Cell type-specific activation of metabolism reveals that β-cell secretion suppresses glucagon release from α-cells in rat pancreatic islets

Abnormal glucagon secretion is often associated with diabetes mellitus. However, the mechanisms by which nutrients modulate glucagon secretion remain poorly understood. Paracrine modulation by β- or δ-cells is among the postulated mechanisms. Herein we present further evidence of the paracrine mechanism. First, to activate cellular metabolism and thus hormone secretion in response to specific secretagogues, we engineered insulinoma INS-1E cells using an adenovirus-mediated expression system. Expression of the Na+-dependent dicarboxylate transporter (NaDC)-1 resulted in 2.5- to 4.6-fold ( P < 0.01) increases in insulin secretion in response to various tricarboxylic acid cycle intermediates. Similarly, expression of glycerol kinase (GlyK) increased insulin secretion 3.8- or 4.2-fold ( P < 0.01) in response to glycerol or dihydroxyacetone, respectively. This cell engineering method was then modified, using the Cre- loxP switching system, to activate β-cells and non-β-cells separately in rat islets. NaDC-1 expression only in non-β-cells, among which α-cells are predominant, caused an increase (by 1.8-fold, P < 0.05) in glucagon secretion in response to malate or succinate. However, the increase in glucagon release was prevented when NaDC-1 was expressed in whole islets, i.e., both β-cells and non-β-cells. Similarly, an increase in glucagon release with glycerol was observed when GlyK was expressed only in non-β-cells but not when it was expressed in whole islets. Furthermore, dicarboxylates suppressed basal glucagon secretion by 30% ( P < 0.05) when NaDC-1 was expressed only in β-cells. These data demonstrate that glucagon secretion from rat α-cells depends on β-cell activation and provide insights into the coordinated mechanisms underlying hormone secretion from pancreatic islets.

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Impact of lack of suppression of glucagon on glucose tolerance in humans

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1999.277.2.e283 ◽

1999 ◽

Vol 277 (2) ◽

pp. E283-E290 ◽

Cited By ~ 23

Author(s):

Pankaj Shah ◽

Ananda Basu ◽

Rita Basu ◽

Robert Rizza

Keyword(s):

Insulin Secretion ◽

Glucose Concentration ◽

Cell Function ◽

Hormone Secretion ◽

Glucose Production ◽

Glucagon Secretion ◽

Β Cell ◽

Β Cell Function ◽

Glucagon Suppression

People with type 2 diabetes have defects in both α- and β-cell function. To determine whether lack of suppression of glucagon causes hyperglycemia when insulin secretion is impaired but not when insulin secretion is intact, twenty nondiabetic subjects were studied on two occasions. On both occasions, a “prandial” glucose infusion was given over 5 h while endogenous hormone secretion was inhibited. Insulin was infused so as to mimic either a nondiabetic ( n = 10) or diabetic ( n = 10) postprandial profile. Glucagon was infused at a rate of 1.25 ng ⋅ kg−1 ⋅ min−1, beginning either at time zero to prevent a fall in glucagon (nonsuppressed study day) or at 2 h to create a transient fall in glucagon (suppressed study day). During the “diabetic” insulin profile, lack of glucagon suppression resulted in a marked increase ( P < 0.002) in both the peak glucose concentration (11.9 ± 0.4 vs. 8.9 ± 0.4 mmol/l) and the area above basal of glucose (927 ± 77 vs. 546 ± 112 mmol ⋅ l−1 ⋅ 6 h) because of impaired ( P < 0.001) suppression of glucose production. In contrast, during the “nondiabetic” insulin profile, lack of suppression of glucagon resulted in only a slight increase ( P< 0.02) in the peak glucose concentration (9.1 ± 0.4 vs. 8.4 ± 0.3 mmol/l) and the area above basal of glucose (654 ± 146 vs. 488 ± 118 mmol ⋅ l−1 ⋅ 6 h). Of interest, when glucagon was suppressed, glucose concentrations differed only minimally during the nondiabetic and diabetic insulin profiles. These data indicate that lack of suppression of glucagon can cause substantial hyperglycemia when insulin availability is limited, therefore implying that inhibitors of glucagon secretion and/or glucagon action are likely to be useful therapeutic agents in such individuals.

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The adaptor protein APPL2 controls glucose-stimulated insulin secretion via F-actin remodeling in pancreatic β-cells

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2016997117 ◽

2020 ◽

Vol 117 (45) ◽

pp. 28307-28315

Author(s):

Baile Wang ◽

Huige Lin ◽

Xiaomu Li ◽

Wenqi Lu ◽

Jae Bum Kim ◽

...

Keyword(s):

Insulin Secretion ◽

Pancreatic Islets ◽

Adaptor Protein ◽

Hek293 Cells ◽

Β Cells ◽

Β Cell ◽

Pancreatic Β Cells ◽

Actin Remodeling ◽

Actin Depolymerization ◽

Glucose Stimulated Insulin Secretion

Filamentous actin (F-actin) cytoskeletal remodeling is critical for glucose-stimulated insulin secretion (GSIS) in pancreatic β-cells, and its dysregulation causes type 2 diabetes. The adaptor protein APPL1 promotes first-phase GSIS by up-regulating solubleN-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein expression. However, whether APPL2 (a close homology of APPL1 with the same domain organization) plays a role in β-cell functions is unknown. Here, we show that APPL2 enhances GSIS by promoting F-actin remodeling via the small GTPase Rac1 in pancreatic β-cells. β-cell specific abrogation of APPL2 impaired GSIS, leading to glucose intolerance in mice. APPL2 deficiency largely abolished glucose-induced first- and second-phase insulin secretion in pancreatic islets. Real-time live-cell imaging and phalloidin staining revealed that APPL2 deficiency abolished glucose-induced F-actin depolymerization in pancreatic islets. Likewise, knockdown of APPL2 expression impaired glucose-stimulated F-actin depolymerization and subsequent insulin secretion in INS-1E cells, which were attributable to the impairment of Ras-related C3 botulinum toxin substrate 1 (Rac1) activation. Treatment with the F-actin depolymerization chemical compounds or overexpression of gelsolin (a F-actin remodeling protein) rescued APPL2 deficiency-induced defective GSIS. In addition, APPL2 interacted with Rac GTPase activating protein 1 (RacGAP1) in a glucose-dependent manner via the bin/amphiphysin/rvs-pleckstrin homology (BAR-PH) domain of APPL2 in INS-1E cells and HEK293 cells. Concomitant knockdown of RacGAP1 expression reverted APPL2 deficiency-induced defective GSIS, F-actin remodeling, and Rac1 activation in INS-1E cells. Our data indicate that APPL2 interacts with RacGAP1 and suppresses its negative action on Rac1 activity and F-actin depolymerization thereby enhancing GSIS in pancreatic β-cells.

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SUN-658 CHL1 Increases Insulin Secretion&Negatively Regulates the Poliferation of Pancreatic β Cell

Journal of the Endocrine Society ◽

10.1210/jendso/bvaa046.1269 ◽

2020 ◽

Vol 4 (Supplement_1) ◽

Author(s):

Tao Yang ◽

Qi Fu ◽

Hemin Jiang

Keyword(s):

Cell Cycle ◽

Insulin Secretion ◽

Pancreatic Islets ◽

High Fat Diet ◽

Nod Mice ◽

Pancreatic Β Cell ◽

High Fat ◽

Β Cells ◽

Β Cell ◽

Min6 Cells

Abstract CHL1 Increases Insulin Secretion & Negatively Regulates The Poliferation Of Pancreatic β Cell Objective: CHL1 belongs to neural recognition molecules of the immunoglobulin superfamily, is mainly expressed in the nervous system. CHL1 is involved in neuronal migration, axonal growth, and dendritic projection. RNA sequencing of single human islet cells confirmed that CHL1 had an expression difference in β cells of type 2 diabetes and healthy controls. However, whether CHL1 gene regulates islet function remained to be explored. Methods: PCR and Western Blot were applied to investigate the tissue distribution of CHL1 in wild-type C57BL/6J mice. The islet expression of CHL1 gene was observed in pancreatic islets of NOD mice and high-fat-diet C57BL/6J mice of different ages. MIN6 cells with siRNA to silence CHL1 or with lentivirus to overexpress CHL1 were constructed. Effects of the gene on proliferation, apoptosis, cell cycle and insulin secretion were determined by using CCK8, EdU, TUNEL, AV/PI, GSIS, electron microscopy and flow cytometry. Results: CHL1 was localized on the cell membrane and expressed in the nervous system, islet of pancreas and gastrointestinal tract. CHL1 was hypoexpressed in the pancreatic islets of obese mice, hyperexpressed in the pancreatic islets of NOD mice and in vitro after treated with cytokines. After silencing CHL1 in MIN6 cells, insulin secretion decreased in 20 mM glucose with down-regulation of INS1, SLC2A2 gene, and transmission electron microscope showed the number of insulin secretary granules <50nm from the cell membrane was significantly reduced. Silencing of CHL1 in MIN6 cells induced cell proliferation, reduced apoptosis rate, prolonged the S phase of cell cycle and shortened the G1 phase with downregulated expression of p21, p53 and up-regulated expression of cyclin D1, opposite results were found in CHL1 over-expressing MIN6 cells. Proliferation induced by silencing of CHL1 was inhibited by ERK inhibitor (PD98059), which indicates that ERK pathway is essential for signaling by these molecules in pancreatic β cell. Conclusion: The expression of CHL1 gene was significantly decreased in the pancreatic islets of obese mice induced by high-fat diet. The low expression of CHL1 gene promotes the proliferation of MIN6 cells through the ERK pathway and affect cell cycle through the p53 pathway. This may be one of the mechanisms that pancreatic β cells compensatory hyperplasia in the stage of obesity-induced pre-diabetes.

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Human pseudoislet system enables detection of differences in G-protein-coupled-receptor signaling pathways between α and β cells

10.1101/842989 ◽

2019 ◽

Author(s):

John T. Walker ◽

Rachana Haliyur ◽

Heather A. Nelson ◽

Matthew Ishahak ◽

Gregory Poffenberger ◽

...

Keyword(s):

Insulin Secretion ◽

Signaling Pathways ◽

G Protein ◽

Intracellular Signaling ◽

Hormone Secretion ◽

Glucagon Secretion ◽

Integrated Approach ◽

Designer Drugs ◽

Β Cells ◽

G Protein Coupled

SUMMARYG-protein-coupled-receptors (GPCRs) modulate insulin secretion from β cells and glucagon secretion from α cells. Here, we developed an integrated approach to study the function of primary human islet cells using genetically modified pseudoislets that resemble native islets across multiple parameters. We studied the Gi and Gq GPCR pathways by expressing the designer receptors exclusively activated by designer drugs (DREADDs) hM4Di or hM3Dq. Activation of Gi signaling reduced insulin and glucagon secretion, while activation of Gq signaling stimulated glucagon secretion but had both stimulatory and inhibitory effects on insulin secretion. Further, we developed a microperifusion system that allowed synchronous acquisition of GCaMP6f biosensor signal and hormone secretory profiles and showed that the dual effects for Gq signaling occur through changes in intracellular Ca2+. By combining pseudoislets with a microfluidic system, we co-registered intracellular signaling dynamics and hormone secretion and demonstrated differences in GPCR signaling pathways between human β and α cells.

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Somatostatin Inhibits Insulin and Glucagon Secretion via Two Receptor Subtypes: An in Vitro Study of Pancreatic Islets from Somatostatin Receptor 2 Knockout Mice*

Endocrinology ◽

10.1210/endo.141.1.7263 ◽

2000 ◽

Vol 141 (1) ◽

pp. 111-117 ◽

Cited By ~ 132

Author(s):

M. Z. Strowski ◽

R. M. Parmar ◽

A. D. Blake ◽

J. M. Schaeffer

Keyword(s):

Insulin Secretion ◽

Pancreatic Islets ◽

Insulin Release ◽

Somatostatin Receptor ◽

Glucagon Secretion ◽

Endocrine Function ◽

Receptor Subtype ◽

Glucagon Release ◽

Glucagon And Insulin

Abstract Somatostatin (SST) potently inhibits insulin and glucagon release from pancreatic islets. Five distinct membrane receptors (SSTR1–5) for SST are known, and at least two (SSTR2 and SSTR5) have been proposed to regulate pancreatic endocrine function. Our current understanding of SST physiology is limited by the receptor subtype selectivity of peptidyl SST analogs, making it difficult to assign a physiological function to an identified SST receptor subtype. To better understand the physiology of SSTRs we studied the in vitro effects of potent subtype-selective nonpeptidyl SST analogs on the regulation of pancreatic glucagon and insulin secretion in wild-type (WT) and in somatostatin receptor 2 knockout (SSTR2KO) mice. There was no difference in basal glucagon and insulin secretion between islets isolated from SSTR2KO and WT mice; however, potassium/arginine-stimulated glucagon secretion was approximately 2-fold higher in islets isolated from SSTR2KO mice. Neither SST nor any SSTR-selective agonist inhibited basal glucagon or insulin release. SST-14 potently inhibited stimulated glucagon secretion in islets from WT mice and much less effectively in islets from SSTR2KO mice. The SSTR2 selective analog L-779,976 inhibited glucagon secretion in islets from WT, but was inactive in islets from SSTR2KO mice. L-817,818, an SSTR5 selective analog, slightly reduced glucagon release in both animal groups, whereas SSTR1, -3, and -4 selective analogs were inactive. SST and L-817,818 inhibited glucose stimulated insulin release in islets from WT and SSTR2KO mice. L-779,976 much less potently reduced insulin secretion from WT islets. In conclusion, our data demonstrate that SST inhibition of glucagon release in mouse islets is primarily mediated via SSTR2, whereas insulin secretion is regulated primarily via SSTR5.

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GIP mediates the incretin effect and glucose tolerance by dual actions on α cells and β cells

Science Advances ◽

10.1126/sciadv.abf1948 ◽

2021 ◽

Vol 7 (11) ◽

pp. eabf1948

Author(s):

K. El ◽

S. M. Gray ◽

M. E. Capozzi ◽

E. R. Knuth ◽

E. Jin ◽

...

Keyword(s):

Insulin Secretion ◽

Nutrient Intake ◽

Cell Communication ◽

Glucagon Secretion ◽

Optimal Level ◽

Β Cells ◽

Β Cell ◽

Postprandial State ◽

Glucagon And Insulin

Glucose-dependent insulinotropic polypeptide (GIP) communicates nutrient intake from the gut to islets, enabling optimal levels of insulin secretion via the GIP receptor (GIPR) on β cells. The GIPR is also expressed in α cells, and GIP stimulates glucagon secretion; however, the role of this action in the postprandial state is unknown. Here, we demonstrate that GIP potentiates amino acid–stimulated glucagon secretion, documenting a similar nutrient-dependent action to that described in β cells. Moreover, we demonstrate that GIP activity in α cells contributes to insulin secretion by invoking paracrine α to β cell communication. Last, specific loss of GIPR activity in α cells prevents glucagon secretion in response to a meal stimulus, limiting insulin secretion and driving glucose intolerance. Together, these data uncover an important axis by which GIPR activity in α cells is necessary to coordinate the optimal level of both glucagon and insulin secretion to maintain postprandial homeostasis.

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Pancreatic β Cells Inhibit Glucagon Secretion from α Cells: An In Vitro Demonstration of α–β Cell Interaction

Nutrients ◽

10.3390/nu13072281 ◽

2021 ◽

Vol 13 (7) ◽

pp. 2281

Author(s):

Wenqian Gu ◽

Camilla Christine Bundgaard Anker ◽

Christine Bodelund Christiansen ◽

Tilo Moede ◽

Per-Olof Berggren ◽

...

Keyword(s):

Cell Line ◽

Cell Lines ◽

Hormone Secretion ◽

Glucagon Secretion ◽

Cell Contact ◽

Β Cells ◽

Β Cell ◽

Gene Expressions ◽

The Impact

Interactions between endocrine α and β cells are critical to their secretory function in vivo. The interactions are highly regulated, although yet to be fully understood. In this study, we aim to assess the impact of α and β cell co-culture on hormone secretion. Mouse clonal cell lines α-TC6-1 (α cell line) and MIN-6 (β cell line) were cultured independently or in combination in a medium containing 5.5, 11.1, or 25 mM glucose, respectively. After 72 h, hormone release was measured using insulin and glucagon secretion assays, the cell distribution was visualized by inverted microscopy and an immunocytochemistry assay, and changes in gene expressions were assessed using the RT-PCR technique. The co-culture of the two cell lines caused a decrease in glucagon secretion from α-TC1-6 cells, while no effect on insulin secretion from MIN-6 cells was revealed. Both types of cells were randomly scattered throughout the culture flask, unlike in mice islets in vivo where β cells cluster in the core and α cells are localized at the periphery. During the α–β cell co-culture, the gene expression of glucagon (Gcg) decreased significantly. We conclude that islet β cells suppress glucagon secretion from α cells, apparently via direct cell-to-cell contact, of which the molecular mechanism needs further verification.

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Inhibition of Cholesterol Biosynthesis Impairs Insulin Secretion and Voltage-Gated Calcium Channel Function in Pancreatic β-Cells

Endocrinology ◽

10.1210/en.2008-0161 ◽

2008 ◽

Vol 149 (10) ◽

pp. 5136-5145 ◽

Cited By ~ 81

Author(s):

Fuzhen Xia ◽

Li Xie ◽

Anton Mihic ◽

Xiaodong Gao ◽

Yi Chen ◽

...

Keyword(s):

Insulin Secretion ◽

Pancreatic Islets ◽

Cholesterol Biosynthesis ◽

Squalene Epoxidase ◽

Β Cells ◽

Β Cell ◽

Pancreatic Β Cells ◽

Min6 Cells ◽

Voltage Gated ◽

Endogenous Cholesterol

Insulin secretion from pancreatic β-cells is mediated by the opening of voltage-gated Ca2+ channels (CaV) and exocytosis of insulin dense core vesicles facilitated by the secretory soluble N-ethylmaleimide-sensitive factor attachment protein receptor protein machinery. We previously observed that β-cell exocytosis is sensitive to the acute removal of membrane cholesterol. However, less is known about the chronic changes in endogenous cholesterol and its biosynthesis in regulating β-cell stimulus-secretion coupling. We examined the effects of inhibiting endogenous β-cell cholesterol biosynthesis by using the squalene epoxidase inhibitor, NB598. The expression of squalene epoxidase in primary and clonal β-cells was confirmed by RT-PCR. Cholesterol reduction of 36–52% was observed in MIN6 cells, mouse and human pancreatic islets after a 48-h incubation with 10 μm NB598. A similar reduction in cholesterol was observed in the subcellular compartments of MIN6 cells. We found NB598 significantly inhibited both basal and glucose-stimulated insulin secretion from mouse pancreatic islets. CaV channels were markedly inhibited by NB598. Rapid photolytic release of intracellular caged Ca2+ and simultaneous measurements of the changes in membrane capacitance revealed that NB598 also inhibited exocytosis independently from CaV channels. These effects were reversed by cholesterol repletion. Our results indicate that endogenous cholesterol in pancreatic β-cells plays a critical role in regulating insulin secretion. Moreover, chronic inhibition of cholesterol biosynthesis regulates the functional activity of CaV channels and insulin secretory granule mobilization and membrane fusion. Dysregulation of cellular cholesterol may cause impairment of β-cell function, a possible pathogenesis leading to the development of type 2 diabetes.

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Inhibition of the malate–aspartate shuttle in mouse pancreatic islets abolishes glucagon secretion without affecting insulin secretion

Biochemical Journal ◽

10.1042/bj20140697 ◽

2015 ◽

Vol 468 (1) ◽

pp. 49-63 ◽

Cited By ~ 7

Author(s):

Jelena A. Stamenkovic ◽

Lotta E. Andersson ◽

Alice E. Adriaenssens ◽

Annika Bagge ◽

Vladimir V. Sharoyko ◽

...

Keyword(s):

Type 2 Diabetes ◽

Insulin Secretion ◽

Pancreatic Islets ◽

Glucagon Secretion ◽

Nutrient Sensing ◽

Β Cells ◽

Mouse Pancreatic Islets ◽

Glucagon And Insulin ◽

Malate Aspartate Shuttle

Secretion of both glucagon and insulin is perturbed in Type 2 diabetes (T2D). In the present study, we identify a difference in mitochondrial shuttling between α- and β-cells that adjusts nutrient sensing and which potentially could be employed to specifically target secretion of either hormone.

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Vitamin B6 deficiency disrupts serotonin signaling in pancreatic islets and induces gestational diabetes in mice

Communications Biology ◽

10.1038/s42003-021-01900-0 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Ashley M. Fields ◽

Kevin Welle ◽

Elaine S. Ho ◽

Clementina Mesaros ◽

Martha Susiarjo

Keyword(s):

Cell Proliferation ◽

Insulin Secretion ◽

Pancreatic Islets ◽

Glucose Intolerance ◽

Vitamin B6 ◽

Serotonin Receptor ◽

Dependent Manner ◽

Β Cell ◽

Vitamin B6 Deficiency ◽

Maternal Glucose

AbstractIn pancreatic islets, catabolism of tryptophan into serotonin and serotonin receptor 2B (HTR2B) activation is crucial for β-cell proliferation and maternal glucose regulation during pregnancy. Factors that reduce serotonin synthesis and perturb HTR2B signaling are associated with decreased β-cell number, impaired insulin secretion, and gestational glucose intolerance in mice. Albeit the tryptophan-serotonin pathway is dependent on vitamin B6 bioavailability, how vitamin B6 deficiency impacts β-cell proliferation during pregnancy has not been investigated. In this study, we created a vitamin B6 deficient mouse model and investigated how gestational deficiency influences maternal glucose tolerance. Our studies show that gestational vitamin B6 deficiency decreases serotonin levels in maternal pancreatic islets and reduces β-cell proliferation in an HTR2B-dependent manner. These changes were associated with glucose intolerance and insulin resistance, however insulin secretion remained intact. Our findings suggest that vitamin B6 deficiency-induced gestational glucose intolerance involves additional mechanisms that are complex and insulin independent.

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