【作者】 涂翔；

【导师】 叶学锋；

【作者基本信息】 成都中医药大学 ， 中西医结合临床， 2010， 博士

【摘要】 糖尿病肾病(DN)是糖尿病的主要慢性并发症之一。蛋白尿不仅是DN的临床表现,而且是DN加重、进展的重要因素。寻求降低DN蛋白尿更加有效的方法对于DN的治疗而言具有重要意义。本论文对口服益气活血中药复方降低糖尿病肾病尿蛋白的随机对照试验(RCT)进行Meta分析。本论文选用清热通络中医治法,重点观察其对链脲佐菌素诱导DN大鼠模型蛋白尿的干预作用及其机制研究。目的：观察清热通络法对实验性糖尿病肾病大鼠蛋白尿的干预作用及其机制研究。方法：采用链脲佐菌素(STZ)腹腔注射法制备1型糖尿病肾病大鼠模型。实验分为4组：正常对照组、模型对照组、清热通络组、厄贝沙坦对照组。造模成功12周后,观察大鼠一般状态、体重、空腹血糖、24小时尿量及尿蛋白定量、肾功能的变化;采用光镜和电镜观察肾小球及肾小管间质形态学改变；采用原位杂交方法检测大鼠肾小球足细胞Nephrin和Podocin mRNA表达的变化,并半定量分析其阳性表达面积和表达强度的变化。结果：实验结束时,①与正常对照大鼠比较,DN模型大鼠体重均下降,空腹血糖、24h尿量均增加,DN大鼠与正常大鼠之间比较,以上指标差异有统计学意义。模型大鼠与正常对照大鼠比较,24h尿蛋白增加,差异有统计学意义；经过厄贝沙坦干预,24h尿蛋白减少与模型对照组对比差异有统计学意义；清热通络法干预后,24h尿蛋白减少,与厄贝沙坦干预比较差异有统计学意义。②HE染色结果各组大鼠均未见明显病理学改变,符合DN4级分类系统中1级水平的病理学改变。③电子显微镜统计学结果显示：DN模型大鼠与正常对照大鼠比较,基底膜厚度增厚,差异有统计学意义；经过厄贝沙坦和清热通络法干预,增厚程度显著减轻,与模型对照组差异有统计学意义。④原位杂交结果显示：与正常对照大鼠比较,DN模型大鼠Nephrin mRNA表达减少,差异有统计学意义；厄贝‘沙坦干预改善DN模型大鼠Nephrin mRNA表达减少,与模型对照组差异有统计学意义；与厄贝沙坦比较,清热通络法改善Nephrin mRNA表达减少的效果有统计学意义。与正常对照大鼠比较,DN模型大鼠Podocin mRNA表达减少,差异有统计学意义；厄贝沙坦与清热通络法干预均能显著改善DN模型大鼠Podocin mRNA表达减少,与模型对照组比较差异有统计学意义。结论：①STZ腹腔内一次性注射造模14周,可用于早期糖尿病肾病造模,动物模型病理学改变符合DN4级分类系统中1级水平。②清热通络法能够减少早期DN大鼠尿蛋白。③清热通络法减少DN持续蛋白尿的机制之一是拮抗糖尿病肾小球足细胞裂孔膜蛋白Nephrin和Podocin表达下调。④清热通络法减少DN持续蛋白尿的机制之一是拮抗糖尿病肾小球基底膜增厚。更多还原

【Abstract】 Diabetic nephropathy (DN) is one of the major complications of diabetes mellitus. Proteinuria is not only the manifestation of DN but an important factor in DN progression. It is important to seek a more effective method to reduce proteinuria in the management of DN. The present paper carried out a meta-analysis for the randomized controlled trials evaluating Chinese herbal combination prescriptions supplementing Qi and activating blood circulation for proteinuria reduction in patients with DN. The present paper also studied the effect of a Chinese herbal combination prescription, namely, Qingre Tonglu, on proteinuria in DN rats induced by Streptozotocin (STZ) and its potential mechanism.Objectives:study the effect of Qingre Tongluo on proteinuria in DN rats induced by STZ and its potential mechanism..Methods:Type 1 diabetic nephropathy rat model was induced by STZ intraperitoneal injection. The rats were randomly assigned into four groups:normal control group, modeling group, Qingre Tongluo group and irbesartan group. Twelve weeks after successful modeling, the changes in the weight, fasting blood glucose (FBG),24 h urine volume and urinary protein and renal function were studied; the morphological changes in the glomeruli, tubules and interstitium were studied by light microscope and electron microscope; the changes in the mRNA expression of podocyte proteins, i.e., Nephrin and Podocin, were studied by in situ hybridization and the changes in positive expression area and intensity were semi-quantified. Results:at the end of experiment,①compared with normal control rats, the weight of DN rats reduced while FBG and 24 h urine volume increased. The differences between normal rats and DN rats about these measures were statistically significant. Compared with normal rats, the 24h urinary protein was statistically increased in DN rats. The rats received irbesartan showed a reduction in 24h urinary protein and the difference between irbesartan and modeling group was statistically significant. The 24h urinary protein reduced in Qingre Tongluo group and the difference between Qingre Tongluo group and irbesartan group was statistically significant.②No obvious pathological changes were found in HE stained sections. This result was in line with the first grade pathological change in the four-grade DN classification system.③The results of electron microscope showed that the glomerular basement membrane (GBM) thickened in DN rats. The difference between normal rats and DN rats was statistically significant. When received irbesartan and Qingre Tongluo, the thickening degree of GBM obviously alleviated and, compared with modeling group, the difference was statistically significant.④The results of in situ hybridization demonstrated that the expression of Nephrin mRNA reduced in DN rats. Compared with normal rats the difference was statistically significant. Irbesartan improved the reduction in Nephrin mRNA in DN rats and the difference between modeling group and irbesartan group was statistically significant. Compared with irbesartan, the difference about Qingre Tongluo improve the reduction in Nephrin mRNA was statistically significant. Compared with normal rats, the expression of Podocin mRNA in DN rats reduced and the difference was statistically significant. Both irbesartan and Qingre Tongluo could ameliorate the reduction in Podocin mRNA in DN rats and compared with modeling group the difference was statistically significant.Conclusions:①The method could be used in early stage DN modeling that a single intraperitoneal injection of STZ after 14 weeks. The pathological change of this model is in line with the first level in the four-grade DN classification system.②Qingre Tongluo could reduce the proteinuria in early stage DN.③One of the mechanisms by which Qingre Tongluo reduces proteinuria is its antagonism against the downregulation of Nephrin and Podocin.④One of the mechanisms by which Qingre Tongluo reduces proteinuria is its antagonism against the thickening of glomerular basement membrane.更多还原