基于miR-155/JAK2/STAT3信号通路探讨仙茅苷减轻大鼠心肌缺血再灌注损伤作用机制

Study on Mechanism of Curculigoside Alleviating Myocardial Ischemia-Reperfusion Injury in Rats Based on miR-155/JAK2/STAT3 Signaling Pathway

目的探讨仙茅苷对心肌缺血再灌注(I/R)大鼠心肌损伤的改善作用及对微小RNA(miR)-155/Janus蛋白酪氨酸激酶2/信号转导及转录激活蛋白3(JAK2/STAT3)信号通路的调节作用。方法将60只大鼠随机分为假手术组、模型组、仙茅苷组、miR-155过表达组和miR-155过表达+仙茅苷组。除假手术组外,其余组大鼠采用冠状动脉左前降支结扎法建立心肌I/R模型,仙茅苷组和miR-155过表达+仙茅苷组大鼠于建模前6 d腹腔注射仙茅苷50 mg/kg,1次/d;miR-155过表达组和miR-155过表达+仙茅苷组大鼠于建模前在左心室上取3个位点注射miR-155 mimic。再灌注24 h后超声心动图检测心功能,TTC染色检测心肌梗死面积,实时荧光定量PCR(qRT-PCR)检测心肌组织中miR-155表达水平,苏木精-伊红(HE)染色观察心肌损伤病理表现,ELISA检测血清中肌酸激酶同工酶MB(CK-MB)、心肌肌钙蛋白T(cTnT)和乳酸脱氢酶(LDH)水平,蛋白质免疫印迹法检测心肌组织中p-JAK2和p-STAT3蛋白相对表达量。结果与模型组比较,仙茅苷组心肌组织miR-155水平降低,心肌梗死面积减小,左室射血分数(LVEF)和左室缩短分数(LVFS)升高,左室舒张末期内径(LVESD)和左室收缩末期内径(LVEDD)减小,血清中CK-MB、cTnT、LDH水平下降,心肌组织中p-JAK2和p-STAT3蛋白相对表达量升高,而miR-155过表达组以上各指标变化趋势相反(均P<0.05);与miR-155过表达+仙茅苷组比较,miR-155过表达组miR-155水平升高,心肌梗死面积增大,LVEF和LVFS降低,LVESD和LVEDD增大,血清中CK-MB、cTnT、LDH水平上升,p-JAK2和p-STAT3蛋白相对表达量降低,而仙茅苷组以上各指标变化呈相反趋势(均P<0.05)。结论仙茅苷可减轻大鼠心肌I/R损伤,改善心功能,其可能通过抑制miR-155表达从而上调JAK2/STAT3信号通路发挥作用。

Objective To investigate the ameliorative effects of curculigoside on myocardial ischemia/reperfusion(I/R)injury in rats and the regulation of microRNA(miR-155)/Janus kinase 2/signal transducer and activator of transcription 3(JAK2/STAT3)signaling pathway.Methods Sixty rats were randomly divided into sham operation group,model group,curculigoside group,miR-155 overexpression group and miR-155 overexpression+curculigoside group.Except for sham operation group,I/R models were established in other groups through ligation of left anterior descending coronary artery.Six days before modeling,the rats in the curculigoside group and the miR-155 overexpression+curculigoside group were introperitoneally injected with curculigoside(dose 50 mg/kg)once a day.The rats in the miR-155 overexpression group and the miR-155 overexpression+curculigoside group were injected with miR-155 mimic at 3 sites in the left ventricle 2 h before modeling.Then 24 h after reperfusion,echocardiography was used to detect cardiac function,TTC staining was used to detect myocardial infarction area,qRT-PCR was used to detect mir-155 expression level,and HE staining was used to observe myocardial pathological injury.The activities of creatine kinase isoenzyme MB(CK-MB),cardiac troponin T(cTnT)and lactate dehydrogenase(LDH)in serum were detected by ELISA,and the relative protein expression levels of p-JAK2 and p-STAT3 were detected by Western blotting.Results Compared with the model group,the miR-155 level was decreased,the myocardial infarction area was decreased,LVEF and LVFS were increased,LVESD and LVEDD were decreased,serum levels of CK-MB,cTnT and LDH were decreased and the relative protein expression levels of p-JAK2 and p-STAT3 were increased in curculigoside group,while the changes of above indexes showed an opposite trend in miR-155 overexpression group(P<0.05).Compared with miR-155 overexpression+curculigoside group,miR-155 level was increased,myocardial infarction size was increased,LVEF and LVFS were decreased,LVESD and LVEDD were increased,serum levels of CK-MB,cTnT and LDH were increased and relative protein expressions of p-JAK2 and p-STAT3 were decreased in miR-155 overexpression group.The changes of above indexes showed an opposite trend in curculigoside group(P<0.05).Conclusion Curculigoside can alleviate myocardial injury and improve cardiac function in I/R rats,which may play a role by inhibiting mir-155 expression and up-regulating JAK2/STAT3 signaling pathway.