Channeling β-cell Maturity: KATP Surface Localization Imparts Glucose Sensing

Abstract An early sign of islet failure in type 2 diabetes (T2D) is the loss of normal patterns of pulsatile insulin release. Disruptions in pulsatility are associated with a left shift in glucose sensing that can cause excessive insulin release in low glucose (relative hyperinsulinemia, a hallmark of early T2D) and β-cell exhaustion, leading to inadequate insulin release during hyperglycemia. Our hypothesis was that reducing excessive glucokinase activity in diabetic islets would improve their function. Isolated mouse islets were exposed to glucose and varying concentrations of the glucokinase inhibitor d-mannoheptulose (MH) to examine changes in intracellular calcium ([Ca2+]i) and insulin secretion. Acutely exposing islets from control CD-1 mice to MH in high glucose (20 mM) dose dependently reduced the size of [Ca2+]i oscillations detected by fura-2 acetoxymethyl. Glucokinase activation in low glucose (3 mM) had the opposite effect. We then treated islets from male and female db/db mice (age, 4 to 8 weeks) and heterozygous controls overnight with 0 to 10 mM MH to determine that 1 mM MH produced optimal oscillations. We then used 1 mM MH overnight to measure [Ca2+]i and insulin simultaneously in db/db islets. MH restored oscillations and increased insulin secretion. Insulin secretion rates correlated with MH-induced increases in amplitude of [Ca2+]i oscillations (R2 = 0.57, P < 0.01, n = 10) but not with mean [Ca2+]i levels in islets (R2 = 0.05, not significant). Our findings show that correcting glucose sensing can restore proper pulsatility to diabetic islets and improved pulsatility correlates with enhanced insulin secretion.

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Mechanisms of glucose sensing in the pancreatic β-cell

Islets ◽

10.4161/isl.3.5.16409 ◽

2011 ◽

Vol 3 (5) ◽

pp. 224-230 ◽

Cited By ~ 6

Author(s):

Leonid E. Fridlyand ◽

Louis H. Phillipson

Keyword(s):

Glucose Sensing ◽

Pancreatic Β Cell ◽

Β Cell

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Major metabolic homocysteine-derivative, homocysteine thiolactone, exerts changes in pancreatic β-cell glucose-sensing, cellular signal transduction and integrity

Archives of Biochemistry and Biophysics ◽

10.1016/j.abb.2007.02.011 ◽

2007 ◽

Vol 461 (2) ◽

pp. 287-293 ◽

Cited By ~ 5

Author(s):

Steven Patterson ◽

Peter R. Flatt ◽

Neville H. McClenaghan

Keyword(s):

Signal Transduction ◽

Glucose Sensing ◽

Pancreatic Β Cell ◽

Β Cell ◽

Homocysteine Thiolactone ◽

Cellular Signal Transduction ◽

Cellular Signal ◽

Cell Glucose

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Human EndoC-βH1 β-cells form pseudoislets with improved glucose sensitivity and enhanced GLP-1 signaling in the presence of islet-derived endothelial cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00272.2017 ◽

2018 ◽

Vol 314 (5) ◽

pp. E512-E521 ◽

Cited By ~ 1

Author(s):

Michael G. Spelios ◽

Lauren A. Afinowicz ◽

Regine C. Tipon ◽

Eitan M. Akirav

Keyword(s):

Endothelial Cells ◽

Insulin Secretion ◽

Cell Biology ◽

Three Dimensional ◽

Cell Types ◽

Human Islets ◽

Glucose Sensing ◽

Β Cells ◽

Β Cell ◽

Glucose Levels

Three-dimensional (3D) pseudoislets (PIs) can be used for the study of insulin-producing β-cells in free-floating islet-like structures similar to that of primary islets. Previously, we demonstrated the ability of islet-derived endothelial cells (iECs) to induce PIs using murine insulinomas, where PI formation enhanced insulin production and glucose responsiveness. In this report, we examined the ability of iECs to spontaneously induce the formation of free-floating 3D PIs using the EndoC-βH1 human β-cell line murine MS1 iEC. Within 14 days, the coculturing of both cell types produced fully humanized EndoC-βH1 PIs with little to no contaminating murine iECs. The size and shape of these PIs were similar to primary human islets. iEC-induced PIs demonstrated reduced dysregulated insulin release under low glucose levels and higher insulin secretion in response to high glucose and exendin-4 [a glucagon-like peptide-1 (GLP-1) analog] compared with monolayer cells cultured alone. Interestingly, iEC-PIs were also better at glucose sensing in the presence of extendin-4 compared with PIs generated on a low-adhesion surface plate in the absence of iECs and showed an overall improvement in cell viability. iEC-induced PIs exhibited increased expression of key genes involved in glucose transport, glucose sensing, β-cell differentiation, and insulin processing, with a concomitant decrease in glucagon mRNA expression. The enhanced responsiveness to exendin-4 was associated with increased protein expression of GLP-1 receptor and phosphokinase A. This rapid coculture system provides an unlimited number of human PIs with improved insulin secretion and GLP-1 responsiveness for the study of β-cell biology.

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Correction to Insulin Promoter-Driven Gaussia Luciferase-Based Insulin Secretion Biosensor Assay for Discovery of β-Cell Glucose-Sensing Pathways

ACS Sensors ◽

10.1021/acssensors.7b00012 ◽

2017 ◽

Vol 2 (2) ◽

pp. 316-316 ◽

Cited By ~ 1

Author(s):

Michael A. Kalwat ◽

Chonlarat Wichaidit ◽

Alejandra Y. Nava Garcia ◽

Melissa K. McCoy ◽

Kathleen McGlynn ◽

...

Keyword(s):

Insulin Secretion ◽

Glucose Sensing ◽

Β Cell ◽

Gaussia Luciferase ◽

Insulin Promoter ◽

Cell Glucose

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In vitro Characterization of Insulin−Producing β-Cell Spheroids

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2020.623889 ◽

2021 ◽

Vol 8 ◽

Author(s):

Yonela Ntamo ◽

Ebrahim Samodien ◽

Joleen Burger ◽

Nolan Muller ◽

Christo J. F. Muller ◽

...

Keyword(s):

Glucose Utilization ◽

Cell Function ◽

3D Culture ◽

Glucose Sensing ◽

Blue Staining ◽

Cell Physiology ◽

Β Cell ◽

3D Spheroids

Over the years, immortalized rodent β-cell lines such as RIN, HIT, MIN, βTC, and INS-1 have been used to investigate pancreatic β-cell physiology using conventional two-dimensional (2D) culture techniques. However, physical and physiological limitations inherent to 2D cell culture necessitates confirmatory follow up studies using sentient animals. Three-dimensional (3D) culture models are gaining popularity for their recapitulation of key features of in vivo organ physiology, and thus could pose as potential surrogates for animal experiments. In this study, we aimed to develop and characterize a rat insulinoma INS-1 3D spheroid model to compare with 2D monolayers of the same cell line. Ultrastructural verification was done by transmission electron microscopy and toluidine blue staining, which showed that both 2D monolayers and 3D spheroids contained highly granulated cells with ultrastructural features synonymous with mature pancreatic β-cells, with increased prominence of these features observed in 3D spheroids. Viability, as assessed by cellular ATP quantification, size profiling and glucose utilization, showed that our spheroids remained viable for the experimental period of 30 days, compared to the limiting 5-day passage period of INS-1 monolayers. In fact, increasing ATP content together with spheroid size was observed over time, without adverse changes in glucose utilization. Additionally, β-cell function, assessed by determining insulin and amylin secretion, showed that the 3D spheroids retained glucose sensing and insulin secretory capability, that was more acute when compared to 2D monolayer cultures. Thus, we were able to successfully demonstrate that our in vitro INS-1 β-cell 3D spheroid model exhibits in vivo tissue-like structural features with extended viability and lifespan. This offers enhanced predictive capacity of the model in the study of metabolic disease, β-cell pathophysiology and the potential treatment thereof.

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Primary cilia control glucose homeostasis via islet paracrine interactions

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2001936117 ◽

2020 ◽

Vol 117 (16) ◽

pp. 8912-8923 ◽

Cited By ~ 10

Author(s):

Jing W. Hughes ◽

Jung Hoon Cho ◽

Hannah E. Conway ◽

Michael R. DiGruccio ◽

Xue Wen Ng ◽

...

Keyword(s):

Insulin Secretion ◽

Glucose Homeostasis ◽

Cross Talk ◽

Primary Cilia ◽

Calcium Influx ◽

Glucose Sensing ◽

Intrinsic Activity ◽

Β Cell ◽

Nutrient Metabolism ◽

Cellular Processes

Pancreatic islets regulate glucose homeostasis through coordinated actions of hormone-secreting cells. What underlies the function of the islet as a unit is the close approximation and communication among heterogeneous cell populations, but the structural mediators of islet cellular cross talk remain incompletely characterized. We generated mice specifically lacking β-cell primary cilia, a cellular organelle that has been implicated in regulating insulin secretion, and found that the β-cell cilia are required for glucose sensing, calcium influx, insulin secretion, and cross regulation of α- and δ-cells. Protein expression profiling in islets confirms perturbation in these cellular processes and reveals additional targets of cilia-dependent signaling. At the organism level, the deletion of β-cell cilia disrupts circulating hormone levels, impairs glucose homeostasis and fuel usage, and leads to the development of diabetes. Together, these findings demonstrate that primary cilia not only orchestrate β-cell–intrinsic activity but also mediate cross talk both within the islet and from islets to other metabolic tissues, thus providing a unique role of cilia in nutrient metabolism and insight into the pathophysiology of diabetes.

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Glucose-sensing neurons of the hypothalamus

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2005.1763 ◽

2005 ◽

Vol 360 (1464) ◽

pp. 2227-2235 ◽

Cited By ~ 172

Author(s):

Denis Burdakov ◽

Simon M Luckman ◽

Alexei Verkhratsky

Keyword(s):

Feeding Behaviour ◽

Arcuate Nucleus ◽

Glucose Sensing ◽

Β Cell ◽

Hypothalamic Neurons ◽

Extracellular Glucose ◽

Hypothalamic Arcuate Nucleus ◽

Hypothalamic Cells ◽

Melanin Concentrating Hormone ◽

Cell Glucose

Specialized subgroups of hypothalamic neurons exhibit specific excitatory or inhibitory electrical responses to changes in extracellular levels of glucose. Glucose-excited neurons were traditionally assumed to employ a ‘β-cell’ glucose-sensing strategy, where glucose elevates cytosolic ATP, which closes K ATP channels containing Kir6.2 subunits, causing depolarization and increased excitability. Recent findings indicate that although elements of this canonical model are functional in some hypothalamic cells, this pathway is not universally essential for excitation of glucose-sensing neurons by glucose. Thus glucose-induced excitation of arcuate nucleus neurons was recently reported in mice lacking Kir6.2, and no significant increases in cytosolic ATP levels could be detected in hypothalamic neurons after changes in extracellular glucose. Possible alternative glucose-sensing strategies include electrogenic glucose entry, glucose-induced release of glial lactate, and extracellular glucose receptors. Glucose-induced electrical inhibition is much less understood than excitation, and has been proposed to involve reduction in the depolarizing activity of the Na + /K + pump, or activation of a hyperpolarizing Cl − current. Investigations of neurotransmitter identities of glucose-sensing neurons are beginning to provide detailed information about their physiological roles. In the mouse lateral hypothalamus, orexin/hypocretin neurons (which promote wakefulness, locomotor activity and foraging) are glucose-inhibited, whereas melanin-concentrating hormone neurons (which promote sleep and energy conservation) are glucose-excited. In the hypothalamic arcuate nucleus, excitatory actions of glucose on anorexigenic POMC neurons in mice have been reported, while the appetite-promoting NPY neurons may be directly inhibited by glucose. These results stress the fundamental importance of hypothalamic glucose-sensing neurons in orchestrating sleep-wake cycles, energy expenditure and feeding behaviour.

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Regulating insulin signaling and β-cell function through IRS proteinsThis paper is one of a selection of papers published in this Special Issue, entitled Second Messengers and Phosphoproteins—12th International Conference.

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y06-008 ◽

2006 ◽

Vol 84 (7) ◽

pp. 725-737 ◽

Cited By ~ 104

Author(s):

Morris F. White

Keyword(s):

Insulin Resistance ◽

Type 2 Diabetes ◽

Insulin Signaling ◽

Cell Function ◽

Second Messengers ◽

Glucose Sensing ◽

Β Cell ◽

Peripheral Insulin Resistance ◽

Chronic Hyperglycemia

Diabetes mellitus is a complex disorder that arises from various causes, including dysregulated glucose sensing and impaired insulin secretion (maturity onset diabetes of youth, MODY), autoimmune-mediated β-cell destruction (type 1), or insufficient compensation for peripheral insulin resistance (type 2). Type 2 diabetes is the most prevalent form that usually occurs at middle age; it afflicts more than 30 million people over the age of 65, but is appearing with greater frequency in children and adolescents. Dysregulated insulin signaling exacerbated by chronic hyperglycemia promotes a cohort of systemic disorders—including dyslipidemia, hypertension, cardiovascular disease, and female infertility. Understanding the molecular basis of insulin resistance can prevent these disorders and their inevitable progression to type 2 diabetes.

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