (CatL) and subsequently upregulated Reg-mediated cellular proliferation in the islets. However, the exact molecular events that link catL activity to upregulated Reg and islet cell proliferation remain unclear. As increased E-cadherin levels during islet formation mediate a decline in beta-cell proliferation, and Reg upregulation has been linked to activation of beta-catenin, we hypothesized that the functional crosstalk between pancreatic ductal epithelium and the islets may involve CatL-mediated activation of the E-cadherin/β-catenin pathway. In support of this we found that E-cadherin was downregulated with a concomitant increase in Reg1 in mice treated with serpin B13 monoclonal antibody (mAb). Likewise, exposure to recombinant CatL or ductal epithelium extract treated with serpin B13 mAb caused decreased E-cadherin expression in the pancreatic islets during in vitro culture. On the other hand, this expression was recovered with CatL inhibitor. Moreover, treatment of MIN-6 insulinoma cells with ductal epithelium extract and serpin B13 mAb stimulated intranuclear accumulation of β-catenin, while Reg gene expression in the islets was markedly reduced in the presence of the β-catenin chemical inhibitor, FH535. These data establish E-cadherin as a potential target of CatL in the regulation of islet cell proliferation by an immunological response to serpin B13, and implicate the E-cadherin/β-catenin pathway as an islet sensing mechanism of the signals that are generated in the pancreatic exocrine milieu.

Disclosure

C. Lo: None. T. Sheu: None. J. Czyzyk: None.
GLP-1 Controls Pancreatic Endocrine Development
Authors: CRAS-MÉNEUR CORENTIN M.H. , SEELEY RANDY J.
Published: June 2018

Glucagon-like peptide 1 (GLP-1) has been successfully leveraged as a treatment of type 2 diabetes mellitus. Its beneficial effects on β-cell growth and function are documented in the adult rodents but its role during pancreatic organogenesis remains unexplored. GLP-1 has numerous targets in vivo and has been shown to regulate key signaling pathways in β-cells. In the mature islets, GLP-1 modulates the MAPK and Akt signaling pathways, controls glucose sensing and responsiveness and increases Pdx-1 levels. All these processes are also known to be instrumental in the regulation of the proper growth, differentiation and maturation of the early endocrine progenitors. A few studies indirectly implied that GLP-1 could also have some impact on pancreatic organogenesis, promoting endocrine development in in vitro models. However, adult GLP-1 receptor (GLP-1R) knock out mice only display mild metabolic impairments mostly triggered by exposure to high-fat diet, but the phenotype of these animals had not been studied during early development.

Using our total body knockout of the GLP-1 receptor, we found that total size of the pancreas of the newborn knocked-out animals appears to be reduced at birth and the β-cell mass is diminished by 89%. These results support that GLP-1R signaling is crucial for normal pancreatic development but that catch up growth can takes place between birth and adulthood to compensate for reduced GLP-1R signaling. These data illustrate that in addition to its insulinotropic effects in mature islets, GLP-1 signaling plays a central role during pancreatic development and the acquisition of a normal β-cell mass.

Disclosure

C.M.H. Cras-Méneur: None. R.J. Seeley: Research Support; Self; Novo Nordisk Inc., Novo Nordisk Inc., Ethicon US, LLC.. Consultant; Self; Janssen Pharmaceuticals, Inc.. Research Support; Self; Zafgen, MedImmune. Consultant; Self; Kallyope, Scohia.
Glis3 Regulates Pancreatic ß-Cell Development in Part through Coordination with Other Islet Transcription Factors
Authors: SCOVILLE DAVID
Published: June 2018

Pancreatic β-cells play a central role in regulating glucose homeostasis. One prominent regulator of both β-cell fate and function is Glis3, a transcription factor with both co-activating and co-repressive roles. Both humans and mice lacking a functional copy of Glis3 are diabetic from birth, with a near complete absence of β-cells. Interestingly, Glis3 is expressed relatively early during pancreas development prior to the differentiation of ductal and endocrine cells from a common lineage, and is maintained in both β-cells as well as ductal cells. Furthermore, postnatal deletion of Glis3 also results in hyperglycemia, suggesting Glis3 as a critical regulator of postnatal β-cell function as well. Yet, while much is known about the phenotype of Glis3 deletions, little is known about its molecular function.

My research focuses on how Glis3 regulates β-cell development and function. To determine which genes are regulated by Glis3, a mouse model with a pancreas-specific deletion of Glis3 was utilized. RNA-seq analysis of isolated islets revealed a large number of both upregulated and downregulated genes, and ChIP-seq analysis of Glis3 in islets revealed many of these genes are direct targets of Glis3 regulation. Interestingly, Glis3 binding largely overlaps with binding of the transcription factor Nkx6.1, and partially overlaps with transcription factors Pdx1 and Nkx2.2, but shows less overlap with other islet-enriched transcription factors. This indicates that Glis3 could function in distinct regulatory complexes, and that better understanding of Glis3 gene regulation within the pancreatic β-cell will lead to a better understanding of how β-cell fate is controlled and maintained, as well as which genes are essential for postnatal function.

Disclosure

D. Scoville: None.
Cdk2-Dependent Activation of ß-Cell Metabolism Is Eclipsed by Its Inhibitory Effect on Plasma Membrane Excitability and Insulin Secretion
Authors: SDAO SOPHIA , POUDEL CHETAN , MORTENSEN KARA M. , MERRINS MATTHEW J.
Published: June 2018

The loss of insulin secretion is a hallmark of diabetes, and it is becoming clear that cell cycle regulators control insulin secretion as well as proliferation. It was recently reported that embryonic deletion of cyclin-dependent kinase 2 (Cdk2) in β-cells (Pdx1-Cre:Cdk2-/-) results in impaired glucose tolerance due to defective insulin secretion (Kim et al., J Biol Chem, 2017), revealing a developmental or perinatal requirement of Cdk2. Here, we show that tamoxifen-inducible deletion of Cdk2 in adult β-cells (MIP-CreER:Cdk2-/-) enhances insulin secretion. Despite these differences, both models of Cdk2 depletion exhibited a strong increase in the glucose dependence of β-cell calcium oscillations. This effect is likely due to the loss of ATP-sensitive K+ (KATP) channels, since kir6.2 transcript was reduced in Cdk2-null β-cells, and pharmacological inhibition of Cdk2 correspondingly reduced KATP channel conductance. These results reveal that Cdk2 inhibits insulin secretion by limiting plasma membrane excitability via KATP. In adult β-cells lacking Cdk2, we also observed a 45% reduction in depolarization-induced exocytosis, indicating that Cdk2 is required for the metabolic amplification of insulin secretion. Within the intermediary metabolic pathways, Cdk2-null β-cells exhibited reductions in glucose-dependent lactate accumulation, mitochondrial membrane potential, and mitochondrial NADH utilization. Together, these studies reveal that Cdk2 transcriptionally limits β-cell membrane excitability and the insulin secretory response, and is required to maintain proper metabolic function.

Disclosure

S. Sdao: None. C. Poudel: None. K.M. Mortensen: None. M.J. Merrins: None.
FSTL3 Inhibition Restores Glucose Responsive Insulin Secretion in Nonfunctional Human Islets
Authors: SCHNEYER ALAN , BROWN MELISSA
Published: June 2018

Type 2 diabetes results from inadequate insulin production to control blood glucose. To address this need for replacing lost insulin-producing beta cells, most therapeutic development strategies are focused on enhancing functional beta cell number. Fairbanks Pharmaceuticals is pursuing therapies that enhance beta cell function and regeneration and is based on prior research in which inactivation of follistatin-like-3 (FSTL3) resulted in enlarged islets, increased beta cell number, and enhanced insulin secretion. At least some of these new beta cells arose from alpha to beta cell transdifferentiation in FSTL3 knockout mice. Fairbanks has produced a monoclonal antibody (FP-101) that can completely block FSTL3 from binding activin A and B, GDF11, and myostatin that could also disrupt pre-formed and otherwise irreversible complexes. FP-101 was also able to restore glucose-responsive insulin secretion in otherwise non-functional islets taken from HFD-fed mice. To determine if FSTL3 neutralization could be effective in humans, human islets from normal donors were first exposed to elevated glucose (25 nM) for 48-72 hours, a commonly utilized model for glucotoxicity. As expected, this caused loss of glucose response to subsequent GSIS. However, when treated for 24 hours with FP-101, GSIS was restored, indicating that neutralization of FSTL3 might be an effective strategy for restoring function to human non-functional islets. This observation agrees with previous research in which activin treatment directly restored GSIS in human islets from diabetic donors. Taken together with the potential for induction of alpha to beta cell transdifferentiation, these results suggest that neutralization of FSTL3 may have a dual benefit with acute restoration of GSIS in non-functional islets and chronic treatment leading to restoration of beta cell number through enhancement of alpha to beta cell transdifferentiation. In vivo studies will be initiated shortly.

Disclosure

A. Schneyer: Stock/Shareholder; Self; Fairbanks Pharmaceuticals, Inc.. Stock/Shareholder; Spouse/Partner; Fairbanks Pharmaceuticals, Inc.. Employee; Self; Fairbanks Pharmaceuticals, Inc.. M. Brown: None.
Substance Use in Adults with Type 1 Diabetes in the T1D Exchange
Authors: PETRY NANCY , FOSTER NICOLE C. , CENGIZ EDA , TAMBORLANE WILLIAM V. , WAGNER JULIE , POLSKY SARIT
Published: June 2018

Substance use can have adverse effects on many health conditions, yet little is known about the prevalence of use in individuals with type 1 diabetes (T1D). We evaluated the frequency of use and problem use of psychoactive substances in adults with T1D. Standardized instruments for assessing tobacco, alcohol, and psychoactive substance use were emailed to 4311 adult participants at 69 T1D Exchange Clinic Registry centers. A total of 936 respondents (61% female, 90% non-hispanic white, age 38±16 years) completed the survey. In the sample, tobacco use was reported by 166 (18%) participants in the past year and 51 (5%) daily (Figure 1). Past year alcohol use was reported by 742 (79%) participants, past month use by 592 (63%) and daily or near daily use by 87 (9%). Use of any other psychoactive substance in the past year was reported by 228 respondents (24%), with marijuana being the most commonly used substance (Figure 2). Past year problem use of these substances was noted in 31 (3%) respondents. Adults with T1D in the U.S. use substances at rates that meet or exceed that of the general population; problematic use occurs at rates similar to the general population. These data delineate the need to inquire about regular, intermittent and problematic use of nicotine, alcohol, and other substances in individuals with T1D. A better understanding of the impact of moderate and occasional use of substances on T1D management and clinical outcomes is also needed.

Disclosure

N. Petry: None. N.C. Foster: None. E. Cengiz: Advisory Panel; Self; Novo Nordisk Inc.. Speaker's Bureau; Self; Novo Nordisk Inc.. Advisory Panel; Self; MannKind Corporation, ADOCIA, Arecor. W.V. Tamborlane: Consultant; Self; AstraZeneca, Boehringer Ingelheim GmbH, Eli Lilly and Company, Medtronic MiniMed, Inc., Novo Nordisk Inc., Sanofi, Takeda Pharmaceuticals U.S.A., Inc.. J. Wagner: None. S. Polsky: Research Support; Self; Dexcom, Inc.. Other Relationship; Self; T1D Exchange. Research Support; Self; National Institute of Diabetes and Digestive and Kidney Diseases.
Glucagon Expression Is Regulated by Exendin-4 via CaMKK/AMPK/FoxO1 Pathway in Alpha TC1.6 Cells
Authors: LYU JINGYA , IMACHI HITOMI , FUKUNAGA KENSAKU , SATO SEISUKE , DONG TAO , IBATA TOMOHIRO , KOBAYASHI TOSHIHIRO , MURAO KOJI
Published: June 2018

Exendin-4, as an analogue of glucagon-like peptide-1, is currently used for the treatment of type 2 diabetes by enhancement of glucose-induced insulin secretion. Preliminary studies showed that secretion of glucagon is inhibited by exendin-4 in vivo and in vitro. However, the detailed mechanism in exendin-4 regulated glucagon expression is not clear.In this study, we firstly proved that exendin-4 decreased the mRNA and protein expression of glucagon in a dose-dependent manner. Following, the promoter activity of glucagon was significantly decreased by exendin-4 and inhibition of Ca2+/calmodulin-dependent protein kinase kinase (CaMKK) signaling pathway by its specific inhibitor, STO-609 rescued this effect. Since glucagon secretion is regulated by AMP-activiated protein kinase (AMPK) and AMPK is able to be activated by CaMKK, we checked the role of AMPK in this process and found exendin-4 activated AMPK at Thr172 site. Additionally, inhibition of AMPK activity by its specific inhibitor or overexpression of domain-negative AMPK canceled the effect of exendin-4 on glucagon promoter activity. As activation of AMPK enhances FoxO transcriptional activity, to further study the transcription of glucagon, we used ChIP assay to demonstrate that transcription factor FoxO1 could bind to glucagon promoter region and exendin-4 decreased glucagon expression via down-regulation of FoxO1. Mutation of FoxO1-binding site in glucagon promoter or silence of FoxO1 canceled the effect of exendin-4 on glucagon expression.

In summary, our results proved that exendin-4 decreases the expression and transcription of glucagon via CaMKK/AMPK/FoxO1 pathway in pancreatic alpha cells, contributing to new knowledge about the regulation of glucagon gene by exendin-4.

Disclosure

J. Lyu: None. H. Imachi: None. K. Fukunaga: None. S. Sato: None. T. Dong: None. T. Ibata: None. T. Kobayashi: None. K. Murao: Research Support; Self; Novartis Pharma K.K..
Profound Hypoglycemia and Corresponding Insulin Levels with Massive Glargine Overdose
Authors: SAEED ZEB I. , ZIA ZAHRA , MARIASH CARY N.
Published: June 2018

Introduction: Despite widespread use of insulin in diabetes, reports of intentional overdose are rare. Insulin kinetics are different in acute poisoning from that in therapeutic doses in part due to delayed absorption from the injection site and reduced clearance. We present a case of insulin glargine overdose where the peak serum insulin levels were observed more than 24 hours after presentation with persistent hypoglycemia for over 100 hours from initial presentation.

Case: An unidentified middle-aged female was found comatose in an abandoned house by emergency medical services. She was hemodynamically stable except hypothermic at 34°C. Initial point of care glucose was undetectable and 25mg/dL after 1mg IM glucagon administration. She was given 75ml of Dextrose 50% intravenously and intubated in the field for airway protection. Serum glucose on hospital arrival was <20mg/dL, necessitating the initiation of dextrose 5%, then 10% and eventually continuous infusion of dextrose 40% (D40) via central venous catheter for persistent hypoglycemia. Evaluation revealed serum insulin level of 362 mCU/mL (4-30) with C-peptide 0.3ng/ml (1.4-4.4) when serum glucose was 51, suspicious for exogenous insulin use. Renal and liver function tests were normal, AM cortisol was appropriate and oral hypoglycemic agent panel was negative. Peak total serum insulin levels were 1395mCU/ml with free insulin levels 1022mCU/ml 36 hours after initial presentation. With exaggerated insulin levels, testing for insulin antibodies was done which was negative. D40 drip was weaned off slowly over the next few days and discontinued at 102 hours after arrival. Patient was extubated soon afterwards and admitted to intentional overdosing with over 5000units (5-6 vials) of Lantus. She was admitted to a psychiatry facility for management of depression and suicidality.

Conclusion: Profound and persistent hypoglycemia can be observed for days due to altered pharmacokinetics after massive doses of long-acting insulins.

Disclosure

Z.I. Saeed: None. Z. Zia: None. C.N. Mariash: None.
A Coronary Artery Calcium Score (CACS) Higher than 100 Agatston Units Is Associated with a Profile of Very High Cardiovascular Risk in the Diabetic Population
Authors: VALENSI PAUL , BANU ISABELA , FYSEKIDIS MARINOS , COSSON EMMANUEL
Published: June 2018

Rationale: CACS can improve the cardio-vascular risk (CVR) estimate and select the patients eligible for further cardio-vascular investigations. We aimed to examine the relationship between a CACS ≥100 Agatston units (AU) and integrators of CVR in the diabetic population.

Patients and Methods: CACS was measured in type 2 diabetic patients without symptoms, history of coronary disease or ECG ischemic changes presenting ≥2 additional CVR factors. We included 338 patients with very high CVR (age 61±9 years, diabetes duration 15 ± 8 years, insulin treated 47%, hypertension 86%, dyslipidemia 88%, smokers 22%, lower limb artery disease 9.5% and/or carotid atheroma 5.5%, chronic kidney disease 48%). Silent myocardial ischemia (SMI) was assessed using stress myocardial scintigraphy. Artery stiffness indexes (pulse wave velocity (PWV) and augmentation index (AIx@75)) and Buckberg index of myocardial perfusion (SEVR) were measured with applanation tonometry (Sphygmocor®).

Results: Compared with the patients with CACS <100 AU, the 139 patients (41%) with CACS ≥100 were less treated by metformin (73 vs. 88%, p<0.001) or GLP-1-receptor agonists (18 vs. 28%, p<0.05), but more treated by anti-hypertensive drugs, statins (79 vs. 70%, p=0.058) or fibrates (8.5 vs. 3.7%, p=0.072), with lower LDL-cholesterol levels (2.1±0.9 vs. 2.4±0.9 mmol/l, p<0.01). SMI prevalence (21.6 vs. 11.5%, OR 2.1[1.2-3.8], p<0.05), NT-proBNP (201±375 vs. 73±136 mg/L, p<0.01) and PWV (11.0±2.7 vs. 9.4±3.6 m/sec, p<0.01) were higher. However, the rate of elevated PWV (compared to normative data considering age and blood pressure), central pulse pressure and SEVR were very close in the two groups while AIx@75 was lower in the patients with CACS≥100 (22.1±9.3 vs. 26.4±10.0, p<0.01).

Conclusion: The CACS ≥100 is a marker of very high CVR associated with a higher SMI prevalence and NT-proBNP levels. Artery stiffness and SEVR were not altered.

Disclosure

P. Valensi: None. I. Banu: None. M. Fysekidis: None. E. Cosson: Consultant; Self; Lilly France. Board Member; Self; Novartis France. Consultant; Self; Roche Diagnostics France. Board Member; Self; Ascensia Diabetes Care. Consultant; Self; Bezins France. Board Member; Self; MSD France.
Next Generation of Spontaneous Diabetic Model of Fatzo Mice with Intact Leptin Signaling Develop Nephropathy after High-Fat and High-Sucrose Induction
Authors: SUN GAO , ZHANG GUODONG , WANG YIXIN
Published: June 2018

Introduction: Progressive albuminuria and bi-phasic changes in glomular filtration rate (GFR) are hall marks of functional deterioration of the kidney in diabetic nephropathy. However, not all diabetic rodent models can capture the development of such changes in the kidney. FATZO, a new generation of diabetic mouse model with intact leptin signaling pathway shows all the metabolic dysfunctions that mimic human patients such as obesity, dyslipidemia, diabetes and NASH etc. Here, we sought to further characterize the model for the evaluation of its development in diabetic nephropathy.

Method: Male FATZO mice on 50diet or on western diet with addition of 10% fructose (WD+fru) from age of 11 weeks were subject to monthly urine collection for albumin and creatinine measurement for 28 weeks, after which animals were treated with Lisinopril (15 mg/kg) and pioglitazone (20 mg/kg) daily for 6 weeks (n=8 for each group). Animals were also monitored for GFR using a subcutaneous monitoring device (Medibeacon, US).

Results: Animals on WD+fru were heavier than those on 50diet. Notably, WD+fru markedly elevated urine albumin levels compared to 50diet in FATZO mice which resulted in higher albumin/creatinine ratio and urine albumin excretion rate (UAE) from 2 weeks on diet, while lisinopril treatment for 7 weeks could reduce the levels respectively. In addition, GFR was slightly elevated after 6 weeks of diet induction, which could be normalized by either pioglitazone or lisinopril treatment.

Conclusion: We have created a mouse model with progressive albuminuria and elevated GFR that can be reversed by angiotensin-converting-enzyme (ACE) inhibitor. The model with associated dysmetabolic phenotypes including obesity and dyslipidemia can be used as an ideal tool for studying antidiabetic nephropathy treatment.

Disclosure

G. Sun: None. G. Zhang: None. Y. Wang: None.
Exendin-4 Ameliorates High-Glucose-Induced Fibrosis by Inhibiting the Secretion of miR-192 from Injured Renal Tubular Epithelial Cells
Authors: JIA YIJIE , XUE YAO-MING
Published: June 2018

Extracellular vesicles (EVs), which contain microRNA (miRNA), constitute a novel means of cell communication that may contribute to the inevitable expansion of renal fibrosis during diabetic kidney disease (DKD). Exendin-4 is effective for treating DKD through its action on GLP-1R. However, the effect of exendin-4 on EVs miRNA expression and renal cell communication during the development of DKD remains unknown. In this study, we found that EVs derived from exendin-4 and high-glucose (Ex-HG)-pretreated HK-2 cells, which was taken up by normal HK-2 cells, resulted in decreased levels of FN and Col-I compared with those treated with HG pretreated HK-2 cells derived EVs. Furthermore, we found this may contribute to a decrease in miR-192 in both HK-2 cells and EVs after exendin-4 treatment in a p53-dependent manner. Besides, we found that the amelioration of renal fibrosis by exendin-4 occurs through a miR-192-GLP-1R pathway, indicating a new pathway by which exendin-4 regulates GLP-1R. The results of this study suggest that exendin-4 can inhibit the transfer of EVs miR-192 from HG-induced renal tubular epithelial cells to normal cells, thus inhibiting GLP-1R down-regulation and protecting renal cells. This study reports a new mechanism by which exendin-4 exerts a protective effect against DKD.

Disclosure

Y. Jia: None. Y. Xue: None.