Compensatory growth of pancreatic beta-cells in adult rats after short-term glucose infusion

The extent to which adult pancreatic beta-cells can respond in vivo to a sustained glucose stimulus by increasing their mass through either hyperplasia or hypertrophy has remained unanswered. Therefore, we studied the in vivo effect of short-term (96-h) hyperglycemia on the growth of beta-cells by infusing adult rats with 35 or 50% glucose or 0.45% saline. After 96 h of glucose infusion, the beta-cell mass, quantified by point-counting morphometrics of immunoperoxidase-stained paraffin sections, showed a 50% increase (9.57 +/- 0.87 mg, n = 5, 50% glucose infused; 9.50 +/- 1.23, n = 7, 35% glucose infused; 6.15 +/- 0.55, n = 6, 0.45% saline infused). This growth was selective for beta-cells; the non-beta-cell mass was unchanged. The mitotic index, measured by accumulated mitotic frequency after a 4-h colchicine treatment, increased fivefold in glucose-infused animals compared to saline-infused animals. This enhanced replication of beta-cells provides evidence for increase in cell number or hyperplasia. In addition, hypertrophy of the beta-cell was also quantified. Mean cell volume, determined from the mean cell cross-sectional area measured planimetrically from low-magnification electron micrographs, increased to 150% of control values after 96 h of 50% glucose infusion. Seven days after the 96-h infusion, in reversal experiments, the beta-cell mass had not returned to saline-infused levels. In addition, the non-beta-cell mass of glucose-infused animals had increased. The mitotic index of the beta-cell of glucose-infused rats was, however, significantly lower than that of the saline controls, but the mean cell volume of the beta-cells remained elevated.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of glucose infusion on cerebral cortical glucose and lactate concentrations during endotoxemia in rats.

Hypoglycemia has been observed in several species before death from endotoxemia. Although several studies have emphasized the importance of maintaining brain glucose at normal concentration during endotoxemia, the effect of glucose infusion on cerebral glucose metabolism has not been studied. Accordingly, the effects of glucose infusion on interstitial glucose and lactate concentrations in the cerebral cortex during endotoxemia were studied in 22 Wistar rats. Cerebral glucose and lactate were measured at 30-min intervals for 4 h using microdialysis. Animals were divided into four groups: 1) saline-infused control (n = 5); 2) saline-infused endotoxemia (n = 7); 3) glucose-infused control (n = 5); and 4) glucose-infused endotoxemia (n = 5). In groups 2 and 4, endotoxemia was induced by intravenous injection with E. coli lipopolysaccharide B (5 mg.100 g-1). One hour after endotoxin administration, saline or 50% glucose was infused at a flow rate of 0.5 ml.100 g-1.h-1 for 3 h. Endotoxin induced a significant increase (P less than 0.05) in blood glucose in the saline-infused rats, which survived for 4 h (n = 5), from 91.6 +/- 15.4 mg.100 ml-1 at baseline to 136.3 +/- 23.3 mg.100 ml-1 (149%) at 1 h, followed by a gradual decrease (to 63% of the basal concentration at 4 h). Similar changes were observed in brain glucose (14.9 +/- 1.9 mg.100 ml-1 baseline, 175% of baseline at 2 h, and 57% of baseline at 4 h).(ABSTRACT TRUNCATED AT 250 WORDS)     

Gastrin stimulates beta-cell neogenesis and increases islet mass from transdifferentiated but not from normal exocrine pancreas tissue

It is still unclear which factors regulate pancreatic regeneration and beta-cell neogenesis and which precursor cells are involved. We evaluated the role of intravenously infused gastrin in regenerating pancreas of duct-ligated rats. The ligation of exocrine ducts draining the splenic half of the pancreas resulted in acinoductal transdifferentiation within the ligated part but not in the unligated part. We found that infusion of gastrin from day 7 to 10 postligation resulted in a doubling of the beta-cell mass in the ligated part as measured by morphometry. This increase in insulin-expressing cells was not associated with increased proliferation, hypertrophy, or reduced cell death of the beta-cells. Furthermore, we found an increased percentage of single, extra-insular beta-cells and small beta-cell clusters induced by gastrin infusion. These changes occurred only in the ligated part of the pancreas, where transdifferentiation of the exocrine acinar cells to ductlike cells (metaplasia) had occurred, and was not found in the normal unaffected pancreatic tissue. In conclusion, we demonstrate that administration of gastrin stimulates beta-cell neogenesis and expansion of the beta-cell mass from transdifferentiated exocrine pancreas.     

Islet neogenesis is stimulated by brief occlusion of the main pancreatic duct.

OBJECTIVE: Current models of islet neogenesis either cause substantial pancreatic damage or continuously stimulate the pancreas, making these models unsuitable for the study of early events that occur in the neogenic process. We aimed to develop a method where the initial events that culminate in increased pancreatic endocrine mass can be studied. DESIGN AND METHODS: Ten 12-week-old female Wistar rats were subjected to a midline laparotomy, the pancreas was isolated and the main pancreatic duct was occluded for 60 seconds. The pancreas was released and carefully relocated within the abdomen. Ten age-, strain- and sex-matched control rats were subjected to a sham operation. The animals were killed 56 days post occlusion, and the pancreata excised and fixed for histological analysis. Body, pancreatic and hepatic weights were noted at termination and serum was taken for analysis. The endocrine-to-exocrine ratio was calculated and the number of endocrine cells in each islet from the sectioned pancreata was counted. RESULTS: Occlusion of the main pancreatic duct for 60 seconds results in an increase in endocrine mass by 80% 56 days post occlusion. This constitutes an increase in endocrine units (1-6 cells), and in small (7-30 cells), medium (31-60 cells) and large (> 60 cells) islets by 85%, 96%, 95% and 71% respectively. CONCLUSION: Brief occlusion of the main pancreatic duct results in an increase in pancreatic endocrine mass. An increase in endocrine units and small islets is indicative of islet neogenesis. Therefore, owing to the briefness of the stimulation, this model can therefore be used to study the initial events that occur during the neogenic process.     

Mechanisms of compensatory beta-cell growth in insulin-resistant rats: roles of Akt kinase

The physiological mechanisms underlying the compensatory growth of beta-cell mass in insulin-resistant states are poorly understood. Using the insulin-resistant Zucker fatty (fa/fa) (ZF) rat and the corresponding Zucker lean control (ZLC) rat, we investigated the factors contributing to the age-/obesity-related enhancement of beta-cell mass. A 3.8-fold beta-cell mass increase was observed in ZF rats as early as 5 weeks of age, an age that precedes severe insulin resistance by several weeks. Closer investigation showed that ZF rat pups were not born with heightened beta-cell mass but developed a modest increase over ZLC rats by 20 days that preceded weight gain or hyperinsulinemia that first developed at 24 days of age. In these ZF pups, an augmented survival potential of beta-cells of ZF pups was observed by enhanced activated (phospho-) Akt, phospho-BAD, and Bcl-2 immunoreactivity in the postweaning period. However, increased beta-cell proliferation in the ZF rats was only detected at 31 days of age, a period preceding massive beta-cell growth. During this phase, we also detected an increase in the numbers of small beta-cell clusters among ducts and acini, increased duct pancreatic/duodenal homeobox-1 (PDX-1) immunoreactivity, and an increase in islet number in the ZF rats suggesting duct- and acini-mediated heightened beta-cell neogenesis. Interestingly, in young ZF rats, specific cells associated with ducts, acini, and islets exhibited an increased frequency of PDX-1+/phospho-Akt+ staining, indicating a potential role for Akt in beta-cell differentiation. Thus, several adaptive mechanisms account for the compensatory growth of beta-cells in ZF rats, a combination of enhanced survival and neogenesis with a transient rise in proliferation before 5 weeks of age, with Akt serving as a potential mediator in these processes.     

A pentadecapeptide fragment of islet neogenesis-associated protein increases beta-cell mass and reverses diabetes in C57BL/6J mice

OBJECTIVE: The objective of this study was to demonstrate that islet neogenesis-associated protein (INGAP) peptide, a pentadecapeptide containing the biologically active portion of native INGAP, increases functional beta-cell mass in normal animals and can be used therapeutically to reverse hyperglycemia in streptozotocin-induced diabetes. SUMMARY BACKGROUND DATA: INGAP, a 175 amino acid pancreatic acinar cell protein, has been suggested to be implicated in beta-cell mass expansion. METHODS: In the first part of this study, normoglycemic hamsters were administered either 500 microg INGAP peptide (n = 30) or saline (n = 20) intraperitoneally daily and sacrificed after 10 or 30 days of treatment. Blood glucose and insulin levels were measured, and a histologic and morphometric analysis of the pancreas was performed to determine the effect of INGAP peptide on the endocrine pancreas. In the second part of the study, 6- to 8-week-old C57BL/6J mice (n = 8) were administered multiple low doses of the beta-cell toxin streptozotocin (STZ) inducing insulitis and hyperglycemia. The mice were then injected with INGAP peptide (n = 4) or saline (n = 4) for 39 days and sacrificed at 48 days. Two additional groups of diabetic mice were administered either a peptide composed of a scrambled sequence of amino acids from INGAP peptide (n = 5) or exendin-4 (n = 5), an incretin that has been associated with amelioration of hyperglycemia. RESULTS: Islet cell neogenesis was stimulated in INGAP-treated hamsters by 10 days. At 30 days, the foci of new endocrine cells had the appearance of mature islets. There was a 75% increase in islet number, with normal circulating levels of blood glucose and insulin. Administration of INGAP peptide to diabetic mice reversed the diabetic state in all animals, and this was associated with increased expression of PDX-1 in duct cells and islet cell neogenesis with a reduction of insulitis in the new islets. Diabetic mice treated with exendin-4 or a scrambled INGAP peptide did not revert from hyperglycemia. CONCLUSION: Because there is a deficiency of beta-cell mass in both type-1 and type-2 diabetes, INGAP peptide stimulation of fully functional neoislet differentiation may provide a novel approach for diabetes therapy.     

Beta-cell mass dynamics in Zucker diabetic fatty rats. Rosiglitazone prevents the rise in net cell death

The evolution of diabetes in the male leptin receptor-deficient (fa/fa) Zucker diabetic fatty (ZDF) rat is associated with disruption of normal islet architecture, beta-cell degranulation, and increased beta-cell death. It is unknown whether these changes precede or develop as a result of the increasing plasma glucose, or whether the increased beta-cell death can be prevented. Early intervention with thiazolidinediones prevents disruption of the islet architecture. To determine the specific effects of rosiglitazone (RSG) on beta-cell mass dynamics, male fa/fa (obese) and +/fa or +/+ (lean) rats age 6 weeks were fed either chow (control group [CN]) or chow mixed with rosiglitazone (RSG group) at a dosage of 10 micromol. kg(-1) body wt.day(-1). Rats were killed after 0, 2, 4, 6, or 10 weeks of treatment (at age 6, 8, 10, 12, or 16 weeks). Plasma glucose increased from 8.9 +/- 0.4 mmol/l at 0 weeks to 34.2 +/- 1.8 mmol/l (P = 0.0001) at 6 weeks of treatment in obese CN rats and fell from 8.0 +/- 0.3 to 6.3 +/- 0.4 mmol/l in obese RSG rats (P = 0.02). beta-cell mass fell by 51% from 2 to 6 weeks of treatment (ages 8-12 weeks) in obese CN rats (6.9 +/- 0.9 to 3.4 +/- 0.5 mg; P < 0.05), whereas beta-cell mass was unchanged in obese RSG rats. At 10 weeks of treatment (age 16 weeks), beta-cell mass in obese CN rats was only 56% of that of obese RSG rats (4.4 +/- 0.4 vs. 7.8 +/- 0.3 mg, respectively; P = 0.0001). The beta-cell replication rate fell from a baseline value of 0.95 +/- 0.12% in lean rats and 0.94 +/- 0.07% in obese rats (at 0 weeks) to approximately 0.3-0.5% in all groups by 6 weeks of treatment (age 12 weeks). After 10 weeks of treatment, beta-cell replication was higher in obese RSG rats than in CN rats (0.59 +/- 0.14 vs. 0.28 +/- 0.05%, respectively; P < 0.02). Application of our mass balance model of beta-cell turnover indicated that net beta-cell death was fivefold higher in obese CN rats as compared with RSG rats after 6 weeks of treatment (age 12 weeks). The increase in beta-cell death in obese CN rats during the 6-week observation period was well correlated with the increase in plasma glucose (r2 = 0.90, P < 0.0001). These results suggest that the development of hyperglycemia in ZDF rats is concomitant with increasing net beta-cell death. beta-cell proliferation compensates for the increased beta-cell loss at a time when plasma glucose is moderately elevated, but compensation ultimately fails and the plasma glucose levels increase beyond approximately 20 mmol/l. Treatment with rosiglitazone, previously shown to reduce insulin resistance, prevents the loss of beta-cell mass in obese ZDF rats by maintaining beta-cell proliferation and preventing increased net beta-cell death.     

Cerebral extracellular glucose and lactate concentrations during and after moderate hypoxia in glucose- and saline-infused rats.

To assess the effect of a glucose infusion on brain extracellular fluid (ECF) during systemic hypoxia, changes in glucose and lactate concentrations in cerebral ECF during and after moderate hypoxic hypoxia were measured in adult, conscious, unrestrained rats, with a microdialysis probe in the posterior hippocampus. The rats were given either saline (n = 6) or 50% glucose solution (n = 6) for 3 h, starting 60 min before the onset of hypoxia. Hypoxia was produced by circulating 7% O2 gas in a plastic chamber for 90 min. In saline-infused animals, brain ECF glucose concentrations decreased slightly during hypoxia, although blood glucose concentrations did not change. Blood lactate concentrations increased to 6.28 +/- 0.91 mM, at 60 min after the onset of hypoxia (P less than 0.05). Brain ECF lactate concentrations increased to 3.53 +/- 0.20 mM and remained constant during 60 min of steady-state hypoxia (P less than 0.05) and decreased to the basal level within 60 min after the end of hypoxia. When sodium lactate solution was infused intravenously for 90 min (n = 4), blood lactate concentrations increased to a level as high as those found during hypoxia. However, the brain ECF lactate concentration increased only to 1.86 +/- 0.09 mM. In glucose-infused animals, the blood glucose concentration reached 339.1 +/- 32.3 mg/dl at the end of the glucose infusion, and the brain ECF glucose concentration increased to 54.7 +/- 7.3 mg/dl.(ABSTRACT TRUNCATED AT 250 WORDS)     

Exendin-4 treatment expands graft beta-cell mass in diabetic mice transplanted with a marginal number of fresh islets

Exendin-4 stimulates insulin secretion, suppresses glucagons secretion, increases beta-cell replication and neogenesis, and reduces beta-cell apoptosis. However, it has been shown that posttransplant exendin-4 treatment did not improve glucose homeostasis in diabetic mice transplanted with a large number of freshly isolated islets. The aim of this study was to test if exendin-4 is beneficial for hyperglycemic recipients with a marginal number of fresh islets. We transplanted 150 C57BL/6 mouse islets under the kidney capsule of inbred streptozotocin-diabetic mice, and then treated the recipients with and without exendin-4 for 6 weeks. Before and after transplantation, recipients' blood glucose, body weight, and intraperitoneal glucose tolerance test were measured. At 6 weeks, the grafts were removed to determine beta-cell mass. Blood glucose levels in both groups decreased progressively after transplantation, and the exendin-4-treated group had had lower blood glucose than controls since day 3. By 6 weeks, euglycemia was achieved more in mice treated with exendin-4 than in controls (100% vs. 62.5%, p = 0.018). The time to obtain normoglycemia was shorter in the exendin-4-treated group than in controls (12 +/- 8 vs. 29 +/- 13 days, p < 0.001). Blood glucose at 6 weeks was 123 +/- 18 and 170 +/- 62 mg/dl in the exendin-4-treated group and controls, respectively (p = 0.008). Additionally, the exendin-4-treated group had better glucose tolerance than controls at 2 and 4 weeks (p <0.02). However, both groups exhibited increased body weight over time, and weight changes did not significantly differ between the two groups throughout the study period. At 6 weeks after transplantation, grafts in the exendin-4-treated group were more prominent and contained more insulin-stained cells than those of controls. They had 2.3-fold beta-cell mass of the graft compared with controls (0.30 +/- 0.11 vs. 0.13 +/- 0.03 mg, p = 0.012). These results indicate posttransplant exendin-4 treatment in the diabetic recipient with a marginal number of fresh islets expands graft beta-cell mass and improves transplantation outcome.     

Insulin secretory response to secretagogues by perifused islets from chronically glucose-infused rats

Perifused islets from rats infused for 7 days with 40% glucose exhibited an altered secretory response to selected stimuli. Both phases of insulin release were blunted when 20 mM L-leucine was tested; the secretory response to a subsequent leucine stimulation was also blunted compared with the control group. The ability of 20 mM alpha-ketoisocaproate to stimulate the release of insulin was also greatly diminished in islets from glucose-infused rats. The secretory response to 50 microM tolbutamide plus 7 mM glucose by perifused islets from glucose-infused rats was 45% lower than in the control group. In addition, the response to a subsequent 10 mM glucose stimulation was lost. On the other hand, islets from glucose-infused rats responded to 20 microM forskolin plus 16.7 mM glucose with on significant change in the amount of insulin released during both phases of stimulation compared with the control group. The response to 100 nM phorbol 12-myristate 13-acetate was 3.1-fold higher in islets from glucose-infused compared with saline-infused rats. The finding that chronic infusions of glucose lead to selective impairment of the secretory response to fuel stimuli and agents such as tolbutamide that act on metabolically regulated K+ channels gives support to the notion that alterations in the generation of metabolic coupling signals might be involved in the phenomenon described here.     

beta-cell turnover: its assessment and implications

The pancreatic beta-cells are responsible for the maintenance of the body's glucose levels within a very narrow range; their population is dynamic and undergoes compensatory changes to maintain euglycemia. The structural parameters that allow mass changes (replication, neogenesis, cell volume changes, and cell death) can now be assessed and have proved to be powerful tools. Changes in one parameter can dramatically affect the beta-cell mass. Unfortunately, conclusions are often drawn on measurements that do not assess beta-cell mass but only relative volumes. Throughout the lifetime of a mammal, low levels of beta-cell replication and apoptosis are balanced and result in a slowly increasing mass. The balance allows gradual replacement of the beta-cell population; thus, beta-cells should be considered a slowly renewed tissue. Two major implications of beta-cell turnover are that 1) at any time, the beta-cells would be at different ages and 2) any limitation on replacement could have dire consequences for glucose homeostasis.     

Combination therapy with glucagon-like peptide-1 and gastrin induces beta-cell neogenesis from pancreatic duct cells in human islets transplanted in immunodeficient diabetic mice

Pancreatic islet transplantation as a treatment for type 1 diabetes is limited by human donor tissue availability. We investigated whether the beta-cell mass in human isolated islets could be expanded by treatments with glucagon-like peptide-1 (GLP-1) and gastrin, peptides reported to stimulate beta-cell growth in mice and rats with deficits in beta-cell mass. Human islets with low endocrine cell purity (7% beta-cells, 4% alpha-cells) and abundant exocrine cells (29% duct cells and 25% acinar cells) were implanted under the renal capsule of nonobese diabetic-severe combined immune deficiency (NOD-scid) mice made diabetic with streptozotocin. The mice were treated with GLP-1 and gastrin, separately and together, daily for 5 weeks. Blood glucose was significantly reduced only in mice implanted with human pancreatic cells and treated with GLP-1 plus gastrin. Correction of hyperglycemia was accompanied by increased insulin content in the human pancreatic cell grafts as well as by increased plasma levels of human C-peptide in the mice. Immunocytochemical examination revealed a fourfold increase in insulin-positive cells in the human pancreatic cell grafts in GLP-1 plus gastrin-treated mice, and most of this increase was accounted for by the appearance of cytokeratin 19-positive pancreatic duct cells expressing insulin. We conclude that combination therapy with GLP-1 and gastrin expands the beta-cell mass in human islets implanted in immunodeficient diabetic mice, largely from pancreatic duct cells associated with the islets, and this is sufficient to ameliorate hyperglycemia in the mice.     

Glucose infusion arrests the decompensatory phase of hemorrhagic shock

Waning of hyperglycemia has been shown to be closely associated with the deterioration of mechanisms supporting homeostasis during hemorrhagic shock. However, the mechanisms which link plasma glucose levels to maintenance of homeostasis during hemorrhagic shock are not clear. The goal of the present study was to evaluate the importance of glucose to maintenance of compensatory mechanisms. This was undertaken by maintaining plasma glucose levels through infusion of hypertonic glucose (2-3 M) starting at the onset of decompensation during persisting hypovolemia. Administration of glucose at a rate of between 60 and 80 mumoles/min X kg arrested the fall in glucose concentration and significantly slowed or arrested the decompensatory phase. All of the saline infused control animals (n = 6) died within 3 hours after reaching their maximum shed blood volume, averaging 145 +/- 25 minutes, while two of the eight animals in the glucose infusion group died less than 4 hours after reaching the maximum shed blood volume. The remaining six animals were sacrificed between 270 and 397 minutes (average, 340 +/- 22 minutes) after reaching the maximum shed blood volume since decompensation was arrested. Compared to the saline-infused control group, animals receiving glucose infusion exhibited a more moderate acidosis, and the hemoconcentration which normally accompanies decompensation was also prevented. Since the increase in plasma osmolality and the fraction of the total osmolality change accounted for by glucose was less in the glucose-infused animals, these results suggest that the effect is not mediated through a glucose-related maintenance of a transcapillary osmotic gradient.(ABSTRACT TRUNCATED AT 250 WORDS)     

Linear correlation between beta-cell mass and body weight throughout the lifespan in Lewis rats: role of beta-cell hyperplasia and hypertrophy

We determined the beta-cell replicative rate, beta-cell apoptosis, cross-sectional beta-cell area, and pancreatic beta-cell mass throughout the entire postweaning lifespan (months 1, 3, 7, 10, 15, and 20) of Lewis rats. Beta-cell replication was progressively reduced in the initial months of life but remained stable after month 7 (month 1, 0.99 +/- 0.10%; month 3, 0.24 +/- 0.04%; month 7, 0.12 +/- 0.02%; month 10, 0.14 +/- 0.02%; month 15, 0.10 +/- 0.03%; month 20, 0.13 +/- 0.03%; analysis of variance [ANOVA], P < 0.001). Beta-cell apoptosis was low and did not change significantly from month 1 to 20 of life. Cross-sectional area of individual beta-cells increased progressively in the initial months, remained stable from month 7 to 15, and increased again on month 20. The estimated number of beta-cells per pancreas, calculated as the ratio of total beta-cell mass to individual beta-cell mass, tripled from month 1 to 7 but did not change significantly thereafter. Beta-cell mass increased approximately 8 times from month 1 to 20 (month 1, 2.04 +/- 0.28 mg; month 20, 15.5 +/- 2.32 mg; ANOVA, P < 0.001) and showed a strong and significant linear correlation with body weight (r = 0.98, P < 0.001). In summary, we have shown that beta-cell replication was maintained throughout the lifespan in normal rats, clearly establishing that the beta-cell birth rate does not fall to 0, even in very old rats. Beta-cell mass increased throughout the lifespan, closely matching the increment in total body weight at any time point. This increment was selective for beta-cells, since the growth of the endocrine non-beta-cell mass was limited to the initial months of life. Both beta-cell hypertrophy and hyperplasia contributed to increased beta-cell mass in young animals, but only beta-cell hypertrophy was responsible for the increased beta-cell mass found in old animals. This study provides a global perspective for understanding the dynamics of beta-cell mass in young, adult, and aged animals.     

Glucose infusion in mice: a new model to induce beta-cell replication

Developing new techniques to induce beta-cells to replicate is a major goal in diabetes research. Endogenous beta-cells replicate in response to metabolic changes, such as obesity and pregnancy, which increase insulin requirement. Mouse genetic models promise to reveal the pathways responsible for compensatory beta-cell replication. However, no simple, short-term, physiological replication stimulus exists to test mouse models for compensatory replication. Here, we present a new tool to induce beta-cell replication in living mice. Four-day glucose infusion is well tolerated by mice as measured by hemodynamics, body weight, organ weight, food intake, and corticosterone level. Mild sustained hyperglycemia and hyperinsulinemia induce a robust and significant fivefold increase in beta-cell replication. Glucose-induced beta-cell replication is dose and time dependent. Beta-cell mass, islet number, beta-cell size, and beta-cell death are not altered by glucose infusion over this time frame. Glucose infusion increases both the total protein abundance and nuclear localization of cyclin D2 in islets, which has not been previously reported. Thus, we have developed a new model to study the regulation of compensatory beta-cell replication, and we describe important novel characteristics of mouse beta-cell responses to glucose in the living pancreas.     

Increased beta-cell apoptosis prevents adaptive increase in beta-cell mass in mouse model of type 2 diabetes: evidence for role of islet amyloid formation rather than direct action of amyloid

Nondiabetic obese humans adapt to insulin resistance by increasing beta-cell mass. In contrast, obese humans with type 2 diabetes have an approximately 60% deficit in beta-cell mass. Recent studies in rodents reveal that beta-cell mass is regulated, increasing in response to insulin resistance through increased beta-cell supply (islet neogenesis and beta-cell replication) and/or decreased beta-cell loss (beta-cell apoptosis). Prospective studies of islet turnover are not possible in humans. In an attempt to establish the mechanism for the deficit in beta-cell mass in type 2 diabetes, we used an obese versus lean murine transgenic model for human islet amyloid polypeptide (IAPP) that develops islet pathology comparable to that in humans with type 2 diabetes. By 40 weeks of age, obese nontransgenic mice did not develop diabetes and adapted to insulin resistance by a 9-fold increase (P < 0.001) in beta-cell mass accomplished by a 1.7-fold increase in islet neogenesis (P < 0.05) and a 5-fold increase in beta-cell replication per islet (P < 0.001). Obese transgenic mice developed midlife diabetes with islet amyloid and an 80% (P < 0.001) deficit in beta-cell mass that was due to failure to adaptively increase beta-cell mass. The mechanism subserving this failed expansion was a 10-fold increase in beta-cell apoptosis (P < 0.001). There was no relationship between the extent of islet amyloid or the blood glucose concentration and the frequency of beta-cell apoptosis. However, the frequency of beta-cell apoptosis was related to the rate of increase of islet amyloid. These prospective studies suggest that the formation of islet amyloid rather than the islet amyloid per se is related to increased beta-cell apoptosis in this murine model of type 2 diabetes. This finding is consistent with the hypothesis that soluble IAPP oligomers but not islet amyloid are responsible for increased beta-cell apoptosis. The current studies also support the concept that replicating beta-cells are more vulnerable to apoptosis, possibly accounting for the failure of beta-cell mass to expand appropriately in response to obesity in type 2 diabetes.     

Beneficial effects of pancreas transplantation: regeneration of pancreatic islets in the spontaneously diabetic Torii rat

AIMS: Type 2 diabetes is characterized by a combination of insulin resistance and pancreatic beta-cell dysfunction. Although pancreas transplantation (PTx) is mainly performed in patients with type 1 disease, both clinical and experimental data have demonstrated that PTx improves insulin sensitivity in type 2 diabetic recipients. However, it remains unclear whether PTx has the potential to induce islet neogenesis in a recipient's native pancreas. METHODS: Nondiabetic 10-week-old and diabetic (defined as blood glucose level >250 mg/dL) 25-week-old (average onset age of diabetes) male spontaneously diabetic Torii (SDT; RT1(a)) rats served as donors and recipients, respectively. RESULTS: In nontreated control SDT rats, beta-cell mass gradually decreased and blood glucose levels progressively increased (>600 mg/dL after 40 weeks of age). In PTx rats, however, the onset of diabetes was significantly delayed (>47.5 +/- 18.2 [graft age] versus 25.2 +/- 3.9 weeks in control rats). On immunohistochemical staining, insulin-secreting islets were observed in the naive pancreata of 40-week-old recipients with PTx (PTx40w), whereas no islets were found in 40-week-old control SDT rats. Moreover, the islets in the native pancreata of PTx40w recipients were located close to ductal structures, and PDX-1 (pancreatic duodenal homeobox-1)-positive cells were more clearly visible. These results indicate the possibility of beta-cell regeneration in the recipient native pancreas by avoiding glucose toxicity under normoglycemic condition achieved by PTx. CONCLUSIONS: Pancreas transplantation has beneficial effects on impaired islet, inducing regeneration in the spontaneously diabetic Torii rat.     

Multiple alterations in insulin responses to glucose in islets from 48-h glucose-infused nondiabetic rats

To examine the biochemical mechanisms by which hyperglycemia produces insulin secretory abnormalities, we studied isolated islets from control rats and rats infused for 48 h with a 50% glucose solution. To preserve the effects of in vivo hyperglycemia during in vitro handling for islet isolation, our standard isolation procedure utilized buffers containing 16.8 mM glucose. Islets from infused rats released similar amounts of insulin in low or high glucose during first incubations at 37 degrees C (92.4 +/- 7.0 ng.10 islets-1.45 min-1 at 2.8 mM, 84.4 +/- 4.1 ng.10 islets-1.45 min-1 at 16.8 mM) in contrast with control (uninfused) islets (18.6 +/- 2.8 ng.10 islets-1.45 min-1 at 2.8 mM and 109.8 +/- 8.0 ng.10 islets-1.45 min-1 at 16.8 mM glucose) (P less than 0.01). Secretion by islets of glucose-infused rats was lower during 60-min second incubations at 28 mM glucose than in first incubations of the same islets in low glucose (P less than 0.01). This phenomenon is comparable to the paradoxical hypersecretion observed during the first 10-15 min of exposure of glucose-infused pancreas to low-glucose perfusions. Paradoxical secretion in low glucose waned rapidly, so that during second incubations at 37 degrees C, little immunoreactive insulin release occurred at 2.8 mM glucose, despite the persistence of two additional lesions. The glucose-insulin dose-response curves in second incubations showed a leftward shift for glucose-infused islets, with two- to threefold higher secretion at 5.6-8.4 mM glucose than control islets. This is termed sensitization to glucose.(ABSTRACT TRUNCATED AT 250 WORDS)     

Recombinant human betacellulin promotes the neogenesis of beta-cells and ameliorates glucose intolerance in mice with diabetes induced by selective alloxan perfusion

Betacellulin (BTC), a member of the epidermal growth factor family, is expressed predominantly in the human pancreas and induces the differentiation of a pancreatic acinar cell line (AR42J) into insulin-secreting cells, suggesting that BTC has a physiologically important role in the endocrine pancreas. In this study, we examined the in vivo effect of recombinant human BTC (rhBTC) on glucose intolerance and pancreatic morphology using a new mouse model with glucose intolerance induced by selective alloxan perfusion. RhBTC (1 microg/g body wt) or saline was injected subcutaneously every day from the day after alloxan treatment. The intraperitoneal glucose tolerance test revealed no difference between rhBTC-treated and rhBTC-untreated glucose-intolerant mice at 2-4 weeks. However, glucose tolerance was significantly improved and body weight was significantly increased in rhBTC-treated mice compared with untreated mice at 8 weeks. Islet-like cell clusters, consisting mainly of beta-cells, were increased in the pancreas and were localized in contact with the ductal lining cells and sometimes with acinar cells. In conclusion, administration of rhBTC improved glucose tolerance in this mouse model by increasing beta-cell volume, primarily through accelerated neogenesis from ductal lining cells.