摘要
目的研究三氟淫羊藿素(ICTF)对大鼠心肌缺血再灌注损伤(MI/RI)的治疗作用,并探讨其是否通过丝氨酸/苏氨酸蛋白激酶/哺乳动物雷帕霉素靶蛋白(Akt/mTOR)信号通路调节自噬起作用。方法采用结扎左冠状动脉前降支45 min,再灌注60 min的方法制作MI/RI模型,雄性SD大鼠分为假手术组、MI/RI模型组、ICTF 0.5,1.0和2.0 mg·kg-1组。标准肢体Ⅱ导联心电图监测T波和ST段变化,TTC染色测定心肌梗死面积,Western印迹法检测心肌组织中微管相关蛋白2/1轻链3(LC3Ⅱ/LC3Ⅰ)比值、Akt和mTOR蛋白磷酸化水平,免疫荧光检测心肌组织LC3蛋白表达水平。结果监测心电图发现,与假手术组相比,MI/RI模型组大鼠再灌注60 min时的T波(P<0.05)和ST段(P<0.01)明显升高;与模型组相比,ICTF 1.0 mg·kg-1组的T波(P<0.05)和ST段(P<0.01)显著降低。TTC染色结果表明,模型组出现明显的心肌梗死灶(P<0.01),ICTF 1.0 mg·kg-1组心肌梗死面积百分数显著降低(P<0.01)。Western印迹结果显示,与假手术组比较,模型组LC3Ⅱ/LC3Ⅰ比值显著升高(P<0.01),Beclin-1蛋白磷酸化水平升高(P<0.05),Akt(P<0.01)和m TOR(P<0.05)蛋白磷酸化水平降低;与模型组相比,ICTF 1.0 mg·kg-1组LC3Ⅱ/LC3Ⅰ比值明显降低(P<0.01),Beclin-1(P<0.01)蛋白磷酸化水平降低,Akt和m TOR蛋白磷酸化水平升高(P<0.01)。免疫荧光结果表明,与假手术组相比,模型组大鼠心肌组织中LC3蛋白表达增多(P<0.01);与模型组比较,ICTF 1.0 mg·kg-1组表达减少(P<0.01)。结论ICTF对大鼠MI/RI具有保护作用,其机制可能与调控Akt/m TOR信号通路抑制细胞过度自噬有关。
OBJECTIVE To investigate the effect of trifluoro-icaritin(ICTF)on myocardial ischemia/reperfusion injury(MI/RI)in rats and to explore whether it plays a role in regulating autophagy through the serine/threonine kinase/mammalian target of rapamycin(Akt/mTOR)signaling pathway.METHODS Male SD rats were ligated for 45 min and reperfused for 60 min to establish an MI/RI model.The rats were divided into sham,MI/RI model and model+ICTF 0.5,1.0 and 2.0 mg·kg-1 groups.Ⅱlead electrocardiogram(ECG)in T wave and ST segment changes were recorded.The area of myocardial infarction was determined by TTC.The protein levels and phosphorylation levels of microtubule-associated protein 2/1 light chain 3(LC3Ⅱ/LC3Ⅰ),Beclin-1,Akt and m TOR in myocardial tissues were detected by Western blotting.The level of LC3 in myocardial tissues was detected by immunofluorescence test.RESULTS The ECG showed that the T wave(P<0.05)and ST segment(P<0.01)of the model group were significantly higher than those of the sham group after 60 min of reperfusion,while the T wave(P<0.05)and ST segment(P<0.01)of the ICTF 1.0 mg·kg-1 group were obviously lower than those of the model group.TTC staining of heart sections showed that the area of myocardial infarction in the model group was larger than in the sham group(P<0.01),while that in the ICTF 1.0 mg·kg-1 group was smaller than in the model group(P<0.01).Western blotting results showed that compared with the sham group,the ratios of LC3Ⅱ/LC3Ⅰ(P<0.01)and p-Beclin-1/Beclin-1(P<0.05)in the model group were significantly increased,while Akt(P<0.01)and mTOR(P<0.05)were decreased.In addition,compared with the model group,the ratio of LC3Ⅱ/LC3Ⅰ(P<0.01)and p-Beclin-1/Beclin-1(P<0.01)in the ICTF 1.0 mg·kg-1 group was reduced,and the phosphorylation levels of Akt(P<0.01)and m TOR(P<0.01)were increased.Immunofluorescence results of frozen sections of myocardial tissues showed that LC3 protein expression increased in the model group compared with the sham group(P<0.01),but decreased in the ICTF1.0 mg·kg-1 group compared with the model group(P<0.01).CONCLUSION ICTF has a protective effect on myocardial ischemia/reperfusion injury in rats,and its mechanism may be related to the regulation of Akt/mTOR signaling pathway to inhibit excessive autophagy.
作者
刁思帏
陈建宁
刘国祥
杨宇博
彭燕婷
汪文略
叶嘉民
陈浩
黎晓
黄志华
DIAO Si-wei;CHEN Jian-ning;LIU Guo-xiang;YANG Yu-bo;PENG Yan-ting;WANG Wen-lue;YE Jia-min;CHEN Hao;LI Xiao;HUANG Zhi-hua(School of Basic Medicine,Gannan Medical University,Ganzhou 341000,China)
出处
《中国药理学与毒理学杂志》
CAS
北大核心
2019年第8期569-575,共7页
Chinese Journal of Pharmacology and Toxicology
基金
江西省自然科学基金（20161BAB205228）
赣南医学院本科生科技创新项目（XS201701）~~
