Table of Contents

Volume 4, Issue 1

  January/February 2012

  Pages 1 - 77

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REVIEWS

Overcoming the spatial barriers of the stimulus secretion cascade in pancreatic β-cells

Peter R. Huypens, Mei Huang and Jamie W. Joseph
Pages 1 - 9
http://dx.doi.org/10.4161/isl.18338
Abstract | Full Text | PDF

The ability of the pancreatic β-cells to adapt the rate of insulin release in accordance to changes in circulating glucose levels is essential for glucose homeostasis. Two spatial barriers imposed by the plasma membrane and inner mitochondrial membrane need to be overcome in order to achieve stringent coupling between the different steps in the stimulus-secretion cascade.

 

The first spatial barrier is overcome by the presence of a glucose transporter (GLUT) in the plasma membrane, whereas a low affinity hexokinase IV (glucokinase, GK) in the cytosol conveys glucose availability into a metabolic flux that triggers and accelerates insulin release. The mitochondrial inner membrane comprises a second spatial barrier that compartmentalizes glucose metabolism into glycolysis (cytosol) and tricarboxylate (TCA) cycle (mitochondrial matrix).

The exchange of metabolites between cytosol and mitochondrial matrix is mediated via a set of mitochondrial carriers, including the aspartate-glutamate carrier (aralar1), α- ketoglutarate carrier (OGC), ATP/ADP carrier (AAC), glutamate carrier (GC1), dicarboxylate carrier (DIC) and citrate/isocitrate carrier (CIC). The scope of this review is to provide an overview of the role these carriers play in stimulus-secretion coupling and discuss the importance of these findings in the context of the exquisite glucose responsive state of the pancreatic β-cell.

Mathematical models of pancreatic islet size distributions

Junghyo Jo, Manami Hara, Ulf Ahlgren, Robert Sorenson and Vipul Periwal
Pages 10 - 19
http://dx.doi.org/10.4161/isl.18660
Abstract | Full Text | PDF

The islets of Langerhans, ranging in size from clusters of a few cells to several thousand cells, are scattered near large blood vessels. While the β-cell mass in mammals is proportional to body weight, the size ranges of islets are similar between species with different body sizes, possibly reflecting an optimal functional size. The large range of islet sizes suggests a stochastic developmental process. It is not fully understood how islets develop to reach such size distributions, and how their sizes change under certain physiological and pathological conditions such as development, pregnancy, aging, obesity, and diabetes. The lack of a high-resolution in vivo imaging technique for pancreatic islets implies that the only data available to elucidate the dynamics of islet development are cross-sectional quantifications of islet size distributions. In this review, we infer biological processes affecting islet morphology in the large by examining changes of islet size distributions. Neonatal islet formation and growth is shown as a particular example of developing a mathematical model of islet size distribution. Application of this modeling to elucidate islet changes under other conditions is also discussed.




SHORT REPORT

GPR54 peptide agonists stimulate insulin secretion from murine, porcine and human islets

James E. Bowe, Victoria L. Foot, Stephanie A. Amiel, Gao Cai Huang, Morgan Lamb, Jonathan Lakey, Peter M. Jones and Shanta J. Persaud
Pages 20 - 23
http://dx.doi.org/10.4161/isl.18261
Abstract | Full Text | PDF

This study was designed to determine the effects of 10 and 13 amino acid forms of kisspeptin on dynamic insulin secretion from mammalian islets since it is not clear from published data whether the shorter peptide is stimulatory while the longer peptide inhibits insulin release. Insulin secretion was measured by radioimmunoassay following perifusion of human, pig, rat and mouse isolated islets with kisspeptin-10 or kisspeptin-13 in the presence of 20 mM glucose. Both peptides stimulated rapid, reversible potentiation of glucose-stimulated insulin secretion from islets of all species tested. These data indicate that both kisspeptin-10 and kisspeptin-13, which is an extension of kisspeptin-10 by three amino acids, act directly at islet β-cells of various species to potentiate insulin secretion, and suggest that inhibitory effects reported in earlier studies may reflect differences in experimental protocols.




RESEARCH PAPERS

Nerve growth factor is associated with islet graft failure following intraportal transplantation

Yukihiko Saito, Nathaniel K. Chan, Naoaki Sakata and Eba Hathout
Pages 24 - 31
http://dx.doi.org/10.4161/isl.18467
Abstract | Full Text | PDF

Nerve growth factor (NGF) has recently been recognized as an angiogenic factor with an important regulatory role in pancreatic β-cell function. We previously showed that treatment of pancreatic islets with NGF improved their quality and viability. Revascularization and survival of islets transplanted under the kidney capsule were improved by NGF. However, the usefulness of NGF in intraportal islet transplantation was not previously tested. To resolve this problem, we transplanted syngeneic islets (360 islet equivalents per recipient) cultured with or without NGF into the portal vein of streptozotocin-induced diabetic BALB/c mice. Analysis revealed that 44.4% (4/9) of control and 12.5% (1/8) of NGF-treated mice attained normoglycemia (≤ 200 mg/dL) (p = 0.195). NGF-treated islets led to worse graft function (area under the curve of intraperitoneal glucose tolerance tests (IPGTT) on post-operative day (POD) 30, control; 35,800 ± 3,960 min*mg/dl, NGF-treated; 47,900 ± 3,220 min*mg/dl: *p = 0.0348). NGF treatment of islets was also associated with increased graft failure [the percentage of TdT-mediated dUTP-biotin nick-end labeling (TUNEL)-positive and necrotic transplanted islets on POD 5, control; 23.8% (5/21), NGF-treated; 52.9% (9/17): p = 0.0650] following intraportal islet transplantation. Nonviable (TUNEL-positive and necrotic) islets in both groups expressed vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α). On the other hand, viable (TUNEL-negative and not necrotic) islets in both groups did not express VEGF and HIF-1α. In the present study, pre-transplant NGF treatment was associated with impaired survival and angiogenesis of intraportal islet grafts. The effect of NGF on islet transplantation may significantly vary according to the transplant site.

Preservation of pancreatic islets in cold UW solution before transplantation

Show Ishii, Takuro Saito, Kazuya Ise, Michitoshi Yamashita, Yoshihiro Sato, Takaharu Saito, Manabu Tsukada, Ikuro Oshibe, Akira Kenjo, Takashi Kimura, Takayuki Anazawa, Shigeya Suzuki and Mitsukazu Gotoh
Pages 32 - 39
http://dx.doi.org/10.4161/isl.18607
Abstract | Full Text | PDF

Culture of islets prior to transplantation needs to be revisited for maintaining functional islet capacity. This study was conducted to compare cold UW (University of Wisconsin) preservation with conventional culture based on insulin secretory capacity in vitro and in vivo. Islets isolated from Wistar rats were either cultured for 24 h at 37°C in RPMI1640 medium or DMEM containing various concentrations of glucose or preserved for the same period in UW solution or in DMEM solution at 4°C. The islet yield in UW group, but not in other groups, was maintained as comparable with that of fresh islets. Insulin secretory capacity in response to glucose was maintained only in the islets of UW group, but not in other groups. SCID mice given 300 IEQ islets of UW group showed gradual restoration of normoglycemia as found in the mice given freshly isolated islets. Meanwhile, those mice given cultured islets for 24 h at 37°C in RPMI1640 medium showed rapid decrease of blood glucose levels on day 1 followed by relatively elevated levels on day 2, suggesting unstable insulin secretory capacity of islets.

 

Morphological staining with anti-HMGB1 (high mobility group B1) antibody revealed central damage of islets in all culture groups regardless of glucose concentration and in islets of cold DMEM group, whereas those in the UW group were quite intact. These results suggest that cold preservation in UW solution is simple and beneficial in protecting islets morphologically and functionally before transplantation.

Short-term effects of INGAP and Reg family peptides on the appearance of small β-cells clusters in non-diabetic mice

Rahul Kapur, Tine Westergaard Højfeldt, Sif Groth Rønn, Allan E. Karlsen and Scott Heller
Pages 40 - 48
http://dx.doi.org/10.4161/isl.18659
Abstract | Full Text | PDF

The Reg3 peptides INGAP-PP and human Reg3α/β (HIP) have been hypothesized to stimulate β-cell neogenesis in the pancreas. Administration of INGAP-PP has been shown to cause an increase in β-cell mass in multiple animal models, reverse streptozotocin (STZ) induced diabetes in mice and reduces HbA1c levels in type 2 diabetic humans. In this study, we have examined the ability of the INGAP-PP and HIP peptides to induce β-cell formation in vivo in normal mice through short-term administration of the peptides. We assessed the peptides ability to induce an increase in extra-islet insulin-positive cell clusters by looking at β-cell number by point counting morphometry on pancreata that had been randomized using the smooth fractionator principle in non-diabetic NMRI mice after short-term injections of the peptides (5 d). Five day continuous BrdU labeling was used to determine if the new β-cells were derived from replicating β-cells. Real time quantitative RT-PCR and immuno-histochemistry was used to analyze changes in pancreatic transcription factor expression. A 1.5- to 2-fold increase in the volume of small extra-islet insulin-positive clusters post 5 d treatment with INGAP-PP and HIP as compared with mice treated with a non-peptide control or scrambled peptide (p < 0.05) (n = 7) was found. Five day continuous BrdU infusion during the 5 d period showed little or no incorporation in islets or small insulin clusters. Five days of treatment with INGAP-PP or HIP, showed a tendency toward increased levels of pancreatic progenitor markers such as Ngn3, Nkx6.1, Sox9 and Ins. These are the first studies to compare and indicate that the human Reg3 α/β (HIP) peptide has similar bioactivity in vivo as INGAP by causing formation of small β-cell clusters in extra-islet pancreatic tissue after only 5 d of treatment. Upregulation of pancreatic transcription factors may be part of the mechanism of action.

Adhesion molecule CADM1 contributes to gap junctional communication among pancreatic islet α-cells and prevents their excessive secretion of glucagon

Akihiko Ito, Naoki Ichiyanagi, Yuki Ikeda, Man Hagiyama, Takao Inoue, Keiko B. Kimura, Minami A. Sakurai, Kazuyuki Hamaguchi and Yoshinori Murakami
Pages 49 - 55
http://dx.doi.org/10.4161/isl.18675
Abstract | Full Text | PDF

Cell adhesion molecule-1 (CADM1) is a recently identified adhesion molecule of pancreatic islet α-cells that mediates nerve–α-cell interactions via trans-homophilic binding and serves anatomical units for the autonomic control of glucagon secretion. CADM1 also mediates attachment between adjacent α-cells. Since gap junctional intercellular communication (GJIC) among islet cells is essential for islet hormone secretion, we examined whether CADM1 promotes GJIC among α-cells and subsequently participates in glucagon secretion regulation. Dye transfer assays using αTC6 mouse α-cells, which endogenously express CADM1, supported this possibility; efficient cell-to-cell spread of gap junction-permeable dye was detected in clusters of αTC6 cells transfected with nonspecific, but not with CADM1-targeting, siRNA. Immunocytochemical analysis of connexin 36, a major component of the gap junction among αTC6 cells, revealed that it was localized exclusively to the cell membrane in CADM1-non-targeted αTC6 cells, but diffusely to the cytoplasm in CADM1-targeted cells. Next, we incubated CADM1-targeted and non-targeted αTC6 cells in a medium containing 1 mM glucose and 200 mM arginine for 30 min to induce glucagon secretion, and found that the targeted cells secreted three times more glucagon than did the non-targeted. We conducted similar experiments using pancreatic islets that were freshly isolated from wild-type and CADM1-knockout mice, and expressed glucagon secretion as ratios relative to baseline values. The increase in ratio was larger in CADM1-knockout islets than in wild-type islets. These results suggest that CADM1 may serve as a volume limiter of glucagon secretion by sustaining α-cell attachment necessary for efficient GJIC.

Insulin-secreting INS-1E cells express functional TRPV1 channels

Amanda Jabin Fågelskiöld, Kristina Kannisto, Anna Boström, Banina Hadrovic, Cecilia Farre, Mohamed Eweida, Kenneth Wester and Md. Shahidul Islam
Pages 56 - 63
http://dx.doi.org/10.4161/isl.18915
Abstract | Full Text | PDF

We have studied whether functional TRPV1 channels exist in the INS-1E cells, a cell type used as a model for β-cells, and in primary β-cells from rat and human. The effects of the TRPV1 agonists capsaicin and AM404 on the intracellular free Ca²⁺ concentration ([Ca²⁺]_i) in the INS-1E cells were studied by fura-2 based microfluorometry. Capsaicin increased [Ca²⁺]_i in a concentration-dependent manner, and the [Ca²⁺]_i increase was dependent on extracellular Ca²⁺. AM404 also increased [Ca²⁺]_i in the INS-1E cells. Capsazepine, a specific antagonist of TRPV1, completely blocked the capsaicin- and AM404-induced [Ca²⁺]_i increases. Capsaicin did not increase [Ca²⁺]_i in the primary β-cells from rat and human. Whole cell patch clamp configuration was used to record currents across the plasma membrane in the INS-1E cells. Capsaicin elicited inward currents that were inhibited by capsazepine. Western blot analysis detected TRPV1 proteins in the INS-1E cells and the human islets. Immunohistochemistry was used to study the expression of TRPV1, but no TRPV1 protein immunoreactivity was detected in the human islet cells and the human insulinoma cells. We conclude that the INS-1E cells, but not the primary β-cells, express functional TRPV1 channels.

Open Access Article

Glucose decreases extracellular adenosine levels in isolated mouse and rat pancreatic islets

Gary K. Yang, Paul E. Squires, Faming Tian, Timothy J. Kieffer, Yin Nam Kwok and Nicholas Dale
Pages 64 - 70
http://dx.doi.org/10.4161/isl.19037
Abstract | Full Text | PDF

The pancreatic islets of Langerhans are responsible for the regulated release of the endocrine hormones insulin and glucagon that participate in the control of glucose homeostasis. Abnormal regulation of these hormones can result in glucose intolerance and lead to the development of diabetes. Numerous efforts have been made to better understand the physiological regulators of insulin and glucagon secretion. One of these regulators is the purine nucleoside, adenosine. Though exogenous application of adenosine has been demonstrated to stimulate glucagon release and inhibit insulin release, the physiological significance of this pathway has been unclear. We used a novel 7 µm enzyme-coated electrode biosensor to measure adenosine levels in isolated rodent islets. In the mouse islets, basal adenosine levels in the presence of 3 mM glucose were estimated to be 5.7 ± 0.6 µM. As glucose was increased, extracellular adenosine diminished. A 10-fold increase of extracellular KCl increased adenosine levels to 16.4 ± 2.0 µM. This release required extracellular Ca²⁺ suggesting that it occurred via an exocytosis-dependent mechanism. We also found that while rat islets were able to convert exogenous ATP into adenosine, mouse islets were unable to do this. Our study demonstrates for the first time the basal levels of adenosine and its inverse relationship to extracellular glucose in pancreatic islets.




ARTICLE ADDENDA

Development of type 2 diabetes caused by a deficiency of a tRNA^lys modification

Fan-Yan Wei and Kazuhito Tomizawa
Pages 71 - 73
http://dx.doi.org/10.4161/isl.18262
Abstract | Full Text | PDF

Genetic variations in the cdk5 regulator associated protein 1-like 1 (cdkal1) gene have been identified in whole genome association studies as a risk factor for the development of type 2 diabetes (T2D). A recent study showed that Cdkal1 was a mammalian methythiotransferase, which specifically synthesizes 2-methylthio-N⁶-threonylcarbamoyladenosine (ms²t⁶A) at position 37 of tRNA^lys(UUU). The ms²t⁶A modification in tRNA^lys(UUU) was important for the accurate decoding of its cognate codon. In pancreatic β-cell-specific Cdkal1 knockout (Cdkal1 KO) mice, a deficiency of ms²t⁶A caused the mistranslation of a Lys codon in proinsulin, resulting in improper processing. The mice showed a decrease in insulin secretion and glucose intolerance. In addition, the mistranslation contributed to the expression of the endoplasmic reticulum (ER) stress response in Cdkal1-deficient β-cells. Furthermore, Cdkal1 KO mice were hypersensitive to high-fat diet-induced glucose intolerance, as well as the ER stress response. These findings might potentially explain the molecular pathogenesis of T2D in patients carrying Cdkal1 variations.

Protein farnesylation is requisite for mitochondrial fuel-induced insulin release: Further evidence to link reactive oxygen species generation to insulin secretion in pancreatic β-cells

Andrea Matti, Chandrashekara Kyathanahalli and Anjaneyulu Kowluru
Pages 74 - 77
http://dx.doi.org/10.4161/isl.19121
Abstract | Full Text | PDF

Several lines of recent evidence implicate regulatory roles for reactive oxygen species (ROS) in islet function and insulin secretion. The phagocyte-like NADPH oxidase (Nox2) has recently been shown to be one of the sources of ROS in the signaling events leading to glucose stimulated insulin secretion (GSIS). We recently reported inhibition of glucose- or mitochondrial fuel-induced Nox2-derived ROS by a specific inhibitor of protein farnesyl transferse (FTase; FTI-277), suggesting that activation of FTase might represent one of the upstream signaling events to Nox2 activation. Furthermore, FTase inhibitors (FTI-277 and FTI-2628) have also been shown to attenuate GSIS in INS 832/13 cells and normal rodent islets. Herein, we provide further evidence to suggest that inhibition of FTase either by pharmacological (e.g., FTI-277) or gene silencing (siRNA-FTase) approaches markedly attenuates mitochondrial fuel-stimulated insulin secretion (MSIS) in INS 832/13 cells. Together, our findings further establish a link between nutrient-induced Nox2 activation, ROS generation and insulin secretion in the pancreatic β-cell.