【作者】 汪凌云；

【导师】 鲁云霞；

【作者基本信息】 安徽医科大学 ， 生物化学与分子生物学， 2011， 硕士

【摘要】 目的研究豹皮樟总黄酮(TFLC,Total Flavonoids of Litsea Coreana)对实验性2型糖尿病大鼠模型的治疗作用及部分机制。方法以改良的造模方法建立2型糖尿病大鼠模型,用口服葡萄糖耐量实验(OGTT)和测定空腹血糖和血胰岛素水平来评价模型是否成功建立,然后将成模大鼠分为模型组和治疗组,治疗组以非诺贝特和吡咯列酮作为阳性对照药,分别采用大、小剂量的TFLC治疗6周,同时设立正常对照组和高脂对照组,治疗结束时进行OGTT和胰岛素耐量实验(ITT)。将所有大鼠处死后,留取大鼠血清测定各项生化指标,留取肝脏组织做油红染色观察脂肪的聚集,提取肝脏的总RNA进行RT Realtime-PCR检测2型糖尿病相关基因PTP1B的mRNA水平表达,留取胰腺做胰岛素的免疫组化分析和促凋亡基因Bak1的mRNA水平表达,Western Blot检测大鼠骨骼肌中PTP1B的蛋白水平表达。结果口服葡萄糖耐量实验和胰岛素敏感指数(ISI)证实改良的造模方法已成功地建立了2型糖尿病大鼠模型,且模型组的体重、尿液葡萄糖含量都显著高于对照组。用药治疗后,治疗组大鼠的多个血清生化指标,如空腹血糖(FBG)、糖化血红蛋白(HbA1c)、空腹胰岛素(Fins)、ISI、甘油三酯(TG)、总胆固醇(TC)等均显著降低。肝脏组织切片的油红染色结果显示治疗组与模型组及高脂对照组相比,脂滴的数目和大小均明显减少。RT Realtime-PCR结果显示模型组PTP1BmRNA水平显著升高,治疗组PTP1B mRNA水平的表达与模型组相比显著降低。模型组胰腺组织的胰岛面积显著小于对照组,相对应地,Bak1基因表达增加,而TFLC可部分恢复减小的胰岛面积,减少Bak1基因的表达。骨骼肌的WesternBlot检测显示与模型组相比,大剂量TFLC治疗组PTP1B的表达可显著降低。结论TFLC具有明显的降脂降糖效应,其作用可能与其促进胰岛的增生,减少胰腺的凋亡,降低肝脏和骨骼肌中PTP1B在mRNA水平和蛋白质水平的表达,进而提高胰岛素的敏感性有关。更多还原

【Abstract】 Objective: To explore the effects and part mechanism of action of TFLC on experimental T2DM rat model.Method: We established type 2 diabetes mellitus rat model in an improved method. With oral glucose tolerance test (OGTT) and determination of fasting blood glucose and blood insulin levels ,the model was assessed whether established successfully. These rat models were then randomly allocated into model group and treatment group which contained Fenofibrate and Pioglitazone as positive controls. It also contained large dose and small dose of TFLC which were fed 6 weeks for therapy. At the same time, we set up normal control group and high fat control group. At the end of treatment, we did OGTT and insulin tolerance test(ITT). As all rats were sacrificed, blood serum of rats were taken to detect kinds of biochemical indices; liver tissues were kept to carry out dyeing of oil red O to survey the accumulation of fat and messenger RNA was extracted from liver tissues to proceed Realtime-PCR experiments to explore the expression of PTP1B, a related gene of type 2 diabetes mellitus ; pancreatic tissues were obtained to do immunohistochemical analysis;skeletal muscles were used to analyze the expression of PTP1B by Western Blot.Result: The OGTT and insulin sensitivity index(ISI) indicateed that type 2 diabetes mellitus rat model was established successfully in an improved method. Body weights and glucose in urine of T2DM model group are higher than normal control group. After medicating, several serum index of rats in treatment group were significantly decreased, such as fasting blood glucose(FBG), glycosylated hemoglobin(HbA1c),fasting insulin(Fins),ISI, triglyceride(TG), total cholesterol(TC) and so on. It was suggested the enhancement of insulin sensitivity. Staining on liver tissue slices with oil red O showed a significant reduction of lipid droplet in treatment groups, compared with T2DM group and high fat control group. Results of RT Realtime-PCR displayed that mRNA of PTP1B in model group increased significantly and mRNA of PTP1B in treatment groups were decreased significantly compared to T2DM group. Pancreatic islet area in model group was much smaller than in normal control. Consistent with this, gene Bak1 was expressed higher in model group. However, pancreatic islet area could be partially restored and expression of Bak1 could be inhibited by TFLC. Detection of Western Blot of skeletal muscles indicates a notable reduction of PTP1B expressions in large dose of TFLC treatment group compared to T2DM group.Conclusion:TFLC could reduce blood fat and glucose apparently. Its role may be related to the proliferation of pancreatic islets, the reduction of apoptosis in pancreas, decrease of PTP1B in mRNA and protein levels, and then enhancement of insulin sensitivity.更多还原