心衰康抑制慢性心衰大鼠心肌自噬及MAPK/ERK1/2信号通路
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摘要
目的:探究心衰康对慢性心衰大鼠心肌自噬的影响及其与MAPK/ERK1/2通路的关系。方法:将雄性Wistar大鼠随机分为假手术(sham)组、模型(model,结扎左冠状动脉前降支法复制大鼠慢性心衰模型)组、心衰康低、中和高剂量(TL、TM和TH)组及卡托普利(captopril)组,每组12只,采用彩色多普勒超声仪评估大鼠心脏功能;HE染色观察心肌组织形态学变化;TUNEL染色法检测心肌细胞凋亡情况;免疫荧光法检测心肌组织中微管相关蛋白1轻链3II(LC3-II)表达情况;Western blot法检测心肌组织p-ERK、p-p38 MAPK、LC3-II、beclin-1和p62的蛋白水平。结果:与sham组相比,model组左心室舒张末期内径(LVEDD)和左心室收缩末期内径(LVESD)升高,舒张末期左室后壁厚度(LVPWTd)、收缩末期左室后壁厚度(LVPWTs)和左心室射血分数(LVEF)降低,心输出量(CO)、左心室舒张压(LVDP)、左心室收缩压(LVSP)和左心室压力最大上升/下降速率(+dp/dt_(max)/-dp/dt_(max))降低(P<0.05);心肌细胞变形、坏死,心肌纤维断裂,伴有炎性细胞浸润;细胞凋亡率升高,LC3-II阳性率升高,p-ERK、p-p38 MAPK、LC3-II/LC3-I和beclin-1的蛋白水平均升高,p62蛋白表达降低(P<0.05)。与model组相比,心衰康各组与卡托普利组的LVEDD和LVESD降低,LVPWTd、LVPWTs和LVEF升高,CO、LVSP、LVDP、+dp/dt_(max)和-dp/dt_(max)升高(P<0.05);心肌细胞形态逐渐恢复正常,炎性细胞浸润减轻;细胞凋亡率降低,LC3-II阳性率降低,p-ERK、p-p38 MAPK、LC3-II/LC3-I和beclin-1的蛋白水平均降低,p62蛋白表达升高(P<0.05)。结论:心衰康能够抑制心肌自噬,发挥心肌保护作用,其机制可能与抑制MAPK/ERK1/2通路有关。
AIM: To explore the effect of Xinshuaikang on myocardial autophagy in the rats with chronic heart failure and its relationship with the MAPK/ERK1/2 signaling pathway. METHODS:The rats were divided into sham group, model group(rat model of chronic heart failure was established by ligation of anterior descending branch of left coronary artery), low-, middle-, and high-dose Xinshuaikang treatment(TL, TM and TH) groups and captopril group(treated with captopril as positive control), with 12 in each group. Doppler echocardiography was used to evaluate the cardiac function. The morphological changes of the myocardium were observed by HE staining. TUNEL staining was used to detect cardiomyocyte apoptosis. The expression of microtubule-associated protein 1 light chain 3-II(LC3-II) in the myocardium was detected by immunofluorescence labeling. The protein levels of p-ERK, p-p38 MAPK, LC3-II, beclin-1 and p62 in the myocardium were determined by Western blot. RESULTS: Compared with sham group, left ventricular end-diastolic dia-meter(LVEDD) and left ventricular end-systolic diameter(LVESD) in model group were increased, while left ventricular posterior wall thickness at end-diastole(LVPWTd), left ventricular posterior wall thickness at end-systole(LVPWTs), left ventricular ejection fraction(LVEF), cardiac output(CO), left ventricular diastolic pressure(LVDP), left ventricular systolic pressure(LVSP) and maximum rate of rise/decrease of left ventricular pressure(+dp/dt_(max)/-dp/dt_(max)) were decreased(P<0.05). The myocardial cells were deformed and necrotic, and the myocardial fibers were broken, with inflammatory cell infiltration. The apoptotic rate, the positive rate of LC3-II, and the protein levels of p-ERK, p-p38 MAPK, LC3-II/LC3-I and beclin-1 were increased, and the protein expression of p62 was decreased(P<0.05). Compared with model group, the levels of LVEDD and LVESD were decreased, LVPWTd, LVPWTs, LVEF, CO, LVSP, LVDP, +dp/dt_(max) and-dp/dt_(max) were increased in Xinshuaikang groups and captopril group(P<0.05). The morphological changes of myocardial cells were gradually returned to normal, and inflammatory cell infiltration, the apoptotic rate and the positive rate of LC3-II were decreased. The protein levels of p-ERK, p-p38 MAPK, LC3-II/LC3-I and beclin-1 were decreased, and the protein expression of p62 was increased(P<0.05). CONCLUSION: Xinshuaikang inhibits myocardial auto-phagy to play a role of cardiac protection in the rats with chronic heart failure, and its mechanism may be related to inhibition of MAPK/ERK1/2 signaling pathway.
AIM: To explore the effect of Xinshuaikang on myocardial autophagy in the rats with chronic heart failure and its relationship with the MAPK/ERK1/2 signaling pathway. METHODS:The rats were divided into sham group, model group(rat model of chronic heart failure was established by ligation of anterior descending branch of left coronary artery), low-, middle-, and high-dose Xinshuaikang treatment(TL, TM and TH) groups and captopril group(treated with captopril as positive control), with 12 in each group. Doppler echocardiography was used to evaluate the cardiac function. The morphological changes of the myocardium were observed by HE staining. TUNEL staining was used to detect cardiomyocyte apoptosis. The expression of microtubule-associated protein 1 light chain 3-II(LC3-II) in the myocardium was detected by immunofluorescence labeling. The protein levels of p-ERK, p-p38 MAPK, LC3-II, beclin-1 and p62 in the myocardium were determined by Western blot. RESULTS: Compared with sham group, left ventricular end-diastolic dia-meter(LVEDD) and left ventricular end-systolic diameter(LVESD) in model group were increased, while left ventricular posterior wall thickness at end-diastole(LVPWTd), left ventricular posterior wall thickness at end-systole(LVPWTs), left ventricular ejection fraction(LVEF), cardiac output(CO), left ventricular diastolic pressure(LVDP), left ventricular systolic pressure(LVSP) and maximum rate of rise/decrease of left ventricular pressure(+dp/dt_(max)/-dp/dt_(max)) were decreased(P<0.05). The myocardial cells were deformed and necrotic, and the myocardial fibers were broken, with inflammatory cell infiltration. The apoptotic rate, the positive rate of LC3-II, and the protein levels of p-ERK, p-p38 MAPK, LC3-II/LC3-I and beclin-1 were increased, and the protein expression of p62 was decreased(P<0.05). Compared with model group, the levels of LVEDD and LVESD were decreased, LVPWTd, LVPWTs, LVEF, CO, LVSP, LVDP, +dp/dt_(max) and-dp/dt_(max) were increased in Xinshuaikang groups and captopril group(P<0.05). The morphological changes of myocardial cells were gradually returned to normal, and inflammatory cell infiltration, the apoptotic rate and the positive rate of LC3-II were decreased. The protein levels of p-ERK, p-p38 MAPK, LC3-II/LC3-I and beclin-1 were decreased, and the protein expression of p62 was increased(P<0.05). CONCLUSION: Xinshuaikang inhibits myocardial auto-phagy to play a role of cardiac protection in the rats with chronic heart failure, and its mechanism may be related to inhibition of MAPK/ERK1/2 signaling pathway.
引文
[1] 施洋,李澜,邢晓雪,等.心肌纤维化与慢性充血性心力衰竭研究进展[J].中国临床药理学杂志,2016,32(1):87-90.
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[5] 杨广胜,孙晓,张小龙.白藜芦醇对早期心衰大鼠模型血流动力学的影响[J].中国新药与临床杂志,2014,33(12):916-918.
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[7] 薛一涛,李秋实,焦华琛,等.复心合剂对心衰大鼠信号通路Raf-MEK-ERK的影响[J].中华中医药学刊,2015,33(4):791-794.
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[11] 苗梦露,李七一,严士海,等.抗心衰颗粒对舒张性心衰大鼠模型心肌细胞内钙离子、ATP酶及心肌线粒体超微结构的影响[J].北京中医药大学学报,2013,36(7):468-471.
[12] Jia Z,Wang J,Shi Q,et al.SOX6 and PDCD4 enhance cardiomyocyte apoptosis through LPS-induced miR-499 inhibition[J].Apoptosis,2016,21(2):174-183.
[13] 王学惠,朱秀英,贵英英,等.神经调节蛋白1β与卡托普利对慢性心衰大鼠心肌纤维化及转化生长因子β1的影响[J].中国病理生理杂志,2018,34(8):1501-1506.
[14] Lin H,Li HF,Chen HH,et al.Activating transcription factor 3 protects against pressure-overload heart failure via the autophagy molecule beclin-1 pathway[J].Mol Pharmacol,2014,85(5):682-691.
[15] Jensen BC,Bultman SJ,Holley D,et al.Upregulation of autophagy genes and the unfolded protein response in human heart failure[J].Int J Clin Exp Med,2017,10(1):1051-1058.
[16] Burotto M,Chiou VL,Lee JM,et al.The MAPK pathway across different malignancies:A new perspective[J].Cancer,2014,120(22):3446-3456.
[17] Kim EK,Choi EJ.Compromised MAPK signaling in human diseases:an update[J].Arch Toxicol,2015,89(6):867-882.
[18] Cipolletta E,Rusciano MR,Maione AS,et al.Targeting the CaMKII/ERK interaction in the heart prevents cardiac hypertrophy[J].PLoS One,2015,10(6):e0130477.
[19] 蒋智,贾中申,吴玥婷,等.EdCC通过MEK-ERK信号通路减轻小鼠心肌缺血再灌注损伤[J].中国病理生理杂志,2016,32(2):221-227.
[20] Gedik N,Thielmann M,Kottenberg E,et al.No evidence for activated autophagy in left ventricular myocardium at early reperfusion with protection by remote ischemic preconditioning in patients undergoing coronary artery bypass grafting[J].PLoS One,2014,9(5):e96567.
[2] Li B,Chi RF,Qin FZ,et al.Distinct changes of myocyte autophagy during myocardial hypertrophy and heart failure:association with oxidative stress[J].Exp Physiol,2016,101(8):1050-1056.
[3] 叶汝萍,胡镜清,杨燕,等.中医药在慢性心衰康复中的应用现状[J].辽宁中医杂志,2018,45(4):868-872.
[4] 官颖,官捷.益气温阳法对老年慢性心衰康复治疗的临床观察[J].辽宁中医杂志,2009,36(5):789-790.
[5] 杨广胜,孙晓,张小龙.白藜芦醇对早期心衰大鼠模型血流动力学的影响[J].中国新药与临床杂志,2014,33(12):916-918.
[6] 谷仿丽,陈光亮.心衰康颗粒对心衰大鼠心室重塑的影响[J].中国实验方剂学杂志,2009,15(4):54-56.
[7] 薛一涛,李秋实,焦华琛,等.复心合剂对心衰大鼠信号通路Raf-MEK-ERK的影响[J].中华中医药学刊,2015,33(4):791-794.
[8] Ahmad S,Panda BP,Kohli K,et al.Folic acid ameliorates celecoxib cardiotoxicity in a doxorubicin heart failure rat model[J].Pharm Biol,2017,55(1):1295-1303
[9] Lee HM,Lee SH,Yang IH,et al.Acromegaly with normal insulin-like growth factor-1 levels and congestive heart failure as the first clinical manifestation[J].Endocrinol Metab,2015,30(3):395-401.
[10] 段学忠,王翔艳,徐静,等.慢衰康颗粒治疗慢性心力衰竭临床疗效观察[J].中华全科医学,2013,11(11):1743-1744.
[11] 苗梦露,李七一,严士海,等.抗心衰颗粒对舒张性心衰大鼠模型心肌细胞内钙离子、ATP酶及心肌线粒体超微结构的影响[J].北京中医药大学学报,2013,36(7):468-471.
[12] Jia Z,Wang J,Shi Q,et al.SOX6 and PDCD4 enhance cardiomyocyte apoptosis through LPS-induced miR-499 inhibition[J].Apoptosis,2016,21(2):174-183.
[13] 王学惠,朱秀英,贵英英,等.神经调节蛋白1β与卡托普利对慢性心衰大鼠心肌纤维化及转化生长因子β1的影响[J].中国病理生理杂志,2018,34(8):1501-1506.
[14] Lin H,Li HF,Chen HH,et al.Activating transcription factor 3 protects against pressure-overload heart failure via the autophagy molecule beclin-1 pathway[J].Mol Pharmacol,2014,85(5):682-691.
[15] Jensen BC,Bultman SJ,Holley D,et al.Upregulation of autophagy genes and the unfolded protein response in human heart failure[J].Int J Clin Exp Med,2017,10(1):1051-1058.
[16] Burotto M,Chiou VL,Lee JM,et al.The MAPK pathway across different malignancies:A new perspective[J].Cancer,2014,120(22):3446-3456.
[17] Kim EK,Choi EJ.Compromised MAPK signaling in human diseases:an update[J].Arch Toxicol,2015,89(6):867-882.
[18] Cipolletta E,Rusciano MR,Maione AS,et al.Targeting the CaMKII/ERK interaction in the heart prevents cardiac hypertrophy[J].PLoS One,2015,10(6):e0130477.
[19] 蒋智,贾中申,吴玥婷,等.EdCC通过MEK-ERK信号通路减轻小鼠心肌缺血再灌注损伤[J].中国病理生理杂志,2016,32(2):221-227.
[20] Gedik N,Thielmann M,Kottenberg E,et al.No evidence for activated autophagy in left ventricular myocardium at early reperfusion with protection by remote ischemic preconditioning in patients undergoing coronary artery bypass grafting[J].PLoS One,2014,9(5):e96567.