Page 108 - 中国全科医学2022-18

http://www.chinagp.net E-mail:zgqkyx@chinagp.net.cn ·2281·

*
Corresponding author：WANG Chunai，Chief physician；E-mail：chunaixx@163.com
【Abstract】 Background There is no safe and effective antidote for cardiotoxicity induced by bupivacaine，an
extensively used anaesthetic. Our previous research shows that acupoint treatment may partially improve bupivacaine-induced
cardiotoxicity，but the mechanism of action needs to be further studied. Objective To observe the effect of acupoint injection
of puerarin on mitochondrial energy metabolism in myocardia in bupivacaine-poisoned rats，and to explore its mechanism
of action. Methods This experiment was conducted from January to December 2019. Forty 6-month-old male SPF Wistar
rats were selected and equally randomized into a blank group （group C），a model group （group B），a treatment group
（group T），and a prevention group （group P）. Interventions were given to the groups as follows：Group P received：0.1 ml
puerarin injection injected into bilateral Neiguan acupoints. Then 5 minutes later，bupivacaine-poisoned model was established
for three groups（except for group C） using intravenous infusion of 0.5% bupivacaine（10 mg/kg） via femoral vein within 2
minutes. Group C received 0.1 ml of 0.9% sodium chloride solution injected into bilateral Neiguan acpoints. ：Immediately after
successful modeling，0.1 ml of 0.9% sodium chloride solution was injected into bilateral Neiguan acupoints for group B，and：
0.1 ml puerarin injection was injected into bilateral Neiguan points for C. The rats in group C were sacrificed at the 8th minute
of intravenous infusion of 0.9% sodium chloride solution，and those in groups B，T，and P were sacrificed at the 8th minute
of intravenous infusion of bupivacaine，then the left ventricular myocardium of all rats were taken out and cardiomyocyte mitochondria
were isolated，the structure of which was observed under the transmission electron microscope. The semi-quantitative analysis
of mitochondrial structure was evaluated by Flameng classification system. The activity of mitochondrial adenosine triphosphate
synthase （ATPase） was measured by ELISA. The content of adenosine triphosphate （ATP） was measured by colorimetric
assay. The mitochondrial membrane potential （MMP） was detected by JC-1 fluorescence staining and colorimetry. The protein
levels of myocardial mitochondrial peroxisome proliferator-activated receptor-γ coactivator-1α （PGC-1α），nuclear
respiratory factor 1（NRF1），and mitochondrial transcription factor A （mtTFA） were measured by Western blotting. Results
Group C had lower Flameng score than did other three groups（P<0.05）. The Flameng score in group P was lower than that of
groups B and T（P<0.05）. Group C had lower activity of ATPase than did groups B and T（P<0.05）. The activity of ATPase
in group P was higher than that in groups B and T（P<0.05）. Group C had higher ATP content and MMP，and protein levels of
PGC-1α，NRF1 and mtTFA than did other three groups（P<0.05）. Group B had lower ATP content，MMP，and PGC-1α
protein level than did groups T and P（P<0.05）. Group B had lower protein level of mtTFA than did groups T and P（P<0.05）.
The protein level of mtTFA in group P was higher than that in group T（P<0.05）. Conclusion Acupoint injection of puerarin
could reduce the damage of myocardial mitochondria caused by bupivacaine，and its mechanism of action may be related to
puerarin's protection of mitochondrial energy metabolism by regulating the content of ATP synthesis-related proteins PGC-1α，
NRF1 and mtTFA in mitochondria.
【Key words】 Cardiotoxicity；Myocardium；Mitochondria；Puerarin；Hydro acupuncture therapy；Bupivacaine；
Energy metabolism；Molecular mechanisms of pharmacological action （TCD）

穴位注射是在传统针灸基础上发展起来的一种疗效 粒体结构和功能损伤的影响及其可能的机制。
显著的中西医结合疗法，其以经络学说为指导，将腧穴、 1 材料与方法
经络、药效有机结合，兼具针刺和药物的治疗作用，具 1.1 实验动物及时间 6 月龄 SPF 级 Wistar 雄性大鼠
有用药剂量小、不良反应少的特点，还能大幅度提升临 40 只，体质量（280±20）g，由兰州兽医研究所实验
床疗效。近年来穴位注射方法在辅助麻醉及心律失常治 动物中心提供（SCXK［甘］2019-0002）。本实验遵循
疗领域上广泛应用 ［1-2］ 。 美国国家卫生研究院《实验动物使用指南》，并经甘肃
布比卡因是一种长效的酰胺类局麻药，因其常规剂 省中医院伦理委员会批准（批件号 2019-005）。实验
量下使用相对安全，且起效快速，常被用于各类手术 ［3］ 。 时间为 2019 年 1—12 月。
然而，过量使用或误入静脉仍会导致心律失常、心肌收 1.2 药品与试剂 盐酸布比卡因注射液购自上海朝辉
缩力降低以及循环衰竭从而引发心脏骤停 ［4］ 。临床常 药业有限责任公司（批号：1404113），葛根素注射
用针刺辅助麻醉以减少布比卡因所需剂量，从而避免其 液购自成都天台山制药有限公司（批号：180201），
［5］
不良反应 。内关穴是治疗心血管疾病的常用穴位之一， JC-1 试剂盒购自 Beyotime 公司，大鼠三磷酸腺苷合成
刺激内关穴具有抗心律失常、减轻心肌损伤的作用 ［6］ 。 酶（ATPase）酶联免疫吸附实验（ELISA）试剂盒购自
大量研究表明葛根素具有明显的心脏保护作用 ［7］ 。本 Elab Science 公司，三磷酸腺苷（ATP）ELISA 试剂盒、
研究旨在探讨穴位注射葛根素对布比卡因诱导的心肌线 Anti- 过氧化物酶体增殖物激活受体 γ 共激活因子 1α