摘要
目的:探讨扶正活血化瘀中药芪红合剂对肾间质纤维化模型大鼠肾组织中基质金属蛋白酶组织抑制剂-1(TIMP-1)表达的影响,及其对肾间质纤维化的作用机理。
方法:采用单侧输尿管结扎(UUO)致肾间质纤维化的动物模型,以目前公认具有肾脏保护作用的血管紧张素转换酶抑制剂(ACEI)作为阳性对照,将40只雄性Wistar大鼠(体重为150g±3g)随机分为4组,每组10只:假手术组(A组)、手术模型组(B组)、芪红合剂组(C组)、依那普利组(D组)。于单侧输尿管结扎术前3天开始给各治疗组灌胃给药,假手术组和手术模型组正常进水进食。术后14天处死各组大鼠,收集血清测定血肌酐(Scr)、尿素氮(Bun)水平,肾组织用石蜡包埋切片,分别用HE、Masson染色后采用计算机图像分析系统进行分析,评定各组肾间质纤维化程度,采用免疫组化方法分别检测肾组织中基质金属蛋白酶组织抑制剂-1(TIMP-1)和纤维连接蛋白(FN)的表达,采用原位杂交的方法测定肾组织中TIMP-1mRNA的表达,并用计算机图像分析系统进行分析。
结果:镜下可见假手术组无明显病理改变,而在模型组大鼠肾组织中,则可见肾小管上皮细胞变性、扩张,部分肾小管萎缩,偶有坏死,肾间质增宽,大量炎细胞浸润,间质纤维化。模型组大鼠肾组织中FN和TIMP-1蛋白的表达较正常组显著升高,差异具有显著性(P<0.01),中药治疗组和西药治疗组大鼠肾组织中FN和TIMP-1的表达较模型组明显降低,差异具有显著性(P<0.01),两治疗组间比较,中药治疗组大鼠肾组织中FN和TIMP-1蛋白的表达较西药治疗组的表达增高,但差异无显著性(P>0.05)。模型组大鼠肾组织中TIMP-1 mRNA的表达较假手术组明显升高,差异具有显著性(P<0.01),两治疗组中TIMP-1 mRNA的表达较模型组明显降低,差异有显著性(P<0.05)(P<0.01),两治疗组间比较无显著性差异(P>0.05)。模型组大鼠血肌酐、尿素氮水平明显增高(P<0.01),两治疗组较模型组下降,有显著性差异(P<0.01),两治疗组间比较无显著性差异(P>0.05)。
结论:在单侧输尿管梗阻导致的肾间质纤维化大鼠模型中,扶正活血化瘀中药芪红合剂可降低大鼠血尿素氮、血肌酐水平,并可通过抑制基质金属蛋白酶组织抑制剂-1(TIMP-1)和FN的表达,而起到减轻肾间质纤维化病变程度的作用,推测其抑制TIMP-1表达上调的作用可能是其抗肾小管损伤及间质纤维化的作用机制之一。
Objective:To investigate the expression of TIMP-1 in renal tubulointerstitial fibrosis in rats with Unilatreal Ureteral Obstruction .To investigate the theraputic effects and its mechanisms of Astragalus and Safflower mixture.
Methods:Rat renal interstitial fibrosis model was produced by unilateral ureteral obstruction,40 Wistar male rats were randomly divided into four groups,10 in each :sham group(A group), UUO group(B group), A&S group(C group) and ACEI group(D group). Astragalus and Safflower mixture,ACEI or the same amount of water were administered by gavage beginning 3 days before UUO preparation and continued through forteen days after UUO.Sera and kidney tissues were collected from each group on the forteenth day.Bun and Scr were measured.It was carried out to measure the level of tubulointerstitial damage by Masson staining. The expression of TIMP-1 and fibronectin(FN) in kidneys was analyzed by immunohistochemical method. TIMP-1mRNA was detected by in-situ hybridization method.
Result: There was no changed in sham group.But in UUO group ,it was shown that there was tubular epithelial cells denaturalization,renal tubular expand or atrophy,interstitial macrophage and monocyt infiltration,interstitial expand,and there was no glomeruli change.While the change in A&S group and ACEI group were lighter than that in UUO group. The protein expression of TIMP-1 and FN and the renal interstitial fibrotic area were highter significantly than that in sham group(P<0.01). Compared with UUO group, those of the treatment groups decreased markedly(P<0.01). There was no significant difference between A&S group and ACEI group(P>0.05). The expression of TIMP-1mRNA were highter significantly than that in sham group(P<0.01). Compared with UUO group, those of the treatment groups decreased markedly(P<0.05)(P<0.01). There was no significant difference between two treatment groups(P>0.05). The level of BUN and Scr in UUO group were significantly highter than that in sham group(P<0.01).Compared with UUO group ,the level of BUN and Scr in A&S group and ACEI group were significantly lower(P<0.01).There was no significantly difference in these two groups (P>0.05).
Conclusion: Astragalus and Safflower mixture can downregulate the level of BUN and Scr in UUO animal model rats, Astragalus and Safflower mixture can suppress the upregulation of TIMP-1 and FN expression in kidneys of UUO animal model rats and decrease their interstitial fibrotic area.It is inferred that it may be one of the mechanisms of Astragalus and Safflower mixture suppressing renal tubular damage and interstitial fibrosis.
引文
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[13] 董建平,陈香美,冯金洲等,反义金属蛋白酶抑制剂-1 对大鼠肾小管间质损害的治疗作用,中华医学杂志,2002,82(9):617-621