Insulin Secretion In Pancreatic Islets Research Articles

Caveolin-1 Deficiency Inhibits Palmitate-Induced Intracellular Lipid Accumulation and Enhances Autophagy in Pancreatic ß Cells via AMPK/mTOR Pathway

Lipotoxicity induced intracellular lipid accumulation in pancreatic β cells, one of the strongest predictors for T2DM. The objective of this study is to explore the role of Cav-1 in intracellular lipid accumulation of β cells under lipotoxicity condition. Our data showed that Cav-1 silencing significantly reduced PA-induced intracellular triglyceride (TG) accumulation, enhanced autophagy and insulin secretion in NIT-1 cells and murine pancreatic islets. Further investigation found that Cav-1 depletion facilitated AMPK phosphorylation, decreased the expression of downstream lipogenic makers (SREBP-1c, FAS and ACC) and increased the expression of fatty acid oxidation maker (CPT-1), leading to the ultimate enhancement of lipid metabolism. Meanwhile, Cav-1 depletion also suppressed mTOR phosphorylation, decreased the expression of p62 protein and increased the expression of LC3-II proteins, resulting in the ultimate increase of autophagy. Subsequent inhibition of AMPK and mTOR pathways abolished Cav-1 depletion mediated autophagy enhancement and TG content reduction, leading to the reduction of insulin secretion. Our findings suggest the potential application of the Cav-1 molecule as a target for effective T2DM treatment through preservation of lipotoxicity-induced β-cell intracellular lipid accumulation and dysfunction. Disclosure W. Zeng: None. K. Liu: None. J. Tang: None. H. Peng: None. C. Lin: None. S. Lin: None. K. Lin: None. Y. Jiang: None. L. Zeng: None.

Aerobic Exercise Training Attenuates Tumor Growth and Reduces Insulin Secretion in Walker 256 Tumor-Bearing Rats.

Aerobic exercise training can improve insulin sensitivity in many tissues; however, the relationship among exercise, insulin, and cancer cell growth is unclear. We tested the hypothesis that aerobic exercise training begun during adolescence can attenuate Walker 256 tumor growth in adult rats and alter insulin secretion. Thirty-day-old male Wistar rats engaged in treadmill running for 8 weeks, 3 days/week, 44 min/day, at 55–65% VO2max until they were 90 days old (TC, Trained Control). An equivalently aged group was kept inactive during the same period (SC, Sedentary Control). Then, half the animals of the SC and TC groups were reserved as the control condition and the other half were inoculated with Walker 256 cancer cells, yielding two additional groups (Sedentary Walker and Trained Walker). Zero mortalities were observed in tumor-bearing rats. Body weight (BW), food intake, plasma glucose, insulin levels, and peripheral insulin sensitivity were analyzed before and after tumor cell inoculation. We also evaluated tumor growth, metastasis and cachexia. Isolated pancreatic islets secretory activity was analyzed. In addition, we evaluated mechanic sensibility. Our results showed improved physical performance according to the final workload and VO2max and reduced BW in trained rats at the end of the running protocol. Chronic adaptation to the aerobic exercise training decreased tumor weight, cachexia and metastasis and were associated with low glucose and insulin levels and high insulin sensitivity before and after tumor cell inoculation. Aerobic exercise started at young age also reduced pancreatic islet insulin content and insulin secretion in response to a glucose stimulus, without impairing islet morphology in trained rats. Walker 256 tumor-bearing sedentary rats also presented reduced pancreatic islet insulin content, without changing insulin secretion through isolated pancreatic islets. The mechanical sensitivity test indicated that aerobic exercise training did not cause injury or trigger inflammatory processes prior to tumor cell inoculation. Taken together, the current study suggests that aerobic exercise training applied during adolescence may mitigate tumor growth and related disorders in Walker 256 tumor-bearing adult rats. Improved insulin sensibility, lower glucose and insulin levels and/or reduced insulin secretion stimulated by glucose may be implicated in this tumor attenuation.

The potent insulin secretagogue effect of betulinic acid is mediated by potassium and chloride channels

Betulinic acid (BA) has been described as an insulin secretagogue which may explain its potent antihyperglycemic effect; however, the exact role of BA as an insulinogenic agent is not clear. The aim of this study was to investigate the mechanism of BA on calcium influx and static insulin secretion in pancreatic islets isolated from euglycemic rats. We found that BA triggers calcium influx by a mechanism dependent on ATP-dependent potassium channels and L-type voltage-dependent calcium channels. Additionally, the voltage-dependent and calcium-dependent chloride channels are also involved in the mechanism of BA, probably due to an indirect stimulation of calcium entry and increased intracellular calcium. Additionally, the downstream activation of PKC, which is necessary for the effect of BA on calcium influx, is involved in the full stimulatory response of the triterpene. BA stimulated the static secretion of insulin in pancreatic islets, indicating that the abrupt calcium influx may be a key step in its secretagogue effect. As such, BA stimulates insulin secretion through the activation of electrophysiological mechanisms, such as the closure of potassium channels and opening of calcium and chloride channels, inducing cellular depolarization associated with metabolic-biochemical effects, in turn activating PKC and ensuring the secretion of insulin.

Critical Roles of Carbon Monoxide and Nitric Oxide in Ca2+ Signaling for Insulin Secretion in Pancreatic Islets.

Glucagon-like peptide-1 (GLP-1) increases intracellular Ca2+ concentrations, resulting in insulin secretion from pancreatic β-cells through the sequential production of Ca2+ mobilizing messengers nicotinic acid adenine dinucleotide phosphate (NAADP) and cyclic ADP-ribose (cADPR). We previously found that NAADP activates the neuronal type of nitric oxide (NO) synthase (nNOS), the product of which, NO, activates guanylyl cyclase to produce cyclic guanosine monophosphate (cGMP), which, in turn, induces cADPR formation. Our aim was to explore the relationship between Ca2+ signals and gasotransmitters formation in insulin secretion in β-cells upon GLP-1 stimulation. We show that NAADP-induced cGMP production by nNOS activation is dependent on carbon monoxide (CO) formation by heme oxygenase-2 (HO-2). Treatment with exogenous NO and CO amplifies cGMP formation, Ca2+ signal strength, and insulin secretion, whereas this signal is impeded when exposed to combined treatment with NO and CO. Furthermore, CO potentiates cGMP formation in a dose-dependent manner, but higher doses of CO inhibited cGMP formation. Our data with regard to zinc protoporphyrin, a HO inhibitor, and HO-2 knockdown, revealed that NO-induced cADPR formation and insulin secretion are dependent on HO-2. Consistent with this observation, the administration of NO or CO donors to type 2 diabetic mice improved glucose tolerance, but the same did not hold true when both were administered concurrently. Our research reveals the role of two gas transmitters, CO and NO, for Ca2+ second messengers formation in pancreatic β-cells. These results demonstrate that CO, the downstream regulator of NO, plays a role in bridging the gap between the Ca2+ signaling messengers during insulin secretion in pancreatic β-cells.

Deletion of the RabGAP TBC1D1 Leads to Enhanced Insulin Secretion and Fatty Acid Oxidation in Islets From Male Mice.

The Rab guanosine triphosphatase-activating protein (RabGAP) TBC1D1 has been shown to be a key regulator of glucose and lipid metabolism in skeletal muscle. Its function in pancreatic islets, however, is not yet fully understood. Here, we aimed to clarify the specific impact of TBC1D1 on insulin secretion and substrate use in pancreatic islets. We analyzed the dynamics of glucose-stimulated insulin secretion (GSIS) and lipid metabolism in isolated islets from Tbc1d1-deficient (D1KO) mice. To further investigate the underlying cellular mechanisms, we conducted pharmacological studies in these islets. In addition, we determined morphology and number of both pancreatic islets and insulin vesicles in β-cells using light and transmission electron microscopy. Isolated pancreatic islets from D1KO mice exhibited substantially increased GSIS compared with wild-type (WT) controls. This was attributed to both enhanced first and second phase of insulin secretion, and this enhanced secretion persisted during repetitive glucose stimuli. Studies with sulfonylureas or KCl in isolated islets demonstrated that TBC1D1 exerts its function via a signaling pathway at the level of membrane depolarization. In line, ultrastructural analysis of isolated pancreatic islets revealed both higher insulin-granule density and number of docked granules in β-cells from D1KO mice compared with WT controls. Like in skeletal muscle, lipid use in isolated islets was enhanced upon D1KO, presumably as a result of a higher mitochondrial fission rate and/or higher mitochondrial activity. Our results clearly demonstrate a dual role of TBC1D1 in controlling substrate metabolism of the pancreatic islet.

CXC chemokine ligand 12α-mediated increase in insulin secretion and survival of mouse pancreatic islets in response to oxidative stress through modulation of calcium uptake

We examined whether CXCL12? improves insulin secretion by influencing the Ca2+ oscillation pattern and Ca2+ influx ([Ca2+]i), thereby enhancing the viability of pancreatic islet cells in oxidative stress. The islets of Langerhans were isolated from male OF1 mice and pretreated with 40 ng/mL of CXCL12? prior to exposure to 7.5 ?M hydrogen peroxide, which served to induce oxidative stress. Incubation of islets with CXCL12? induced pancreatic ?-cell proliferation and improved the ability of ?-cells to withstand oxidative stress. Consecutive treatments of isolated islets with hydrogen peroxide caused a decline in ?-cell functioning over time, while the CXCL12? pretreatment of islets exhibited a physiological response to high glucose that was comparable to control islets. The attenuated response of islets to a high D-glucose challenge was observed as a partial to complete abolishment of [Ca2+]i. Treatments with increasing concentrations of CXCL12? decreased the number of Ca2+ oscillations that lasted longer, thus pointing to an overall increase in [Ca2+]i, which was followed by increased insulin secretion. In addition, treatment of islets with CXCL12? enhanced the transcription rate for insulin and the CXCR4 gene, pointing to the importance of CXCL12/CXCR4 signaling in the regulation of Ca2+ intake and insulin secretion in pancreatic islet cells. We propose that a potential treatment with CXCL12? could help to remove preexisting glucotoxicity and associated temporary ?-cell stunning that might be present at the time of diabetes diagnosis in vivo.

Delta-like Ligand-4-Notch Signaling Inhibition Regulates Pancreatic Islet Function and Insulin Secretion

Although Notch signaling has been proposed as a therapeutic target for type-2 diabetes, liver steatosis, and atherosclerosis, its direct effect on pancreatic islets remains unknown. Here, we demonstrated a function of Dll4-Notch signaling inhibition on the biology of insulin-producing cells. We confirmed enhanced expression of key Notch signaling genes in purified pancreatic islets from diabetic NOD mice and showed that treatment with anti-Dll4 antibody specifically abolished Notch signaling pathway activation. Furthermore, we showed that Notch inhibition could drive proliferation of β-islet cells and confer protection from the development of STZ-induced diabetes. Importantly, inhibition of the Dll4 pathway in WT mice increased insulin secretion by inducing the differentiation of pancreatic β-islet cell progenitors, as well asthe proliferation of insulin-secreting cells. These findings reveal a direct effect of Dll4-blockade on pancreatic islets that, in conjunction with its immunomodulatory effects, could be used for unmet medical needs hallmarked by inefficient insulin action.

Metformin Inhibits Mouse Islet Insulin Secretion and Alters Intracellular Calcium in a Concentration-Dependent and Duration-Dependent Manner near the Circulating Range.

Metformin is considered the first-line treatment for type 2 diabetes. While metformin primarily increases insulin sensitivity, evidence also suggests that metformin affects the activity of insulin-secreting pancreatic islets. This study was designed to systematically examine the direct effects of metformin by measuring insulin secretion and the kinetics of the calcium response to glucose stimulation in isolated mouse islets using varying concentrations (20 μM, 200 μM, and 1 mM) and durations (~1, 2, and 3 days) of metformin exposure. We observed both concentration- and duration-dependent inhibitory effects of metformin. Concentrations as little as 20 μM (nearing circulating therapeutic levels) were sufficient to reduce insulin secretion following 3-day treatment. Concentrations of 200 μM and 1 mM produced more pronounced effects more rapidly. With 1 mM metformin, islets showed severe impairments in calcium handling, inhibition of insulin secretion, and increased cell death. No stimulatory effects of metformin were observed for any experimental endpoint. We conclude that the direct effects of metformin on islets are inhibitory at near-physiological concentrations. Beneficial effects of metformin observed on islets under various stressors may occur by “resting” fatigued cellular processes. However, metformin may have unintended consequences on normally functioning islets within the circulating range that require further evaluation.

Guanylin and uroguanylin mRNA expression is increased following Roux-en-Y gastric bypass, but guanylins do not play a significant role in body weight regulation and glycemic control

AimTo determine whether intestinal expression of guanylate cyclase activator 2A (GUCA2A) and guanylate cyclase activator 2B (GUCA2B) genes is regulated in obese humans following Roux-en-Y gastric bypass (RYGB), and to evaluate the corresponding guanylin (GN) and uroguanylin (UGN) peptides for potentially contributing to the beneficial metabolic effects of RYGB. MethodsEnteroendocrine cells were harvested peri- and post-RYGB, and GUCA2A/GUCA2B mRNA expression was compared. GN, UGN and their prohormones (proGN, proUGN) were administered subcutaneously in normal-weight mice to evaluate effects on food intake and glucose regulation. The effect of pro-UGN or UGN overexpression, using adeno-associated virus (AAV) vectors, was assessed in diet-induced obese (DIO) mice. Intracerebroventricular administration of GN and UGN was performed in rats for assessment of putative centrally mediated effects on food intake. GN and UGN, as well as their prohormones, were evaluated for effects on glucose-stimulated insulin secretion (GSIS) in rat pancreatic islets and perfused rat pancreas. ResultsGUCA2A and GUCA2B mRNA expression was significantly upregulated in enteroendocrine cells after RYGB. Peripheral administration of guanylins or prohormones did not influence food intake, oral glucose tolerance, and GSIS. Central administration of GN and UGN did not affect food intake in rats. Chronic AVV-mediated overexpression of UGN and proUGN had no effect on body weight or glucose homeostasis in DIO mice. ConclusionGN and UGN, as well as their prohormones, do not seem to play a significant role in body weight regulation and glycemic control, suggesting that guanylin-family peptides do not show promise as targets for the treatment of obesity or diabetes.

Exendin-4 protects mice from D-galactose-induced hepatic and pancreatic dysfunction.

ABSTRACTInvestigations into pharmaceutical intervention of pancreatic and hepatic dysfunction associated with metabolic disturbances have received relatively little attention. The aim of this study was to investigate the protective effects of exendin-4 in mice receiving D-galactose, a reducing sugar that triggers ROS production and inflammatory mediators affecting the pancreas and liver. Exendin-4 is an United States Food and Drug Administration (FDA) approved glucagon-like peptide that increases insulin dependent glycogen synthesis and glucose uptake. Male NMRI mice (20–25 g), 3 months of age, were randomly divided into 6 groups of 12 mice each: control, exendin-4 (1 nmol/kg), exendin-4 (10 nmol/kg), D-galactose, D-galactose + exendin-4 (1 nmol/kg) and D-galactose + exendin-4 (10 nmol/kg). D-galactose (500 mg/kg) was given daily by oral gavage for 6 weeks. During the last 10 days, exendin-4 (1 and 10 nmol/kg) was injected intraperitoneally daily. Glucose, insulin, insulin resistance, lipid profiles, and hepatic enzyme levels significantly increased in the D-galactose group (p < 0.05), along with a significant decrease in superoxide dismutase activity and pancreatic islet insulin secretion (p < 0.05). Exendin-4 decreased D-galactose-induced increases in serum glucose and insulin, insulin resistance, lipid profiles, and hepatic enzymes, and improved pancreatic islet insulin secretion and antioxidant defense status. The results show that exendin-4 can prevent complications in mice with compromised pancreatic and hepatic function. Long term administration of D-galactose in mice may be a useful model to study insulin resistance, metabolic syndrome, and aging.

Protective effect of metformin on toxicity of butyric acid and arsenic in isolated liver mitochondria and langerhans islets in male mice: an in vitro study.

Objective(s):Arsenic, a toxic metal in drinking water and butyric acid (BA) is a free fatty acid found in many foods. These two can induce oxidative stress in some tissues. The present study investigated the protective effect of metformin against toxicity induced by Arsenic (As) and BA in isolated mice liver mitochondria and pancreatic islets.Materials and Methods:In this study, liver mitochondria were isolated by adopting different centrifugation methods and pancreatic islets isolated by a collagenase method. Mitochondria were incubated by BA (75 μM), As (100 μM) and metformin (0, 0.5, 1, 3, 10 mM) and the islets also incubated by BA (1000 μM), As (100 μM) and metformin (0, 1, 3, 10 mM) for 1 hr. At the end of study, mitochondrial viability (MTT), mitochondrial membrane potential (MMP), reactive oxygen species (ROS), malondial- dehyde (MDA), glutathione (GSH) and islets insulin secretion were measured employing specific relevant methods.Results:As and BA significantly increased ROS, MDA and ΔΨm levels and decreased GSH level, succinate dehydrogenase activity and insulin secretion. On the other hand, pretreatment with metformin, returned mitochondrial complex II activity, reduced ROS, MDA and ΔΨm levels and increased GSH level and insulin secretion of pancreatic islets.Conclusion:As and BA in combination or in isolation induce oxidative stress in liver mitochondria and decrease insulin secretion of pancreatic islets. Metformin has a protective effect probably caused by its antioxidant feature. The findings suggest the potential role of metformin in mitochondria therapy and insulin secretion in many diseases.

Phenolic Substances from Ocimum Species Enhance Glucose-Stimulated Insulin Secretion and Modulate the Expression of Key Insulin Regulatory Genes in Mice Pancreatic Islets

Ocimum gratissimum and Ocimum basilicum are plants ethnopharmacologically used to treat diabetes mellitus, a life-threatening disease that affects millions of people worldwide. In order to further understand their antidiabetic potential, which has been previously demonstrated in animal models, we aimed to investigate the acute and chronic effects of major phenolic substances from both plants on insulin secretion and gene expression in pancreatic islets isolated from NMRI mice. Insulin secretion was measured after acute (1 h) and long-term (72 h) incubation of islets with one of four cinnamic acid derivatives (caftaric, caffeic, chicoric, and rosmarinic acids) or a C-glucosylated flavonoid (vicenin-2). All substances acutely enhanced glucose-stimulated insulin secretion (GSIS) from islets at concentrations from 10-10 to 10-6 M. They also increased GSIS after chronic incubation (10-8 M). None of them increased insulin secretion in the presence of low glucose concentration. Furthermore, these substances markedly changed the gene expression profile of key insulin regulatory genes INS1, INS2, PDX1, INSR, IRS1, and proliferative genes as well as glucose transporter 2 (GLUT2), in treated islets. Thus, they may play an important role in diabetes treatment. This is the first report on the insulin-secretory activity of caftaric acid, rosmarinic acid, and vicenin-2.

Sirolimus induces depletion of intracellular calcium stores and mitochondrial dysfunction in pancreatic beta cells

Sirolimus (rapamycin) is an immunosuppressive drug used in transplantation. One of its major side effects is the increased risk of diabetes mellitus; however, the exact mechanisms underlying such association have not been elucidated. Here we show that sirolimus impairs glucose-stimulated insulin secretion both in human and murine pancreatic islets and in clonal β cells in a dose- and time-dependent manner. Importantly, we demonstrate that sirolimus markedly depletes calcium (Ca2+) content in the endoplasmic reticulum and significantly decreases glucose-stimulated mitochondrial Ca2+ uptake. Crucially, the reduced mitochondrial Ca2+ uptake is mirrored by a significant impairment in mitochondrial respiration. Taken together, our findings indicate that sirolimus causes depletion of intracellular Ca2+ stores and alters mitochondrial fitness, eventually leading to decreased insulin release. Our results provide a novel molecular mechanism underlying the increased incidence of diabetes mellitus in patients treated with this drug.

220 - iPLA2γ Ablation Alters Glucose Homeostasis and Insulin Secretion in Response to Fatty Acids

Calcium-independent phospholipases (iPLA2s) are a family of enzymes participating in cellular signaling by simultaneously producing free fatty acids and lysophospholipids. Phospholipase iPLA2γ (PNPLA8) is targeted to mitochondria and has been shown to augment GPR40-dependent insulin secretion in β-cell model insulinoma INS1-E cells [1]. Here, we investigated the participation of iPLA2γ on the blood glucose homeostasis and pancreatic β-cell insulin secretion using iPLA2γ-KO mice. Wild type (wt) controls and iPLA2γ-KO mice showed similar cholesterol, triglyceride and basal glucose levels after 6 hours of starving. However, the uric acid levels were increased in iPLA2γ-KOmice. Following the intraperitoneal injection of Intralipid, iPLA2γ-KO mice exhibited a prolonged hyperglycemic state compared to wt mice. Glucose-stimulated insulin release in isolated pancreatic islets (PI) was moderately decreased in iPLA2γ-KO PI. Physiologically relevant concentrations of palmitic acid stimulated insulin secretion in PI from wt mice, and this stimulation was absent in iPLA2γ-KO PI. In conclusion, the data are consistent with iPLA2γ ablation causing impairment of GPR40-dependent insulin secretion, resulting in prolonged glycemia. Thus, our results support the role of iPLA2γ in regulating insulin secretion in vivo.

Tocotrienols Stimulate Insulin Secretion of Rat Pancreatic Isolated Islets in a Dynamic Culture.

Tocotrienols (T3) are the naturally occurring vitamin E derivatives that possess antioxidant properties and therapeutic potential in diabetic complications. The bioactivities of the derivatives are determined by the number and arrangement of methyl substitution on the structure. The objective of this study was to determine the effects of T3 derivatives, σ-T3, γ-T3 and α-T3 on insulin secretion of rat pancreatic islets in a dynamic culture. Pancreatic islets isolated from male Wistar rats were treated with T3 for 1 h at 37°C in a microfluidic system with continuous operation that provided a stable cell culture environment. Glucose (2.8 mM and 16.7 mM, as basal and stimulant, respectively) and potassium chloride (KCl) (30 mM) were added to the treatment in calcium free medium. The supernatant was collected for insulin measurements. Short-term exposure (1 h) of σ-T3 to β cells in the stimulant glucose condition significantly potentiated insulin secretion in a dose-dependent manner. γ-T3 and α-T3 also displayed dosedependent effect but were less effective in the activation of insulin secretion. Essentially, KCl, a pancreatic β cell membrane depolarizing agent, added into the treatment further enhanced the insulin secretion of σ-T3, γ-T3 and α-T3 with ED50 values of 504, 511 and 588 µM, respectively. The findings suggest the potential of σ-T3 in regulating glucose-stimulated insulin secretion (GSIS) in response to the intracellular calcium especially in the presence of KCl.

Transcriptomic and Functional Analyses on the Effects of Dioxin on Insulin Secretion of Pancreatic Islets and β-Cells.

In this study, transcriptomic and Ingenuity Pathway Analysis (IPA) underlined that an ex-vivo TCDD treatment (0.1 nM) stimulated insulin-release in mouse pancreatic islets via the effect on the Akt-mTOR-p70S6K, AMPK and ERK1/2 pathways. Functional studies using both ex-vivo islets and the mouse β-cell-line (Min-6) validated the stimulatory effects of TCDD (0.1 and 1 nM) on basal-insulin secretion. At 0.1 nM TCDD treatment on Min-6, Western blot analysis showed activation of ERK1/2 and decreased expression of pyruvate dehydrogenase kinase (PDK). A reduction of PDK expression is associated with an increase of pyruvate dehydrogenase flux. This observation was supported by the detection of significantly higher cellular ATP levels, an increase of glucose-stimulated-insulin-secretion (GSIS), and an inhibition of the AMPK pathway. At 1 nM TCDD treatment on Min-6, significant inhibitions of the Akt-mTOR pathway, cellular ATP production, and GSIS were evident. The experimental studies in Min-6 supported the IPA of transcriptomic data in pancreatic islets. Collectively, TCDD treatment caused an elevated basal-insulin release in both islets and β-cell cultures. Moreover, our data revealed that the modulation of the Akt-mTOR-p70S6K, AMPK and ERK1/2 pathways might be an important component of the mechanism for the TCDD-perturbing effects on ATP production in β-cells in affecting insulin secretion.

Glucose-stimulated insulin release: Parallel perifusion studies of free and hydrogel encapsulated human pancreatic islets.

To explore the effects immune-isolating encapsulation has on the insulin secretion of pancreatic islets and to improve our ability to quantitatively describe the glucose-stimulated insulin release (GSIR) of pancreatic islets, we conducted dynamic perifusion experiments with isolated human islets. Free (unencapsulated) and hydrogel encapsulated islets were perifused, in parallel, using an automated multi-channel system that allows sample collection with high temporal resolution. Results indicated that free human islets secrete less insulin per unit mass or islet equivalent (IEQ) than murine islets and with a less pronounced first-phase peak. While small microcapsules (d = 700 µm) caused only a slightly delayed and blunted first-phase insulin response compared to unencapsulated islets, larger capsules (d = 1,800 µm) completely blunted the first-phase peak and decreased the total amount of insulin released. Experimentally obtained insulin time-profiles were fitted with our complex insulin secretion computational model. This allowed further fine-tuning of the hormone-release parameters of this model, which was implemented in COMSOL Multiphysics to couple hormone secretion and nutrient consumption kinetics with diffusive and convective transport. The results of these GSIR experiments, which were also supported by computational modeling, indicate that larger capsules unavoidably lead to dampening of the first-phase insulin response and to a sustained-release type insulin secretion that can only slowly respond to changes in glucose concentration. Bioartificial pancreas type devices can provide long-term and physiologically desirable solutions only if immunoisolation and biocompatibility considerations are integrated with optimized nutrient diffusion and insulin release characteristics by design.

ID: 1079 Improved glucose homeostasis effect of conditioned media from adipose-derived stem cells in type 1 diabetic mice

Background: Many studies suggested adipose derived stem cell (ASC) transplantation as a new approach to control hyperglycemia in type 1 diabetes mellitus. It is proposed that the effects of these cells could be not only based on the direct cell-cell interaction but also the secretion of cytokines. This study aimed to demonstrate the effect of adipose stem cell-derived conditioned medium (CM) on the treatment of STZ-induced diabetic mice.&#x0D; Methods: CM was obtained from 24-hours-cultured medium of ASCs and centrifuged to remove the debris. Type 1 diabetic mice were intraperitoneally injected CM for 30 consecutive days. Therapeutic efficacy of CM was assessed by survival rate, blood glucose level, serum insulin level, histological structure of pancreatic islets.&#x0D; Results: The results showed that CM treatment could decrease mortality rate (from 33,33% to 0%) as well as blood glucose level (from 425,667±65,753 mg/dl to 203,500 mg ±20,350 mg/dl) and enhance insulin secretion, improve size and function of pancreatic islets of diabetic mice.&#x0D; Conclusion: Conditioned medium maybe a promising therapy for type 1 diabetes mellitus.

S-Equol Activates cAMP Signaling at the Plasma Membrane of INS-1 Pancreatic β-Cells and Protects against Streptozotocin-Induced Hyperglycemia by Increasing β-Cell Function in Male Mice

Background:S-equol, which is enantioselectively produced from daidzein by gut microbiota, has been suggested as a chemopreventive agent against type 2 diabetes mellitus (T2DM), but the underlying mechanisms remain unclear.Objective: We investigated the effects of S-equol on pancreatic β-cell function.Methods: β-Cell growth and insulin secretion were evaluated with male Institute of Cancer Research mice and isolated pancreatic islets from the mice, respectively. The mechanisms by which S-equol stimulated β-cell response were examined in INS-1 β-cells. The effect of S-equol treatment on β-cell function was assessed in low-dose streptozotocin-treated mice. S-equol was used at 10 μmol/L for in vitro and ex vivo studies and was administered by oral gavage (20 mg/kg, 2 times/d throughout the experimental period) for in vivo studies.Results:S-equol administration for 7 d increased Ki67-positive β-cells by 27% (P < 0.01) in mice. S-equol enantioselectively enhanced glucose-stimulated insulin secretion in mouse pancreatic islets by 41% (P < 0.001). In INS-1 cells, S-equol exerted stronger effects than daidzein on cell growth, insulin secretion, and cAMP-response element (CRE)-mediated transcription. These S-equol effects were diminished by inhibiting protein kinase A. The effective concentration of S-equol for stimulating cAMP production at the plasma membrane was lower than that for phosphodiesterase inhibition. S-equol-stimulated CRE activation was negatively controlled by the knockdown of G-protein α subunit group S (stimulatory) and positively controlled by that of G-protein-coupled receptor kinase-3 and -6. Compared with vehicle-treated controls, S-equol gavage treatment resulted in an increase in β-cell mass of 104% (P < 0.05), a trend toward high plasma insulin concentrations (by 118%; P = 0.06), and resistance to hyperglycemia after streptozotocin treatment (78% of AUC after glucose challenge; P < 0.01). S-equol administration significantly increased the number of Ki67-positive proliferating β-cells by 62% (P < 0.01) and decreased that of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive apoptotic β-cells by 75% (P < 0.05).Conclusions: Our results show that S-equol boosts β-cell function and prevents hypoglycemia in mice, suggesting its potential for T2DM prevention.

Ultrastructural analysis of insulin secretory granule biology by super resolution and transmission electron microscopy

Insulin secretion in pancreatic islets occurs in two phases: a short and rapid first phase followed by a sustained second phase, with younger insulin secretoy granules (SGs) being preferentially poised for exocytosis relative to older SGs. Here we labeled insulin SGs of different age in beta cells of isolated pancreatic islets from SOFIA (Study of insulin aging) mice, in which an insulin2-SNAP allele had been knocked-in in the Ins2 locus. Combining structured illumination microscopy and transmission electron microscopy (TEM) for correlative light-electron microscopy (CLEM) we precisely tracked age-distinct insulin SGs in Tokuyasu sections of beta cells. Ultrastructural analysis followed by mathematical modeling revealed a significant decrease in the number of labeled SGs starting from 2.7 days of age with a reduction of ˜60% at an SG age of 5 days, indicating a half life of 4.6 days. The reduction was accompanied by the detection of SNAP-labeled older SGs been disposed in multigranular bodies. Furthermore, we could show for the first time the preservation of fluorescence directly associated with protein tags in epoxy resin. This allowed for CLEM of age-distinct insulin SGs in SOFIA islets fixed by high pressure freezing (HPF) and embedded in epoxy resin, providing optimal ultrastructural preservation and contrast. Exploitation of this approach for morphometric analysis of labeled and unlabeled SGs will provide quantitative information about the life cycle of insulin SGs in different metabolic conditions. CLEM in Epon sections after HPF will allow for investigating the interaction of insulin SGs with the cytoskeleton at nanometer-resolution.