Bladder and erectile dysfunctions in the Type 2 diabetic Goto-Kakizaki rat

Elevated Expression and Activity of Protein-Tyrosine Phosphatase 1B in Skeletal Muscle of Insulin-Resistant Type II Diabetic Goto-Kakizaki Rats

Interrelationships between carbohydrate metabolism and melatonin have been discussed for a long time. The melatonin(MT1)-receptor and Gi-mediated decrease of insulin secretion by melatonin is a well known fact. Recently, we found reduced melatonin plasma levels in type 2 diabetic patients and diabetic Goto Kakizaki (GK) rats, a model of non insulin dependent diabetes mellitus (NIDDM). Several mechanisms, such as diabetic neuropathy and oxidative stress were discussed by others to be the basis for this observation. We reported the expression of insulin receptor (InsR)-mRNA in the rat pineal gland. Earlier studies showed the InsR as being located at the cell membrane of the pinealocytes. The aim of this study was to investigate glucose, insulin and melatonin plasma levels by radioimmunoassay, regarding age and metabolic state of metabolic healthy Wistar and diabetic GK rats. Diurnal pineal InsR-mRNA expression was monitored by real-time RT-PCR. The influence of insulin and glucose on norepinephrine (NE) stimulated pineal melatonin secretion. Here we show that GK rats are permanently hyperglycemic and hyperinsulinemic at night-time. Hyperglycemia is stronger in adult than in young animals. The pineal expression level of InsR-mRNA is significantly lower in young, but higher in adult pineal glands of GK rats. No diurnal rhythmicity of expression was observed. Insulin-stimulated pineals from batch culture supernatants of both, GK- and Wistar rats, showed decreased melatonin production. First perifusion experiments revealed a lower melatonin output when stimulating pineals with NE and insulin or NE and glucose than with NE alone. The effects were not cAMP-mediated. Our results suggest that there is an insulin/InsR-mediated interrelationship between beta cells and pinealocytes. A contribution of hyperglycemia and hyperinsulinemia to reduced melatonin plasma levels seems to be likely and should be further investigated.

Regenerating (REG) proteins are associated with islet development, β-cell damage, diabetes and pancreatitis. Particularly, REG-1 and REG-3-beta are involved in cell growth/survival and/or inflammation and the Reg1 promoter contains interleukin-6 (IL-6)-responsive elements. We showed by transcriptome analysis that islets of Goto-Kakizaki (GK) rats, a model of spontaneous type 2 diabetes, overexpress Reg1, 3α, 3β and 3γ, vs Wistar islets. Goto-Kakizaki rat islets also exhibit increased cytokine/chemokine expression/release, particularly IL-6. Here we analyzed Reg1 and Reg3β expression and REG-1 immuno-localization in the GK rat pancreas in relationship with inflammation. Isolated pancreatic islets and acinar tissue from male adult Wistar and diabetic GK rats were used for quantitative RT-PCR analysis. REG-1 immunohistochemistry was performed on paraffin sections with a monoclonal anti-rat REG-1 antibody. Islet cytokine/chemokine release was measured after 48 h-culture. Islet macrophage-positive area was quantified on cryostat sections using anti-CD68 and major histocompatibility complex (MHC) class II antibodies. Pancreatic exocrine-to-endocrine Reg1 and Reg3β mRNA ratios were markedly increased in Wistar vs GK rats. Conversely, both genes were upregulated in isolated GK rat islets. These findings were unexpected, because Reg genes are expressed in the pancreatic acinar tissue. However, we observed REG-1 protein labeling in acinar peri-ductal tissue close to islets and around large, often disorganized, GK rat islets, which may retain acinar cells due to their irregular shape. These large islets also showed peri-islet macrophage infiltration and increased release of various cytokines/chemokines, particularly IL-6. Thus, IL-6 might potentially trigger acinar REG-1 expression and secretion in the vicinity of large diabetic GK rat islets. This increased acinar REG-1 expression might reflect an adaptive though unsuccessful response to deleterious microenvironment.

Reduced Innervation and Delayed Re-Innervation After Epithelial Wounding in Type 2 Diabetic Goto-Kakizaki Rats

The Goto-Kakizaki (GK) rat represents a spontaneous animal model of non-insulin-dependent diabetes mellitus (NIDDM) characterized by impaired glucose-stimulated insulin release from the pancreatic beta cells. To study whether an alteration in their islet beta-cell numbers occurs in parallel with the impairment of insulin secretion in this model, the relative volume density of beta cells was determined by means of conventional point sampling in immunostained 4-microns-thick sections of the pancreata from 8-week-old GK rats. The pancreata of nondiabetic Wistar rats were used as control parenchyma. In the GK pancreata the majority of islets was found to have a normal structure; only a few of the islets demonstrated an irregular shape (starfish-shaped islets) with fibrosis. The relative volume of the total endocrine parenchyma was found to be 2.0 +/- 0.6% (mean +/- SEM) of the whole pancreatic parenchyma in GK rats. In the control rats the corresponding value was 2.3 +/- 0.5%. The islet beta-cell density was also similar in GK and control rat islets, amounting to 75.2 +/- 8.5 and 66.9 +/- 6.6%, respectively. Thus, the total relative volume of beta cells was 1.5 +/- 0.5% in GK rats and 1.6 +/- 0.4% in controls. In conclusion, the density of beta cells is preserved in the pancreata of the young, diabetic GK rats, suggesting the absence of a causal relationship between the relative pancreatic beta-cell volume and the impaired glucose-induced insulin secretion in this NIDDM animal model.

Glucagon-like peptide-1 (GLP-1) is postprandially secreted from enteroendocrine L-cells and enhances insulin secretion. Currently, it is still controversial whether postprandial GLP-1 responses are altered in obesity and diabetes. To address the issue and to find out possible factors related, we compared postprandial GLP-1 responses in normal rats and in diabetic rats chronically fed an obesogenic diet. Male Wistar rats and diabetic Goto-Kakizaki (GK) rats were fed either a control diet or a high-fat/high-sucrose (HFS, 30 % fat and 40 % sucrose) diet for 26 weeks. Meal tolerance tests were performed for monitoring postprandial responses after a liquid diet administration (62·76 kJ/kg body weight) every 4 or 8 weeks. Postprandial glucose, GLP-1 and insulin responses in Wistar rats fed the HFS diet (WH) were higher than Wistar rats fed the control diet (WC). Although GK rats fed the HFS diet (GH) had higher glycaemic responses than GK rats fed the control diet (GC), these groups had similar postprandial GLP-1 and insulin responses throughout the study. Jejunal and ileal GLP-1 contents were increased by the HFS diet only in Wistar rats. Furthermore, mRNA expression levels of fatty acid receptors (Ffar1) in the jejunum were mildly (P = 0·053) increased by the HFS diet in Wistar rats, but not in GK rats. These results demonstrate that postprandial GLP-1 responses are enhanced under an obesogenic status in normal rats, but not in diabetic rats. Failure of adaptive enhancement of GLP-1 response in GK rats could be partly responsible for the development of glucose intolerance.

To investigate the influence and significance of gastric bypass surgery on hepatic gluconeogenesis in type 2 diabetic Goto Kakizaki(GK) rats. Forty GK rats were randomly divided into Roux-en-Y gastric bypass group(group A) and sham operation group(group B). Differences in glucose tolerance experiment(OGTT) at preoperative and postoperative 1, 2 and 4 weeks were compared and weight was recorded. Glycated hemoglobin levels were measured preoperatively and 4 weeks postoperatively. The animals were sacrificed 4 weeks after surgery and liver tissues were harvested to detect the relative expression of mRNA and protein of glucose 6 phosphatase(G-6-P) and phosphoenol pyruvate kinase(PEPCK) with RT-PCR and Western blot. Fasting blood glucose levels were 6.5, 4.9, and 4.7 mmol/L in group A, and were 10.3, 10.4, and 12.5 mmol/L in group B, and the differences between two groups were statistically significant(P<0.05). The blood glucose level at 2 h after stomach lavage were 8.3, 6.4 and 5.5 mmol/L in group A, and were 21.4, 23.8 and 24.7 mmol/L in group B at postoperative 1, 2, 4 weeks, and the differences between two groups were statistically significant(P<0.05). The glycosylated hemoglobin at postoperative 4 weeks was(6.8±1.0)%, significantly lower than that in group B[(7.9±0.8)%, P<0.05]. Hepatic G-6-P and PEPCK mRNA relative expression at postoperative 4 weeks was reduced by 21.0% and 25.9% respectively as compared to group B, and the protein expression reduced as well. Immunohistochemistry showed that hepatic glycogen sedimentary in group A increased significantly. The relative mRNA and protein level of key enzymes of hepatic gluconeogenesis are significantly decreased after Roux-en-Y gastric bypass surgery and hepatic gluconeogenesis is reduced, which may be a potential mechanism of the decrease of blood glucose.

Metformin has been reported to increase the expression of the glucagon-like peptide-1 (GLP-1) receptor in pancreatic beta cells in a peroxisome proliferator-activated receptor (PPAR)-α-dependent manner. We investigated whether a PPARα agonist, fenofibrate, exhibits an additive or synergistic effect on glucose metabolism, independent of its lipid-lowering effect, when added to metformin. Non-obese diabetic Goto-Kakizaki (GK) rats were divided into four groups and treated for 28 days with metformin, fenofibrate, metformin plus fenofibrate or vehicle. The random blood glucose levels, body weights, food intake and serum lipid profiles were not significantly different among the groups. After 4 weeks, metformin, but not fenofibrate, markedly reduced the blood glucose levels during oral glucose tolerance tests, and this effect was attenuated by adding fenofibrate. Metformin increased the expression of the GLP-1 receptor in pancreatic islets, whereas fenofibrate did not. During the intraperitoneal glucose tolerance tests with the injection of a GLP-1 analog, metformin and/or fenofibrate did not alter the insulin secretory responses. In conclusion, fenofibrate did not confer any beneficial effect on glucose homeostasis but reduced metformin's glucose-lowering activity in GK rats, thus discouraging the addition of fenofibrate to metformin to improve glycemic control.

In the present report, we examined the responses of diabetic Goto-Kakizaki (GK) rats and control Wistar-Kyoto (WKY) rats fed either a standard chow or high-fat diet (HFD) from weaning to 20 weeks of age. This comparison included gene expression profiling of skeletal muscle using Affymetrix gene array chips. The expression profiling is interpreted within the context of a wide array of physiological measurements. Genes whose expressions are different between the 2 strains regardless of diet, as well as genes that differ between strains only with HFD, were identified. In addition, genes that were regulated by diet in 1 or both strains were identified. The results suggest that both strains respond to HFD by an increased capacity to oxidize lipid fuels in the musculature but that this adaptation occurs more rapidly in WKY rats. The results also demonstrated an impaired cytokine signalling and heightened inflammatory status in the GK rats.

&lt;p dir="ltr"&gt;Roux-en-Y gastric bypass (RYGB) has been shown to inhibit β-cell apoptosis, but the underlying mechanisms are not yet fully understood. Cytochrome c oxidase subunit 6A2 (COX6A2) is expressed in β-cells. Here, we sought to investigate the role of COX6A2 in β-cell apoptosis, especially following RYGB. We found that RYGB significantly reduced β-cell apoptosis, accompanied by decreased COX6A2 expression in islets from diabetic Goto-Kakizaki (GK) rats. It is noteworthy that overexpression of COX6A2 promoted β-cell apoptosis, whereas COX6A2 deficiency suppressed it, suggesting the pro-apoptotic role of COX6A2 in β-cells. Mechanistically, increased COX6A2 interacted with and upregulated the expression of cyclophilin D (CypD), facilitating the release of cytochrome c from mitochondria to the cytoplasm, thereby promoting β-cell apoptosis. Furthermore, high-glucose-activated ChREBP epigenetically regulated COX6A2 expression by recruiting histone acetyltransferase p300 to augment histone H3 acetylation at the &lt;i&gt;Cox6a2&lt;/i&gt; promoter, a process inhibited by GLP-1 signaling. Given that RYGB enhances GLP-1 signaling, RYGB is likely to deactivate ChREBP by boosting GLP-1/PKA signaling, thereby reducing COX6A2 expression in islets from GK rats. These findings highlight the crucial role of the GLP-1/PKA/ChREBP axis-controlled COX6A2 in β-cell apoptosis, revealing a previously unrecognized mechanism underlying the reduction in β-cell apoptosis induced by RYGB.&lt;/p&gt;

Accumulated evidence links an important signal involved in glucose-stimulated insulin release to the activation of the islet lysosomal glycogenolytic enzyme acid glucan-1,4-alpha-glucosidase. We have analyzed the function of the lysosomal system/lysosomal enzyme activities in pancreatic islets of young (6-8 weeks), spontaneously diabetic, GK (Goto-Kakizaki) rats and Wistar control rats in relation to glucose-induced insulin release. The insulin secretory response to glucose was markedly impaired in the GK rat, but was restored by the adenylate cyclase activator forskolin. Islet activities of classical lysosomal enzymes, e.g.. acid phosphatase, N-acetyl-beta-D-glucosaminidase, beta-glucuronidase, and cathepsin D, were reduced by 20-35% in the GK rat compared with those in Wistar controls. In contrast, the activities of the lysosomal alpha-glucosidehydrolases, i.e.. acid glucan-1,4-alpha-glucosidase and acid alpha-glucosidase, were increased by 40-50%. Neutral alpha-glucosidase (endoplasmic reticulum) was unaffected. Comparative analysis of liver tissue showed that lysosomal enzyme activities were of the same magnitude in GK and Wistar rats. Notably, in Wistar rats, the activities of acid glucan-1,4-alpha-glucosidase and acid alpha-glucosidase were approximately 15-fold higher in islets than in liver. Other lysosomal enzymes did not display such a difference. Normalization of glycemia in GK rats by phlorizin administered for 9 days did not influence either the lysosomal alpha-glucosidehydrolase activities or other lysosomal enzyme activities in GK islets. Finally, the pseudotetrasaccharide acarbose, which accumulates in the lysosomal system, inhibited acid glucan-1,4-alpha-glucosidase activity in parallel with its inhibitory action on glucose-induced insulin release in intact Wistar islets, whereas no effect was recorded for either parameter in intact GK islets. In contrast, acarbose inhibited the enzyme activity equally in islet homogenates from both GK and Wistar rats, showing that the catalytic activity of the enzyme itself in disrupted cells was unaffected. We propose that dysfunction of the islet lysosomal/vacuolar system is an important defect impairing the transduction mechanisms for glucose-induced insulin release in the GK rat.

Hypertension superimposed on type II diabetes in Goto Kakizaki rats induces progressive nephropathy

Bone histomorphometric and biomechanical analyses in Type 2 diabetic Goto-Kakizaki rats

Physical activity exerts positive effects on glycemic control in type 2 diabetes (T2D), which is mediated in part by extensive metabolic and molecular remodeling of skeletal muscle in response to exercise, while many regulators of skeletal muscle remain unclear. In the present study, we investigated the effects of acute exercise on skeletal muscle transcriptomic responses in the Goto-Kakizaki (GK) rats which can spontaneously develop T2D. The transcriptomes of skeletal muscle from both 8-week-old GK and Wistar rats that underwent a single exercise session (60 min running using an animal treadmill at 15 m/min) or remained sedentary were analyzed by next-generation RNA sequencing. We identified 819 differentially expressed genes in the sedentary GK rats compared with those of the sedentary Wistar rats. After a single bout of running, we found 291 and 598 genes that were differentially expressed in the exercise GK and exercise Wistar rats when compared with the corresponding sedentary rats. By integrating our data and previous studies including RNA or protein expression patterns and transgenic experiments, the downregulated expression of Fasn and upregulated expression of Tbc1d1, Hk2, Lpin1, Ppargc1a, Sorbs1, and Hmox1 might enhance glucose uptake or improve insulin sensitivity to ameliorate hyperglycemia in the exercise GK rats. Our results provide mechanistic insight into the beneficial effects of exercise on hyperglycemia and insulin action in skeletal muscle of diabetic GK rats.

Effects of KIOM-79 on hyperglycemia and diabetic nephropathy in type 2 diabetic Goto-Kakizaki rats