Overexpression of a dominant negative GIP receptor in transgenic mice results in disturbed postnatal pancreatic islet and beta-cell development

The expression of a dominant negative glucose-dependent insulinotropic polypeptide receptor (GIPRdn) under the control of the rat pro-insulin gene promoter induces severe diabetes mellitus in transgenic mice. This study aims to gain further insight into the effect of the expression of a dominant negative GIPR on glucose homeostasis and postnatal development of the endocrine pancreas. The diabetic phenotype of GIPRdn transgenic animals was first observed between 14 and 21 days of age (urine glucose>1000 mg/dl). After onset of diabetes, serum glucose was significantly higher and insulin values were significantly lower in GIPRdn transgenic mice vs. non-transgenic littermate controls. Morphometric studies of pancreatic islets and their endocrine cell types were carried out at 10, 30 and 90 days of age. The total islet and total beta-cell volume of transgenic mice was severely reduced as compared to control mice, irrespective of the age at sampling (p<0.05). The total volume of isolated insulin positive cells that were not contained within established islets was significantly reduced in transgenic mice, indicating disturbed islet neogenesis. These findings demonstrate in vivo evidence that intact signaling of G-protein coupled receptors is involved in postnatal islet and beta-cell development and neogenesis of the pancreatic islets.     

Ex vivo gene transfer of viral interleukin-10 to BB rat islets: no protection after transplantation to diabetic BB rats

Allogeneic and autoimmune islet destruction limits the success of islet transplantation in autoimmune diabetic patients. This study was designed to investigate whether ex vivo gene transfer of viral interleukin-10 (vIL-10) protects BioBreeding (BB) rat islets from autoimmune destruction after transplantation into diabetic BB recipients. Islets were transduced with adenoviral constructs (Ad) expressing the enhanced green fluorescent protein (eGFP), alpha-1 antitrypsin (AAT) or vIL-10. Transduction efficiency was demonstrated by eGFP-positive cells and vIL-10 production. Islet function was determined in vitro by measuring insulin content and insulin secretion and in vivo by grafting AdvIL-10-transduced islets into syngeneic streptozotocin (SZ)-diabetic, congenic Lewis (LEW.1 W) rats. Finally, gene-modified BB rat islets were grafted into autoimmune diabetic BB rats. Ad-transduction efficiency of islets increased with virus titre and did not interfere with insulin content and insulin secretion. Ad-transduction did not induce Fas on islet cells. AdvIL-10-transduced LEW.1 W rat islets survived permanently in SZ-diabetic LEW.1 W rats. In diabetic BB rats AdvIL-10-transduced BB rat islets were rapidly destroyed. Prolongation of islet culture prior to transplantation improved the survival of gene-modified islets in BB rats. Several genes including those coding for chemokines and other peptides associated with inflammation were down-regulated in islets after prolonged culture, possibly contributing to improved islet graft function in vivo. Islets transduced ex vivo with vIL-10 are principally able to cure SZ-diabetic rats. Autoimmune islet destruction in diabetic BB rats is not prevented by ex vivo vIL-10 gene transfer to grafted islets. Graft survival in autoimmune diabetic rats may be enhanced by improvements in culture conditions prior to transplantation.     

Hormone secretion and glucose metabolism in islets of Langerhans of the isolated perfused pancreas from normal and streptozotocin diabetic rats.

The glucose responsiveness of alpha- and beta-cells of normal as well as untreated and insulin-treated streptozotocin diabetic rats was tested in the extracorporeal perfusion system. Also assessed was the possible in vitro effect of added insulin on the glucose sensitivity of islets from untreated diabetic animals. Insulin and glucose responsiveness of the two cell types. The rate of glucose entry islet tissue was estimated, and the effect of glucose on the tissue supply of ATP and lactate and the cyclic 3':5'-AMP level of islets was measured under the above in vitro conditions. It was demonstrated that beta-cells are more accessible to glucose than alpha-cells, that glucose entry into islet cells is not significantly modified by insulin and that glucose had no effect on ATP, lactate and cyclic 3':5'-AMP levels of islet tissue under any of the conditions investigated. High insulin in vitro elevated ATP levels of alpha-cell islets independent of extracellular glucose. Glucose caused insulin release from normal but not from diabetic islets and rapidly and efficiently suppressed stimulated glucagon secretion of the pancreas from normal and insulin treated diabetic rats. Glucose was less effective in inhibiting stimulated glucagon secretion by the pancreas from untreated diabetic rats whether insulin was added to the perfusion media or not. Therefore, profound differences of glucose responsiveness of alpha-cells fail to manifest themselves in alterations of basic parameters of glucose and energy metabolism in contrast to what had been postulated in the literature. It is however, apparent that the glucose responsiveness of alpha-cells is modified by insuling by an as yet undefined mechanism.     

CART knock out mice have impaired insulin secretion and glucose intolerance, altered beta cell morphology and increased body weight

CART peptides are anorexigenic and are widely expressed in the central and peripheral nervous systems, as well as in endocrine cells in the pituitary, adrenal medulla and the pancreatic islets. To study the role of CART in islet function, we used CART null mutant mice (CART KO mice) and examined insulin secretion in vivo and in vitro, and expression of islet hormones and markers of beta-cell function using immunocytochemistry. We also studied CART expression in the normal pancreas. In addition, body weight development and food intake were documented. We found that in the normal mouse pancreas, CART was expressed in numerous pancreatic nerve fibers, both in the exocrine and endocrine portion of the gland. CART was also expressed in nerve cell bodies in the ganglia. Double immunostaining revealed expression in parasympathetic (vasoactive intestinal polypeptide (VIP)-containing) and in fewer sensory fibers (calcitonin gene-related peptide (CGRP)-containing). Although the expression of islet hormones appeared normal, CART KO islets displayed age dependent reduction of pancreatic duodenal homeobox 1 (PDX-1) and glucose transporter-2 (GLUT-2) immunoreactivity, indicating beta-cell dysfunction. Consistent with this, CART KO mice displayed impaired glucose-stimulated insulin secretion both in vivo after an intravenous glucose challenge and in vitro following incubation of isolated islets in the presence of glucose. The impaired insulin secretion in vivo was associated with impaired glucose elimination, and was apparent already in young mice with no difference in body weight. In addition, CART KO mice displayed increased body weight at the age of 40 weeks, without any difference in food intake. We conclude that CART is required for maintaining normal islet function in mice.     

Reduced beta-cell mass and altered glucose sensing impair insulin-secretory function in betaIRKO mice

Pancreatic beta-cell-restricted knockout of the insulin receptor results in hyperglycemia due to impaired insulin secretion, suggesting that this cell is an important target of insulin action. The present studies were undertaken in beta-cell insulin receptor knockout (betaIRKO) mice to define the mechanisms underlying the defect in insulin secretion. On the basis of responses to intraperitoneal glucose, approximately 7-mo-old betaIRKO mice were either diabetic (25%) or normally glucose tolerant (75%). Total insulin content was profoundly reduced in pancreata of mutant mice compared with controls. Both groups also exhibited reduced beta-cell mass and islet number. However, insulin mRNA and protein were similar in islets of diabetic and normoglycemic betaIRKO mice compared with controls. Insulin secretion in response to insulin secretagogues from the isolated perfused pancreas was markedly reduced in the diabetic betaIRKOs and to a lesser degree in the nondiabetic betaIRKO group. Pancreatic islets of nondiabetic betaIRKO animals also exhibited defects in glyceraldehyde- and KCl-stimulated insulin release that were milder than in the diabetic animals. Gene expression analysis of islets revealed a modest reduction of GLUT2 and glucokinase gene expression in both the nondiabetic and diabetic mutants. Taken together, these data indicate that loss of functional receptors for insulin in beta-cells leads primarily to profound defects in postnatal beta-cell growth. In addition, altered glucose sensing may also contribute to defective insulin secretion in mutant animals that develop diabetes.     

Inactivation of the dual Bmp/Wnt inhibitor Sostdc1 enhances pancreatic islet function

Current endeavors in the type 2 diabetes (T2D) field include gaining a better understanding of extracellular signaling pathways that regulate pancreatic islet function. Recent data suggest that both Bmp and Wnt pathways are operative in pancreatic islets and play a positive role in insulin secretion and glucose homeostasis. Our laboratory found the dual Bmp and Wnt antagonist Sostdc1 to be upregulated in a mouse model of islet dysmorphogenesis and nonimmune-mediated lean diabetes. Because Bmp signaling has been proposed to enhance β-cell function, we evaluated the role of Sostdc1 in adult islet function using animals in which Sostdc1 was globally deleted. While Sostdc1-null animals exhibited no pancreas development phenotype, a subset of mutants exhibited enhanced insulin secretion and improved glucose homeostasis compared with control animals after 12-wk exposure to high-fat diet. Loss of Sostdc1 in the setting of metabolic stress results in altered expression of Bmp-responsive genes in islets but did not affect expression of Wnt target genes, suggesting that Sostdc1 primarily regulates the Bmp pathway in the murine pancreas. Furthermore, our data indicate that removal of Sostdc1 enhances the downregulation of the closely related Bmp inhibitors Ctgf and Gremlin in islets after 8-wk exposure to high-fat diet. These data imply that Sostdc1 regulates expression of these inhibitors and provide a means by which Sostdc1-null animals show enhanced insulin secretion and glucose homeostasis. Our studies provide insights into Bmp pathway regulation in the endocrine pancreas and reveal new avenues for improving β-cell function under metabolic stress.     

Miglitol (BAY m 1099) Treatment of Diabetic Hypothalamic-Dietary Obese Rats Improves Islet Response to Glucose

AXEN, KATHLEEN V., XUE LI, AND ANTHONY SCLAFANI. Miglitol (BAY m 1099) treatment of diabetic hypothalamic-dietary obese rats improves islet response to glucose. Obes Res. 1999;7:83–89. Objective : The well-absorbed α-glucosidase inhibitor, miglitol (BAY m 1099), was included in the diets of hypothalamic-dietary obese diabetic rats to investigate its ability to improve glycemia and thereby reverse glucotoxic effects on islet secretory response. Research Methods and Procedures : Female rats received bilateral electrolytic lesions of the ventromedial hypothalamus and were fed high-fat, sucrosesupplemented diets until hyperinsulinemia and hyperglycemia were observed after 3 hours of food deprivation (nonfed). Diabetic animals were assigned to miglitol-treated (40 mg/17 g of diet) or untreated groups for 3 weeks; pancreatic islets were isolated for incubation experiments. Results : No differences in food intake, body weights, or nonfed plasma glucose or insulin levels were seen between treated and untreated diabetic rats. Islets isolated from untreated diabetic rats showed elevated basal insulin release and no insulin secretory response to an elevation in glucose concentration. In contrast, islets obtained from miglitol-treated rats showed more normal basal release and a significant insulin secretory response to glucose. Incubation of islets, obtained from normal control rats or untreated diabetic rats, in media containing miglitol at levels estimated to exist in plasma of treated rats had no effect on islet insulin secretory responses to glucose. Discussion : Islet secretory response was improved despite continued hyperglycemia and severe insulin resistance. Miglitol treatment may improve islet sensitivity to glucose either through effects on islet metabolism requiring prolonged exposure or by improvement in postmeal glycemia, despite persistent hyperglycemia.     

Persistent expression of HNF6 in islet endocrine cells causes disrupted islet architecture and loss of beta cell function

We used transgenesis to explore the requirement for downregulation of hepatocyte nuclear factor 6 (HNF6) expression in the assembly, differentiation, and function of pancreatic islets. In vivo, HNF6 expression becomes downregulated in pancreatic endocrine cells at 18. 5 days post coitum (d.p.c.), when definitive islets first begin to organize. We used an islet-specific regulatory element (pdx1(PB)) from pancreatic/duodenal homeobox (pdx1) gene to maintain HNF6 expression in endocrine cells beyond 18.5 d.p.c. Transgenic animals were diabetic. HNF6-overexpressing islets were hyperplastic and remained very close to the pancreatic ducts. Strikingly, alpha, delta, and PP cells were increased in number and abnormally intermingled with islet beta cells. Although several mature beta cell markers were expressed in beta cells of transgenic islets, the glucose transporter GLUT2 was absent or severely reduced. As glucose uptake/metabolism is essential for insulin secretion, decreased GLUT2 may contribute to the etiology of diabetes in pdx1(PB)-HNF6 transgenics. Concordantly, blood insulin was not raised by glucose challenge, suggesting profound beta cell dysfunction. Thus, we have shown that HNF6 downregulation during islet ontogeny is critical to normal pancreas formation and function: continued expression impairs the clustering of endocrine cells and their separation from the ductal epithelium, disrupts the spatial organization of endocrine cell types within the islet, and severely compromises beta cell physiology, leading to overt diabetes.     

Cyproheptadine and beta cell function in the rat: insulin secretion from pancreas segments in vitro.

Pancreatic islet cell vacuolization, hyperglycemia, and glucose intolerance develop in rats after oral administration of cyproheptadine (CPH). In order to determine whether these effects were associated with abnormal insulin secretion, pancreas segments from CPH-treated and control rats were compared for their ability to secrete insulin in response to several stimuli. Oral administration of CPH (45 mg/kg/day) to rats for 1 or 8 days inhibited glucose-mediated insulin secretion from pancreas segments obtained 3 and 24 hr after the last dose of the drug. Insulin secretion had returned to normal by 48 hr after drug administration. Intraperitoneal administration of the drug was less effective than oral administration in inhibiting in vitro insulin secretion. Other stimuli for insulin secretion (tolbutamide, glucagon, L-leucine, and dibutyryl 3',5'cyclic AMP), like glucose, were incapable of releasing normal amounts of insulin from pancreas segments of CPH-treated rats. CPH and a metabolite, desmethyl-CPH, inhibited glucose-stimulated insulin secretion when added in vitro to pancreas segments from control rats. This suggests that the inhibition of insulin secretion in pancreas segments taken from animals treated with CPH could be due, at least in part, to the presence of drug and its metabolite in the tissue. A previously observed reduction in the pancreatic content of insulin in CPH-treated rats may also contribute to the abnormal insulin release in animals given the drug.     

Distribution of vasoactive intestinal polypeptide, neuropeptide-Y and substance P and their effects on insulin secretion from the in vitro pancreas of normal and diabetic rats

This study examined the pattern of distribution of vasoactive intestinal polypeptide (VIP), neuropeptide-Y (NPY) and substance P (SP) in the pancreas of diabetic rat to determine whether there are changes in the number and pattern of distribution of these neuropeptides after the onset of diabetes. Moreover, the effect of VIP, NPY and SP on insulin secretion from the pancreas of normal and diabetic rats was also examined. Diabetes mellitus (DM) was induced by a single dose of streptozotocin (STZ) given intraperitoneally (i.p.) (60 mg kg body weight(-1)). Four weeks after the induction of DM, diabetic (n = 6) and normal (n = 6) rats were anesthetized with chloral hydrate and their pancreases removed and processed for immunohistochemistry and insulin secretion. The number of insulin-positive cells in the islets of Langerhans was reduced while that of VIP and NPY increased significantly after the onset of diabetes. The pattern of distribution of VIP, NPY and SP in the nerves innervating the pancreas was similar in both normal and diabetic rats. VIP-evoked large and significant (P < 0.02) increases in insulin secretion from the pancreas of normal and diabetic rats. NPY also induced a marked (P < 0.005) increase in insulin release from pancreatic tissue fragments of normal rat. Stimulation of pancreatic tissue fragments of diabetic rat with NPY resulted in a slight but not significant increase in insulin release. SP induced a large and significant (P < 0.005) increase in insulin secretion from the pancreas of normal rat but inhibited insulin secretion significantly (P < 0.03) from isolated pancreas of diabetic rat. In summary, VIP and NPY can stimulate insulin secretion from the pancreas after the onset of diabetes. The stimulatory effect of SP on insulin secretion is reversed to inhibitory in diabetic rats.     

The role of old age in the effects of glucose on insulin secretion, pentosephosphate shunt activity, pyridine nucleotides and glutathione of rat pancreatic islets

In pancreatic islets of adult (three month) and old (24 month) rats the effect of glucose on glucose oxidation, pyridine nucleotides, glutathione and insulin secretion was studied. DNA content was similar in both groups of animals; however, islets of old rats exhibited 30% less insulin content. While glucose-induced (16.7 mM) insulin secretion in islets of old rats was approximately 50% less than in islets of adults, no significant difference was observed in the insulin releasing effect of theophylline (1 mM). Although islet production of 14CO2 in the presence of 16.7 mM glucose increased equally in both groups, elevation of glucose failed to increase the percentage of total glucose oxidation via the pentose phosphate shunt in islets of old rats. Elevation of glucose increased the NADPH/NADP and the NADH/NAD ratio in both groups of islets in a similar manner. The effect of glucose on the GSH/GSSG ratio revealed a dose-related increase in the islets of adult rats, whereas islets of old rats did not respond to elevation of glucose. Our data seem to indicate that the lower secretory response of islets of old rats is related to the failure of glucose to increase the GSH/GSSG ratio. In contrast the insulin release induced by theophylline does not appear to depend on islet thiols.     

Expression and Regulation of Nampt in Human Islets

Nicotinamide phosphoribosyltransferase (Nampt) is a rate-limiting enzyme in the mammalian NAD+ biosynthesis of a salvage pathway and exists in 2 known forms, intracellular Nampt (iNampt) and a secreted form, extracellular Nampt (eNampt). eNampt can generate an intermediate product, nicotinamide mononucleotide (NMN), which has been reported to support insulin secretion in pancreatic islets. Nampt has been reported to be expressed in the pancreas but islet specific expression has not been adequately defined. The aim of this study was to characterize Nampt expression, secretion and regulation by glucose in human islets. Gene and protein expression of Nampt was assessed in human pancreatic tissue and isolated islets by qRT-PCR and immunofluorescence/confocal imaging respectively. Variable amounts of Nampt mRNA were detected in pancreatic tissue and isolated islets. Immunofluorescence staining for Nampt was found in the exocrine and endocrine tissue of fetal pancreas. However, in adulthood, Nampt expression was localized predominantly in beta cells. Isolated human islets secreted increasing amounts of eNampt in response to high glucose (20 mM) in a static glucose-stimulated insulin secretion assay (GSIS). In addition to an increase in eNampt secretion, exposure to 20 mM glucose also increased Nampt mRNA levels but not protein content. The secretion of eNampt was attenuated by the addition of membrane depolarization inhibitors, diazoxide and nifedipine. Islet-secreted eNampt showed enzymatic activity in a reaction with increasing production of NAD+/NADH over time. In summary, we show that Nampt is expressed in both exocrine and endocrine tissue early in life but in adulthood expression is localized to endocrine tissue. Enzymatically active eNampt is secreted by human islets, is regulated by glucose and requires membrane depolarization.     

The duct-ligated pancreas transplant and its effect on the islet cellular composition of the host pancreas

Summary Rats rendered diabetic by streptozotocin were subjected to pancreas transplantation. After twenty weeks, the duct-ligated pancreas transplant was studied morphometrically to determine the effect of duct occlusion on the various cell populations of the islets. Concomitantly, the streptozotocin-treated host pancreas was examined for a possible influence of the graft on the diabetic pattern of islet cell population. Twenty weeks after pancreas transplantation, the volume fractions of insulin, glucagon, somatostatin and pancreatic polypeptide cells in the graft islets did not differ from those of the normal control pancreas. In the pancreas of nontransplanted diabetic rats, insulin-positive B cells were reduced from 60–65% to less than 10% of the islet volume, whereas non-B cells were significantly increased in volume density. The changes in fractional volume of the various islet cells correlated fairly well with changes in plasma concentration of the corresponding pancreas hormones. In the recipient's own pancreas, the relative volumes of glucagon and somatostatin cells were unaffected by the pancreas transplant. However, the insulin cell mass was significantly increased, and comprised about 20% of the islet volume, while cells containing pancreatic polypeptide were found only sporadically.Supported by Nordic Insulin Fund, The Swedish Diabetes Association, and MFR, proj. no. 4499. The technical assistance by M. Maxe and M. Carlesson is gratefully acknowledged     

Oral administration of soybean peptide Vglycin normalizes fasting glucose and restores impaired pancreatic function in Type 2 diabetic Wistar rats

Vglycin, a natural 37-residue polypeptide isolated from pea seeds in which six half-cysteine residues are embedded in three pairs of disulfide bonds, is resistant to digestive enzymes and has antidiabetic potential. To investigate the pharmacological activity of Vglycin in vivo and to examine the mechanisms involved, the therapeutic effect of Vglycin in diabetic rats was examined. Diabetes was induced in Wistar rats by high-fat diet and multiple streptozotocin intraperitoneal injections. Diabetic rats were treated daily with Vglycin for 4 weeks. Body weight, food intake, fasting plasma glucose and insulin levels were assayed weekly. Glucose and insulin tolerance tests were conducted on Day 29. Subsequently, levels of p-Akt in the liver and pancreas and cleaved PARP, Pdx-1 and insulin in the pancreas were detected by immunoblotting. The morphology of the pancreas and the insulin expression in the pancreas were analyzed by hematoxylin–eosin staining and immunohistochemistry, respectively. Furthermore, human liver-derived cell lines were used to explore the in vitro effects of Vglycin on insulin sensitivity and glucose uptake. Chronic treatment with Vglycin normalized fasting glucose levels in diabetic rats. The improvement in glucose homeostasis and the increased insulin sensitivity mediated by restored insulin signaling likely contributed to decreased food intake and reduced body weight. Vglycin protected pancreatic cells from damage by streptozotocin. Although insulin synthesis and secretion in impaired β-cell were not significantly elevated, islets morphology was improved in the Vglycin-treated groups. These results suggest that Vglycin could be useful in Type 2 diabetes for restoring impaired insulin signaling, glucose tolerance and pancreatic function.     

Ghrelin is expressed in a novel endocrine cell type in developing rat islets and inhibits insulin secretion from INS-1 (832/13) cells.

Ghrelin is produced mainly by endocrine cells in the stomach and is an endogenous ligand for the growth hormone secretagogue receptor (GHS-R). It also influences feeding behavior, metabolic regulation, and energy balance. It affects islet hormone secretion, and expression of ghrelin and GHS-R in the pancreas has been reported. In human islets, ghrelin expression is highest pre- and neonatally. We examined ghrelin and GHS-R in rat islets during development with immunocytochemistry and in situ hybridization. We also studied the effect of ghrelin on insulin secretion from INS-1 (832/13) cells and the expression of GHS-R in these cells. We found ghrelin expression in rat islet endocrine cells from mid-gestation to 1 month postnatally. Islet expression of GHS-R mRNA was detected from late fetal stages to adult. The onset of islet ghrelin expression preceded that of gastric ghrelin. Islet ghrelin cells constitute a separate and novel islet cell population throughout development. However, during a short perinatal period a minor subpopulation of the ghrelin cells co-expressed glucagon or pancreatic polypeptide. Markers for cell lineage, proliferation, and duct cells revealed that the ghrelin cells proliferate, originate from duct cells, and share lineage with glucagon cells. Ghrelin dose-dependently inhibited glucose-stimulated insulin secretion from INS-1 (832/13) cells, and GHS-R was detected in the cells. We conclude that ghrelin is expressed in a novel developmentally regulated endocrine islet cell type in the rat pancreas and that ghrelin inhibits glucose-stimulated insulin secretion via a direct effect on the beta-cell.     

A general and islet cell-enriched overexpression of IGF-I results in normal islet cell growth, hypoglycemia, and significant resistance to experimental diabetes

Insulin-like growth factor I (IGF-I) is normally produced from hepatocytes and various other cells and tissues, including the pancreas, and is known to stimulate islet cell replication in vitro, prevent Fas-mediated beta-cell destruction and delay the onset of diabetes in nonobese diabetic mice. Recently, however, the notion that IGF-I stimulates islet cell growth has been challenged by the results of IGF-I and receptor gene targeting. To test the effects of a general, more profound increase in circulating IGF-I on islet cell growth and glucose homeostasis, we have characterized MT-IGF mice, which overexpress the IGF-I gene under the metallothionein I promoter. In early reports, a 1.5-fold-elevated serum IGF-I level caused accelerated somatic growth and pancreatic enlargement. We demonstrated that the transgene expression, although widespread, was highly concentrated in the beta-cells of the pancreatic islets. Yet, islet cell percent and pancreatic morphology were unaffected. IGF-I overexpression resulted in significant hypoglycemia, hypoinsulinemia, and improved glucose tolerance but normal insulin secretion and sensitivity. Pyruvate tolerance test indicated significantly suppressed hepatic gluconeogenesis, which might explain the severe hypoglycemia after fasting. Finally, due to a partial prevention of beta-cell death against onset of diabetes and/or the insulin-like effects of IGF-I overexpression, MT-IGF mice (which overexpress the IGF-I gene under the metallothionein I promoter) were significantly resistant to streptozotocin-induced diabetes, with diminished hyperglycemia and prevention of weight loss and death. Although IGF-I might not promote islet cell growth, its overexpression is clearly antidiabetic by improving islet cell survival and/or providing insulin-like effects.     

Functional rest through intensive treatment with insulin and potassium channel openers preserves residual beta-cell function and mass in acutely diabetic BB rats.

Diazoxide and the diazoxide-analogue, NNC 55-0118, are potassium channel openers that interfere with insulin secretion from beta-cells. In vitro, we show that these two drugs inhibit insulin release from diabetes-resistant BB rat islets cultured at either low or high glucose concentration and cause an intracellular accumulation of insulin with high glucose. Preservation of beta-cells was investigated in newly diabetic BB rats treated with insulin implants from day 0-8 under oral diazoxide, NNC 55-0118 or solvent gavage once a day from day 0-7. Three of eight rats (37.5%) treated with diazoxide and three of ten (30%) treated with NNC 55-0118 retained near normal C-peptide responses when challenged with glucose/arginine on day 9, whereas none of eight (0%) solvent-treated rats showed a C-peptide response. Immunohistochemical staining for insulin and glucagon showed that all the C-peptide responding rats had insulin-positive cells in their islets. In contrast, islets from non-responding rats displayed marked inflammation or end-stage lesions. Furthermore, rats with C-peptide response and treated with NNC 55-0118 exhibited only minimal signs of islet inflammation, whereas C-peptide responding diazoxide-treated rats had low level islet inflammation. These results imply that it is conceivable to preserve residual beta-cells at diabetes onset by induction of target cell rest with potassium channel openers and continuous insulin treatment.     

Islet neuronal abnormalities associated with impaired insulin secretion in type 2 diabetes in the Chinese hamster.

This study examined the relationship between islet neurohormonal characteristics and the defective glucose-stimulated insulin secretion in genetic type 2 diabetic Chinese hamsters. Two different sublines were studied: diabetes-prone CHIG hamsters and control CHIA hamsters. The CHIG hamsters were divided into three subgroups, depending on severity of hyperglycemia. Compared to normoglycemic CHIG hamsters and control CHIA hamsters, severely hyperglycemic CHIG hamsters (glucose > 15 mmol/l) showed marked glucose intolerance during i.p. glucose tolerance test and 75% impairment of glucose-stimulated insulin secretion from isolated islets. Mildly hyperglycemic CHIG animals (glucose 7.2-15 mmol/l) showed only moderate glucose intolerance and a 60% impairment of glucose-stimulated insulin secretion from the islets. Immunostaining for neuropeptide Y and tyrosine hydroxylase (markers for adrenergic nerves) and for vasoactive intestinal peptide (marker for cholinergic nerves) revealed significant reduction in immunostaining of islets in the severely but not in the mildly hyperglycemic animals, compared to control CHIA hamsters. The study therefore provides evidence that in this model of type 2 diabetes in Chinese hamsters, severe hyperglycemia is accompanied not only by marked glucose intolerance and islet dysfunction but also by reduced islet innervation. This suggests that islet neuronal alterations may contribute to islet dysfunction in severe but not in mild diabetes.     

Orexin-1 Receptor Co-Localizes with Pancreatic Hormones in Islet Cells and Modulates the Outcome of Streptozotocin-Induced Diabetes Mellitus

Recent studies have shown that orexins play a critical role in the regulation of sleep/wake states, feeding behaviour, and reward processes. The exocrine and endocrine pancreas are involved in the regulation of food metabolism and energy balance. This function is deranged in diabetes mellitus. This study examined the pattern of distribution of orexin-1 receptor (OX₁R) in the endocrine cells of the pancreas of normal and diabetic Wistar (a model of type 1 diabetes), Goto-Kakizaki (GK, a model of type 2 diabetes) rats and in orexin-deficient (OX^−/−) and wild type mice. Diabetes mellitus (DM) was induced in Wistar rats and mice by streptozotocin (STZ). At different time points (12 h, 24 h, 4 weeks, 8 months and 15 months) after the induction of DM, pancreatic fragments of normal and diabetic rats were processed for immunohistochemistry and Western blotting. OX₁R-immunoreactive nerves were observed in the pancreas of normal and diabetic Wistar rats. OX₁R was also discernible in the pancreatic islets of normal and diabetic Wistar and GK rats, and wild type mice. OX₁R co-localized with insulin (INS) and glucagon (GLU) in the pancreas of Wistar and GK rats. The number of OX₁R-positive cells in the islets increased markedly (p<0.0001) after the onset of DM. The increase in the number of OX₁R-positive cells is associated with a high degree of co-localization with GLU. The number of GLU- positive cells expressing OX₁R was significantly (p<0.0001) higher after the onset of DM. The tissue level of OX₁R protein increased with the duration of DM especially in type 1 diabetes where it co-localized with cleaved caspase 3 in islet cells. In comparison to STZ-treated wild type mice, STZ-treated OX^−/− animals exhibited reduced hyperglycemia and handled glucose more efficiently in glucose tolerance test. The findings suggest an important role for the OX-OX₁R pathway in STZ-induced experimental diabetes.     

Viability criteria of islets of Langerhans isolated and purified using Ficoll

Rejection of islet allografts is generally explained by immunologic problems, due to both cellular and antibody mechanisms. But another great problem is in the isolation of intact and viable islets of Langerhans: it is necessary to use a good method of pancreas distention, to determine the optimal concentration of collagenase for digestion, to select an effective technique for purifying the islets. This study correlates the morphology of isolated pancreatic islets of rats and dogs with secretion of insulin. The islets are incubated in a perifusion system and are tested during four periods; the glucose concentrations of the perifusion fluid are: 5.5 mM during the initial 70 min. period, 16.5 mM during the second 60 min. period, 5.5 mM during the third 60 min. period and 16.5 during the fourth 50 min. period. This "double glucose stimulation" is a good test of islet viability. The intact, viable isolated islets showed a significant increase of insulin secretion during the two 16.5 mM glucose periods. Damaged islets with some little morphologic alterations after showed a good insulin release during the first glucose stimulation, but a very poor insulin response to glucose during the second stimulation period.