Insulin Secretion In Pancreatic Β-cells Research Articles

Saponins of Momordica charantia increase insulin secretion in INS-1 pancreatic β-cells via the PI3K/Akt/FoxO1 signaling pathway

Saponins are the main bioactive substances with anti-hyperglycemic activities of Momordica charantia. This study aimed to verify the effects of M. charantia saponins on insulin secretion and explore the potential underlying mechanisms in INS-1 pancreatic β-cells. We injured INS-1 cells with 33.3mM glucose and then treated them with saponins. Saponins improved cell morphology and viability as demonstrated by inverted microscopy and CCK8 detection and significantly increased insulin secretion in a concentration-dependent manner as shown by ELISA. Thus, we obtained the optimal concentration for the subsequent experiments. Potential mechanisms were explored by immunofluorescence, western blotting, and RT-qPCR techniques. First, saponins increased the mRNA and protein levels of IRS-2 but decreased the serine 731 phosphorylation level of IRS-2. Moreover, saponins increased the phosphorylation of Akt protein and decreased the protein level of FoxO1, which were both reversed by the PI3K inhibitor ly294002. Furthermore, saponins increased the protein level of the downstream molecule and insulin initiating factor PDX-1, which was also reversed by ly294002. Saponins also increased Akt and PDX-1 mRNA and decreased FoxO1 mRNA, which were both reversed by ly294002. Saponins increased glucose-stimulated insulin secretion (GSIS) and intracellular insulin content, which were reversed by ly294002, as determined by ELISA. The immunofluorescence results also confirmed this tendency. In conclusion, our findings improve our understanding of the function of saponins in INS-1 pancreatic β-cells and suggest that saponins may increase insulin secretion via the PI3K/Akt/FoxO1 signaling pathway.

Effects of Polyphenols on Insulin Resistance.

Insulin resistance (IR) is apparent when tissues responsible for clearing glucose from the blood, such as adipose and muscle, do not respond properly to appropriate signals. IR is estimated based on fasting blood glucose and insulin, but some measures also incorporate an oral glucose challenge. Certain (poly)phenols, as supplements or in foods, can improve insulin resistance by several mechanisms including lowering postprandial glucose, modulating glucose transport, affecting insulin signalling pathways, and by protecting against damage to insulin-secreting pancreatic β-cells. As shown by intervention studies on volunteers, the most promising candidates for improving insulin resistance are (−)-epicatechin, (−)-epicatechin-containing foods and anthocyanins. It is possible that quercetin and phenolic acids may also be active, but data from intervention studies are mixed. Longer term and especially dose-response studies on mildly insulin resistant participants are required to establish the extent to which (poly)phenols and (poly)phenol-rich foods may improve insulin resistance in compromised groups.

Elevated Circulating LINC-P21 Serves as a Diagnostic Biomarker of Type 2 Diabetes Mellitus and Regulates Pancreatic β-cell Function by Sponging miR-766-3p to Upregulate NR3C2.

The purpose of this study was to evaluate the clinical value and biological function of long non-coding RNA (lncRNA) LINC-P21 in type 2 diabetes mellitus (T2DM), and explore the underlying mechanisms. The expression of LINC-P21 was estimated using quantitative real-time PCR. The functional role of LINC-P21 was explored by gain- and loss-of-function experiments. INS-1 cell proliferation was analyzed using a cell counting kit-8 (CCK-8)assay, and the glucose-stimulated insulin secretion was measured using an ELISA kit. The miRNAs that might be sponged by LINC-P21 were analyzed, and the subsequent target genes were predicted and assessed in INS-1 cells. Serum expression of LINC-P21 was elevated in T2DM patients, which was correlated with fasting blood glucose levels and disease diagnosis. The glucose-stimulated insulin secretion and the proliferation of INS-1 cells were enhanced by LINC-P21 knockdown, but the overexpression of LINC-P21 led to opposite effects. miR-766-3p could be directly inhibited by LINC-P21 in INS-1 cells and reverse the effects of LINC-P21 on β-cell function. Additionally, NR3C2 was determined as a target of miR-766-3p, which could be positively regulated by LINC-P21 and had same effects with LINC-P21 on INS-1 cell proliferation and insulin secretion. All the data demonstrated that serum elevated LINC-P21 and decreased miR-766-3p serve as candidate diagnostic biomarkers in T2DM patients. LINC-P21 acts as a potential regulator in insulin secretion and proliferation of pancreatic β-cells through targeting miR-766-3p to upregulate NR3C2.

Tetraspanin-7 regulation of L-type voltage-dependent calcium channels controls pancreatic β-cell insulin secretion.

Tetraspanin (TSPAN) proteins regulate many biological processes, including intracellular calcium (Ca2+ ) handling. TSPAN-7 is enriched in pancreatic islet cells; however, the function of islet TSPAN-7 has not been identified. Here, we characterize how β-cell TSPAN-7 regulates Ca2+ handling and hormone secretion. We find that TSPAN-7 reduces β-cell glucose-stimulated Ca2+ entry, slows Ca2+ oscillation frequency and decreases glucose-stimulated insulin secretion. TSPAN-7 controls β-cell function through a direct interaction with L-type voltage-dependent Ca2+ channels (CaV 1.2 and CaV 1.3), which reduces channel Ca2+ conductance. TSPAN-7 slows activation of CaV 1.2 and accelerates recovery from voltage-dependent inactivation; TSPAN-7 also slows CaV 1.3 inactivation kinetics. These findings strongly implicate TSPAN-7 as a key regulator in determining the set-point of glucose-stimulated Ca2+ influx and insulin secretion. Glucose-stimulated insulin secretion (GSIS) is regulated by calcium (Ca2+ ) entry into pancreatic β-cells through voltage-dependent Ca2+ (CaV ) channels. Tetraspanin (TSPAN) transmembrane proteins control Ca2+ handling, and thus they may also modulate GSIS. TSPAN-7 is the most abundant islet TSPAN and immunostaining of mouse and human pancreatic slices shows that TSPAN-7 is highly expressed in β- and α-cells; however, the function of islet TSPAN-7 has not been determined. Here, we show that TSPAN-7 knockdown (KD) increases glucose-stimulated Ca2+ influx into mouse and human β-cells. Additionally, mouse β-cell Ca2+ oscillation frequency was accelerated by TSPAN-7 KD. Because TSPAN-7 KD also enhanced Ca2+ entry when membrane potential was clamped with depolarization, the effect of TSPAN-7 on CaV channel activity was examined. TSPAN-7 KD enhanced L-type CaV currents in mouse and human β-cells. Conversely, heterologous expression of TSPAN-7 with CaV 1.2 and CaV 1.3 L-type CaV channels decreased CaV currents and reduced Ca2+ influx through both channels. This was presumably the result of a direct interaction of TSPAN-7 and L-type CaV channels because TSPAN-7 coimmunoprecipitated with both CaV 1.2 and CaV 1.3 from primary human β-cells and from a heterologous expression system. Finally, TSPAN-7 KD in human β-cells increased basal (5.6 mM glucose) and stimulated (45 mM KCl + 14 mM glucose) insulin secretion. These findings strongly suggest that TSPAN-7 modulation of β-cell L-type CaV channels is a key determinant of β-cell glucose-stimulated Ca2+ entry and thus the set-point of GSIS.

Leonurine protects against dexamethasone-induced cytotoxicity in pancreatic β-cells via PI3K/Akt signaling pathway

Glucocorticoid excess induces pancreatic β-cell apoptosis and insulin secretion impairment, which may lead to hyperglycemia and steroid diabetes. Leonurine is a natural alkaloid extracted from the Herba leonuri, which has been widely used in the treatment of obstetric and gynecological diseases. However, whether leonurine performs a protective role in pancreatic β-cells remains unknown. In this study, we evaluated the effect of leonurine on dexamethasone -treated β-cells. Our data showed that leonurine inhibited dexamethasone-induced INS-1 cell apoptosis and facilitated cell proliferation. Moreover, leonurine attenuated dexamethasone-impaired insulin secretion in mice islets. Leonurine ameliorated dexamethasone-induced dephosphorylation of Akt, Bad and GSK-3β. Importantly, the protective role of leonurine on dexamethasone-induced cytotoxicity was blocked by LY294002 in INS-1 cells. Our findings revealed for the first time that leonurine could protect against dexamethasone-induced cytotoxicity in pancreatic β-cells via PI3K/Akt signaling pathway, suggesting leonurine may be a promising therapeutic agent for steroid diabetes.

Protein Kinase CK2 Controls CaV2.1-Dependent Calcium Currents and Insulin Release in Pancreatic β-Cells.

The regulation of insulin biosynthesis and secretion in pancreatic β-cells is essential for glucose homeostasis in humans. Previous findings point to the highly conserved, ubiquitously expressed serine/threonine kinase CK2 as having a negative regulatory impact on this regulation. In the cell culture model of rat pancreatic β-cells INS-1, insulin secretion is enhanced after CK2 inhibition. This enhancement is preceded by a rise in the cytosolic Ca2+ concentration. Here, we identified the serine residues S2362 and S2364 of the voltage-dependent calcium channel CaV2.1 as targets of CK2 phosphorylation. Furthermore, co-immunoprecipitation experiments revealed that CaV2.1 binds to CK2 in vitro and in vivo. CaV2.1 knockdown experiments showed that the increase in the intracellular Ca2+ concentration, followed by an enhanced insulin secretion upon CK2 inhibition, is due to a Ca2+ influx through CaV2.1 channels. In summary, our results point to a modulating role of CK2 in the CaV2.1-mediated exocytosis of insulin.

NUPR1 preserves insulin secretion of pancreatic β-cells during inflammatory stress by multiple low-dose streptozotocin and high-fat diet.

Obesity is associated with dyslipidemia and subclinical inflammation that promotes metabolic disturbances including insulin resistance and pancreatic β-cell dysfunction. The nuclear protein, transcriptional regulator 1 (NUPR1) responds to cellular stresses and features tissue protective properties. To characterize the role of NUPR1 in endocrine pancreatic islets during inflammatory stress, we generated transgenic mice with β-cell-specific Nupr1 overexpression (βNUPR1). Under normal conditions, βNUPR1 mice did not differ from wild type (WT) littermates and display normal glucose homeostasis and β-cell mass. For induction of inflammatory conditions, mice were treated with multiple low-dose streptozotocin (mld-STZ) and/or fed a high-fat diet (HFD). All treatments significantly worsened glycaemia in WT mice, while βNUPR1 mice substantially preserved insulin secretion and glucose tolerance. HFD increased β-cell mass in all animals, with βNUPR1 mice tending to show higher values. The improved outcome of βNUPR1 mice was accompanied by decreased NF-κB activation and lymphocyte infiltration in response to mld-STZ. In vitro, isolated βNUPR1 islets preserved insulin secretion and content with insignificantly low apoptosis during culture stress and IL-1β exposure. These findings suggest that NUPR1 plays a vital role in the protection of β-cells from apoptosis, related degradation of insulin storages and subsequent secretion during inflammatory and obesity-related tissue stress.

Costus pictus D. Don leaf extract stimulates GLP-1 secretion from GLUTag L-cells and has cytoprotective effects in BRIN-BD11 β-cells

Costus pictus D. Don, commonly known as insulin plant, is a traditional Indian antidiabetic herbal medicine with glucose-lowering and insulin secretory effects having been reported in animal models and humans with Type 2 diabetes. However, its effects on GLP-1 secretion from intestinal endocrine L-cells and potential metabolic and protective effects in insulin secreting pancreatic β-cells are not yet fully understood. This study is aimed to elucidate the effects of Costus pictus D. Don leaf extract (CPE) on L-cell function and GLP-1 secretion using the established murine GLUTag L-cell model and to investigate its potential cytoprotective effects against detrimental effects of palmitate and cytokines in pancreatic β-cells using BRIN-BD11cells. Costus pictus D. Don dried leaf powder was extracted by soxhlet method. Cell viability was determined by MTT assay. Changes in gene and protein expression were quantified by qPCR and western blotting, respectively. GLP-1 and insulin secretion were measured by ELISA. CPE significantly enhanced the percentage of viable BRIN-BD11 and GLUTag cells and protected BRIN-BD11cells against palmitate- and proinflammatory cytokine-induced toxicity. CPE enhanced acute GLP-1 secretion 6.4-16.3-fold from GLUTag cells at both low (1.1mM) and high (16.7mM) glucose (P<0.01) concentrations. Antioxidant (Nrf2, Cat & Gpx1) and pro-proliferative (Erk1 and Jnk1) gene expression were upregulated by 24h culture with CPE, while proinflammatory transcription factor NF-κB was downregulated. Diminished postprandial GLP-1 secretion and loss of insulin secreting β-cells are known contributors of T2DM. Our data suggests that CPE acutely stimulates GLP-1 secretion from L-cells. Long term exposure of the BRIN-BD11cells to CPE enhances cell number and may protect against palmitate and proinflammatory cytokines by activating multiple pathways. Thus, the current study suggests that the possible antidiabetic properties of CPE may be linked to enhanced GLP-1 secretion and β-cell protection which could be beneficial in the management of T2DM.

Coloured Rice Phenolic Extracts Increase Expression of Genes Associated with Insulin Secretion in Rat Pancreatic Insulinoma β-cells.

Glucose-induced oxidative stress is associated with the overproduction of reactive oxygen species (ROS), which may dysregulate the expression of genes controlling insulin secretion leading to β-cell dysfunction, a hallmark of type 2 diabetes mellitus (T2DM). This study investigated the impact of coloured rice phenolic extracts (CRPEs) on the expression of key genes associated with β-cell function in pancreatic β-cells (INS-1E). These genes included glucose transporter 2 (Glut2), silent mating type information regulation 2 homolog 1 (Sirt1), mitochondrial transcription factor A (Tfam), pancreatic/duodenal homeobox protein 1 (Pdx-1) and insulin 1 (Ins1). INS-1E cells were cultured in high glucose (25 mM) to induce glucotoxic stress conditions (HGSC) and in normal glucose conditions (NGC-11.1 mM) to represent normal β-cell function. Cells were treated with CRPEs derived from two coloured rice cultivars, Purple and Yunlu29-red varieties at concentrations ranged from 50 to 250 µg/mL. CRPEs upregulated the expression of Glut2, Sirt1 and Pdx-1 significantly at 250 µg/mL under HGSC. CRPEs from both cultivars also upregulated Glut2, Sirt1, Tfam, Pdx-1 and Ins1 markedly at 250 µg/mL under NGC with Yunlu29 having the greatest effect. These data suggest that CRPEs may reduce β-cell dysfunction in T2DM by upregulating the expression of genes involved in insulin secretion pathways.

Muscadine or amla extracts standardized to ellagic acid content ameliorate glucolipotoxicity associated \u03b2-cell dysfunction via inhibition of IL-1\u03b2 and improved insulin secretion

Glucolipotocixity induces IL-1 β secretion which impairs pancreatic β-cell insulin secretion. Ellagic acid and urolithin A have strong anti-inflammatory effect on cells. Muscadine and amla are very good sources of ellagic acid. The present study examined the effect of ellagic acid, ellagic acid-rich muscadine or amla extract, or urolothin A on inflammation in β cells under glucolipotoxic conditions. Rat NIT-1 β cells were incubated in glucolipotoxic conditions (33.3 mM glucose, 250 μM palmitic acid or 33.3 mM glucose + 250 μM palmitic acid with or without ellagic acid, ellagic acid-rich muscadine or amla extracts standardized to its ellagic acid content, or urolithin A). Inflammatory status was evidenced by ELISA analysis of insulin and IL-1β secretion. Ellagic acid-rich muscadine or amla extracts dose-dependently stimulated insulin secretion and down-regulated IL-1β better than pure ellagic acid, or urolithin A. Urolithin A did not statistically stimulate insulin secretion and did not inhibit IL-1β.

Investigation of Ultrasound-Mediated Intracellular Ca2+ Oscillations in HIT-T15 Pancreatic β-Cell Line.

In glucose-stimulated insulin secretion (GSIS) of pancreatic β-cells, the rise of free cytosolic Ca2+ concentration through voltage-gated calcium channels (VGCCs) triggers the exocytosis of insulin-containing granules. Recently, mechanically induced insulin secretion pathways were also reported, which utilize free cytosolic Ca2+ ions as a direct regulator of exocytosis. In this study, we aimed to investigate intracellular Ca2+ responses on the HIT-T15 pancreatic β-cell line upon low-intensity pulsed ultrasound (LIPUS) stimulation and found that ultrasound induces two distinct types of intracellular Ca2+ oscillation, fast-irregular and slow-periodic, from otherwise resting cells. Both Ca2+ patterns depend on the purinergic signaling activated by the rise of extracellular ATP or ADP concentration upon ultrasound stimulation, which facilitates the release through mechanosensitive hemichannels on the plasma membrane. Further study demonstrated that two subtypes of purinergic receptors, P2X and P2Y, are working in a competitive manner depending on the level of glucose in the cell media. The findings can serve as an essential groundwork providing an underlying mechanism for the development of a new therapeutic approach for diabetic conditions with further validation.

Placental Lactogen as a Marker of Maternal Obesity, Diabetes, and Fetal Growth Abnormalities: Current Knowledge and Clinical Perspectives.

Placental lactogen (PL) is a peptide hormone secreted throughout pregnancy by both animal and human specialized endocrine cells. PL plays an important role in the regulation of insulin secretion in pancreatic β-cells, stimulating their proliferation and promoting the expression of anti-apoptotic proteins. Cases of pregnancy affected by metabolic conditions, including obesity and diabetes, are related to alterations in the PL secretion pattern. Whereas obesity is most often associated with lower PL serum concentrations, diabetes results in increased PL blood levels. Disruptions in PL secretion are thought to be associated with an increased prevalence of gestational complications, such as placental dysfunction, diabetic retinopathy, and abnormalities in fetal growth. PL is believed to be positively correlated with birth weight. The impaired regulation of PL secretion could contribute to an increased incidence of both growth retardation and fetal macrosomia. Moreover, the dysregulation of PL production during the intrauterine period could affect the metabolic status in adulthood. PL concentration measurement could be useful in the prediction of fetal macrosomia in women with normal oral glucose tolerance test (OGTT) results or in evaluating the risk of fetal growth restriction, but its application in standard clinical practice seems to be limited in the era of ultrasonography.

Exposure to inorganic arsenic and its methylated metabolites alters metabolomics profiles in INS-1 832/13 insulinoma cells and isolated pancreatic islets.

Inorganic arsenic (iAs) is an environmental diabetogen, but mechanisms underlying its diabetogenic effects are poorly understood. Exposures to arsenite (iAsIII) and its methylated metabolites, methylarsonite (MAsIII) and dimethylarsinite (DMAsIII), have been shown to inhibit glucose-stimulated insulin secretion (GSIS) in pancreatic β-cells and isolated pancreatic islets. GSIS is regulated by complex mechanisms. Increase in ATP production through metabolism of glucose and other substrates is the ultimate trigger for GSIS in β-cells. In the present study, we used metabolomics to identify metabolites and pathways perturbed in cultured INS-1 832/13 rat insulinoma cells and isolated murine pancreatic islets by exposures to iAsIII, MAsIII and DMAsIII. We found that the exposures perturbed multiple metabolites, which were enriched primarily in the pathways of amino acid, carbohydrate, phospholipid and carnitine metabolism. However, the effects of arsenicals in INS-1 832/13 cells differed from those in the islets and were exposure specific with very few overlaps between the three arsenicals. In INS-1 832/13 cells, all three arsenicals decreased succinate, a metabolite of Krebs cycle, which provides substrates for ATP synthesis in mitochondria. Acetylcarnitine was decreased consistently by exposures to arsenicals in both the cells and the islets. Acetylcarnitine is usually found in equilibrium with acetyl-CoA, which is the central metabolite in the catabolism of macronutrients and the key substrate for Krebs cycle. It is also thought to play an antioxidant function in mitochondria. Thus, while each of the three trivalent arsenicals perturbed specific metabolic pathways, which may or may not be associated with GSIS, all three arsenicals appeared to impair mechanisms that support ATP production or antioxidant defense in mitochondria. These results suggest that impaired ATP production and/or mitochondrial dysfunction caused by oxidative stress may be the mechanisms underlying the inhibition of GSIS in β-cells exposed to trivalent arsenicals.

Loss of the voltage-gated proton channel Hv1 decreases insulin secretion and leads to hyperglycemia and glucose intolerance in mice

Insulin secretion by pancreatic islet β-cells is regulated by glucose levels and is accompanied by proton generation. The voltage-gated proton channel Hv1 is present in pancreatic β-cells and extremely selective for protons. However, whether Hv1 is involved in insulin secretion is unclear. Here we demonstrate that Hv1 promotes insulin secretion of pancreatic β-cells and glucose homeostasis. Hv1-deficient mice displayed hyperglycemia and glucose intolerance because of reduced insulin secretion but retained normal peripheral insulin sensitivity. Moreover, Hv1 loss contributed much more to severe glucose intolerance as the mice got older. Islets of Hv1-deficient and heterozygous mice were markedly deficient in glucose- and K+-induced insulin secretion. In perifusion assays, Hv1 deletion dramatically reduced the first and second phase of glucose-stimulated insulin secretion. Islet insulin and proinsulin content was reduced, and histological analysis of pancreas slices revealed an accompanying modest reduction of β-cell mass in Hv1 knockout mice. EM observations also indicated a reduction in insulin granule size, but not granule number or granule docking, in Hv1-deficient mice. Mechanistically, Hv1 loss limited the capacity for glucose-induced membrane depolarization, accompanied by a reduced ability of glucose to raise Ca2+ levels in islets, as evidenced by decreased durations of individual calcium oscillations. Moreover, Hv1 expression was significantly reduced in pancreatic β-cells from streptozotocin-induced diabetic mice, indicating that Hv1 deficiency is associated with β-cell dysfunction and diabetes. We conclude that Hv1 regulates insulin secretion and glucose homeostasis through a mechanism that depends on intracellular Ca2+ levels and membrane depolarization.

Characterization of the taste receptor-related G-protein, α-gustducin, in pancreatic β-cells.

Aims/IntroductionTaste receptors, T1rs and T2rs, and the taste‐selective G‐protein, α‐gustducin, are expressed outside the taste‐sensing system, such as enteroendocrine L cells. Here, we examined whether α‐gustducin also affects nutrition sensing and insulin secretion by pancreatic β‐cells.Materials and MethodsThe expression of α‐gustducin and taste receptors was evaluated in β‐cell lines, and in rat and mouse islets either by quantitative polymerase chain reaction or fluorescence immunostaining. The effects of α‐gustducin knockdown on insulin secretion and on cyclic adenosine monophosphate and intracellular Ca2+ levels in rat INS‐1 cells were estimated. Sucralose (taste receptor agonist)‐induced insulin secretion was investigated in INS‐1 cells with α‐gustducin suppression and in islets from mouse disease models.ResultsThe expression of Tas1r3 and α‐gustducin was confirmed in β‐cell lines and pancreatic islets. Basal levels of cyclic adenosine monophosphate, intracellular calcium and insulin secretion were significantly enhanced with α‐gustducin knockdown in INS‐1 cells. The expression of α‐gustducin was decreased in high‐fat diet‐fed mice and in diabetic db/db mice. Sucralose‐induced insulin secretion was not attenuated in INS‐1 cells with α‐gustducin knockdown or in mouse islets with decreased expression of α‐gustducin.Conclusionsα‐Gustducin is involved in the regulation of cyclic adenosine monophosphate, intracellular calcium levels and insulin secretion in pancreatic β‐cells in a manner independent of taste receptor signaling. α‐Gustducin might play a novel role in β‐cell physiology and the development of type 2 diabetes.

Chronic stimulation induces adaptive potassium channel activity that restores calcium oscillations in pancreatic islets in vitro.

Insulin pulsatility is important to hepatic response in regulating blood glucose. Growing evidence suggests that insulin-secreting pancreatic β-cells can adapt to chronic disruptions of pulsatility to rescue this physiologically important behavior. We determined the time scale for adaptation and examined potential ion channels underlying it. We induced the adaptation both by chronic application of the ATP-sensitive K+ [K(ATP)] channel blocker tolbutamide and by application of the depolarizing agent potassium chloride (KCl). Acute application of tolbutamide without pretreatment results in elevated Ca2+ as measured by fura-2AM and the loss of endogenous pulsatility. We show that after chronic exposure to tolbutamide (12-24 h), Ca2+ oscillations occur with subsequent acute tolbutamide application. The same experiment was conducted with potassium chloride (KCl) to directly depolarize the β-cells. Once again, following chronic exposure to the cell stimulator, the islets produced Ca2+ oscillations when subsequently exposed to tolbutamide. These experiments suggest that it is the chronic stimulation, and not tolbutamide desensitization, that is responsible for the adaptation that rescues oscillatory β-cell activity. This compensatory response also causes islet glucose sensitivity to shift rightward following chronic tolbutamide treatment. Mathematical modeling shows that a small increase in the number of K(ATP) channels in the membrane is one adaptation mechanism that is compatible with the data. To examine other compensatory mechanisms, pharmacological studies provide support that Kir2.1 and TEA-sensitive channels play some role. Overall, this investigation demonstrates β-cell adaptability to overstimulation, which is likely an important mechanism for maintaining glucose homeostasis in the face of chronic stimulation.

Klf6 protects β-cells against insulin resistance-induced dedifferentiation

ObjectivesIn the pathogenesis of type 2 diabetes, development of insulin resistance triggers an increase in pancreatic β-cell insulin secretion capacity and β-cell number. Failure of this compensatory mechanism is caused by a dedifferentiation of β-cells, which leads to insufficient insulin secretion and diabetic hyperglycemia. The β-cell factors that normally protect against dedifferentiation remain poorly defined. Here, through a systems biology approach, we identify the transcription factor Klf6 as a regulator of β-cell adaptation to metabolic stress. MethodsWe used a β-cell specific Klf6 knockout mouse model to investigate whether Klf6 may be a potential regulator of β-cell adaptation to a metabolic stress. ResultsWe show that inactivation of Klf6 in β-cells blunts their proliferation induced by the insulin resistance of pregnancy, high-fat high-sucrose feeding, and insulin receptor antagonism. Transcriptomic analysis showed that Klf6 controls the expression of β-cell proliferation genes and, in the presence of insulin resistance, it prevents the down-expression of genes controlling mature β-cell identity and the induction of disallowed genes that impair insulin secretion. Its expression also limits the transdifferentiation of β-cells into α-cells. ConclusionOur study identifies a new transcription factor that protects β-cells against dedifferentiation, and which may be targeted to prevent diabetes development.

Geniposide Balances the Redox Signaling to Mediate Glucose-Stimulated Insulin Secretion in Pancreatic β-Cells.

PurposeTo investigate the effect of geniposide on the biosynthesis of insulin and the expression protein disulfide isomerase (PDI) and endoplasmic reticulum oxidoreductin 1 (ERO1) in the presence of low (5 mM) and high (25 mM) glucose in pancreatic β cells.MethodsThe content of insulin was measured by ELISA, the number of SH groups was determined with the classical chromogenic reagent, 5,5ʹ-dithiobis-(2-nitrobenzoic) acid (DTNB; also known as Ellman’s reagent), the expressions of PDI and ERO1 were analyzed by Western blot.ResultsGeniposide played contrary roles on the accumulation of H2O2, the ratio of GSH/GSSG and the thiol–disulfide balance in the presence of low (5 mM) and high (25 mM) glucose in rat pancreatic INS-1 cells. Geniposide also regulated the protein levels of protein disulfide isomerase (PDI) and endoplasmic reticulum oxidoreductin1 (ERO1), the two key enzymes for the production of H2O2 during the biosynthesis of insulin in INS-1 cells.ConclusionGeniposide affects glucose-stimulated insulin secretion by modulating the thiol–disulfide balance that is controlled by the redox signaling in pancreatic β cells.

Ablating the Rab-GTPase activating protein TBC1D1 predisposes rats to high-fat diet-induced cardiomyopathy.

Although the role of TBC1D1 within the heart remains unknown, expression of TBC1D1 increases in the left ventricle following an acute infarction, suggesting a biological importance within this tissue. We investigated the mechanistic role of TBC1D1 within the heart, aiming to establish the consequences of attenuating TBC1D1 signalling in the development of diabetic cardiomyopathy, as well as to determine potential sex differences. TBC1D1 ablation increased plasma membrane fatty acid binding protein content and myocardial palmitate oxidation. Following high-fat feeding, TBC1D1 ablation dramatically increased fibrosis and induced end-diastolic dysfunction in both male and female rats in the absence of changes in mitochondrial bioenergetics. Altogether, independent of sex, ablating TBC1D1 predisposes the left ventricle to pathological remodelling following high-fat feeding, and suggests TBC1D1 protects against diabetic cardiomyopathy. TBC1D1, a Rab-GTPase activating protein, is involved in the regulation of glucose handling and substrate metabolism within skeletal muscle, and is essential for maintaining pancreatic β-cell mass and insulin secretion. However, the function of TBC1D1 within the heart is largely unknown. Therefore, we examined the role of TBC1D1 in the left ventricle and the functional consequence of ablating TBC1D1 on the susceptibility to high-fat diet-induced abnormalities. Since mutations within TBC1D1 (R125W) display stronger associations with clinical parameters in women, we further examined possible sex differences in the predisposition to diabetic cardiomyopathy. In control-fed animals, TBC1D1 ablation did not alter insulin-stimulated glucose uptake, or echocardiogram parameters, but increased accumulation of a plasma membrane fatty acid transporter and the capacity for palmitate oxidation. When challenged with an 8week high-fat diet, TBC1D1 knockout rats displayed a four-fold increase in fibrosis compared to wild-type animals, and this was associated with diastolic dysfunction, suggesting a predisposition to diet-induced cardiomyopathy. Interestingly, high-fat feeding only induced cardiac hypertrophy in male TBC1D1 knockout animals, implicating a possible sex difference. Mitochondrial respiratory capacity and substrate sensitivity to pyruvate and ADP were not altered by diet or TBC1D1 ablation, nor were markers of oxidative stress, or indices of overt heart failure. Altogether, independent of sex, ablation of TBC1D1 not only increased the susceptibility to high-fat diet-induced diastolic dysfunction and left ventricular fibrosis, independent of sex, but also predisposed male animals to the development of cardiac hypertrophy. These data suggest that TBC1D1 may exert cardioprotective effects in the development of diabetic cardiomyopathy.

Overexpression of native IF1 downregulates glucose-stimulated insulin secretion by pancreatic INS-1E cells

We have previously reported that transient knock-down of ATPase inhibitory factor 1 (IF1) by siRNA upregulates ATP levels and subsequently augments insulin secretion in model pancreatic β-cells INS-1E. Here we investigated how long-term IF1-overexpression impacts pancreatic β-cell bioenergetics and insulin secretion. We generated INS-1E cell line stably overexpressing native IF1. We revealed that IF1 overexpression leads to a substantial decrease in ATP levels and reduced glucose-stimulated insulin secretion. A decrease in total cellular ATP content was also reflected in decreased free ATP cytosolic and mitochondrial levels, as monitored with ATeam biosensor. Consistently, cellular respiration of IF1-overexpressing cells was decreased. 3D structured illumination microscopy (SIM) revealed a higher amount of insulin granules with higher volume in IF1-overexpressing cells. Similar effects occurred when cells were incubated at low glucose concentrations. Noteworthy, activation of PKA by dibutyryl cAMP entirely abolished the inhibitory effect of IF1 overexpression on ATP production and insulin secretion. Mitochondrial network morphology and cristae ultrastructure in INS-1E overexpressing IF1 remained mostly unchanged. Finally, we show that INS-1E cells decrease their IF1 protein levels relative to ATP synthase α-subunit in response to increased glucose. In conclusion, IF1 actively downregulates INS-1E cellular metabolism and reduces their ability to secrete insulin.