Enlarged Islets Research Articles

Marginal Zinc Deficiency Promotes Pancreatic Islet Enlargement While Zinc Supplementation Improves the Pancreatic Insulin Response in Zucker Diabetic Fatty Rats.

Zinc deficiency has been associated with the worsening of diabetes while zinc supplementation has been proposed to ameliorate diabetes. This study examined the effects of marginal zinc deficiency (MZD) and zinc supplementation (ZS) on obesity, glycemic control, pancreatic islets, hepatic steatosis and renal function of Zucker diabetic fatty (ZDF) rats. Male ZDF rats were fed an MZD, zinc control (ZC) or ZS diet (4, 30 and 300 mg Zn/kg diet, respectively), and lean Zucker rats were fed a ZC diet for 8 weeks. MZD and ZS did not alter body weight or whole-body composition in ZDF rats. MZD ZDF rats had reduced zinc concentrations in the femur and pancreas, a greater number of enlarged pancreatic islets and a diminished response to an oral glucose load based on a 1.8-fold greater incremental area-under-the-curve (AUC) for glucose compared to ZC ZDF. ZS ZDF rats had elevated serum, femur and pancreatic zinc concentrations, unchanged pancreatic parameters and a 50% reduction in the AUC for insulin compared to ZC ZDF rats, suggesting greater insulin sensitivity. Dietary zinc intake did not alter hepatic steatosis, creatinine clearance, or levels of proteins that contribute to insulin signaling, inflammation or zinc transport in epididymal fat. Potential adverse effects of ZS were suggested by reduced hepatic copper concentrations and elevated serum urea compared to ZC ZDF rats. In summary, ZS improved the pancreatic insulin response but not the glucose handling. In contrast, reduced zinc status in ZDF rats led to impaired glucose tolerance and a compensatory increase in the number and size of pancreatic islets which could lead to β-cell exhaustion.

GPR92 activation in islet macrophages controls β cell function in a diet-induced obesity model.

The molecular mechanisms underlying obesity-induced increases in β cell mass and the resulting β cell dysfunction need to be elucidated further. Our study revealed that GPR92, expressed in islet macrophages, is modulated by dietary interventions in metabolic tissues. Therefore, we aimed to define the role of GPR92 in islet inflammation by using a high-fat diet-induced (HFD-induced) obese mouse model. GPR92-KO mice exhibited glucose intolerance and reduced insulin levels - despite the enlarged pancreatic islets - as well as increased islet macrophage content and inflammation level compared with WT mice. These results indicate that the lack of GPR92 in islet macrophages can cause β cell dysfunction, leading to disrupted glucose homeostasis. Alternatively, stimulation with the GPR92 agonist farnesyl pyrophosphate results in the inhibition of HFD-induced islet inflammation and increased insulin secretion in WT mice, but not in GPR92-KO mice. Thus, our study suggests that GPR92 can be a potential target to alleviate β cell dysfunction via the inhibition of islet inflammation associated with the progression of diabetes.

Morphological alteration of the pancreatic islet in ovariectomized rats fed a high-fat high-fructose diet.

Diabetes and its complications are major causes of mortality worldwide. Type 2 diabetes coexists with insulin resistance and β-cell dysfunction, which are aggravated by overconsumption and estrogen-deprived conditions. However, the morphology of pancreatic islets in a combined condition of excessive caloric intake and estrogen deficiency has never been described. Herein, we examined morphological changes in the pancreatic islets of ovariectomized (OVX) rats fed a high-fat high-fructose diet (HFFD) for 12weeks. The histological changes in the size and number of pancreatic islets were assessed by hematoxylin-eosin and immunohistochemical staining. Enlarged pancreatic islets with fat deposition in OVX rats were accompanied by whole-body insulin resistance and hyperglycemia. The addition of a HFFD to OVX rats (OVX + HFFD) further aggravated insulin resistance, with a substantial increase in the density of enlarged pancreatic islets and fat accumulation. The augmented number of enlarged islets was correlated with elevated plasma glucose and insulin levels. Intriguingly, unlike the HFFD and OVX alone, the OVX + HFFD markedly expanded the area of insulin-producing β-cells and glucagon-producing α-cells. Importantly, enlarged islets, pancreatic fat deposits, and diabetic states developing in OVX + HFFD conditions were resolved by estrogen replacement. Collectively, the morphological characteristics of pancreatic islets were influenced in an insulin-resistant state caused by estrogen deficiency and HFFD consumption and were distinct from each factor alone. A combination of estrogen deficiency with HFFD consumption worsened the integrity of pancreatic islets, ultimately resulting in disease progression. These findings expand our understanding of the causal relationship between pancreatic morphology and diabetes development and suggest therapeutic strategies.

Efficient Restoration of Beta Cell Dedifferentiation by Calorie Restriction With High Fat/Low Carbohydrate Diet in Obese Diabetes Model and the Possible Role of GLP-1

In type 2 diabetes, pancreatic beta cells are gradually ‘exhausted’ and fall into beta cell dysfunction, which proceeds more severe insulin dependence. Among the proposed mechanisms of beta cell dysfunction such as endoplasmic reticulum stress and oxidative stress, the beta cell heterogeneity has attracted the researcher’s interest recently. In 2012, Talchai et al. revealed that the beta cells were dedifferentiated in diabetic mice model, and nowadays it is considered as one form of the beta cell heterogeneity and is observed broadly among diabetic animal models and human patients. Previously we showed that food restriction had the best effect to restore beta cell gene expression in obese diabetic model mice, among the known diabetic treatments which we tested. In the current study, we aimed to unveil the molecular basis in the improvement of beta cell dedifferentiation during the calorie restriction. First, we utilized the high-fat/low carbohydrate diet (HF) or low-fat/high carbohydrate (HC) diet, to determine whether fat restriction or sugar restriction reduces the beta cell dedifferentiation in obese mice. When calorie intake was restricted evenly, both HF diet and HC diet decreased the body weight and hyperglycemia in db/db mice equally. Albeit the same metabolic profile, db/db group fed with HC diet had more enlarged islets and more dedifferentiated beta cell features than db/dbs fed with HF diet, which indicated the compensatory beta cell response in HC diet group. Moreover, HC diet group showed more severe fatty liver than HF diet group, along with the elevated synthesis and accumulation of triglycerides and cholesterol in liver. It is speculated that the insulin resistance in liver might impact on the beta cell dedifferentiation. Next, we analyzed the effect of glucagon-like peptide 1 (GLP-1) on beta cell dedifferentiation, since GLP-1 is secreted more from intestine by protein and fat intake, rather than by sugar intake. Also, increasing number of reports have suggested the improving effect of GLP-1 on beta cell dysfunction and fatty liver. Indeed, GLP-1 administration altered the reduced beta cell/alpha cell ratio in db/db mice, which indicated the restoration of beta cell heterogeneity. We are now investigating if GLP-1 administration reimburse the beta cell dedifferentiation in db/db mice fed with HC diet, to illuminate the role of incretins in beta cell dedifferentiation induced by unbalanced nutrition during diet. Also, we will present the RNA sequencing data of the liver in db/db mice fed with HF and HC diet, to elucidate the key molecules and genes which connect the beta cell function and metabolic state in liver.

Islet Dysfunction in a Novel Transgenic Model of T Cell Insulitis.

The newly established CD3FLAG-mIR transgenic mouse model on a C57Bl/6 background has a FLAG tag on the mouse Insulin Receptor (mIR), specifically on T cells, as the FLAG-tagged mIR gene was engineered behind CD3 promoter and enhancer. The IR is a chemotactic molecule for insulin and the Flag-tagged mIR T cells in the BL/6-CD3FLAGmIR transgenic mice can migrate into the pancreas, as shown by immunofluorescent staining. While the transgenic mice do not become diabetic, there are phenotypic and metabolic changes in the islets. The transgenic islets become enlarged and disorganized by 15 weeks and those phenotypes continue out to 35 weeks of age. We examined the islets by RT-PCR for cell markers, ER stress markers, beta cell proliferation markers, and cytokines, as well as measuring serum insulin and insulin content in the pancreas at 15, 25, and 35 weeks of age. In transgenic mice, insulin in serum was increased at 15 weeks of age and glucose intolerance developed by 25 weeks of age. Passage of transgenic spleen cells into C57Bl/6 RAG−/− mice resulted in enlarged and disorganized islets with T infiltration by 4 to 5 weeks post-transfer, replicating the transgenic mouse studies. Therefore, migration of non-antigen-specific T cells into islets has ramifications for islet organization and function.

Peripancreatic adipose tissue protects against high-fat-diet-induced hepatic steatosis and insulin resistance in mice

Background/objectivesVisceral adiposity is associated with increased diabetes risk, while expansion of subcutaneous adipose tissue may be protective. However, the visceral compartment contains different fat depots. Peripancreatic adipose tissue (PAT) is an understudied visceral fat depot. Here, we aimed to define PAT functionality in lean and high-fat-diet (HFD)-induced obese mice.Subjects/methodsFour adipose tissue depots (inguinal, mesenteric, gonadal, and peripancreatic adipose tissue) from chow- and HFD-fed male mice were compared with respect to adipocyte size (n = 4–5/group), cellular composition (FACS analysis, n = 5–6/group), lipogenesis and lipolysis (n = 3/group), and gene expression (n = 6–10/group). Radioactive tracers were used to compare lipid and glucose metabolism between these four fat depots in vivo (n = 5–11/group). To determine the role of PAT in obesity-associated metabolic disturbances, PAT was surgically removed prior to challenging the mice with HFD. PAT-ectomized mice were compared to sham controls with respect to glucose tolerance, basal and glucose-stimulated insulin levels, hepatic and pancreatic steatosis, and gene expression (n = 8–10/group).ResultsWe found that PAT is a tiny fat depot (~0.2% of the total fat mass) containing relatively small adipocytes and many “non-adipocytes” such as leukocytes and fibroblasts. PAT was distinguished from the other fat depots by increased glucose uptake and increased fatty acid oxidation in both lean and obese mice. Moreover, PAT was the only fat depot where the tissue weight correlated positively with liver weight in obese mice (R = 0.65; p = 0.009). Surgical removal of PAT followed by 16-week HFD feeding was associated with aggravated hepatic steatosis (p = 0.008) and higher basal (p < 0.05) and glucose-stimulated insulin levels (p < 0.01). PAT removal also led to enlarged pancreatic islets and increased pancreatic expression of markers of glucose-stimulated insulin secretion and islet development (p < 0.05).ConclusionsPAT is a small metabolically highly active fat depot that plays a previously unrecognized role in the pathogenesis of hepatic steatosis and insulin resistance in advanced obesity.

1875-P: The Impact of Carbohydrate/Fat Intake Balance on Beta-Cell Dedifferentiation and Fatty Liver in Obese Mouse Model

In clinical, the ’fatigue’ of the pancreatic beta cells is a famous phenomenon in type 2 diabetes. Nowadays, researchers describe the several mechanisms of beta cell dysfunction, such as glucolipotoxicity, endoplasmic reticulum stress and oxidative stress. In 2012, Talchai et al. revealed that the beta cells were dedifferentiated in diabetes, and previously we showed that food restriction had the most manifold good effects to restore beta cell gene expression, among the known diabetic treatments that we tested. In the current study, we aimed to unveil which nutrients should be cut down to protect beta cell differentiation, sugar or fat. First, we performed the pair-feeding with db/+ and db/db to reduce food intake of db/db mice. We utilized the high-fat/low carbohydrate diet (HF) or low-fat/high carbohydrate (HC) diet which were adjusted to the same total calorie intake. Pair fed-db/db mice had better body weight and glucose tolerance control than ad libitum db/db group, as previously we reported, and there was no significant difference between HF and HC diet. Albeit the same metabolic profile, HC diet group had more enlarged islets and more dedifferentiated beta cell features, which indicated the compensatory beta cell response in HC diet group. Moreover, HC diet group showed more severe fatty liver than HF diet group, along with the elevated synthesis and accumulation of triglycerides and cholesterol in liver. We are now investigating the relationship between sugar load, insulin resistance in liver and beta cell dedifferentiation. Disclosure E. Ishida: None. X. Lei: None. E. Yamada: None. S. Okada: None. M. Yamada: None. Funding Japan Society for the Promotion of Science (18K16222)

Tumor-like features of gene expression and metabolic profiles in enlarged pancreatic islets are associated with impaired incretin-induced insulin secretion in obese diabetes: A study of Zucker fatty diabetes mellitus rat.

Aims/IntroductionPancreatic islets are heterogenous. To clarify the relationship between islet heterogeneity and incretin action in the islets, we studied gene expression and metabolic profiles of non‐large and enlarged islets of the Zucker fatty diabetes mellitus rat, an obese diabetes model, as well as incretin‐induced insulin secretion (IIIS) in these islets.Materials and MethodsPancreatic islets of control (fa/+) and fatty (fa/fa) rats at 8 and 12 weeks‐of‐age were isolated. The islets of fa/fa rats at 12 weeks‐of‐age were separated into non‐large islets (≤200 μm in diameter) and enlarged islets (>300 μm in diameter). Morphological analyses, insulin secretion experiments, transcriptome analysis, metabolome analysis and oxygen consumption analysis were carried out on these islets.ResultsThe number of enlarged islets was increased with age in fatty rats, and IIIS was significantly reduced in the enlarged islets. Markers for β‐cell differentiation were markedly decreased in the enlarged islets, but those for cell proliferation were increased. Glycolysis was enhanced in the enlarged islets, whereas the tricarboxylic acid cycle was suppressed. The oxygen consumption rate under glucose stimulation was reduced in the enlarged islets. Production of glutamate, a key signal for IIIS, was decreased in the enlarged islets.ConclusionsThe enlarged islets of Zucker fatty diabetes mellitus rats, which are defective for IIIS, show tumor cell‐like metabolic features, including a dedifferentiated state, accelerated aerobic glycolysis and impaired mitochondrial function. The age‐dependent increase in such islets could contribute to the pathophysiology of obese diabetes.

OR14-01 FSTL3 Neutralizing Antibodies Restore Function to Diabetic Mouse and Human Islets: A New Approach for Treating Diabetes

Activin, GDF11 and myostatin are structurally related members of the TGFbeta superfamily of growth factors with many biological roles in animal models and humans. Their actions are neutralized by extracellular proteins such as follistatin and follistatin like-3 (FSTL3). We have previously demonstrated that genetic inactivation of Fstl3 results in enlarged pancreatic islets containing increased numbers of beta cells that produce more insulin in response to glucose compared to wild type litter mates. We further discovered that at least some of these new beta cells arise via transdifferentiation from alpha cells. We also demonstrated that functional human islets from normal donors produce very high levels of activin. In contrast, activin biosynthesis is vastly reduced and FSTL3 synthesis is significantly increased in human islets from diabetic donors suggesting that activin is critical for normal insulin production. This was substantiated by direct treatment of human diabetic islets with activin which restored their response to glucose. These observations support the hypothesis that an FSTL3 neutralizing antibody would constitute a novel therapeutic approach to curing diabetes through restoring beta cell function as well as accelerating generation of new beta cells through transdifferentiation. To test this hypothesis, we produced a mouse monoclonal antibody that neutralized hFSTL3 (FP-101), thereby releasing bioactive activin, GDF11, and myostatin. We have now tested this antibody for biological activity in vitro on mouse and human islets. We used islets from high fat diet (HFD) treated mice to model diabetes-inducing effects of obesity as well as 24-hour incubation in hyperglycemic (33 mM glucose) medium to create human islets that lose responsiveness to high glucose as a model for human diabetes. In mouse islets we found that stimulation of normal (chow diet) islets by high glucose produced a stimulation index (SI) of 3.5 that was reduced to 2 in HFD islets. Treatment with activin, FP-101, or a commercial polyclonal antibody to mFSTL3 all increased response of HFD islets to elevated glucose and partially restored SI to normal levels. In human islets, hyperglycemia eliminated the normal (2.5 SI) response to high glucose while activin or FP-101 treatments dose-responsively restored this response. These results demonstrate that anti-FSTL3 therapy can restore function to compromised beta cells from mouse and human diabetes models. The observation that activin has the same action as anti-FSTL3 antibody indicates that FP-101 works through enhancing the activin signaling pathway. Finally, these results demonstrate that the FSTL3-activin pathway is an important regulator of beta cell function in humans as well as mice, supporting further development of this therapy as a diabetes treatment.

2187-P: Gsα Deficiency in Pancreatic a Cells Leads to Enlarged Islets

Pancreatic β-cell dysfunction leads to diabetes with reduced β-cell mass and impaired glucose-stimulated insulin secretion. The G protein α subunit Gsα couples receptors to generate cAMP, which mediates the insulinotropic effects of incretin hormones, such as GLP-1. We previously reported that β-cell-specific Gsα deficiency causes insulin-deficient diabetes due to reduced islet size and β-cell mass, and that mice with whole pancreatic Gsα deficiency develop insulin deficient diabetes and reduced β-cell mass, but also showed increased proliferation of α-cells, indicating that Gsα plays divergent roles in pancreatic α-and β-cell growth. In the present study, we generated mice with α-cell-specific Gsα deficiency (αGsKO) by mating Gsα-floxed mice with glucagon-cre mice and studied the metabolic consequences. Deletion of Gsα in α-cells did not affect growth, blood glucose, or serum insulin or glucagon levels. However, αGsKO mice displayed enlarged islet size of normal architecture with significant increases in both α- and β-cell numbers, accompanied with increased β-cell proliferation and reduced apoptosis. This increased β-cell proliferation could be detected as early as at 3 days after birth. Consistent with this, αGsKO mice showed increased insulin response to acute glucose loading (3mg/g i.p.), although they had unaltered glucose tolerance when given a lower dose of glucose (2 mg/g i.p.). The results of RT-PCR on pancreatic islets isolated from aged mice (9-10 months old) showed that expression of Maf B and Pax6, transcription factors required for β-cell development, tended to be increased; whereas cell cycle factors, such as Cyclin D1, D2, and CDK4, tended to be decreased. Taken together, our results show that Gsα/cAMP signaling pathway in pancreatic α-cells is critical for both α- and β-cell growth as well as for maintenance of β-cell mass. Disclosure A. Taira: None. H. Sun: None. C. Le: None. O. Gavrilova: None. M. Chen: None. L. Weinstein: None. Funding National Institutes of Health

FSTL3 Inhibition Restores Glucose Responsive Insulin Secretion in Nonfunctional Human Islets

Type 2 diabetes results from inadequate insulin production to control blood glucose. To address this need for replacing lost insulin-producing beta cells, most therapeutic development strategies are focused on enhancing functional beta cell number. Fairbanks Pharmaceuticals is pursuing therapies that enhance beta cell function and regeneration and is based on prior research in which inactivation of follistatin-like-3 (FSTL3) resulted in enlarged islets, increased beta cell number, and enhanced insulin secretion. At least some of these new beta cells arose from alpha to beta cell transdifferentiation in FSTL3 knockout mice. Fairbanks has produced a monoclonal antibody (FP-101) that can completely block FSTL3 from binding activin A and B, GDF11, and myostatin that could also disrupt pre-formed and otherwise irreversible complexes. FP-101 was also able to restore glucose-responsive insulin secretion in otherwise non-functional islets taken from HFD-fed mice. To determine if FSTL3 neutralization could be effective in humans, human islets from normal donors were first exposed to elevated glucose (25 nM) for 48-72 hours, a commonly utilized model for glucotoxicity. As expected, this caused loss of glucose response to subsequent GSIS. However, when treated for 24 hours with FP-101, GSIS was restored, indicating that neutralization of FSTL3 might be an effective strategy for restoring function to human non-functional islets. This observation agrees with previous research in which activin treatment directly restored GSIS in human islets from diabetic donors. Taken together with the potential for induction of alpha to beta cell transdifferentiation, these results suggest that neutralization of FSTL3 may have a dual benefit with acute restoration of GSIS in non-functional islets and chronic treatment leading to restoration of beta cell number through enhancement of alpha to beta cell transdifferentiation. In vivo studies will be initiated shortly. Disclosure A. Schneyer: Stock/Shareholder; Self; Fairbanks Pharmaceuticals, Inc.. Stock/Shareholder; Spouse/Partner; Fairbanks Pharmaceuticals, Inc.. Employee; Self; Fairbanks Pharmaceuticals, Inc.. M. Brown: None.

Impaired incretin-induced insulin secretion in enlarged pancreatic islets in a novel animal model of obese type 2 diabetes

Impaired incretin-induced insulin secretion in enlarged pancreatic islets in a novel animal model of obese type 2 diabetes

Effects of fungal pancreatic enzymes on the function of islet cells in Syrian golden hamsters.

Our previous studies showed that porcine pancreatic enzymes in Syrian golden hamsters with peripheral insulin resistance normalizes the plasma insulin level, reduces the size of enlarged islets and inhibits the increased DNA synthesis in the beta-cell of islets. In order to exclude the possibility that these effects was attributed to some contaminants of this crude material, we tested the effect of purified fungal pancreatic enzyme (FPE) that contains primarily amylase and lipase without (FPE) and with addition of chymotrypsin (FPE+chy). In a pilot study we tested the effect of different doses of FPE given in drinking water on insulin level, islet size and DNA synthesis of islet cells in hamsters with induced peripheral insulin resistance by a high fat diet. The most effective dose of FPE on these parameters was used in a long-term experiment with FPE and FPE+chy in hamsters fed a high-fat diet for 36 or 40 weeks. In the pilot study a dose of 2 g/kg body weight was found to be optimal for controlling the body weight, normalizing plasma insulin level, the size of islets, the DNA synthesis and the number of insulin cells in the islets. These data were produced in the long-term study, where steatorrhea was also inhibited. Addition of chymotrypsin had no effects on these parameters. Pancreatic lipase and amylase appear to be responsible for the observed effects and offer a safe and effective natural product for the treatment of pancreatic diseases, including acute pancreatitis, chronic pancreatic, cystic fibrosis and any conditions associated with peripheral insulin resistance, including obesity and type 2 diabetes. The possible mechanism of the action is discussed.

Adipose Tissue-Specific Deletion of 12/15-Lipoxygenase Protects Mice from the Consequences of a High-Fat Diet

Type 2 diabetes is associated with obesity, insulin resistance, and inflammation in adipose tissue. 12/15-Lipoxygenase (12/15-LO) generates proinflammatory lipid mediators, which induce inflammation in adipose tissue. Therefore we investigated the role of 12/15-LO activity in mouse white adipose tissue in promoting obesity-induced local and systemic inflammatory consequences. We generated a mouse model for fat-specific deletion of 12/15-LO, aP2-Cre; 12/15-LO loxP/loxP, which we call ad-12/15-LO mice, and placed wild-type controls and ad-12/15-LO mice on a high-fat diet for 16 weeks and examined obesity-induced inflammation and insulin resistance. High-fat diet-fed ad-12/15-LO exhibited improved fasting glucose levels and glucose metabolism, and epididymal adipose tissue from these mice exhibited reduced inflammation and macrophage infiltration compared to wild-type mice. Furthermore, fat-specific deletion of 12/15-LO led to decreased peripheral pancreatic islet inflammation with enlarged pancreatic islets when mice were fed the high-fat diet compared to wild-type mice. These results suggest an interesting crosstalk between 12/15-LO expression in adipose tissue and inflammation in pancreatic islets. Therefore, deletion of 12/15-LO in adipose tissue can offer local and systemic protection from obesity-induced consequences, and blocking 12/15-LO activity in adipose tissue may be a novel therapeutic target in the treatment of type 2 diabetes.

Differential synthesis and action of TGFβ superfamily ligands in mouse and rat islets

Members of the TGFβ superfamily, including activins and TGFβ, modulate glucose-stimulated insulin secretion (GSIS) in vitro using rat islets while genetic manipulations that reduce TGFβ superfamily signaling in vivo in mice produced hypoplastic islets and/or hyperglycemia. Moreover, deletion of Fstl3, an antagonist of activin and myostatin, resulted in enlarged islets and β-cell hyperplasia. These studies suggest that endogenous TGFβ superfamily ligands regulate β-cell generation and/or function. To test this hypothesis, we examined endogenous TGFβ ligand synthesis and action in isolated rat and mouse islets. We found that activin A, TGFβ1, and myostatin treatment enhanced rat islet GSIS but none of the ligands tested enhanced GSIS in mouse islets. However, follistatin inhibited GSIS, consistent with a role for endogenous TGFβ superfamily ligands in regulating insulin secretion. Endogenous expression of TGF∆ superfamily members was different in rat and mouse islets with myostatin being highly expressed in mouse islets and not detectable in rats. These results indicate that TGFβ superfamily members directly regulate islet function in a species-specific manner while the ligands produced by islets differ between mice and rats. The lack of in vitro actions of ligands on mouse islets may be mechanical or result from species-specific actions of these ligands.

Reconstitution of Insulin Action in Muscle, White Adipose Tissue, and Brain of Insulin Receptor Knock-out Mice Fails to Rescue Diabetes

Type 2 diabetes results from an impairment of insulin action. The first demonstrable abnormality of insulin signaling is a decrease of insulin-dependent glucose disposal followed by an increase in hepatic glucose production. In an attempt to dissect the relative importance of these two changes in disease progression, we have employed genetic knock-outs/knock-ins of the insulin receptor. Previously, we demonstrated that insulin receptor knock-out mice (Insr(-/-)) could be rescued from diabetes by reconstitution of insulin signaling in liver, brain, and pancreatic β cells (L1 mice). In this study, we used a similar approach to reconstitute insulin signaling in tissues that display insulin-dependent glucose uptake. Using GLUT4-Cre mice, we restored InsR expression in muscle, fat, and brain of Insr(-/-) mice (GIRKI (Glut4-insulin receptor knock-in line 1) mice). Unlike L1 mice, GIRKI mice failed to thrive and developed diabetes, although their survival was modestly extended when compared with Insr(-/-). The data underscore the role of developmental factors in the presentation of murine diabetes. The broader implication of our findings is that diabetes treatment should not necessarily target the same tissues that are responsible for disease pathogenesis.

Sustained glucagon-like peptide 1 expression from encapsulated transduced cells to treat obese diabetic rats

Obesity and type 2 diabetes (T2D) are two prevalent chronic diseases that have become a major public health concern in industrialized countries. T2D is characterized by hyperglycemia and islet beta cell dysfunction. Glucagon-like peptide 1 (GLP-1) promotes β cell proliferation and neogenesis and has a potent insulinotropic effect. Leptin receptor deficient male rats are obese and diabetic and provide a model of T2D. We hypothesized that their treatment by sustained expression of GLP-1 using encapsulated cells may prevent or delay diabetes onset. Vascular smooth muscle cells (VSMC) retrovirally transduced to secrete GLP-1 were seeded into TheraCyte(TM) encapsulation devices, implanted subcutaneously and rats were monitored for diabetes. Rats that received cell implants showed mean plasma GLP-1 level of 119.3 ± 10.2pM that was significantly elevated over control values of 32.4 ± 2.9pM (P<0.001). GLP-1 treated rats had mean insulin levels of 45.9 ± 2.3ng/ml that were significantly increased over control levels of 7.3±1.5ng/ml (P<0.001). In rats treated before diabetes onset elevations in blood glucose were delayed and rats treated after onset became normoglycemic and showed improved glucose tolerance tests. Untreated diabetic rats possess abnormal islet structures characterized by enlarged islets with α-cell infiltration and multifocal vacuolization. GLP-1 treatment induced normalization of islet structures including a mantle of α-cells and increased islet mass. These data suggest that encapsulated transduced cells may offer a potential long term treatment of patients.

Segmentation of the Pathophysiological Stages of Diabetic Changes in the db/db Mouse

The db/db mouse is one of the diabetes mellitus animal models and if the pathophysiological stages of diabetic changes in the mouse model could simulate the stages in human diabetes, the db/db mouse could be used to better evaluate drug candidates. Blood insulin, HbA1c levels and morphological features of pancreatic islets in db/db mice were evaluated to determine the pathophysiological stage. At 6 weeks of age, db/db mice showed the highest level of plasma insulin and lowest level of HbA1c, and histopathological examination revealed enlarged islets with a circular shape and hypertrophic islet cells. By 9 and 12 weeks of age, the plasma insulin levels had decreased to mid levels and HbA1c had increased to mid to high levels; histopathological examination at this time revealed two types of islets coexisting, enlarged circular islets and small irregular-shaped islets. By 15 and 22 weeks of age, plasma insulin had decreased further to low levels and HbA1c was at its highest level; the histopathological examination at this time revealed an increase in irregular-shaped and small islets. Based on blood insulin levels, HbA1c levels and histopathology findings in the db/db mice in this study, the clinical staging of diabetic changes were recognized. The pathophysiological stages of diabetes mellitus in this animal model were similar to the stages in humans.

Effects of chromium picolinate on glycemic control and kidney of the obese Zucker rat

BackgroundChromium picolinate (Cr(pic)3) is advocated as adjuvant therapy for impaired glycemic control, despite concerns for DNA damage. Potential toxicity of Cr(pic)3 should be greater for the kidney that accumulates chromium. Therefore, we tested the hypothesis that Cr(pic)3 treatment of obese Zucker rats (OZR) exacerbates renal abnormalities associated with dysglycemia.MethodsMale OZR were treated with diets lacking or containing 5 and 10 mg/kg of chromium, as Cr(pic)3, for 20 weeks; lean Zucker rats (LZR) served as controls. Glycemic and renal effects of Cr(pic)3 were determined in the context of indices of oxidative stress and inflammation.ResultsThe OZR displayed increased fasting plasma glucose and insulin in association with enlarged pancreatic islets exhibiting collagen and periodic acid Schiff-positive deposits compared to LZR; Cr(pic)3 treatment did not affect these parameters. The OZR, irrespective of Cr(pic)3, excreted more albumin than LZR. Also, other indices of renal function or histopathology were not affected by Cr(pic)3 treatment. Urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), an index of oxidative DNA damage, was greater in the OZR than LZR; dietary Cr(pic)3 treatment attenuated 8-OHdG excretion. However, immunostaining of kidney for 8-OHdG revealed similar staining pattern and intensity, despite significant renal accumulation of chromium in Cr(pic)3-treated groups. Finally, increased renal nitrotyrosine and cyclooxygenase-2 levels and urinary excretion of monocyte chemoattractant protein-1 of OZR were partially reversed by Cr(pic)3 treatment.ConclusionDietary Cr(pic)3 treatment of OZR does not beneficially influence glycemic status or increase the risk for oxidative DNA damage; rather, the treatment attenuates indices of oxidative stress and inflammation.

Plasma Membrane-associated Sialidase as a Crucial Regulator of Transmembrane Signalling

Mammalian sialidases, glycosidases responsible for the removal of sialic acids from glycoproteins and glycolipids, has been implicated to participate in many biological processes as well as in lysosomal catabolism. Among those forms identified to date, plasma membrane-associated sialidase, Neu3, is a key enzyme in degradation of gangliosides, for which it exhibits a special substrate preference. This sialidase has been shown to control transmembrane signalling for many cellular processes, including cell differentiation, cell growth and apoptosis, and human orthologue NEU3 is markedly up-regulated in various cancers. It is known to suppress apoptosis in cancer cells. Furthermore, its overexpression causes impaired glucose tolerance and hyper-insulinaemia together with overproduction of insulin in enlarged islets in the transgenic mice. The present review primarily summarizes our recent results, focusing on Neu3 as a regulator of transmembrane signalling.