Reduced Β-cell Mass Research Articles

Molecular imaging of β-cells: diabetes and beyond.

Since diabetes is becoming a global epidemic, there is a great need to develop early β-cell specific diagnostic techniques for this disorder. There are two types of diabetes (i.e., type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM)). In T1DM, the destruction of pancreatic β-cells leads to reduced insulin production or even absolute insulin deficiency, which consequently results in hyperglycemia. Actually, a central issue in the pathophysiology of all types of diabetes is the relative reduction of β-cell mass (BCM) and/or impairment of the function of individual β-cells. In the past two decades, scientists have been trying to develop imaging techniques for noninvasive measurement of the viability and mass of pancreatic β-cells. Despite intense scientific efforts, only two tracers for positron emission tomography (PET) and one contrast agent for magnetic resonance (MR) imaging are currently under clinical evaluation. β-cell specific imaging probes may also allow us to precisely and specifically visualize transplanted β-cells and to improve transplantation outcomes, as transplantation of pancreatic islets has shown promise in treating T1DM. In addition, some of these probes can be applied to the preoperative detection of hidden insulinomas as well. In the present review, we primarily summarize potential tracers under development for imaging β-cells with a focus on tracers for PET, SPECT, MRI, and optical imaging. We will discuss the advantages and limitations of the various imaging probes and extend an outlook on future developments in the field.

Pancreatic ß-Cell Mass Is Determined by ß-Cell Size after Parturition in Mice

During pregnancy, β-cell mass is increased through β-cell hypertrophy and proliferation to adapt to insulin resistance. While β-cell mass is known to return toward non-pregnant levels after parturition, the cellular mechanisms by which β-cell mass is regulated during this period remains unclear. To address this issue in mice, we first quantified β-cell mass throughout the perinatal period using C57BL/6J female mice, and found that increased β-cell mass at gestational day 18 (G18) was significantly reduced at postpartum days 7 (P7), to comparable level with that in age-matched, non-pregnant (NP) female mice. Co-immunostaining for insulin and E-cadherin, which enabled us to precisely measure individual β-cell size, revealed that the mean β-cell size was significantly increased during perinatal periods at G18 and P1 than during non-pregnant state, and robustly decreased at P7. Intriguingly, the mean β-cell size at P7 was slightly but significantly smaller than that in the non-pregnant mice, and then returned to the non-pregnant levels until P21. On the other hand, quantification of apoptosis by TUNEL assay demonstrated that there was no significant difference in the number of apoptotic cells among the groups. To further investigate the contribution of dedifferentiation or transdifferentiation in reduced β-cell mass after parturition, we performed genetic lineage tracing using Ins1-Cre; ROSA26lacZ mice, which resulted in no evidence for transition into other cell types. Taken together, these findings demonstrate that the reduction of β-cell mass after parturition is mainly through reduced β-cell size rather than increased apoptosis or β-to-non-β transition. Disclosure M. Takahashi: None. T. Miyatsuka: Research Support; Self; Eli Lilly and Company, Japan Society for the Promotion of Science, Daiichi Sankyo Company, Limited, Novartis Pharma K.K., Astellas Pharma Inc., MSD K.K., Takeda Pharmaceutical Company Limited.. M. Himuro: None. L. Suzuki: None. M. Miura: Research Support; Self; Novo Nordisk Inc.. Speaker's Bureau; Self; Sanofi K.K.. Research Support; Self; Kowa Pharmaceuticals America, Inc.. T. Katahira: None. Y. Nishida: Research Support; Self; Novo Nordisk Inc.. Speaker's Bureau; Self; Daiichi Sankyo Company, Limited, AstraZeneca. A. Itakura: None. S. Takeda: None. H. Watada: Advisory Panel; Self; AstraZeneca. Consultant; Self; Astellas Pharma US, Inc., AstraZeneca, Boehringer Ingelheim GmbH, Daiichi Sankyo Company, Limited, Sumitomo Dainippon Pharma Co., Ltd., Eli Lilly and Company, Kissei Pharmaceutical Co., Ltd., Kowa Pharmaceuticals America, Inc., Kyowa Hakko Kirin Co., Ltd., Merck Sharp & Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Novo Nordisk A/S, Novartis AG, Ono Pharmaceutical Co., Ltd., Sanofi, Sanwa Kagaku Kenkyusho Co., Ltd., Takeda Development Center Asia, Pte. Ltd.. Research Support; Self; Abbott, Astellas Pharma US, Inc., AstraZeneca, Bayer AG, Benefit One Health Care Co., Ltd., Boehringer Ingelheim GmbH, Bristol-Myers Squibb Company, Daiichi Sankyo Company, Limited, Sumitomo Dainippon Pharma Co., Ltd., Eli Lilly and Company, Kissei Pharmaceutical Co., Ltd., Kowa Pharmaceuticals America, Inc., Kyowa Hakko Kirin Co., Ltd., Johnson & Johnson Diabetes Institute, LLC., Merck Sharp & Dohme Corp., Mitsubishi Tanabe Pharma Corporation, Mochida Pharmaceutical Co., Ltd., Nitto Boseki Co., Ltd., Novartis AG, Novo Nordisk A/S, Ono Pharmaceutical Co., Ltd., Pfizer Inc., Sanofi, Sanwa Kagaku Kenkyusho Co., Ltd., Taisho Pharmaceutical Co., Ltd., Takeda Development Center Asia, Pte. Ltd., Terumo Medical Corporation.

Critical role for the Tsc1-mTORC1 pathway in β-cell mass in Pdx1-deficient mice.

Mutations in the pancreatic duodenal homeobox (PDX1) gene are associated with diabetes in humans. Pdx1-haploinsufficient mice also develop diabetes, but the molecular mechanism is unknown. To this end, we knocked down Pdx1 gene expression in mouse MIN6 insulinoma cells. Pdx1 suppression not only increased apoptotic cell death but also decreased cell proliferation, which was associated with a decrease in activity of mechanistic target of rapamycin complex 1 (mTORC1). We found that in Pdx1-deficient mice, tuberous sclerosis 1 (Tsc1) ablation in pancreatic β-cells restores β-cell mass, increases β-cell proliferation and size, decreases the number of TUNEL-positive cells and restores glucose tolerance after glucose challenge. In addition, Tsc1 ablation in pancreatic β-cells increases phosphorylation of initiation factor 4E-binding protein 1 (4E-BP1) phosphorylation and 40S ribosomal protein S6, two downstream targets of mTORC1 indicating that Tsc1 mediates mTORC1 downregulation induced by Pdx1 suppression. These results suggest that the Tsc1-mTORC1 pathway plays an important role in mediating the decrease in β-cell proliferation and growth and the reduction in β-cell mass that occurs in Pdx1-deficient diabetes. Thus, mTORC1 may be target for therapeutic interventions in diabetes associated with reductions in β-cell mass.

Inhibition of Human Amylin Aggregation and Cellular Toxicity by Lipoic Acid and Ascorbic Acid.

More than 30 human degenerative diseases result from protein aggregation such as Alzheimer's disease (AD) and type 2 diabetes mellitus (T2DM). Islet amyloid deposits, a hallmark in T2DM, are found in pancreatic islets of more than 90% of T2DM patients. An association between amylin aggregation and reduction in β-cell mass was also established by post-mortem studies. A strategy in preventing protein aggregation-related disorders is to inhibit the protein aggregation and associated toxicity. In this study, we demonstrated that two inhibitors, lipoic acid and ascorbic acid, significantly inhibited amylin aggregation. Compared to amylin (15 μM) as 100%, lipoic acid and ascorbic acid reduced amylin fibril formation to 42.1 ± 17.2% and 42.9 ± 12.8%, respectively, which is confirmed by fluorescence and TEM images. In cell viability tests, both inhibitors protected RIN-m5f β-cells from the toxicity of amylin aggregates. At 10:1 molar ratio of lipoic acid to amylin, lipoic acid with amylin increased the cell viability to 70.3%, whereas only 42.8% RIN-m5f β-cells survived in amylin aggregates. For ascorbic acid, an equimolar ratio achieved the highest cell viability of 63.3% as compared to 42.8% with amylin aggregates only. Docking results showed that lipoic acid and ascorbic acid physically interact with amylin amyloidogenic region (residues Ser20-Ser29) via hydrophobic interactions; hence reducing aggregation levels. Therefore, lipoic acid and ascorbic acid prevented amylin aggregation via hydrophobic interactions, which resulted in the prevention of cell toxicity in vitro.

Analysis of the Role of the Conserved Disulfide in Amyloid Formation by Human Islet Amyloid Polypeptide in Homogeneous and Heterogeneous Environments.

Human islet amyloid polypeptide (hIAPP) is a hormone secreted from β-cells in the Islets of Langerhans in response to the same stimuli that lead to insulin secretion. hIAPP plays an adaptive role in glucose homeostasis but misfolds to form insoluble, fibrillar aggregates in type II diabetes that are associated with the disease. Along the misfolding pathway, hIAPP forms species that are toxic to β-cells, resulting in reduced β-cell mass. hIAPP contains a strictly conserved disulfide bond between residues 2 and 7, which forms a small loop at the N-terminus of the molecule. The loop is located outside of the cross β-core in all models of the hIAPP amyloid fibrils. Mutations in this region are rare, and the disulfide loop plays a role in receptor binding; however, the contribution of this region to the aggregation of hIAPP is not well understood. We define the role of the disulfide by analyzing a collection of analogues that remove the disulfide, by mutation of Cys to Ser, by reduction and modification of the Cys residues, or by deletion of the first seven residues. The cytotoxic properties of hIAPP are retained in the Cys to Ser disulfide-free mutant. Removal of the disulfide bond accelerates amyloid formation in all constructs, both in solution and in the presence of model membranes. Removal of the disulfide weakens the ability of hIAPP to induce leakage of vesicles consisting of POPS and POPC. Smaller effects are observed with vesicles that contain 40 mol % cholesterol, although N-terminal truncation still reduces the extent of leakage.

GLP-1 receptor agonist liraglutide exerts central action to induce β-cell proliferation through medulla to vagal pathway in mice

Endogenous GLP-1 and GLP-1 receptor agonists (GLP-1RAs) regulate glucose metabolism via common and distinct mechanisms. Postprandial release of GLP-1 is modest and it is degraded by DPP-4 within 2 min, and hence it cannot enter the brain in substantial amount. In contrast, DPP-4-resistant GLP-1RAs are administered at 10 times higher concentration than endogenous GLP-1 level, which enables them to reach several brain regions including ARC and AP, the areas implicated in glucose metabolism. Hence, some of the effects of GLP-1RAs observed clinically and experimentally, including pancreatic β-cell proliferation, are thought to involve the brain. However, the effects of centrally acting GLP-1/GLP-1RAs on glucose metabolism and underlying neural mechanism are unclear. This study aimed to establish the link of central GLP-1/GLP-1RA action to pancreatic β-cell proliferation. Both subcutaneous (SC) and intracerebroventricular (ICV) injections of liraglutide increased the number of pancreatic β-cells expressing Ki67 and PCNA, proliferation markers, in C57BL/6J mice. This effect was induced by single ICV administration of liraglutide at relatively low dose that was incapable of suppressing food intake. These SC and ICV liraglutide-induced effects were inhibited by 50% and 70%, respectively, by pretreatment with atropine, a muscarinic receptor blocker. ICV liraglutide induced c-Fos expression in the area postrema (AP), nucleus tractus solitaries (NTS), and dorsal motor nucleus of the vagus (DMX) of the brain stem. These results demonstrate that central action of liraglutide induces pancreatic β-cell proliferation via the pathway involving the brain stem AP/NTS/DMX area and vagus nerve. This route is highly sensitive to GLP-1/GLP-1RA. Hence, this brain-pancreatic β-cell pathway may operate in type 2 diabetic patients treated with GLP-RAs and serve to counteract the reduction of β-cell mass.

Optical clearing of the pancreas for visualization of mature β-cells and vessels in mice

ABSTRACTGlucose metabolism is regulated by insulin, which is produced from β-cells in the pancreas. Because insulin is secreted into vessels in response to blood glucose, vascular structures of the pancreas, especially the relationship between vessels and β-cells, are important for physiological and pathological glucose metabolism. Here, we developed a system to visualize vessels surrounding mature β-cells expressing transcription factor MafA in a three-dimensional manner. Optical clearing of the pancreas prevented light scattering of fluorescence driven by the bacterial artificial chromosome (BAC)-mafA promoter in β-cells. Reconstruction of confocal images demonstrated mature β-cells and the glomerular-like structures of β-cell vasculatures labeled with DyLight 488-conjugated lectin in normal mice as well as in low-dose streptozotocin-injected diabetes model mice with reduced β-cell mass. This technological innovation of organ imaging can be used to investigate morphological changes in vascular structures during transplantation, regeneration and diabetes development.

Caveolin-1 deficiency protects pancreatic β cells against palmitate-induced dysfunction and apoptosis

Lipotoxicity leads to insulin secretion deficiency, which is among the important causes for the onset of type 2 diabetes mellitus. Thus, the restoration of β-cell mass and preservation of its endocrine function are long-sought goals in diabetes research. Previous studies have suggested that the membrane protein caveolin-1 (Cav-1) is implicated in β-cell apoptosis and insulin secretion, however, the underlying mechanisms still remains unclear. Our objective is to explore whether Cav-1 depletion protects pancreatic β cells from lipotoxicity and what are the underlying mechanisms. In this study, we found that Cav-1 silencing significantly promoted β-cell proliferation, inhibited palmitate (PA)-induced pancreatic β-cell apoptosis and enhanced insulin production and secretion. These effects were associated with enhanced activities of Akt and ERK1/2, which in turn downregulated the expression of cell cycle inhibitors (FOXO1, GSK3β, P21, P27 and P53) and upregulated the expression of Cyclin D2 and Cyclin D3. Subsequent inhibition of PI3K/Akt and ERK/MAPK pathways abolished Cav-1 depletion induced β-cell mass protection. Furthermore, under PA induced endoplasmic reticulum (ER) stress, Cav-1 silencing significantly reduced eIF2α phosphorylation and the expression of ER stress-responsive markers BiP and CHOP, which are among the known sensitizers of lipotoxicity. Our findings suggest Cav-1 as potential target molecule in T2DM treatment via the preservation of lipotoxicity-induced β-cell mass reduction and the attenuation of insulin secretion dysfunction.

An SCFFBXO28 E3 Ligase Protects Pancreatic β-Cells from Apoptosis.

Loss of pancreatic β-cell function and/or mass is a central hallmark of all forms of diabetes but its molecular basis is incompletely understood. β-cell apoptosis contributes to the reduced β-cell mass in diabetes. Therefore, the identification of important signaling molecules that promote β-cell survival in diabetes could lead to a promising therapeutic intervention to block β-cell decline during development and progression of diabetes. In the present study, we identified F-box protein 28 (FBXO28), a substrate-recruiting component of the Skp1-Cul1-F-box (SCF) ligase complex, as a regulator of pancreatic β-cell survival. FBXO28 was down-regulated in β-cells and in isolated human islets under diabetic conditions. Consistently, genetic silencing of FBXO28 impaired β-cell survival, and restoration of FBXO28 protected β-cells from the harmful effects of the diabetic milieu. Although FBXO28 expression positively correlated with β-cell transcription factor NEUROD1 and FBXO28 depletion also reduced insulin mRNA expression, neither FBXO28 overexpression nor depletion had any significant impact on insulin content, glucose-stimulated insulin secretion (GSIS) or on other genes involved in glucose sensing and metabolism or on important β-cell transcription factors in isolated human islets. Consistently, FBXO28 overexpression did not further alter insulin content and GSIS in freshly isolated islets from patients with type 2 diabetes (T2D). Our data show that FBXO28 improves pancreatic β-cell survival under diabetogenic conditions without affecting insulin secretion, and its restoration may be a novel therapeutic tool to promote β-cell survival in diabetes.

Sex Difference In the Effect of Fetal Exposure to Maternal Diabetes on Insulin Secretion.

We previously showed that fetal exposure to maternal type 1 diabetes (T1D) is associated with altered glucose-stimulated insulin secretion in adult offspring. Here, we investigated whether this β-cell defect displays a sex dimorphism. Twenty-nine adult nondiabetic offspring of T1D mothers (ODMs) were compared with 29 nondiabetic offspring of T1D fathers. We measured early insulin secretion in response to oral glucose and insulin secretion rate in response to intravenous glucose ramping. Insulin sensitivity and body composition were assessed by a euglycemic, hyperinsulinemic clamp and dual-energy X-ray absorptiometry, respectively. In response to oral glucose, male and female ODMs displayed a reduced insulin secretion. In contrast, in response to graded intravenous glucose infusion, only female ODMs (not males) exhibited decreased insulin secretion. There was no defect in response to combined intravenous arginine and glucose, suggesting that male and female ODMs exhibit a functional β-cell defect rather than a reduced β-cell mass. In conclusion, fetal exposure to maternal diabetes predisposes to β-cell dysfunction in adult male and female offspring. This β-cell defect is characterized by a sexual dimorphism following intravenous glucose stimulation.

Loss of Deubiquitinase USP1 Blocks Pancreatic β-Cell Apoptosis by Inhibiting DNA Damage Response

SummaryImpaired pancreatic β-cell survival contributes to the reduced β-cell mass in diabetes, but underlying regulatory mechanisms and key players in this process remain incompletely understood. Here, we identified the deubiquitinase ubiquitin-specific protease 1 (USP1) as an important player in the regulation of β-cell apoptosis under diabetic conditions. Genetic silencing and pharmacological suppression of USP1 blocked β-cell death in several experimental models of diabetes in vitro and ex vivo without compromising insulin content and secretion and without impairing β-cell maturation/identity genes in human islets. Our further analyses showed that USP1 inhibition attenuated DNA damage response (DDR) signals, which were highly elevated in diabetic β-cells, suggesting a USP1-dependent regulation of DDR in stressed β-cells. Our findings highlight a novel function of USP1 in the control of β-cell survival, and its inhibition may have a potential therapeutic relevance for the suppression of β-cell death in diabetes.

Amyloid formation reduces protein kinase B phosphorylation in primary islet β-cells which is improved by blocking IL-1β signaling.

Amyloid formation in the pancreatic islets due to aggregation of human islet amyloid polypeptide (hIAPP) contributes to reduced β-cell mass and function in type 2 diabetes (T2D) and islet transplantation. Protein kinase B (PKB) signaling plays a key role in the regulation of β-cell survival, function and proliferation. In this study, we used human and hIAPP-expressing transgenic mouse islets in culture as two ex vivo models of human islet amyloid formation to: 1. Investigate the effects of amyloid formation on PKB phosphorylation in primary islet β-cells; 2. Test if inhibition of amyloid formation and/or interleukin-1β (IL-1β) signaling in islets can restore the changes in β-cell phospho-PKB levels mediated by amyloid formation. Human and hIAPP-expressing mouse islets were cultured in elevated glucose with an amyloid inhibitor (Congo red) or embedded within collagen matrix to prevent amyloid formation. To block the IL-1β signaling, human islets were treated with an IL-1 receptor antagonist (anakinra) or a glucagon-like peptide-1 agonist (exenatide). β-cell phospho-PKB levels, proliferation, apoptosis, islet IL-1β levels and amyloid formation were assessed. Amyloid formation in both cultured human and hIAPP-expressing mouse islets reduced β-cell phospho-PKB levels and increased islet IL-1β levels, both of which were restored by prevention of amyloid formation either by the amyloid inhibitor or embedding islets in collagen matrix, resulting in improved β-cell survival. Furthermore, inhibition of IL-1β signaling by treatment with anakinra or exenatide increased β-cell phospho-PKB levels, enhanced proliferation and reduced apoptosis in amyloid forming human islets during 7-day culture. These data suggest that amyloid formation leads to reduced PKB phosphorylation in β-cells which is associated with elevated islet IL-1β levels. Inhibitors of amyloid or amyloid-induced IL-1β production may provide a new approach to restore phospho-PKB levels thereby enhance β-cell survival and proliferation in conditions associated with islet amyloid formation such as T2D and clinical islet transplantation.

Transcriptomic Analysis Reveals Novel Mechanisms Mediating Islet Dysfunction in the Intrauterine Growth-Restricted Rat.

Intrauterine growth restriction (IUGR) increases the risk of type 2 diabetes developing in adulthood. In previous studies that used bilateral uterine artery ligation in a rat model of IUGR, age-associated decline in glucose homeostasis and islet function was revealed. To elucidate mechanisms contributing to IUGR pathogenesis, the islet transcriptome was sequenced from 2-week-old rats, when in vivo glucose tolerance is mildly impaired, and at 10 weeks of age, when rats are hyperglycemic and have reduced β-cell mass. RNA sequencing and functional annotation with Ingenuity Pathway Analysis revealed temporal changes in IUGR islets. For instance, gene expression involving amino acid metabolism was significantly reduced primarily at 2 weeks of age, but ion channel expression, specifically that involved in cell-volume regulation, was more disrupted in adult IUGR islets. Additionally, we observed alterations in the microenvironment of IUGR islets with extracellular matrix genes being significantly increased at 2 weeks of age and significantly decreased at 10 weeks. Specifically, hyaluronan synthase 2 expression and hyaluronan staining were increased in IUGR islets at 2 weeks of age (P < 0.05). Mesenchymal stromal cell-derived factors that have been shown to preserve islet allograft function, such as Anxa1, Cxcl12, and others, also were increased at 2 weeks and decreased in adult islets. Finally, comparisons of differentially expressed genes with those of type 2 diabetic human islets support a role for these pathways in human patients with diabetes. Together, these data point to new mechanisms in the pathogenesis of IUGR-mediated islet dysfunction in type 2 diabetes.

Islet α-cell Inflammation Induced By NF-κB inducing kinase (NIK) Leads to Hypoglycemia, Pancreatitis, Growth Retardation, and Postnatal Death in Mice.

Islet α-cell dysfunction has been shown to contribute to type 2 diabetes; however, whether islet α-cell inflammation is involved in the occurrence of pancreatitis is largely unknown. The aims of this study were to investigate how NF-κB inducing kinase (NIK) regulates pancreatic α-cell function, both in vitro and in vivo, and to assess how islet α-cell inflammation induced by NIK affects the development of pancreatitis.Methods: We utilized adenovirus-mediated NIK overexpression, ELISA, qPCR, RNA-seq, and Western blot analyses to study the role of NIK in islet α cells in vitro. Islet α-cell-specific NIK overexpressing (α-NIK-OE) mice were generated, and pancreatic α/β-cell function and the occurrence of pancreatitis in these mice were assessed via ELISA, qPCR, and immunohistochemical analyses.Results: The LTβR/noncanonical NF-κB signaling pathway is present in islet α cells. Overexpression of NIK in αTC1-6 cells induces inflammation and cell death, contributing to a decrease in the expression and secretion of glucagon. Additionally, α-cell specific overexpression of NIK (α-NIK-OE) results in α-cell death, lower serum glucagon levels, and hypoglycemia in mice. Strikingly, α-NIK-OE mice also display a reduced β-cell mass, growth retardation, pancreatitis, and postnatal death.Conclusions: Islet α-cell specific overexpression of NIK results in islet α-cell dysfunction and causes islet β-cell death and pancreatitis, which are most likely due to paracrine secretion of cytokines and chemokines from islet α cells, thus leading to hypoglycemia, growth retardation, and postnatal death in mice.

GRP94 Is an Essential Regulator of Pancreatic β-Cell Development, Mass, and Function in Male Mice.

Deficiencies in pancreatic β-cell mass contribute to both type 1 and type 2 diabetes. We investigated the role of the glucose-regulated protein (GRP) 94, an endoplasmic reticulum protein abundantly expressed in the pancreatic acini and islets, in β-cell development, survival, and function. We used a conditional knockout (KO) mouse in which the GRP94 gene, Hsp90b1, was specifically deleted in pancreatic and duodenal homeobox 1 (Pdx1)-expressing cells. These Hsp90b1 flox/flox;Pdx1Cre KO mice exhibited pancreatic hypoplasia at embryonic day (E) 16.5 to E18.5 and had significantly reduced β-cell mass at 4 weeks after birth. Further mechanistic studies showed that deletion of GRP94 reduced β-cell proliferation with increased cell apoptosis in both Pdx1+ endocrine progenitor cells and differentiated β cells. Although Hsp90b1 flox/flox;Pdx1Cre KO mice remained euglycemic at 8 weeks of age, they exhibited impaired glucose tolerance. In aggregate, these findings indicate that GRP94 is an essential regulator of pancreatic β-cell development, mass, and function.

The prolyl isomerase Pin1 increases β-cell proliferation and enhances insulin secretion

The prolyl isomerase Pin1 binds to the phosphorylated Ser/Thr-Pro motif of target proteins and enhances their cis-trans conversion. This report is the first to show that Pin1 expression in pancreatic β cells is markedly elevated by high-fat diet feeding and in ob/ob mice. To elucidate the role of Pin1 in pancreatic β cells, we generated β-cell-specific Pin1 KO (βPin1 KO) mice. These mutant mice showed exacerbation of glucose intolerance but had normal insulin sensitivity. We identified two independent factors underlying impaired insulin secretion in the βPin1 KO mice. Pin1 enhanced pancreatic β-cell proliferation, as indicated by a reduced β-cell mass in βPin1 KO mice compared with control mice. Moreover, a diet high in fat and sucrose failed to increase pancreatic β-cell growth in the βPin1 KO mice, an observation to which up-regulation of the cell cycle protein cyclin D appeared to contribute. The other role of Pin1 was to activate the insulin-secretory step: Pin1 KO β cells showed impairments in glucose- and KCl-induced elevation of the intracellular Ca2+ concentration and insulin secretion. We also identified salt-inducible kinase 2 (SIK2) as a Pin1-binding protein that affected the regulation of Ca2+ influx and found Pin1 to enhance SIK2 kinase activity, resulting in a decrease in p35 protein, a negative regulator of Ca2+ influx. Taken together, our observations demonstrate critical roles of Pin1 in pancreatic β cells and that Pin1 both promotes β-cell proliferation and activates insulin secretion.

Temporal Transcriptomic and Proteomic Landscapes of Deteriorating Pancreatic Islets in Type 2 Diabetic Rats.

Progressive reduction in β-cell mass and function comprise the core of the pathogenesis mechanism of type 2 diabetes. The process of deteriorating pancreatic islets, in which a complex network of molecular events is involved, is not yet fully characterized. We used RNA sequencing and tandem mass tag-based quantitative proteomics technology to measure the temporal mRNA and protein expression changes of pancreatic islets in Goto-Kakizaki (GK) rats from 4 to 24 weeks of age. Our omics data set outlines the dynamics of the molecular network during the deterioration of GK islets as two stages: The early stage (4-6 weeks) is characterized by anaerobic glycolysis, inflammation priming, and compensation for insulin synthesis, and the late stage (8-24 weeks) is characterized by inflammation amplification and compensation failure. Further time course analysis allowed us to reveal 5,551 differentially expressed genes, a large portion of which have not been reported before. Our comprehensive and temporal transcriptome and proteome data offer a valuable resource for the diabetes research community and for quantitative biology.

Amelioration of hyperglycemia and associated metabolic abnormalities by a combination of fenugreek (Trigonella foenum-graecum) seeds and onion (Allium cepa) in experimental diabetes.

Fenugreek (Trigonella foenum-graecum) seeds and onion (Allium cepa) are independently known to have antidiabetic effects through different mechanisms. The beeneficial influence of a combination of dietary fenugreek seeds and onion on hyperglycemia and its associated metabolic abnormalities were evaluated in streptozotocin-induced diabetic rats. Diabetes was experimentally induced with streptozotocin and diabetic rats were fed with 10% fenugreek or 3% onion or their combination for 6 weeks. These dietary interventions significantly countered hyperglycemia, partially improved peripheral insulin resistance and impaired insulin secretion, reduced β-cell mass and markedly reversed the abnormalities in plasma albumin, urea, creatinine, glycated hemoglobin and advanced glycation end products in diabetic rats. These beneficial effects were highest in the fenugreek+onion group. Diabetic rats with these dietary interventions excreted lesser glucose, albumin, urea and creatinine, which were accompanied by improved body weights compared with the diabetic controls. These dietary interventions produced ameliorative effects on pancreatic pathology as reflected by near-normal islet cells, restored glycogen and collagen fiber deposition in diabetic rats. This study documented the hypoglycemic and insulinotropic effects of dietary fenugreek and onion, which were associated with countering of metabolic abnormalities and pancreatic pathology. It may be strategic to derive maximum nutraceutical antidiabetic benefits from these functional food ingredients by consuming them together.

VMP1-related autophagy induced by a fructose-rich diet in β-cells: its prevention by incretins.

The aim of the present study was to demonstrate the role of autophagy and incretins in the fructose-induced alteration of β-cell mass and function. Normal Wistar rats were fed (3weeks) with a commercial diet without (C) or with 10% fructose in drinking water (F) alone or plus sitagliptin (CS and FS) or exendin-4 (CE and FE). Serum levels of metabolic/endocrine parameters, β-cell mass, morphology/ultrastructure and apoptosis, vacuole membrane protein 1 (VMP1) expression and glucose-stimulated insulin secretion (GSIS) were studied. Complementary to this, islets isolated from normal rats were cultured (3days) without (C) or with F and F + exendin-4 or chloroquine. Expression of autophagy-related proteins [VMP1 and microtubule-associated protein light chain 3 (LC3)], apoptotic/antiapoptotic markers (caspase-3 and Bcl-2), GSIS and insulin mRNA levels were measured. F rats developed impaired glucose tolerance (IGT) and a significant increase in plasma triacylglycerols, thiobarbituric acid-reactive substances, insulin levels, homoeostasis model assessment (HOMA) for insulin resistance (HOMA-IR) and β-cell function (HOMA-β) indices. A significant reduction in β-cell mass was associated with an increased apoptotic rate and morphological/ultrastructural changes indicative of autophagic activity. All these changes were prevented by either sitagliptin or exendin-4. In cultured islets, F significantly enhanced insulin mRNA and GSIS, decreased Bcl-2 mRNA levels and increased caspase-3 expression. Chloroquine reduced these changes, suggesting the participation of autophagy in this process. Indeed, F induced the increase of both VMP1 expression and LC3-II, suggesting that VMP1-related autophagy is activated in injured β-cells. Exendin-4 prevented islet-cell damage and autophagy development. VMP1-related autophagy is a reactive process against F-induced islet dysfunction, being prevented by exendin-4 treatment. This knowledge could help in the use of autophagy as a potential target for preventing progression from IGT to type2 diabetes mellitus.

Loss of Cyclin-dependent Kinase 2 in the Pancreas Links Primary β-Cell Dysfunction to Progressive Depletion of β-Cell Mass and Diabetes

The failure of pancreatic islet β-cells is a major contributor to the etiology of type 2 diabetes. β-Cell dysfunction and declining β-cell mass are two mechanisms that contribute to this failure, although it is unclear whether they are molecularly linked. Here, we show that the cell cycle regulator, cyclin-dependent kinase 2 (CDK2), couples primary β-cell dysfunction to the progressive deterioration of β-cell mass in diabetes. Mice with pancreas-specific deletion of Cdk2 are glucose-intolerant, primarily due to defects in glucose-stimulated insulin secretion. Accompanying this loss of secretion are defects in β-cell metabolism and perturbed mitochondrial structure. Persistent insulin secretion defects culminate in progressive deficits in β-cell proliferation, reduced β-cell mass, and diabetes. These outcomes may be mediated directly by the loss of CDK2, which binds to and phosphorylates the transcription factor FOXO1 in a glucose-dependent manner. Further, we identified a requirement for CDK2 in the compensatory increases in β-cell mass that occur in response to age- and diet-induced stress. Thus, CDK2 serves as an important nexus linking primary β-cell dysfunction to progressive β-cell mass deterioration in diabetes.