【作者】 唐庆；

【导师】 王小琴；

【作者基本信息】 湖北中医药大学 ， 中医内科学， 2010， 博士

【摘要】 目的：观察肾安提取液对糖尿病肾病(DN)小鼠肾脏的保护作用,对肾脏MASSON染色、超微结构等组织形态学的影响,对肾脏Ⅳ型胶原(Col-Ⅳ)、血管紧张素Ⅱ(AngⅡ)、血管紧张素Ⅱ1型受体(AT1R)、血管紧张素受体相关蛋白(AGTRAP)、结缔组织生长因子(CTGF)、超氧化物歧化酶(SOD)、丙二醛(MDA)、糖基化终产物(AGEs)、糖基化终产物受体(RAGEs)的影响。方法：腹腔注射链脲佐菌素(STZ)建立小鼠DN模型,模型成功后随机分为6组：正常对照组、DN模型组、厄贝沙坦组、肾安高剂量组、肾安中剂量组、肾安低剂量组,药物干预4周后,检测相对肾重、血糖、24h尿蛋白量、血肌酐等；MASSON染色法观察小鼠肾脏一般组织形态改变及胶原纤维变化,透射电镜观察小鼠肾脏超微病理改变；放射免疫法测定肾脏AngⅡ的含量,Realtime-PCR检测肾皮质AT1R、AGTRAP、CTGF、RAGEs mRNA表达,免疫组化法检测肾皮质Col-Ⅳ、AT1R、AGTRAP、CTGF、RAGEs表达；黄嘌呤氧化酶法测定肾组织SOD活性,硫代巴比妥酸反应法检测肾组织MDA含量；分光光度法检测肾组织AGEs含量。结果：DN组小鼠自主活动明显减少,行动迟缓,偶有颤抖,尿量增加明显,多饮多食明显,体型较正常对照组瘦小,MASSON染色法观察发现正常对照组肾小球、肾小管结构正常,毛细血管清晰,呈网状。模型组小鼠肾小球肥大、纤维化明显。透射电镜下观察,糖尿病肾病模型鼠肾脏肾小球基底膜增厚,同一肾小球内不同部位基底膜厚薄差异明显,足细胞肿胀,部分足突融合、甚至消失,部分足突肥大。模型小鼠相对肾重、血糖、24 h尿蛋白蛋量、血肌酐、肾组织AngⅡ、MDA、AGEs、肾皮质CTGF、RAGEs、Col-Ⅳ表达等显著升高(P<0.05或P<0.01),肾组织SOD活性、肾皮质AT1R、AGTRAP表达显著降低(P<0.01)。各药物干预对DN模型小鼠肾脏有保护作用,肾安提取液各个治疗组、厄贝沙坦组有多饮多食现象,尿量增多不明显,活动正常。MASSON染色法观察,EB组、SAG组肾小球、肾小管结构基本正常,内部间隙存在但不规则,呈轻度球囊扩张状。SAZ组、SAD组肾小球肥大,毛细血管网结构塌陷,球囊扩张明显。药物干预组肾皮质CTGF、RAGEs、Col-Ⅳ表达降低(P<0.05或P<0.01),肾皮质AT1R受体、AGTRAP表达升高(P<0.05或P<0.01),肾组织SOD活性增加(P<0.05或P<0.01), AngⅡ、MDA含量降低(P<0.01),DN小鼠蛋白尿、肾功能得到改善。结论：肾安提取液能保护DN小鼠肾脏,其机制可能与降低DN模型小鼠肾组织AngⅡ、MDA、AGEs含量,抑制肾皮质CTGF、RAGEs、Col-Ⅳ表达,增加肾皮质AT1受体、AGTRAP表达,提高肾组织SOD活性有关。更多还原

【Abstract】 Objective:To investigate the protective effect of Shen’an Extraction on diabetic nephropathy mouse’s kidney and effect on renal morphous. And the effect on renal Col-Ⅳ、AngⅡ、AT1R、AGTRAP、CTGF、SOD、MDA、AGEs、RAGEs. Method:The diabetic nephropathy mouse models were induced by intraperitoneal injection of streptozotocin(STZ). The mouse were randomly divided into 6 groups: normal control group; diabetic nephropathy model group; Irbesartan group; high-dose of Shenan group; middle-dose of Shenan group; low-dose of Shenan group. Drug intervention term was 4 weeks. The renal pathological morphous was observed by Masson staining and transmission electron microscope. Relative kidney weight, blood glucose level, serum creatinine content, excretion of the 24 hour urine protein were measured. Renal AngⅡcontent was measured by radioimmunity. The expression of AT1R、AGTRAP、CTGF、RAGE mRNA in renal cortex was detected by Realtime-PCR. The expression of AT1R、AGTRAP、CTGF、RAGE in renal cortex was measured by immunohistochemistry. SOD activity of nephridial tissue was measured by xanthine oxidase method. MDA content of nephridial tissue was measured by thiobarbituric acid reactive substance assay. AGEs content of nephridial tissue was measured by absorption spectrometry.Result:Autonomic activities of diabetic nephropathy mouse reduced notably. Urinary volume of diabetic nephropathy mouse obviously increased.diabetic nephropathy mouse’ glomcrulus got hypertrophia and fibrous degeneration. When observed by transmission electron microscope the glomerular basement membrane of diabetic nephropathy mouse became thick, the podocytes swelled, part of foot process coalesced or overgrow. Relative kidney weight, blood glucose level, serum creatinine、excretion of the 24 hour urine protein, AngⅡMDA、AGEs content and expression of CTGF、RAGEs、Col-Ⅳin cortex renis increased obviously (P<0.05 or P<0.01). SOD activity in nephridial tissue. and expression of AT1R、AGTRAP in cortex renis decreased notably (P<0.01). After Intervention of drugs AngⅡ、MDA、AGEs content and expression of CTGF、RAGEs、Col-Ⅳin cortex renis decreased obviously (P<0.05 or P<0.01).. SOD activity in nephridial tissue.and expression of AT1R、AGTRAP in cortex renis increased markly (P<0.05 or P<0.01). Conclusion: Shen’an Extraction could protect kidney of diabetic nephropathy mouse. The mechanism was related to its inhibiting effect on CTGF、RAGEs、Col-Ⅳexpression in renal cortex and decreasing the AngⅡ、MDA、AGEs content in nephridial tissue and promoting effect on AT1R、AGTRAP expression and elevating the SOD activity in nephridial tissue of diabetic nephropathy mouse.Subject words Diabetic nephropathy/TCM therapy; Diabetic nephropathy/empirical study; Shen’an Extraction; Renin-angiotensin system; Oxidative stress;Advanced glycosylation end prodrcts更多还原