bluuebunny/crossref_metadata_2025_split huggingface dataset:
Metabolic Insufficiency Caused By Cellular Stresses Is Implicated in Beta-Cell Dedifferentiation in the Mouse Model of Wolfram Syndrome
Authors: AMO-SHIINOKI KIKUKO , TANABE KATSUYA , HATANAKA MASAYUKI , TANIZAWA YUKIO 
Published: June 2018

Pancreatic beta cell loss is associated with augmented ER and oxidative stresses in Wolfram syndrome caused by mutations in the WFS1. In the Wfs1-/- mice, beta cells become dedifferentiated and revert to endocrine progenitor-like cells, and a subset of them takes alpha cell fate. Such beta cell plasticity appears after nursing, independently of hyperglycemia, and becomes more apparent along with diabetes progression accompanied with no significant increase in apoptosis. We have found that genetic inhibition of Txnip, which is a stress response molecule involving in various cellular processes, preserved beta cell mass and maintained glucose homeostasis in the Wfs1-/- mice. This suggests its roles in the regulation of beta cell plasticity in the setting of Wfs1 deficiency. One clue to the mechanisms underlying beta cell dedifferentiation was the paradoxical reduction of acetyl-CoA, citrate and ATP content in the Wfs1-/- islets in spite of the enhanced glycolysis with an increase in pyruvate. This metabolic dissociation was correlated with an increase in phosphorylated pyruvate dehydrogenase (PDH). Importantly, Txnip directly interacts with both PDH kinase and PDH, indicating its involvement in the regulation of PDH activity. Indeed, islets of Wfs1-/- mice lacking Txnip demonstrated a robust reduction of phosphorylated PDH and a restoration of capabilities of ATP production in response to glucose. Thus, these finding illustrate impaired energy metabolism in beta cells under the chronic stress conditions and suggest that beta cells may possibly become dedifferentiated to adapt to metabolic insufficiency caused by unresolvable stresses. This provides new insights into molecular mechanisms underlying beta cell loss in diabetes related to cellular stresses, such as Wolfram syndrome.

Disclosure
K. Amo-Shiinoki: None. K. Tanabe: None. M. Hatanaka: None. Y. Tanizawa: None.
MODY Signal Pathway in the Endoplasmic Reticulum Stress and the Role of Nkx6.1 in the Glucolipotoxicity of INS-1-3 Cells
Authors: DONG YANAN , LI YUKUN , XIAO JIANZHONG 
Published: June 2018

Metabolic disturbances induce endoplasmic reticulum stress (ERS) in pancreatic beta cells. This study aims to investigate whether a common pathway exists in the ERS induced by various chemicals, including high levels of glucose and palmitate. ERS in INS-1-3 cells was induced by exposure cells to thapsigargin (TG), tunicamycin (TM) or palmitic acid (PA) +high glucose (HG). Digital gene expression (DGE) profiling technique was used to detect differentially expressed genes. The profile of gene expression was detected by gene oncology (GO) function and pathway enrichment analysis. On the basis of DGE results, Nkx6.1 over-expression was established in INS-1-3 cell lines by lentivirus infection to explore the importance of the findings. Real time reverse transcription polymerase chain reaction (RT-PCR) was used to verify the expression changes of key genes. Cell viability was measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. And the apoptosis was detected by flow cytometry. Glucose stimulates insulin secretion (GSIS) was used to measure the INS-1-3 cell function. DGE demonstrated that there were 135, 57 and 74 differentially expressed genes in TM, TG and HG+PA groups, respectively. Genes that were differentially expressed were enriched to endoplasmic reticulum stress, antigen processing and presentation, and protein export pathways, and surprisingly, the maturity onset diabetes of the young (MODY) pathway. Nkx6.1 is a common abnormal-expressed gene in MODY signaling pathway among TM, TG and HG+PA groups. Apoptosis was ameliorated by 45.4% and proliferation was increased by 40.9% in the Nkx6.1 over-expression cells exposed to high level of glucose and palmitate. At the same time, both basal and glucose stimulated insulin secretion increased by 2.09 and 1.82 folds, respectively. ERS changed the expression of MODY pathway genes. Over-expression of Nkx6.1 protected the INS-1-3 cells from glucolipotoxicity.

Disclosure
Y. Dong: None. Y. Li: None. J. Xiao: None.
Deletion of SLC4A4 in Pancreatic ß Cells Protects from High-Fat Diet–Induced Glucose Intolerance and ß-Cell Dysfunction
Authors: BROWN MATTHEW , HOLMES HEATHER L. , RAKSHIT KUNTOL , ROMERO MICHAEL F. , MATVEYENKO ALEKSEY 
Published: June 2018

Islet failure in type 2 diabetes (T2DM) is characterised by loss of glucose-stimulated insulin secretion (GSIS) attributed in part to β-cell dedifferentiation (e.g., mis-expression of alpha cell-specific genes). Recent single cell transcriptomics studies identified SLC4A4 (a gene encoding an electrogenic Na+-nHCO3- cotransporter, NBCe1) as one of the mis-expressed genes in β-cells of patients with T2DM. Thus, we set out to test the hypothesis that mis-expression of SLC4A4 in β-cells contributes to loss of GSIS and impaired glucose homeostasis in T2DM. To address this hypothesis, we first confirmed induction of robust SLC4A4 protein expression in β-cells of patients with T2DM utilizing immunofluorescence staining of autopsy-derived human pancreas specimens. We next generated β-cell-specific slc4a4 knockout (β-slc4a4-/-) and corresponding control (β-slc4a4+/+) mice through crossing of slc4a4 loxP/loxP and Ins2-Cre mouse models. Adult (3 month old) β-slc4a4-/- mice displayed normal body weight, glycaemia, glucose tolerance and insulin sensitivity under standard chow fed conditions (p&gt;0.vs. β-slc4a4+/+). In contrast, β-slc4a4-/- mice exhibited enhanced glucose tolerance and in vivo glucose-stimulated insulin response under conditions of metabolic stress induced by ad libitum intake of 60% high fat diet (HFD) for 8 weeks (p&lt;0.vs. β-slc4a4+/+ HFD). Interestingly, improved glucose tolerance in β-slc4a4-/-mice was also associated with a tendency for expansion of β-cells mass (82%, p=0.09 vs. β-slc4a4+/+ HFD) and increased β-cells proliferation (26%, p=0.11 vs. β-slc4a4+/+ HFD). These results suggest that β-cell mis-expression of the Na+-nHCO3- cotransporter encoded by SLC4A4 contributes to induction of β-cell failure under conditions of metabolic stress and T2DM.

Disclosure
M. Brown: None. H.L. Holmes: None. K. Rakshit: None. M.F. Romero: None. A. Matveyenko: None.
Investigation on the Protective Effect of Canagliflozin on Pancreatic Beta-Cell Mass Using SPECT/CT Imaging with 111In-Labeled Exendin-4
Authors: HAMAMATSU KEITA , FUJIMOTO HIROYUKI , FUJITA NAOTAKA , MURAKAMI TAKAAKI , SHIOTANI MASAHARU , SAJI HIDEO , INAGAKI NOBUYA 
Published: June 2018

Background: Defects in insulin secretory function can be divided into two elements, the dysfunction of individual beta cells and a decrease of beta cell mass (BCM). Non-invasive methods to quantify BCM have not yet been established, and thus it is not possible to observe longitudinal changes in BCM. We developed [Lys12(111In-BnDTPA-Ahx)]exendin-4 ([111In]Ex4) as a radioactive probe targeting to the glucagon-like peptide-1 receptor with the aim of quantifying BCM by the measurement of probe accumulation in the pancreas. In this study, we investigated the protective effect of sodium-glucose co-transporter-2 (SGLT2) inhibitor on BCM using in-vivo SPECT/CT imaging with this probe.
Aim: We tried to investigate longitudinal change in BCM of diabetic model mice administered with SGLT2 inhibitor using in-vivo SPECT/CT imaging with [111In]Ex4.
Method: Male db/db mice were assigned into two groups: canagliflozin-administered group (group A) and non-administration group (group B). In-vivo SPECT/CT scans were performed before and after the intervention. The pancreatic uptake of [111In]Ex4 was evaluated using ROI with &gt; 40% of the volume of the whole pancreas with the exception of the perirenal space.
Result: The pancreatic uptake was significantly decreased in group B, on the other hand, no significant change was observed in group A. After the intervention, the pancreatic uptake was significantly higher in group A than group B.
Discussion: SPECT/CT imaging analysis indicated that BCM was significantly larger in group A than group B after the intervention. It is considered that this difference was made by the protective effect of canagliflozin on BCM.
Conclusion: It is considered that the protective effect of canagliflozin on BCM could be investigated using in-vivo SPECT/CT imaging with [111In]Ex4.

Disclosure
K. Hamamatsu: None. H. Fujimoto: None. N. Fujita: None. T. Murakami: None. M. Shiotani: Employee; Self; Mitsubishi Tanabe Pharma Corporation. H. Saji: Research Support; Self; Nihon Medi-Physics Co.,Ltd. N. Inagaki: Research Support; Self; Daiichi Sankyo Company, Limited, AstraZeneca, Ono Pharmaceutical Co., Ltd., Sanofi K.K., Mitsubishi Tanabe Pharma Corporation, Takeda Pharmaceutical Company Limited, Japan Tobacco Inc., MSD K.K., Sumitomo Dainippon Pharma Co., Ltd., Nippon Boehringer Ingelheim Co. Ltd., Novartis Pharma K.K., Kissei Pharmaceutical Co., Ltd., Novo Nordisk Pharma Ltd., Eli Lilly and Company, Sanwa Chemical Industrial Co., Ltd., Teijin Pharma Limited.
A Low-Frequency Amerindian Specific Variant in PALD1, a Negative Regulator of Insulin Signaling, Associates with Type 2 Diabetes
Authors: NAIR ANUP K. , SUTHERLAND JEFF R. , ELSON GREGORY R. , PIAGGI PAOLO , KOBES SAYUKO , HANSON ROBERT L. , BOGARDUS CLIFTON , BAIER LESLIE 
Published: June 2018

We recently completed a genome wide association study for type 2 diabetes (T2D) in 7659 American Indians who are predominantly of Pima Indian heritage using a custom designed Axiom array. A novel intronic variant (hg19 chr10:72276664 G&gt;C) in PALD1 was identified that associated with T2D (risk allele frequency (RAF)=0.01, OR=2.19[1.49-3.22] per risk allele [C], P=6.6×10-5 adjusted for age, sex, birthyear, PCs 1-5 and accounting for genetic relatedness). Further analysis in a dataset of Urban American Indians (N=3029) representing several tribal groups living in the Phoenix area, identified a directionally consistent association (OR=2.32, P=0.12), but this variant was rare among the entire Urban American Indian sample (RAF=0.003). A meta-analysis of both datasets resulted in a summary OR of 2.15[1.49-3.13], P=4.9×10-5. In Pima Indians, this variant is highly concordant (r2=0.92) with a novel missense variant (hg19 chr10:72298980 C&gt;T, p.[P568S]) in PALD1. The missense variant also had a significant association with T2D in Pima Indians (RAF=0.02, OR=1.80[1.25-2.60], P=0.002) and in Urban American Indians (RAF=0.008, OR=2.29[1.03-5.12], P=0.04). However, after conditional analysis, only the intronic variant remains significant in the Pima Indian dataset. PALD1 is a predicted phosphatase and acts as a negative regulator of insulin signaling. Overexpression of PALD1 leads to lower insulin stimulated AKT phosphorylation and insulin receptor abundance. For preliminary functional studies, a short fragment (100bp) containing each allele of the PALD1 intronic variant (C or G) was cloned upstream of a SV40 promoter luciferase vector and transfected into C2C12 myoblasts. The T2D risk allele (C) fragment had significantly (P=0.004) higher SV40 promoter mediated luciferase activity as compared to the non-risk (G) allele. This result is consistent with the possible role of PALD1 in T2D pathogenesis. Further mechanistic studies are currently ongoing.

Disclosure
A.K. Nair: None. J.R. Sutherland: None. G.R. Elson: None. P. Piaggi: None. S. Kobes: None. R.L. Hanson: None. C. Bogardus: None. L. Baier: None.
Synergistic Effects of Long-Term Combination Therapy of DPP-4 Inhibitor and SGLT2 Inhibitor on the Preservation of Beta-Cell Volume in Rats with Type 2 Diabetes
Authors: MIZUKAMI HIROKI , GUO DANYANG , TAKAHASHI KAZUHISA , OSONOI SHO , OGASAWARA SAORI , INABA WATARU , YAGIHASHI SOROKU 
Published: June 2018

One of the main pathogenetic features of type 2 diabetes (T2D) is reduction of β cell volume (Vβ). Several antidiabetic oral agents were shown to improve Vβ in experimental studies. While DPP-4 inhibitor (DPi) ameliorated Vβ possibly via the action of GLP-1, recent studies reported that SGLT2 inhibitor (SGi) may facilitate GLP-1 secretion from L cells. These findings raised a hypothesis that the combination therapy (CT) of DPi and SGi may potentiate the effects of DPi on Vβ. To explore this possibility, we recruited non-obese spontaneous T2D Goto-Kakizaki (GK) rats and control Wistar rats (W) at 5 weeks of age. GK was divided into DPi treated group (GTe) (10mg/kg teneligliptine), SGi-treated group (GCa) (10mg/kg canagliflozin) and CT group (GTeCa). During experimental period, diabetic state was evaluated by glycated Hb (gHb), 2g/kg oral glucose tolerance test (OGTT), glucose-stimulated insulin secretion test (GSIS) and serum active GLP-1. At end, pancreata were underwent for the pathological evaluation of each endocrine cell volume by tetra-immnohistochemistry. Over 24 weeks of treatment, GK showed higher gHb and deteriorated OGTT (p&lt;0.01 vs. W). Although DPi and SGi in each significantly improved gHb and OGTT compared to GK, the effect was the most marked in GTeCa. Of note, active GLP-1 was the highest in GCaTe among all groups (p&lt;0.vs. GTe), while the level in GTe was higher than in W, GK, and GCa (p&lt;0.01). GK showed attenuated GSIS (p&lt;0.01 vs. W). It was significantly improved in GTe and GCaTe compared to GK (p&lt;0.vs. GTe, p&lt;0.01 vs. GCaTe), although it was comparable between GK and GCa. Pathological evaluation disclosed a significant decrease in Vβ in GK (p&lt;0.01 vs. W). Vβ was the most well-preserved in GCaTe (p&lt;0.vs. GTe), followed by GTe (p&lt;0.vs. GK), while it was comparable between GK and GCa. Our findings suggest that CT of DPi and SGi is more effective for β cell function and pathology in T2D than the monotherapy of DPi or SGi.

Disclosure
H. Mizukami: Research Support; Self; Sanwa Kagaku Kenkyusho Co., Ltd., Mitsubishi Tanabe Pharma Corporation. D. Guo: None. K. Takahashi: None. S. Osonoi: None. S. Ogasawara: None. W. Inaba: None. S. Yagihashi: None.
No Compensatory Increase in Non-ß Hormone Expressing Cells in the Pancreatic Duct Glands in Type 1 Diabetes
Authors: KULKARNI SHWETA S. , ATKINSON MARK A. , BUTLER ALEXANDRA 
Published: June 2018

Longstanding type 1 diabetes (T1D) entails a near total loss of β cells secondary to autoimmunity, fibrosis and decreased total organ mass, but accumulating evidence suggests there may be an endogenous attempt to regenerate β cells. Pancreatic duct glands (PDGs) represent a distinct anatomical compartment of the pancreas, forming blind ending outpouchings from the main pancreatic duct and its branches. The PDG is purported to be a potential stem/progenitor cell niche. PDGs respond to inflammation in T1D by becoming activated, evidenced by a high frequency of epithelial cell replication and an increase in progenitor-like cells. Hence, we addressed the hypothesis that in T1D, perturbation of hormone-expressing endocrine cells in the interlobular ducts (ILD) and PDGs exist. Pancreatic sections from 10 T1D (disease duration 8-58 year) and 11 similarly aged control (ND) donors were evaluated for replication and expression of known pancreatic endocrine hormones in ducts and PDG. We confirmed increased duct and PDG replication in T1D (T1D vs. ND— ILD: 4.5 ± 1.8 vs. 0.8 ± 0.2%, p&lt;0.05; PDG: 3.4 ± 1.3 vs. 0.4 ± 0.2%, p&lt;0.05) and reduced frequency of insulin expressing cells (T1D vs. ND— ILD: 0.0 ± 0.0 vs. 0.4 ± 0.1%, p&lt;0.001; PDGs: 0.± 0.07 vs. 2.4 ± 0.8%, p&lt;0.05). No differences were found in the frequency of other known endocrine cell types (T1D vs. ND— ILD: Glucagon: 1.5 ± 0.7 vs. 2.1 ± 0.6%, Somatostatin: 1.2 ± 0.4 vs. 1.6 ± 0.3%, Pancreatic Polypeptide: 0.4 ± 0.2 vs. 1.3 ± 0.3%; PDGs: Glucagon: 0.9 ± 0.9 vs. 1.6 ± 1.0%, Somatostatin: 1.96 ± 0.8 vs. 3.7 ± 1.9%, Pancreatic Polypeptide: 1.7 ± 1.1 vs. 2.1 ± 0.8%; p=ns, all). These data suggest that increased cell replication and the deficiency of insulin expressing cells in the PDG compartment of subjects with T1D are not associated with a corresponding increase in non-insulin hormone-expressing endocrine cells. Furthermore, the consistent observation of increased cell replication in the ducts and PDGs in T1D may implicate regenerative mechanisms towards an exocrine cell fate.

Disclosure
S.S. Kulkarni: None. M.A. Atkinson: Other Relationship; Self; Patent Issued. A. Butler: None.
Identification of Candidate Targets in Transcriptome of Islets from Offspring Exposed to High Glucose In Utero
Authors: CASASNOVAS JOSE , RAO XI , KUA KOK LIM 
Published: June 2018

Offspring of diabetic mothers have higher risk to develop type 2 diabetes. Though exposure to hyperglycemia in-utero is an inciting event, the mechanism by which it impacts islet function and leads to increased risk of diabetes is unknown. We hypothesize that high glucose at late gestation alters pancreatic islet transcriptome during early life and impairs offspring islet function in adulthood. To test this, gestation day (GD) 20 fetal rats in left uterine horn were exposed to 48 hours of hyperglycemia (HG) by infusing 4mg/min glucose directly into maternal left uterine artery. Right uterine pups remained euglycemic as internal controls (Con). On GD22, HG and Con pups were either euthanized or cross-fostered to healthy dams. Glucose tolerance testing was performed on weaning and at 2month old (2mo) offspring. Islets of GD22 and weanling pups were isolated for RNAseq. While pregnant dams and Con fetal pups remained euglycemic during infusion, HG pups had higher blood glucose (88±9 mg/dL vs. 51±5mg/dL, p&lt;0.05, n=20/20) and serum insulin levels (4.24±0.8 ng/mL vs. 3.04 ±0.6 ng/mL, p&lt;0.05, 6/6 pups). HG pups also had higher beta cell area (31±12% increase, p&lt;0.05, n=5/5 pups) but lower neonatal blood glucose level (72±9 mg/dL vs. 129±20 mg/dL, p&lt;0.05, n=9/12 pups). Subsequently, weaning and 2mo HG offspring had impaired glucose tolerance (55±23% higher iAUC, n = 8/10 pups, p&lt;0.05). 2mo pups also had lower insulin release during GTT. RNAseq identified 131 differentially expressed genes (DEG) in GD22 islets (fdr&lt;0.1, n=3/3,) and 77 DEG in weanling islets (fdr&lt;0.1, n=4/4,); with 10 common DEG. IPA analysis on these DEGs identified "cell signaling", "cellular movement", and "cell to cell signaling and interaction" as relevant biological processes. Weanling and young adult offspring exposed to late gestation hyperglycemia had impaired islet function with change expression in genes regulating critical biological processes. These genes will serve as target for future mechanistic studies.

Disclosure
J. Casasnovas: None. X. Rao: None. K. Kua: None.
Impact of Mig6 on ß-Cell Regeneration and Repair
Authors: BAUER BRANDON M. , EL KIMBERLEY , FUEGER PATRICK T. 
Published: June 2018

Following autoimmune-mediated destruction of insulin secreting β-cells in type 1 diabetes (T1D), regenerative and repair pathways aim to restore β-cell mass and maintain euglycemia. However, these pathways are often unable to compensate for the loss of β-cell mass, resulting in insufficient insulin secretion and chronic hyperglycemia. Our lab previously demonstrated that this inability to compensate for the loss of β-cell mass may be at least in part due to mitogen inducible gene 6 (Mig6), an anti-proliferative negative feedback inhibitor of epithelial growth factor receptor signaling, which is induced by proinflammatory cytokines central to macrophage-mediated β-cell destruction in T1D. Both Mig6 haploinsufficient mice (Mig6+/-) and mice with pancreas-specific Mig6 knockout (PKO) have increased β-cell mass and improved glucose tolerance when compared to littermate controls in a β-cell ablation model. We sought to determine the extent to which β-cell specific loss of Mig6 confers resistance to depletion of β-cell mass by streptozotocin (STZ, 35 mg/kg). We hypothesized that mice with β-cell specific loss of Mig6 (βKO) would have increased β-cell mass and improved glucose tolerance following STZ treatment when compared to control mice. We treated βKO and littermate controls with multiple low doses of STZ and measured both glucose tolerance and β-cell mass for up to 20 days following STZ treatment. At baseline, Mig6 expression was reduced by 63 ± 3% in βKO mice, and βKO mice had normal glucose homeostasis, insulin sensitivity, and islet architecture compared to littermate control. Unlike Mig6+/- and PKO mice, βKO mice had no improvements in β-cell mass or glucose tolerance compared to STZ-treated control littermates. Therefore, whereas both Mig6+/- and Mig6 PKO mice are able to preserve or restore β-cells with STZ treatment, loss of Mig6 in β-cells alone does not confer the same level of protection, suggesting a potential role for Mig6 in beta β-cell transdifferentiation or neogenesis.

Disclosure
B.M. Bauer: None. K. El: None. P.T. Fueger: None.
Impaired Glucagon and Insulin Responses in Mice Lacking the Cl-Loader Nkcc2a
Authors: KELLY LISA E. , DIFULVIO MAURICIO 
Published: June 2018

In pancreatic β-cells, glucose metabolism and plasma membrane depolarization are accountable for the early secretory response, a mechanism regulated primarily, though not exclusively, by ATP-sensitive K+ channels. In most cells, including β-cells, the net functional balance between Cl- loaders e.g., Nkcc1 and Nkcc2a and extruders like Kcc2 keeps the intracellular Cl- concentration above equilibrium creating an outwardly directed anionic gradient, which is electrogenically dissipated by Cl- channel opening. This makes possible the additional driving force for depolarization, which in β-cells prolongs the action potentials necessary for sustained insulin secretion.β-cells express low levels of the ’kidney-specific’ Na+K+2Cl- transporter Nkcc2a. This protein and Nkcc1 are targeted by bumetanide, which by blocking Cl- loading into β-cells, impairs insulin secretion provoking intermittent hyperglycemia, as seen in patients treated with this diuretic. Since the chronic functional implications of Nkcc2a in fuel homeostasis in vivo remain unexplored, we subjected Nkcc2aKO mice to dynamic tests of glucose homeostasis and determined their secretory responses in vivo and in vitro using islets. We found that Nkcc2aKO mice exhibit: i) increased steady-state blood glucose, plasma insulin, glucagon and de novo hepatic glucose production, ii) absent glucagon responses to physiological (6h) or prolonged (16h) fasting, iii) glucose intolerance and delayed in vivo insulin responses to glucose, iv) hyperplastic and hypertrophic α- and β-cells, v) conserved islet insulin responses to glucose, and vi) normal and reduced expression levels of Nkcc1 and Kcc2, the main Cl- loader and extruder in β-cells, respectively. Together, our results suggest for the first time that mice lacking Nkcc2a exhibit a complex metabolic phenotype related to unrestrained plasma glucagon, impaired insulin and glucagon responses in vivo in spite of compensatory mechanisms related to the regulation of Cl- transport in the islet.

Disclosure
L.E. Kelly: None. M. DiFulvio: None.