降尿酸方对尿酸性肾病大鼠肾脏细胞凋亡的影响

Effect and Mechanism of Jiangniaosuan Prescription on Renal Cell Apoptosis in Rats with Uric Acid Nephropathy

目的观察降尿酸方对尿酸性肾病(UAN)大鼠肾脏细胞凋亡的影响,探讨其可能的作用机制。方法32只SD大鼠随机分为正常组、模型组、别嘌醇组和降尿酸方组,每组8只。采用氧嗪酸钾(1500 mg/kg)和腺嘌呤(100 mg/kg)混合灌胃制备UAN大鼠模型,造模同时别嘌醇组和降尿酸方组分别予别嘌醇(25 mg/kg)和降尿酸方(16 g/kg)灌胃,正常组和模型组予等体积生理盐水灌胃,连续4周。HE染色观察肾组织病理变化,六胺银染色检测肾组织尿酸盐结晶含量,透射电镜观察肾小管上皮细胞超微结构,Western blot检测肾组织p53、Caspase-3、Bax蛋白表达,免疫荧光检测肾组织Bcl-2蛋白表达,TUNEL染色检测肾小管上皮细胞凋亡。结果与正常组比较,模型组大鼠肾小管管壁变薄,管腔扩张,有炎症细胞浸润,肾小管损伤评分明显增加(P<0.001),尿酸盐结晶含量明显增加(P<0.001),肾小管上皮细胞损伤,p53、Caspase-3、Bax蛋白表达明显升高(P<0.05,P<0.01),Bcl-2蛋白表达有降低趋势,肾小管上皮细胞凋亡率明显升高(P<0.001);与模型组比较,降尿酸方组和别嘌醇组大鼠肾小管上皮细胞轻度萎缩,肾小管管腔稍扩张,肾小管损伤评分明显降低(P<0.05,P<0.01),尿酸盐结晶含量明显减少(P<0.01),肾小管上皮细胞损伤减轻,p53、Caspase-3、Bax蛋白表达明显降低(P<0.05,P<0.01),Bcl-2蛋白表达有升高趋势,肾小管上皮细胞凋亡率明显降低(P<0.01)。结论降尿酸方能改善UAN大鼠肾损伤,其机制可能与减少肾脏尿酸盐结晶沉积,抑制p53/Caspase-3信号通路,从而减少肾小管上皮细胞凋亡有关。

Objective To observe the effects of Jiangniaosuan Prescription on renal cell apoptosis in rats with uric acid nephropathy(UAN);To discuss its possible mechanism of action.Methods Totally 32 SD rats were randomly divided into normal group,model group,allopurinol group,and Jiangniaosuan Prescription group,with 8 rats in each group.The UAN rat model was established by gavage with the mixture of potassium oxazinate(1500 mg/kg)and adenine(100 mg/kg).The allopurinol group and Jiangniaosuan Prescription group were gavaged with allopurinol(25 mg/kg)and Jiangniaosuan Prescription(16 g/kg)respectively,the normal group and model group were gavaged with the same volume of normal saline,for 4 weeks.The pathlogical changes of renal tissue was observed by HE staining;the content of uric acid crystallization in renal tissue were detected by silver hexamine method;the ultrastructure of renal tubular epithelial cells was observed by transmission electron microscope;Western blot was used to detect the expressions of p53,Caspase-3 and Bax protein in renal tissue;immunofluorescence was used to detect the expression of Bcl-2 protein in renal tissue;TUNEL was used to detect the apoptosis of renal tubular epithelial cells.Results Compared with the normal group,the renal tubular wall became thinner in the model group,the lumen expanded,there was inflammatory cell infiltration,and the renal tubular injury score increased significantly(P<0.001),the content of uric acid crystallization increased significantly(P<0.001),renal tubular epithelial cells were injured,the expressions of p53,Caspase-3 and Bax protein increased significantly(P<0.05,P<0.01),the expression of Bcl-2 protein decreased,and the apoptosis rate of renal tubular epithelial cells increased significantly(P<0.001).Compared with the model group,the renal tubular epithelial cells in Jiangniaosuan Prescription group and allopurinol group shrank slightly,the renal tubular lumen expanded slightly,the renal tubular injury score decreased significantly(P<0.05,P<0.01),the content of uric acid crystallization reduced significantly(P<0.01),the injury of renal tubular epithelial cells was eased,the expressions of p53,Caspase-3 and Bax decreased significantly(P<0.05,P<0.01),the expression of Bcl-2 protein increased,and the apoptosis rate of renal tubular epithelial cells decreased significantly(P<0.01).Conclusion Jiangniaosuan Prescription can improve renal injury in UAN rats.Its mechanism may be related to reducing renal uric acid crystallization deposition,inhibiting p53/Caspase-3 signaling pathway and reducing renal tubular epithelial cell apoptosis.