罗格列酮对2型糖尿病大鼠肝脏线粒体功能障碍的影响

Effect of rosiglitazone on the hepatic mitochondrial dysfunction in type 2 diabetic rats

目的探讨罗格列酮对2型糖尿病大鼠肝脏线粒体功能障碍的影响,为阐明糖尿病的发病机制以及罗格列酮治疗糖尿病的药理作用提供新的实验依据。方法采用高脂饲养加腹腔注射小剂量链脲佐菌素(35 mg.kg-1)诱导2型糖尿病大鼠;检测肝脏线粒体琥珀酸脱氢酶及细胞色素C氧化酶活性、线粒体膜电位、肝ATP含量等指标以评价线粒体功能;测定过氧化物酶体增殖物激活受体γ辅激活因子1α(PGC-1α)的mRNA水平及线粒体基因细胞色素C氧化酶亚基Ⅰ(COXⅠ)与核基因-βactin的拷贝数之比来反映线粒体的生物合成;并检测解偶联蛋白2(UCP2)的基因转录、NO含量及NOS活性、脂质过氧化产物MDA含量和抗氧化酶SOD活性等指标以探讨糖尿病大鼠肝脏线粒体功能障碍的可能机制。结果糖尿病大鼠血糖、血胰岛素水平显著升高,胰岛素敏感性指数明显降低,表明2型糖尿病大鼠模型建立成功。与正常大鼠相比,2型糖尿病大鼠肝脏线粒体琥珀酸脱氢酶及细胞色素C氧化酶活性下降、线粒体膜电位降低、ATP生成减少并伴有肝脏线粒体生物合成抑制,提示线粒体功能损害;此外,肝脏UCP2转录上调,同时伴肝MDA含量增加,SOD及NOS活性降低,NO含量减少。罗格列酮治疗8周后,不仅明显改善糖尿病大鼠肝脏线粒体功能的损害,而且增加肝脏线粒体生物合成。进一步研究揭示罗格列酮对糖尿病大鼠肝脏线粒体功能的保护作用可能与下调UCP2 mRNA水平,上调PGC-1α转录表达,降低体内氧化应激水平有关。结论罗格列酮对糖尿病大鼠肝脏线粒体功能障碍具有保护作用。

Objective To explore the role of rosiglitazone in the development of hepatic mitochondrial dysfunction of type 2 diabetic rats and to provide novel experimental evidence for the pathogenesis of diabetes mellitus and the pharmacological mechanism of rosiglitazone for type 2 diabetes. Methods Male Sprague-Dawley rats were divided into a control, a diabetic, a rosiglitazone-treated diabetic, and a rosiglitazone-treated control group. Type 2 diabetic rats were induced by first feeding high-fat diet for 4 weeks and second injecting streptozotocin （35 mg· kg^-1 , i. p. ） in the bolus. After being continuously fed high-fat diet for 8 weeks, the activities of sueeinate dehydrogenase and cytochrome C oxidase in the hepatic mitoehondria of rats were measured by colorimetry, the mitoehondrial membrane potential was detected by fluorospeetrophotometry, and ATP levels in the liver were assessed by luciferin/luciferase reaction. These indexes were used for evaluating the mitochondrial function. RT- PCR was employed to detect PGC -1α mRNA levels, PCR was used to assess the copy numbers of mitoehondrial genes such as COX Ⅰ, and nuclear genes, such as β-actin, were used for reflecting the mitochondrial biogenesis. In addition, the mRNA level of UCP2, the activities of NOS and SOD, and the concentrations of NO and MDA were assayed to approach the role of rosiglitazone in the development of hepatic mitochondrial dysfunction in diabetic rats and its potential mechanism. Results Plasma glucose and insulin levels in the diabetic rats were significantly elevated while insulin sensitivity index was obviously decreased, suggesting that the type 2 diabetic rat model was successfully established. Activities of suecinate dehydrogenase and cytochrome C oxidase were suppressed in the hepatic mitochondria from diabetic rats. Mitochondrial membrane potential and ATP levels were decreased in type 2 diabetic rats. In parallel to these changes, hepatic mitochondrial biogenesis was inhibited with the down-regulation of PGC-1α mRNA levels and mitochondrial DNA contents. These results suggest that the mitochondrial function in the liver of diabetic rats was impaired. The transcription of UCP2 was upregulated in the liver of diabetic rats. Inhibited NOS activity, decreased NO contents, increased MDA levels, and suppressed SOD activity were also found in the liver of type 2 diabetic rats. After treatment with rosiglitazone for 8 weeks, not only was mitochondrial function improved but also mitochondrial biogenesis was increased in the liver of type 2 diabetic rats. Further studies revealed that the improvement of rosiglitazone on the mitochondrial dysfunction in the liver of diabetic rats may be associated with downregulating UCP2 transcription, upregulating PGC-1α mRNA levels, and decreasing oxidative stress. Conclusion Rosiglitazone has protective effect on the hepatic mitochondrial dysfunction in type 2 diabetic rats.