真武汤对慢性心力衰竭大鼠心肌细胞ERK5及线粒体自噬关键蛋白的影响

别明珂 曾昭文 陈新宇

〔摘要〕 目的 探討真武汤对慢性心力衰竭（chronic heart failure， CHF）大鼠心肌细胞细胞外信号调节激酶5（extracellular signal regulated kinase 5， ERK5）蛋白及线粒体自噬关键蛋白（PINK1、Parkin、Prohibitin2、LC3-Ⅱ/LC3-Ⅰ）的影响。方法 SPF级SD大鼠52只，随机抽取13只为正常对照组，剩余39只注射阿霉素（1.5 mg/mL溶液，2.0 mL/kg体质量）6周造模。在首次注射阿霉素20 天后，对造模大鼠进行心脏彩超检查，以评估造模的均衡性。按左室射血分数将造模大鼠进行分层，并按随机区组设计分为模型组、依那普利组和真武汤组，每组13只。药物干预4周后，采用Western blot法检测各组大鼠心肌细胞ERK5、p-ERK5蛋白及线粒体自噬关键蛋白（PINK1、Parkin、Prohibitin2、LC3-Ⅱ/LC3-Ⅰ）表达水平。结果 造模6周后，模型组ERK5蛋白表达水平明显低于正常对照组，线粒体自噬关键蛋白（PINK1、Parkin、Prohibitin2、LC3-Ⅱ/LC3-Ⅰ）表达水平明显高于正常对照组（P<0.05）;各治疗组经药物干预4周后，线粒体自噬关键蛋白（PINK1、Parkin、Prohibitin2、LC3-Ⅱ/LC3-Ⅰ）表达水平较模型组均有下降趋势，ERK5蛋白表达水平较模型组均有上升趋势（P<0.05），其中真武汤组与依那普利组比较，差异无统计学意义（P>0.05）。结论 真武汤可能通过上调心肌细胞p-ERK5表达水平，同时下调线粒体自噬关键蛋白的表达水平，从而发挥治疗CHF作用。

〔关键词〕 慢性心力衰竭;真武汤;细胞外信号调节激酶5;线粒体自噬;心肌细胞

〔中图分类号〕R285.5       〔文献标志码〕A        〔文章编号〕doi：10.3969/j.issn.1674-070X.2021.12.005

Effects of Zhenwu Decoction on ERK5 and Key Proteins of Mitochondrial

Autophagy in Rats with Chronic Heart Failure

BIE Mingke1， ZENG Zhaowen2， CHEN Xinyu3*

（1. Hunan University of Chinese Medicine， Changsha， Hunan 410208， China; 2. The Second Affiliated Hospital of Integrated Traditional Chinese and Western Medicine of Hunan University of Chinese Medicine， Changsha， Hunan 410300， China;

3. The First Affiliated Hospital of Hunan University of Chinese Medicine， Changsha， Hunan 410007， China）

〔Abstract〕 Objective To investigate the effects of Zhenwu Decoction on extracellular signal regulated kinase 5 （ERK5） and key proteins of mitochondrial autophagy （PINK1， Parkin， Prohibitin2， LC3-Ⅱ/LC3-Ⅰ） in myocardial cells of rats with chronic heart failure （CHF）. Methods A total of 52 SPF degree SD rats were enrolled into the study， 13 rats were randomly selected as normal control group， and the remaining 39 rats were injected with doxorubicin （1.5 mg/mL solution， 2.0 mL/kg body mass） for 6 weeks for modeling. The model rats were subjected to a cardiac ultrasound examination in order to assess the equilibrium established in the animal model on the 20th day after the first doxorubicin injection. Model rats were underwent CHF model stratification according to left ventricular ejection fraction， and then were divided into 3 groups （model group， enalapril group and Zhenwu Decoction group） using a random block design， with 13 rats in each group. After treated with drugs for 4 weeks， the expression levels of ERK5， p-ERK5 and key proteins of mitochondrial autophagy （PINK1， Parkin， Prohibitin2， LC3-Ⅱ/LC3-Ⅰ） in cardiac myocytes of rats in each group were detected by Western blot. Results After 6 weeks of modeling， the expression of ERK5 in the myocardial cells of the model group was significantly lower than that of the normal control group， while the expression levels of key proteins of mitochondrial autophagy （PINK1， Parkin， Prohibitin2， LC3-Ⅱ/LC3-Ⅰ） were significantly higher than that of the normal control group （P<0.05）; after 4 weeks of drug intervention in each treatment group， the expression of key proteins of mitochondrial autophagy （PINK1， Parkin， Prohibitin2， LC3-Ⅱ/LC3-Ⅰ） had a downward trend compared with the model group， and the expression of ERK5 protein had an upward trend compared with the model group （P<0.05）. There was no significant difference between the naplepril group and Zhenwu Decoction group （P>0.05）. Conclusion Zhenwu Decoction may exert therapeutic effects on CHF by upregulating the expression level of p-ERK5 and downregulating the expression levels of key proteins of mitochondrial autophagy in cardiomyocytes.

〔Keywords〕 chronic heart failure; Zhenwu Decoction; extracellular signal regulated kinase 5; mitochondrial autophagy; cardiomyocytes

慢性心力衰竭（chronic heart failure， CHF）是各种心血管疾病的终末期表现和最主要的死亡原因[1]。随着老龄化进程的加速，CHF患病率显著升高[2]。研究[3]发现，心肌细胞线粒体自噬失衡而引起的心肌细胞死亡是CHF过程中的重要机制。正常范围的线粒体自噬能够及时清除受损的线粒体，对心肌细胞具有保护作用;病理状态下，心肌细胞线粒体自噬失衡，自噬小体过度激活，可直接诱导心肌细胞死亡，加重CHF病情[4]。……