基于PI3K/Akt通路的冠心病痰瘀互结证大鼠发病机制研究

Pathogenesis research of coronary heart disease with phlegm-stasis syndrome rat model based on PI3K/Akt pathway

关键词:  冠心病  痰瘀互结证  PI3K/Akt  细胞凋亡  动物模型

Key words:  coronary heart disease  phlegm and blood-stasis syndrome  PI3K/Akt  apoptosis  animal model

摘要:

[目的] 基于磷脂酰肌醇-3激酶（PI3K）/丝氨酸/苏氨酸激酶（Akt）通路探讨冠心病痰瘀互结证大鼠模型的发病机制。[方法] Wistar雄性大鼠，随机分为空白组、假手术组、痰浊组、血瘀组、痰瘀互结组。第54天，血瘀组和痰瘀互结组结扎冠状动脉左前降支，假手术组只穿线不结扎。其中空白组、血瘀组、假手术组采用普通饲料喂养8周，痰浊组和痰瘀互结组采用高脂饲料喂养8周。采用末端脱氧核苷酸转移酶介导的缺口末端标记测定（Tunel）染色的方法观察心肌细胞凋亡情况，采用蛋白免疫印迹（Western blot）方法检测Akt、磷酸化Akt（p-Akt）、B淋巴细胞瘤-2（Bcl-2）、Bcl-2相关的促凋亡蛋白（Bax）、半胱氨酸蛋白酶-3（Caspase-3）蛋白表达情况。[结果] Tunel染色结果显示：痰浊组、血瘀组和痰瘀互结组细胞凋亡明显增多。Western blot结果显示，与空白组比较，血瘀组和痰瘀互结组Akt、p-Akt和Bcl-2蛋白表达水平明显降低（P<0.05或P<0.01）；Bax和Caspase-3蛋白表达水平明显升高（P<0.05或P<0.01）；痰浊组p-Akt蛋白表达明显升高（P<0.05），Akt、Bcl-2、Bax和Caspase-3蛋白表达无统计学差异（P>0.05）。[结论] 心肌细胞Akt、p-Akt和Bcl-2蛋白的下调，Bax和Caspase-3蛋白的上调，可以激活PI3K/Akt通路，促进细胞凋亡，此过程可能是冠心病痰瘀互结证的发病机制之一。

Abstract:

[Objective] To investigate the pathogenesis of the rat model of coronary heart disease with phlegm and blood-stasis syndrome based on PI3K/Akt pathway.[Methods] Wistar male rats were randomly divided into blank group,sham operation group,turbidity group,blood stasis group and phlegm and blood-stasis group. On the 54th day,the left anterior descending coronary artery of the rats in blood-stasis group and the phlegm and blood-stasis group were ligated,and the rats in the sham operation group only threaded without ligation. The rats in the blank group,the blood-stasis group and the sham operation group were fed with normal feed for 8 weeks,while the phlegm group and the phlegm and blood-stasis group were fed with high fat diet for 8 weeks. The apoptosis of cardiomyocytes was observed by Tunel staining. The expressions of Akt,p-Akt,Bax,Bcl-2 and Caspase-3 were detected by Western blot.[Results] The results of Tunel staining showed that the apoptosis of phlegm group,blood-stasis group and phlegm and blood-stasis group increased significantly. Western blot results showed that the expression levels of Akt,p-Akt and Bcl-2 protein in blood-stasis group and phlegm and blood-stasis group were significantly lower than those in the blank group (P<0.05 or P<0.01). The expression levels of Bax and Caspase-3 were significantly increased (P<0.05 or P<0.01). The expression of p-Akt protein in the phlegm group was significantly increased (P<0.05),and there was no significant difference in the expression of Akt,Bcl-2,Bax and Caspase-3.[Conclusion] It is suggested that PI3K/Akt pathway maybe one of the pathogenesis of coronary heart disease with phlegm and blood-stasis syndrome by down-regulating Akt,p-Akt and Bcl-2 proteins in cardiomyocytes,up-regulating Bax and Caspase-3 proteins.