Preferential alteration of oxidative relative to total glycolysis in pancreatic islets of two rat models of inherited or acquired Type 2 (non-insulin-dependent) diabetes mellitus

Summary In islets from both adult rats injected with streptozotocin during the neonatal period and spontaneously diabetic rats obtained by repeated selective breedings (GK rats), the ratio between d-[3, 4-¹⁴C]glucose oxidation and d-[5-³H]glucose conversion to ³HOH was 25% lower than in islets from control rats, indicating an impaired contribution of oxidative to total glycolysis. No primary defect in the Krebs cycle was found in the islets of diabetic rats, as judged from the ratio between either d-[2-¹⁴C]glucose or d-[6-¹⁴C]glucose and d-[3, 4-¹⁴C]glucose oxidation. Therefore, we propose that a preferential alteration of oxidative glycolysis in the pancreatic beta cell may contribute to the impairment of glucose-induced insulin release not only in a cytotoxic but also in a spontaneous model of non-insulin-dependent diabetes mellitus.     

参芪复方对实验性2型糖尿病大血管病变胰岛素抵抗的干预作用

目的：观察参芪复方对2型糖尿病（T2DM）大血管病变大鼠胰岛素抵抗的干预作用。方法：GK大鼠连续腹腔注射L-N-硝基精氨酸甲酯（L-NAME）,同时喂饲高脂饮食,复制2型糖尿病大血管病变模型。入选大鼠随机分为正常Wistar对照、模型、雷米普利（阳性对照）及参芪复方低、高剂量5个组。造模同时开始给药,连续28 d。实验结束时放射免疫法测血清胰岛素（INS）;免疫印迹（Western Blot）法测骨骼肌葡萄糖转运蛋白4（GLUT4）水平。结果：参芪复方高剂量可降低GK大鼠空腹血糖（P〈0.05）,参芪复方低、高剂量和雷米普利均可改善胰岛素抵抗,增加胰岛素敏感性（P〈0.01）。雷米普利、参芪复方高剂量可上调GLUT4蛋白水平（P〈0.05）。结论：参芪复方具有改善胰岛素抵抗作用,上调骨骼肌GLUT4蛋白水平可能为其机制之一。     

高热量高蛋白饮食诱导GK大鼠糖尿病肾病模型的建立

目的 运用高热量高蛋白饮食诱导GK大鼠2型糖尿病肾病模型的建立,并探讨其可能的作用机制.方法 28周龄GK大鼠24只,随机分成对照组、模型组,每组各12只,模型组给予高热量高蛋白饮食,对照组给予正常饮食,共8周.于第0、4、8周观察24 h尿微量白蛋白、24h尿蛋白、尿肌酐、尿微量白蛋白/尿肌酐比值水平;于第0、8周观察空腹血糖和血清肌酐、尿素氮、总胆固醇、甘油三脂、一氧化氮水平;实验结束时取双肾称重并计算肾肥大指数,取肾组织观察病理形态学变化,检测肾组织钠钾ATP酶活性.结果 与对照组比,模型组大鼠24 h尿微量白蛋白、24 h尿蛋白、尿微量白蛋白/尿肌酐比值、空腹血糖、总胆固醇、甘油三脂、一氧化氮、肾肥大指数水平和肾组织钠钾ATP酶活性显著提高,模型组肾小球体积增大,系膜基质增生,基底膜增厚明显.结论 运用高热量高蛋白饮食诱导GK大鼠可成功建立2型糖尿病肾病模型.血糖血脂的上升是糖尿病肾病形成的重要因素,同时钠钾ATP酶活性增强进一步损伤肾小管功能,一氧化氮升高促使肾小球高灌注、高滤过,也是加速GK大鼠肾病形成的原因.     

Binding properties of [H]gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) enantiomers in the rat central nervous system

Gacyclidine (cis(pip/me)-1-[1-(2-thienyl)-2-methylcyclohexyl]piperidine) is a TCP derivative, which exhibits potent neuroprotective properties against glutamate-induced neurotoxicity in vitro and in vivo. In order to better understand gacyclidine pharmacological properties, the binding parameters of its enantiomers ((−) and (+)[³H]GK11) were determined in the rat central nervous system (CNS). An autoradiographic study has shown that their binding distributions are correlated with those of N-methyl- -aspartate (NMDA) receptors throughout the CNS. Globally, the labeling was the highest with (−)[³H]GK11. In the cerebellum, both radioligands similarly labeled the molecular layer. For both radioligands, on telencephalic, cerebellum and spinal cord homogenates, the association and dissociation kinetics were accounted for by multiphasic process. In all regions, (−)[³H]GK11 exhibited the highest affinity in the nanomolar range. The pharmacological study revealed that both enantiomers labeled both high and low affinity sites in all regions. The pharmacological profile of high affinity sites was correlated with those of NMDA receptors. Those of low affinity sites were different in telencephalic and cerebellar homogenates. Overall, this study showed that low affinity sites might constitute a heterogeneous population, which could include σ receptors in the cerebellum. The autoradiographic study has shown that these sites may be located in the molecular layer. The contribution of low affinity sites to the neuroprotective properties of gacyclidine remains to be investigated.     

Src activation generates reactive oxygen species and impairs metabolism–secretion coupling in diabetic Goto–Kakizaki and ouabain-treated rat pancreatic islets

AIMS/HYPOTHESIS: Na(+)/K(+)-ATPase inhibition by ouabain suppresses ATP production by generating reactive oxygen species (ROS) and impairs glucose-induced insulin secretion from pancreatic islets. To clarify the signal-transducing function of Na(+)/K(+)-ATPase in decreasing ATP production by the generation of ROS in pancreatic islets, the involvement of Src was examined. In addition, the significance of Src activation in diabetic islets was examined. METHODS: Isolated islets from Wistar rats and diabetic Goto-Kakizaki (GK) rats (a model for diabetes) were used. ROS was measured by 5-(and 6)-chloromethyl-2',7'-dichlorofluorescein fluorescence using dispersed islet cells. After lysates were immunoprecipitated by anti-Src antibody, immunoblotting was performed. RESULTS: Ouabain caused a rapid Tyr(418) phosphorylation, indicating activation of Src in the presence of high glucose. The specific Src inhibitor 4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine (PP2) restored the ouabain-induced decrease in ATP content and the increase in ROS production. Both PP2 and ROS scavenger restored the impaired insulin release and impaired ATP elevation in GK islets, but had no such effect in control islets. PP2 reduced the high glucose-induced increase in ROS generation in GK islet cells but had no effect on that in control islet cells. Moreover, ouabain had no effect on ATP content and ROS production in the presence of high glucose in GK islets. CONCLUSIONS/INTERPRETATION: These results indicate that Src plays a role in the signal-transducing function of Na(+)/K(+)-ATPase, in which ROS generation decreases ATP production in control islets. Moreover, ROS generated by Src activation plays an important role in impaired glucose-induced insulin secretion in GK islets, in which Src is endogenously activated independently of ouabain.     

改良胆胰转流术对Goto-Kakizaki大鼠血糖的影响

目的 观察改良胆胰转流术对非肥胖性2型糖尿病模型鼠(GK大鼠)的降糖作用.方法将16只GK大鼠随机分为A、B组,分别对A组行改良胆胰转流术,对B组行假手术.检测术前1周及术后第1、8周两组大鼠体质量、空腹血糖、随机血糖、糖耐量.结果 术后早期A组大鼠体质量由术前的(373.100±9.859)g降至(343.260±12.399)g,与B组比较,差异有统计学意义.术后第8周A组大鼠随机血糖降至( 10.500±1.509) mmol/L,明显低于B组的(19.480±1.281) mmol/L,A组大鼠糖耐量明显优于B组.结论 改良胆胰转流术能改善GK大鼠糖耐量,胆胰液在外科手术治疗2型糖尿病的机制中可能起重要作用.     

Programmed disorders of β‐cell development and function as one cause for type 2 diabetes? The GK rat paradigm

Now that the reduction in beta-mass has been clearly established in humans with type 2 diabetes mellitus (T2DM) 1-4, the debate focuses on the possible mechanisms responsible for decreased beta-cell number and impaired beta-cell function and their multifactorial etiology. Appropriate inbred rodent models are essential tools for identification of genes and environmental factors that increase the risk of abnormal beta-cell function and of T2DM. The information available in the Goto-Kakizaki (GK) rat, one of the best characterized animal models of spontaneous T2DM, are reviewed in such a perspective. We propose that the defective beta-cell mass and function in the GK model reflect the complex interactions of three pathogenic players: (1) several independent loci containing genes causing impaired insulin secretion; (2) gestational metabolic impairment inducing a programming of endocrine pancreas (decreased beta-cell neogenesis) which is transmitted to the next generation; and (3) secondary (acquired) loss of beta-cell differentiation due to chronic exposure to hyperglycemia (glucotoxicity). An important message is that the 'heritable' determinants of T2DM are not simply dependant on genetic factors, but probably involve transgenerational epigenetic responses.     

Reduced sensitivity of dihydroxyacetone on ATP-sensitive K+ channels of pancreatic beta cells in GK rats

Summary In the GK (Goto-Kakizaki) rat, a genetic model of non-insulin-dependent diabetes mellitus, glucose-induced insulin secretion is selectively impaired. In addition, it has been suggested by previous studies that impaired glucose metabolism in beta cells of the GK rat results in insufficient closure of ATP-sensitive K⁺ channels (K_ATP channels) and a consequent decrease in depolarization, leading to a decreased insulin release. We have recently reported that the site of disturbed glucose metabolism is probably located in the early stages of glycolysis or in the glycerol phosphate shuttle. In the present study, in order to identify the impaired metabolic step in diabetic beta cells, we have investigated insulin secretory capacity by stimulation with dihydroxyacetone (DHA), which is known to be directly converted to DHA-phosphate and to preferentially enter the glycerol phosphate shuttle. In addition, using the patch-clamp technique, we also have studied the sensitivity of DHA on the K_ATP channels of beta cells in GK rats. The insulin secretion in response to 5 mmol/l DHA with 2.8 mmol/l glucose was impaired, and DHA sensitivity of the K_ATP channels was reduced in beta cells of GK rats. From these results, we suggest that the intracellular site responsible for impaired glucose metabolism in pancreatic beta cells of GK rats is located in the glycerol phosphate shuttle.Abbreviations DHA Dihydroxyacetone - K_ATP channel ATP-sensitive K⁺ channel - GK rat Goto-Kakizaki rat - KRBB Krebs Ringer bicarbonate buffer - BSA bovine serum albumin - NIDDM non-insulin-dependent diabetes     

偏二甲基肼和四氧化二氮吸入性肺损伤中肺泡表面活性物质的变化

目的研究火箭液体推进剂偏二甲基肼（UDMH）和四氧化二氮（N2O4）吸入性急性肺损伤（ALI）中肺泡表面活性物质（PS）的变化及其与ALI的关系。方法84只大鼠分为对照组和5个实验组。实验组在静式染毒柜内分别吸入UDMH（0．98g／m^3）和N2O4（0．19g／ms）各10min，间隔10min。实验组在吸入后2、6、12、24、48h分别处死。测定各组大鼠常压下支气管肺泡灌洗液（BALF）的表面张力（γ）、加压下最低表面张力（γmin）、磷脂酰胆碱（PC）和磷脂酰乙醇胺（PE）。同时测定肺组织湿／于重比（W／D）、BALF中总蛋白和乳酸脱氢酶（LDH）等肺损伤指标，评价病理学改变。结果UDMH-N2O4吸入性ALI表现为肺W／D比值增加、BALF中总蛋白和LDH升高，以及肺水肿、肺间隔增厚等病理变化。PS的异常表现为：①在UDMH—N2O4吸入后2hBALF中γ和γmin。即明显升高，至24-48h尚未恢复到对照组水平；②吸入后24hBALF中PC明显低于对照组，PE含量明显高于对照组。以BALF中总蛋白作为肺损伤指标与PS异常的指标γ和γmin分别做直线相关分析，相关系数分别为0．435（P〈0．01）和0．419（P〈0．01）。结论UDMH—N2O4吸入性ALI中存在PS异常，PS异常可能参与了UDMH—N2O4吸入性ALI的发生发展过程。     

Anomalies of fetal development in GK rats

Fetal development was investigated in Goto-Kakizaki (GK) rats. Between days 15 and 20 after identification of a positive sperm plug, the GK rats gained less weight than control animals. The number of conceptuses in each litter was not significantly different in control and GK rats. The incidence of abortive fetal development, however, averaged 39.7%±9.1% in GK rats, compared with only 5.6%±0.2% in control animals. The placental weight in living fetuses was slightly lower in GK rats than control rats. The crown-rump length was identical in the fetuses of control and diabetic mothers. The number of ossification points in the lumbosacral spine, pelvic girdle and anterior and posterior limbs was significantly lower in fetuses of GK rats than control animals. These anomalies could not be blamed on a lower plasma insulin concentration in GK than control animals, whether before or during (days 15–20) pregnancy. Moreover, in both control and GK rats, the insulinogenic index was raised during pregnancy. These findings indicate that GK rats represent a new model for the study of diabetes-related fetal anomalies, their pathogenesis and prevention. Received: 30 August 1996 / Accepted in revised form: 5 February 1997