Sequential morphological changes in pancreatic islets of spontaneously diabetic rats

This study describes the sequential morphological changes in pancreatic islets from 1-, 6-, and 18-month-old male eSS rats, as compared to aged-matched control animals. Spontaneous diabetes mellitus was confirmed in 6- and 18-month-old eSS rats after an oral glucose tolerance test. Light microscopic immunocytochemical and morphometric techniques were used to study islet-cell populations. The pancreas was normal, and the morphometric methods did not reveal significant changes in islets from 1-month-old rats. However, 6-month-old eSS animals showed disruption of islet architecture and fibrosis in the stroma. The volume density (Vvi) of endocrine tissue and the Vvi and percentage of B cells were increased, whereas the Vvi of exocrine tissue and the Vvi and percentage of A cells were diminished. Eighteen-month-old eSS rats also exhibited conspicuous islet lesions. Nevertheless, the Vvi of endocrine tissue and the Vvi and percentage of B cells were diminished, while the Vvi of exocrine tissue and the Vvi and percentage of D cells were increased. Our results provide further quantitative evidence for the sequential morphological events occurring in the pancreatic islets of a useful animal model of diabetes mellitus.     

Increased expression of adenylyl cyclase 3 in pancreatic islets and central nervous system of diabetic Goto-Kakizaki rats

Adenylyl cyclase 3 (AC3) is expressed in pancreatic islets of the Goto-Kakizaki (GK) rat, a spontaneous animal model of type 2 diabetes (T2D), and also exerts genetic effects on the regulation of body weight in man. In addition to pancreatic islets, the central nervous system (CNS) plays an important role in the pathogenesis of T2D and obesity by regulating feeding behavior, body weight and glucose metabolism. In the present study, we have investigated AC3 expression in pancreatic islets, striatum and hypothalamus of GK rats to evaluate its role in the regulation of glucose homeostasis. GK and Wistar rats at the age of 2.5 mo were used. A group of GK rats were implanted with sustained insulin release chips for 15 d. Plasma glucose and serum insulin levels were measured. AC3 gene expression levels in pancreatic islets, striatum and hypothalamus were determined by using real-time RT-PCR. Results indicated that plasma glucose levels in Wistar rats were found to be similar to insulin-treated GK rats, and significantly lower compared with non-treated GK rats. AC3 expression levels in pancreatic islets, striatum and hypothalamus of GK rats were higher compared with Wistar rats, while the levels were intermediate in insulin-treated GK rats. The AC3 expression display patterns between pancreatic islets and striatum-hypothalamus were similar. The present study thus provides the first evidence that AC3 is overexpressed in the regions of striatum and hypothalamus of brain, and similarly in pancreatic islets of GK rats suggesting that AC3 plays a role in regulation of glucose homeostasis via CNS and insulin secretion.     

Global profiling of coxsackievirus- and cytokine-induced gene expression in human pancreatic islets

Aims/hypothesis It is thought that enterovirus infections initiate or facilitate the pathogenetic processes leading to type 1 diabetes. Exposure of cultured human islets to cytolytic enterovirus strains kills beta cells after a protracted period, suggesting a role for secondary virus-induced factors such as cytokines.Methods To clarify the molecular mechanisms involved in virus-induced beta cell destruction, we analysed the global pattern of gene expression in human islets. After 48 h, RNA was extracted from three independent human islet preparations infected with coxsackievirus B5 or exposed to interleukin 1 (50 U/ml) plus interferon (1,000 U/ml), and gene expression profiles were analysed using Affymetrix HG-U133A gene chips, which enable simultaneous analysis of 22,000 probe sets.Results As many as 13,077 genes were detected in control human islets, and 945 and 1293 single genes were found to be modified by exposure to viral infection and the indicated cytokines, respectively. Four hundred and eighty-four genes were similarly modified by the cytokines and viral infection.Conclusions/interpretation The large number of modified genes observed emphasises the complex responses of human islet cells to agents potentially involved in insulitis. Notably, both cytokines and viral infection significantly (p<0.02) increased the expression of several chemokines, the cytokine IL-15 and the intercellular adhesion molecule ICAM-1, which might contribute to the homing and activation of mononuclear cells in the islets during infection and/or an early autoimmune response. The present results provide novel insights into the molecular mechanisms involved in viral- and cytokine-induced human beta cell dysfunction and death.Electronic Supplementary Material Supplementary material is available for this article at .     

Increased expression of adenylyl cyclase 3 in pancreatic islets and central nervous system of diabetic Goto-Kakizaki rats: A possible regulatory role in glucose homeostasis

Adenylyl cyclase 3 (AC3) is expressed in pancreatic islets of the Goto-Kakizaki (GK) rat, a spontaneous animal model of type 2 diabetes (T2D), and also exerts genetic effects on the regulation of body weight in man. In addition to pancreatic islets, the central nervous system (CNS) plays an important role in the pathogenesis of T2D and obesity by regulating feeding behavior, body weight and glucose metabolism. In the present study, we have investigated AC3 expression in pancreatic islets, striatum and hypothalamus of GK rats to evaluate its role in the regulation of glucose homeostasis. GK and Wistar rats at the age of 2.5 mo were used. A group of GK rats were implanted with sustained insulin release chips for 15 d. Plasma glucose and serum insulin levels were measured. AC3 gene expression levels in pancreatic islets, striatum and hypothalamus were determined by using real-time RT-PCR. Results indicated that plasma glucose levels in Wistar rats were found to be similar to insulin-treated GK rats, and significantly lower compared with non-treated GK rats. AC3 expression levels in pancreatic islets, striatum and hypothalamus of GK rats were higher compared with Wistar rats, while the levels were intermediate in insulin-treated GK rats. The AC3 expression display patterns between pancreatic islets and striatum-hypothalamus were similar. The present study thus provides the first evidence that AC3 is overexpressed in the regions of striatum and hypothalamus of brain, and similarly in pancreatic islets of GK rats suggesting that AC3 plays a role in regulation of glucose homeostasis via CNS and insulin secretion.     

替米沙坦对糖尿病大鼠胰岛硫氧还蛋白与硫氧还蛋白相互作用蛋白表达的影响

目的探讨替米沙坦对糖尿病大鼠胰岛硫氧还蛋白（TRX）与硫氧还蛋白相互作用蛋白（TXNIP）表达的影响。方法SD大鼠分为正常对照（NC）、糖尿病（DM）和替米沙坦干预（TM）组。DM组和TM组大鼠高脂高糖喂养4周后,腹腔注射链脲佐菌素（STZ）（30mg／kg）构建2型糖尿病（T2DM）模型,TM组予替米沙坦10mg·kg^-1·d^-1干预,8周后测定血浆生化指标、胰岛中胰岛素、TRX和TXNIP的表达。结果DM组和TM组空腹血糖、丙二醛以及胰岛中TXNIP水平均较NC组高（P〈0．05）,TM组血糖与DM组差异无统计学意义,但TM组上述其余指标水平较DM组低（P〈0．05）;DM组和TM组空腹胰岛素、过氧化氢酶、超氧化物歧化酶及胰岛中胰岛素和TRX水平低于NC组（P〈0．05）,但TM组高于DM组（P〈O．05）。结论T2DM大鼠胰岛TRX表达较正常大鼠降低,而TX—NIP升高;氧化应激水平和低胰岛素血症的改善可能与替米沙坦干预了这种变化有关。     

Inhibition of progressive reduction of islet beta-cell mass in spontaneously diabetic Goto-Kakizaki rats by alpha-glucosidase inhibitor

The Goto-Kakizaki (GK) rat, an animal model of type 2 diabetes, exhibits mild hyperglycemia with a reduction of beta-cell mass. The mechanism for islet structural changes in this model and whether the changes are affected by metabolic control are not known. In the present study, we examined the process of islet changes in male GK rats aged 6, 8, 12, 24, and 36 weeks. Treatment effects with an alpha-glucosidase inhibitor (Voglibose; Takeda, Osaka, Japan) for 24 weeks (12 to 36 weeks of age) were also evaluated. The beta-cell mass increased until 8 weeks of age in both GK and control rats, but the increase was significantly (P < .01) smaller in GK rats versus at 8 weeks of age. Thereafter, the beta-cell mass decreased in GK rats, whereas it remained constant in controls. Voglibose treatment significantly (P < .01) inhibited the reduction of beta-cell mass in GK rats. Proliferative activity of beta cells as measured by bromodeoxyuridine (BrdU) uptake was significantly (P < .05) lower in GK rats versus control rats at 6 and 8 weeks, but the difference disappeared after 12 weeks of age, regardless of Voglibose treatment. The present study thus demonstrates a progressive loss of beta cells in GK rats that was mitigated by Voglibose treatment. We consider that the beta-cell loss in GK rats was due to an early impairment in proliferative activity and reduced survival. Voglibose did not appear to stimulate beta-cell proliferation, but exerted its effect via a reduction of hyperglycemia.     

Vascular and renal effects of vasopeptidase inhibition and angiotensin-converting enzyme blockade in spontaneously diabetic Goto-Kakizaki rats

BACKGROUND: The renin-angiotensin system plays an important role in the pathogenesis of diabetes-induced vascular and renal complications. Vasopeptidase inhibitors simultaneously inhibit angiotensin-converting enzyme (ACE) and neutral endopeptidase. OBJECTIVE: To compare the effectiveness of vasopeptidase inhibition and ACE inhibition in preventing hypertension, endothelial dysfunction and diabetic nephropathy in spontaneously diabetic Goto-Kakizaki (GK) rats. METHODS: Eight-week-old GK rats received omapatrilat (40 mg/kg) or enalapril (30 mg/kg) for 12 weeks, either during a normal-sodium or high-sodium diet (7% w/w). Blood pressure, arterial functions and renal morphology were determined. RESULTS: Blood pressure and albuminuria were increased in GK rats compared to non-diabetic Wistar controls. Endothelium-dependent vascular relaxation in response to acetylcholine (ACh) and endothelium-independent vascular relaxation in response to sodium nitroprusside (SNP) were impaired in GK rats. Experiments with N-nitro-L-arginine methyl ester (L-NAME), diclofenac, and L-NAME + diclofenac suggested that cyclooxygenase and endothelium-derived hyperpolarizing factor components of endothelium-dependent vascular relaxation were also impaired. A high-sodium diet aggravated hypertension and diabetes-induced vascular and renal complications. Omapatrilat and enalapril normalized blood pressure and albuminuria during the normal-sodium diet, and effectively ameliorated diabetes-induced renal complications also during the high-sodium diet. However, omapatrilat improved endothelium-dependent relaxation to ACh to a greater extent (85 +/- 5%) than enalapril (68 +/- 6%, P < 0.05). Diclofenac pre-incubation eliminated this difference between omapatrilat and enalapril in ACh-induced vascular relaxation, suggesting that it was mediated, at least in part, via the cyclooxygenase pathway. CONCLUSIONS: Despite comparable blood pressure-lowering and renoprotective properties, omapatrilat may be more effective in preventing vascular dysfunction during diabetes compared to enalapril in GK rats.     

Abnormal insulin secretion and glucose metabolism in pancreatic islets from the spontaneously diabetic GK rat

Summary Insulin secretion and islet glucose metabolism were compared in pancreatic islets isolated from GK/Wistar (GK) rats with spontaneous Type 2 (non-insulin-dependent) diabetes mellitus and control Wistar rats. Islet insulin content was 24.5±3.1 U/ng islet DNA in GK rats and 28.8±2.5 U/ng islet DNA in control rats, with a mean (±SEM) islet DNA content of 17.3±1.7 and 26.5±3.4 ng (p < 0.05), respectively. Basal insulin secretion at 3.3 mmol/l glucose was 0.19±0.03 · ng islet DNA^–1· h^–1 in GK rat islets and 0.40±0.07 in control islets. Glucose (16.7 mmol/l) stimulated insulin release in GK rat islets only two-fold while in control islets five-fold. Glucose utilization at 16.7 mmol/l glucose, as measured by the formation of ³H₂O from [5-³ H]glucose, was 2.4 times higher in GK rat islets (3.1±0.7 pmol · ng islet DNA^–1 · h^–1) than in control islets (1.3±0.1 pmol · ng islet DNA^–1 · h^–1; p<0.05). In contrast, glucose oxidation, estimated as the production of ¹⁴CO₂ from [U-¹⁴C]glucose, was similar in both types of islets and corresponded to 15±2 and 30±3 % (p<0.001) of total glucose phosphorylated in GK and control islets, respectively. Glucose cycling, i. e. the rate of dephosphorylation of the total amount of glucose phosphorylated, (determined as production of labelled glucose from islets incubated with ³H₂O) was 16.4±3.4% in GK rat and 6.4±1.0% in control islets, respectively (p<0.01). We conclude that insulin secretion stimulated by glucose is markedly impaired in GK rat islets. Glucose metabolism is also altered in GK rat islets, with diminished ratio between oxidation and utilization of glucose, and increased glucose cycling, suggesting links between impaired glucose-induced insulin release and abnormal glucose metabolism.     

硝基酪氨酸在糖尿病大鼠胰岛中的表达及普罗布考的干预作用

目的观察硝基酪氨酸（NT）在糖尿病大鼠胰岛中的表达,以及普罗布考的干预对其表达和对胰岛β细胞的影响。方法大鼠高脂高糖饲料喂养4周后,腹腔注射链脲佐菌素30mg／kg复制2型糖尿病大鼠模型,普罗布考组（PB组）每天同时灌胃普罗布考500mg／kg,10周后测定空腹血糖（FPG）、空腹胰岛素（Fins）、总胆固醇（TC）、甘油三酯（TG）、丙二醛（MDA）、超氧化物歧化酶（SOD）,计算胰岛素敏感指数（ISI）,用免疫组化的方法观察NT在胰岛中的表达。结果糖尿病组（DM组）及PB组大鼠的FPG、TG、TC及MDA水平较正常对照组（NC组）明显增加（P〈0．01）,NT的平均光密度也明显高于NC组（P〈0．01）,但PB组上述指标的测定显著低于DM组（P〈0．01）;DM组和PB组大鼠的SOD水平、ISI明显低于NC组（P〈0．01）,PB组这些指标的测定显著高于DM组（P〈0．01）;NC组和PB组的Fins比较差异没有统计学意义,但都明显高于DM组（P〈0．01）。结论NT在胰岛的表达增强,普罗布考作为抗氧化剂,对于STZ引起的或糖尿病引起的氧化应激,均在一定程度上减轻其损伤,保护了胰岛功能。     

Improvement in glucose tolerance as a result of enhanced insulin sensitivity during electroacupuncture in spontaneously diabetic Goto-Kakizaki rats

We studied whether electroacupuncture (EA) applied on the abdomen improved glucose tolerance in the Goto-Kakizaki (GK) rat, a genetic model of type 2 diabetes mellitus. Male GK rats and nondiabetic Wistar rats were studied under pentobarbital anesthesia. Blood samples were drawn from the ventral tail artery during the fasting stage and after a glucose load (0.5 g/kg). Electroacupuncture (15 Hz, 10 mA) was performed for 90 minutes during both the fasting and intravenous glucose tolerance test (IVGTT) periods. A hyperinsulinemic euglycemic clamp was also carried out to assess glucose uptake during EA. A significant decrease in fasting blood glucose and an increase in plasma insulin levels were observed during the fasting period in GK rats treated with EA. Blood glucose levels after glucose load were also significantly lower in GK rats treated with EA compared with controls. The homeostasis model assessment index during IVGTT indicated an improvement in insulin sensitivity in GK rats treated with EA, whereas glucose infusion rate during hyperinsulinemic clamp was increased significantly during EA. The present study demonstrated that EA improved hyperglycemia in the fasting stage with a marked increase in plasma insulin levels. Electroacupuncture also restored impaired glucose tolerance during an IVGTT in GK rats by enhancing insulin sensitivity.     

The pancreatic islets in spontaneously hypertensive rats: islet blood flow and insulin production

The aim of the study was to investigate if hypertension affects pancreatic islet blood flow and endocrine function. For this purpose, spontaneously hypertensive rats (SHR) were compared with normotensive control Wistar-Kyoto rats (WKY). Both islet size and islet cell replication in 4-month-old SHR was increased compared with WKY. The (pro)insulin biosynthesis was reduced in islets isolated from SHR, whereas the insulin content was unchanged. A hyperinsulinemic response to glucose in vivo was observed in 4- and 12-month-old SHR. Pancreatic blood flow, measured using a microsphere technique, was lower in SHR than in WKY in rats aged 5 weeks, 4 months or 1 year. Islet blood flow was lower in 4-month-old and 1-year-old SHR. In 4-month-old animals, islet blood flow was unaffected by administration of enalaprilate and prazosin in both strains, but was markedly decreased by the administration of N(G)-methyl-L-arginine. It was concluded that the islets of SHR have a decreased insulin production in vitro and a decreased islet blood perfusion. The reasons for this are likely to be multifactorial. Because SHR maintained an essentially normal glucose tolerance, an adaptation of the beta-cells to the metabolic and hemodynamic changes imposed by hypertension occurred.     

Skeletal changes in type-2 diabetic Goto-Kakizaki rats

We characterized appendicular and axial bones in rats with type-2 diabetes in five female Goto-Kakizaki (GK) rats, a strain developed from the Wistar rat showing spontaneous type-2 diabetes, and five age- and sex-matched non-diabetic Wistar rats. The humerus, tibia, metatarsals and vertebral bodies were analysed by peripheral quantitative computerized tomography (pQCT). In diabetic rats, the height of the vertebral bodies and length of the humerus were decreased while the length of the metatarsals was increased. A decreased cross-sectional area was found in the vertebral end-plate region and the tibial metaphysis. Notably, the diaphysis in all long bones showed expansion of periosteal and endosteal circumference. In tibia this resulted in increased cortical thickness, whereas in humerus and metatarsal it was unchanged. Areal moment of inertia was increased in all diaphyses suggesting greater bending strength. The most conspicuous finding in diabetic rats pertained to trabecular osteopenia. Thus, trabecular bone mineral density was significantly reduced in all bones examined, by 33-53%. Our pQCT study of axial and appendicular bones suggests that the typical feature of diabetic osteopathy in the GK rat is loss of trabecular bone and expansion of the diaphysis. The loss of metaphyseal trabecular bone if also present in diabetic patients may prove to underlie the susceptibility to periarticular fracture and Charcot arthropathy. The findings suggest that the risk of fracture in diabetes varies according to the specific sub-regions of a bone. The approach described may prove to be useful in the early detection of osteopathy in diabetic patients who may be amenable to preventive treatment.     

Differential muscle gene expression as a function of disease progression in Goto-Kakizaki diabetic rats

The Goto-Kakizaki (GK) rat, a polygenic non-obese model of type 2 diabetes, is a useful surrogate for study of diabetes-related changes independent of obesity. GK rats and appropriate controls were killed at 4, 8, 12, 16 and 20 weeks post-weaning and differential muscle gene expression along with body and muscle weights, plasma hormones and lipids, and blood cell measurements were carried out. Gene expression analysis identified 204 genes showing 2-fold or greater differences between GK and controls in at least 3 ages. Array results suggested increased oxidative capacity in GK muscles, as well as differential gene expression related to insulin resistance, which was also indicated by HOMA-IR measurements. In addition, potential new biomarkers in muscle gene expression were identified that could be either a cause or consequence of T2DM. Furthermore, we demonstrate here the presence of chronic inflammation evident both systemically and in the musculature, despite the absence of obesity.     

Immunocytochemical studies of pancreatic acinar cells from spontaneously diabetic BB Wistar rats

Amylase in pancreatic tissue from normal and spontaneously diabetic BB Wistar rats was assessed by immunocytochemistry and analyzed by biochemical approach. Amylase immunofluorescence, in pancreatic tissues from control "non-BB" Wistar rats, gave a positive reaction. By electron microscopy, it was detected in the rough endoplasmic reticulum, Golgi apparatus, immature, and mature secretory zymogen granules of the acinar cells. Quantitative evaluations of the intensity of labeling have demonstrated the presence of an increasing gradient which followed precisely the secretory pathway. In the spontaneously diabetic BB Wistar rats, concomitant with the disappearance of insulin containing cells in the islets of Langerhans, no reaction for amylase was found in the acinar cells. Labeling for amylase was markedly reduced in the cellular organelles and the gradient along the secretory pathway was altered. In insulin-treated diabetic rats, labeling for amylase was restored. These results were in agreement with those obtained by the biochemical approach and demonstrated that, in diabetic conditions, secretion of amylase by pancreatic acinar cells is selectively impaired. This alteration, found also in pancreatic tissue from streptozotocin-diabetic rats, demonstrates that the exocrine parenchyma is under the influence of islet hormones and that both the pancreatic exocrine and endocrine tissues are closely related forming an integrated organ.     

Fatty acid pattern of pancreatic islet lipids in Goto-Kakizaki rats

Perturbations of fatty acid content and pattern were recently documented in epididymal and parametrial lipids, as well as plasma, liver, spleen, and brain phospholipids and triglycerides of Goto-Kakizaki rats (GK). This study extends such an investigation to pancreatic islets from both control and GK rats. Groups of 5,500-14,560 islets were obtained from either control or GK rats (n = 3 in each case) and examined for their lipid fatty acid content. In the islet triglycerides, the major difference between control and GK rats, i.e., a higher C18:2ω6 content in GK rats, was similar to that found in liver triglycerides. In the islet phospholipids, however, a number of differences between control and GK rats, concerning saturated, monodesaturated, and long-chain polyunsaturated ω3 and ω6 fatty acids, were often not similar to those found in liver phospholipids. The present study reveals a number of anomalies in the fatty acid pattern of islet phospholipids in GK rats, often differing from those encountered in liver phospholipids. Such a tissue specificity was borne out by the finding that, even in control animals, the situation found in islet phospholipids differed from that recorded in liver phospholipids.     

Protective effect of rhIL-1β on pancreatic islets of alloxan-induced diabetic rats

AIM: To observe the protective effect of rhIL-1β on pancreatic islets of alloxan-induced diabetic rats.METHODS: Protection of rhIL-1β on pancreatic islets of alloxan-induced diabetic rats (n = 5) was demonstrated with methods of immunohistochemistry and stereology. The concentration of serum glucose was measured by GOD method and that of serum insulin by RIA.RESULTS: The concentration of serum glucose increased but that of insulin decreased after administration of alloxan(150 mg/kg), and the volume density and numerical density of the islets were zero. In rhIL-1β pretreated rats,although the concentration of serum insulin decreased (from11.9±3.0 mIU/L to 6.1±1.6 mIU/L, P＜0.05), that of glucose was at normal level compared with the control group.As compared with alloxan group, the concentration of serum glucose in rhIL-1β pretreated rats decreased (from19.4+8.9 mmol/L to 12.0±4.0 mmol/L, P＜0.05) and the volume density increased(0/L to. 1/L, P＜0.05).CONCLUSION: rhIL-1β pretreatment may have protective effect on the islets of alloxan-induced diabetic rats.     

Protective effect of rhIL-1β on pancreatic islets of alloxan-induced diabetic rats

AIM: To observe the protective effect of rhIL-1β on pancreatic islets of alloxan-induced diabetic rats. METHODS: Protection of rhIL-1β on pancreatic islets of alloxan-induced diabetic rats (n=5) was demonstrated with methods of immunohistochemistry and stereology. The concentration of serum glucose was measured by GOD method and that of serum insulin by RIA. RESULTS: The concentration of serum glucose increased but that of insulin decreased after administration of alloxan (150mg/kg), and the volume density and numerical density of the islets were zero. In rhIL-1β pretreated rats, although the concentration of serum insulin decreased (from 11.9&#177;3.0mIU/L to 6.1&#177;1.6mIU/L,P&lt;0.05), that of glucose was at normal level compared with the control group. As compared with alloxan group, the concentration of serum glucose in rhIL-1β pretreated rats decreased (from 19.4&#177;8.9mmol/L to 12.0&#177;4.0mmol/L, P&lt;0.05) and the volume density increased(0/L to. 1/L, P&lt;0.05). CONCLUSION: rhIL-1β pretreatment may have protective effect on the islets of alloxan-induced diabetic rats.