【摘要】 背景:结缔组织生长因子对肾细胞的分化、增生及细胞外基质的合成和降解具有很强的调控作用,其异常表达在各种类型肾脏疾病包括糖尿病肾病纤维化进程中起着至关重要的作用。目的:观察解毒通络保肾胶囊对糖尿病大鼠肾脏病变的保护作用及其对肾脏结缔组织生长因子mRNA表达的影响。设计:以动物为观察对象,随机对照实验。单位:长春中医学院中医研究所,吉林大学再生医学科学研究所病理研究室。材料:选用清洁级Wistar大鼠46只,雌雄各半,体质量150～200g。解毒通络保肾胶囊由吉林省中医院制剂室提供(批号:2002101,由人参、黄芪、生地、枸杞子、知母、地骨皮、黄连、榛花、金银花、大黄、丹参、益母草等药物组成,0.25g/粒);苯那普利(北京诺华制药有限公司提供,批号01046)。方法:实验于2002-05/2003-01在吉林大学再生医学科学研究所病理研究室完成。①选用Wistar大鼠46只,按随机数字表法分为4组:正常对照组(10只),模型组(12只),苯那普利治疗组(12只),中药治疗组(12只)。正常对照组:腹腔内注射等体积枸橼酸钠缓冲液;其余大鼠单次腹腔注射链脲佐菌素50mg/kg,临用前用0.1mol/L枸橼酸缓冲液溶解,pH4.2。造模72h后,尾静脉采血测空腹血糖并同时测尿糖,血糖≥16.7mmol/L,尿糖以上者为糖尿病模型造模成功。苯那普利治疗组:灌胃苯那普利混悬液15mg/(kg·d);中药治疗组:灌胃解毒通络保肾胶囊混悬液5g/(kg·d);正常对照组和模型组每日上午灌胃等体积自来水,1次/d,共12周。②用药12周后检测血糖、尿素氮、血肌酐、尿白蛋白排泄率;用放免法检测肾组织血管紧张素Ⅱ;免疫组化法检测肾脏纤维连接蛋白和Ⅳ型胶原蛋白表达水平;应用反转录聚合酶链反应检测肾皮质结缔组织生长因子基因表达,并进行肾脏组织学观察。③组间比较采用方差分析。主要观察指标:各组大鼠血和尿生化学指标和肾组织血管紧张素Ⅱ、病理形态、纤维连接蛋白和兔抗鼠Ⅳ型胶原蛋白表达、结缔组织生长因子mRNA表达比较。结果:实验过程中模型组、苯那普利治疗组、中药治疗组分别死亡5,4,3只,最终进入结果分析正常对照组、模型组、苯那普利治疗组、中药治疗组分别为10,7,8,9只。①体质量与肾质量比值:模型组明显高于正常对照组(P<0.05);苯那普利治疗组和中药治疗组明显低于模型组(P<0.01)。②空腹血糖:模型组明显高于正常对照组(P<0.01),中药治疗组明显低于模型组和苯那普利治疗组(P<0.01)。③尿素氮、血肌酐、内生肌酐清除率、尿白蛋白排泄率和肾组织中血管紧张素Ⅱ含量:模型组均明显高于正常对照组(P<0.01),苯那普利治疗组和中药治疗组明显低于模型组(P<0.05～0.01)。④肾组织病理形态学改变:模型组较严重,苯那普利治疗组和中药治疗组均较轻。⑤纤维连接蛋白、兔抗鼠Ⅳ型胶原蛋白表达:模型组明显高于正常对照组(P<0.01),苯那普利治疗组和中药治疗组明显低于模型组,高于正常对照组(P<0.01)。⑥结缔组织生长因子mRNA表达:与正常对照组(0.43±0.10)比较,模型组肾皮质中结缔组织生长因子mRNA表达为其2.65倍。苯那普利治疗组和中药治疗组较模型组结缔组织生长因子mRNA表达下降23.68%和26.32%(P<0.05,0.01)。结论:解毒通络保肾胶囊对糖尿病肾脏病变有保护作用,该作用发挥可能是减少肾组织中血管紧张素Ⅱ含量,下调糖尿病大鼠肾皮质结缔组织生长因子mRNA表达,从而减少细胞外基质的沉积实现的。更多还原

【Abstract】 BACKGROUND: Connective tissue growth factor (CTGF) has very strong regulative effect on the differentiation and proliferation of renal cells as well as the synthesis and degradation of extracellular matrix. Its abnormal expression plays a key role in the process of fibrous degeneration of various renal diseases including diabetic nephropathy. OBJECTIVE: To observe the protective effect of jiedu tongluo baoshen gelatin capsule on renal disease in rats with diabetes mellitus and its effect on the expression of renal CTGF mRNA. DESIGN: A randomized controlled experiment taking animals as subjects. SETTING: Institute of Traditional Chinese Medicine, Changchun College of Traditional Chinese Medicine; Department of Pathology, Institute of Regenerated Medical Sciences, Jilin University MATERIALS: Totally 46 Wistar rats, of clean degree, male and female in half, weighing 150 to 200 g, were used in the experiment. Jiedu tongluo baoshen gelatin capsule was provided by Preparation Room, Traditional Chinese Medicine Hospital of Jilin Province (Batch No. 2002101, composed of Renshen, Huangqi, Shengdi, Gouqizi, Zhimuv Digupiv Huanglian, Zhenhua, Jinyinhua, Dahuang, Danshen, Yimucao and so on, 0.25 g/pill) and benazepril (Provided by Beijing Nuohua Pharmaceutical Company, Batch No. 01046) were used in the experiment. METHODS: This experiment was carried out in the Department of Pathology, Institute of Regenerated Medical Sciences, Jilin University from May 2002 to January 2003. (1) Totally 46 Wistar rats used in the experiment were randomly divided into 4 groups: normal control group (n=10), model group(n=12), benazepril-treated group (n=12) and traditional Chinese medicine-treated group (n=12).Rats of normal control group were intraperitoneally injected of natrium citricum buffer solution of same volume; Rats of other groups were intraperitoneally injected of 50 mg/kg streptozotocin which was dissolved by 0.1 mol/L citric acid buffer solution before use(pH 4.2). Seventy-two hours after modeling, venous blood was collected from the tails to measure the levels of fasting blood glucose and urine glucose. Blood glucose ≥16.7 mmol/L and urine glucose over suggested diabetic models were successful. Rats of benazepril-treated group were performed intragastric administration of benazepril suspension of 15 mg/(kg·d);Rats of traditional Chinese medicine-treated group were performed intragastric administration of jiedu tongluo baoshen gelatin capsule suspension of 5 g/(kg·d); Rats of normal control group and model group were performed intragastric administration of the same volume of running water on each morning, once a day, 12 weeks in total. (2) Levels of blood glucose, urea nitrogen, serum creatinine and urinary albumin excretion rate were measured 12 weeks after administration; Level of angiotensin Ⅱ of renal tissue was measured with radioimmunoassay; Levels of renal fibronectin and type Ⅳ collagen protein expression were measured with immunohistochemical method; Connective tissue growth factor gene expression of renal cortex was measured with reverse transcription-polymerase chain reaction and renal histological observation was also performed. (3) Analysis of variance was used in the intergroup comparison. MAIN OUTCOME MEASURES: Biochemical index of blood and urine, level of angiotensin Ⅱ of renal tissue, pathological morphology, expression of fibronectin and type Ⅳ rabbit anti-rat collagen protein and expression of connective tissue growth factor mRNA RESULTS: Five, four and three rats died in the process of experiment respectively in model group, benazepril-treated group and traditional Chinese medicine-treated group, finally, 10,7,8 and 9 rats entered the stage of result analysis respectively in normal control group, model group, benazepril-treated group and traditional Chinese medicine-treated group. (1) The ratio between body mass and renal mass was significantly higher in the model group than in the normal control group(P < 0.05),but was significantly lower in the benazepril-treated group and traditional Chinese medicine-treated group than in the model group (P < 0.01). (2) Fasting blood glucose was significantly higher in the model group than in the normal control group (P < 0.01), but was significantly lower in the traditional Chinese medicine-treated group than in the model group and benazepril-treated group (P < 0.01). (3) Levels of urea nitrogen, serum creatinine, creatinine clearance, urinary albumin excretion rate and angiotensin Ⅱ of renal tissue were significantly higher in the model group than in the normal control group (P < 0.01), but significantly lower in the benazepril-treated group and traditional Chinese medicine-treated group than in the model group (P < 0.05-0.01). (4)Patho-morphological change of renal tissue was very severe in the model group, but very mild in the benazepril-treated group and traditional Chinese medicine-treated group. (5)Fibronectin and type Ⅳ rabbit anti-rat collagen protein expression were significantly higher in the model group than in the normal control group (P < 0.01), those were significantly lower in the benazepril-treated group and traditional Chinese medicine-treated group than in the model group, but higher than normal control group (P < 0.01). (6) Expression of CTGF mRNA of renal cortex of model group was 2.65 times that of normal control group. Expression of CTGF mRNA of benazepril-treated group and traditional Chinese medicine-treated group was decreased by 23.68% and 26.32% , respectively (P < 0.05 or 0.01). CONCLUSION: Jiedu tongluo baoshen gelatin capsule has protective effect on diabetic nephropathy, which might be implemented by reducing the level of angiotensin Ⅱ of renal tissue and down-regulating the expression of CTGF mRNA of renal cortex of rats with diabetes mellitus, so as to reduce the deposition of extracellular matrix.更多还原