蛇床子素对大鼠肾脏缺血再灌注损伤的保护作用研究

Protective effect of Osthole in renal ischemia-reperfusion injury in rats

目的研究蛇床子素(Osthole)预处理对大鼠肾缺血再灌注损伤的保护作用及相关机制。方法通过夹闭大鼠左侧肾蒂45 min,去除右肾,再灌注24 h构建大鼠肾缺血再灌注损伤模型。并将30只SD大鼠随机均分成假手术组、肾缺血再灌注损伤组和Osthole预处理组(40 mg/kg)。再灌注24 h后利用全自动生化分析仪检测血清中肌酐(Cr)和尿素氮(BUN)的表达水平; Western blot检测肾脏CleavedCaspase3、Caspase3、Cleaved-Caspase9、Caspase9、Bax、BCL-2以及线粒体胞质中细胞色素c(Cyt C)的表达水平;流式细胞仪检测线粒体膜电位;利用酶法测定氧自由基(ROS)含量和ATP酶活性;苏木精-伊红(HE)染色检测肾组织病理改变; TUNEL检测细胞凋亡。结果肾脏缺血再灌注损伤可明显增加Cr、BUN、Cleaved-Caspase3、Cleaved-Caspase9、Bax、ROS、TUNEL和线粒体胞质Cyt C的表达水平以及肾组织病理损伤,但可明显减少Caspase3、Caspase9和BCL-2的表达水平以及线粒体中线粒体膜电位和ATP酶的活性。而Osthole预处理可明显减少Cr、BUN、Cleaved-Caspase9、CleavedCaspase3、Bax、ROS、TUNEL和线粒体胞质中Cyt C的表达水平以及肾组织病理损伤,增加Caspase3、Caspase9和BCL-2的表达水平以及线粒体膜电位和ATP酶的活性。结论Osthole减轻大鼠肾脏缺血再灌注损伤与抑制ROS介导的线粒体凋亡信号途径相关。

Objective To explore the protective effect of Osthole preconditioning on renal ischemia-reperfusion injury in rats and its related mechanism.Methods The model of renal ischemia-reperfusion injury on rats were constructed by clamping the left renal pedicle of 45 min,removing the right kidney,and reperfusion 24 h.30 SD rats were randomly divided into sham operated group(Sham),renal ischemia-reperfusion injury group(renal,ischemia,reperfusion,injury,RIRI)and Osthole(40 mg/kg).The expression of creatinine(Cr),urea nitrogen(BUN)in serum were evaluated by automatic biochemical analyzer;Western blot was used to measure the expression of Cleaved-Caspase3,Caspase3,Cleaved-Caspase9,Caspase9,Bax,BCL-2 and cytochrome C(CytC)in the cytoplasm;The mitochondrial membrane potential was detected by flow cytometry;The content of reactive oxygen species(ROS)and ATP enzyme activity were determined by enzyme method;Hematoxylin eosin staining(HE staining)was used to examine renal pathological injury;The cell apoptosis was detected by TUNEL.Results Renal ischemia reperfusion injury could increase the pathological damage of kidney and the expression of Cr,BUN,Cleaved-Caspase3,Cleaved-Caspase9,Bax,ROS,TUNEL andcytosolic CytC with decreasing the expression of Caspase3,Caspase9 and BCL-2,the mitochondrial membrane potential and ATP enzyme activity.However,Osthole pretreatment can significantly reduce the pathological damage of kidney and the expression of Cr,BUN,Cleaved-Caspase3,Cleaved-Caspase9,Bax,ROS,TUNEL and cytosolic CytC with increasing the expression of Caspase3,Caspase9 and BCL-2,the mitochondrial membrane potential and ATP enzyme activity.Conclusion Osthole can alleviate renal ischemia-reperfusion injury in rats which is associated with inhibiting ROS mediating mitochondrial apoptotic signaling pathway.