电针“足三里”对脾虚大鼠骨骼肌线粒体自噬相关蛋白表达的影响  

Effects of electroacupuncture at “Zusanli”(ST 36) on expression of mitophagy-related proteins in skeletal muscle in rats with spleen deficiency syndrome

作　　者：董佳梓 张妤[1] 魏云涛[2] 许环宇 刘路 邓婷月 张立德[1] DONG Jiazi;ZHANG Yu;WEI Yuntao;XU Huanyu;LIU Lu;DENG Tingyue;ZHANG Lide(Liaoning University of TCM, Shenyang 110847, China;Liaoning Cancer Hospital ＆ Institute;Wu Zhongpei Memorial Hospital of Shunde District, Foshan City, Guangdong Province)

机构地区：[1]辽宁中医药大学,沈阳110847 [2]辽宁省肿瘤医院 [3]广东省佛山市顺德区伍仲珮纪念医院

出　　处：《中国针灸》2018年第7期741-746,共6页Chinese Acupuncture & Moxibustion

基　　金：国家自然科学基金项目:81603704;辽宁省教育厅一般项目:L 201708

摘　　要：目的:观察脾虚大鼠骨骼肌线粒体自噬相关蛋白表达的变化,以期阐释针刺足三里治疗脾虚证的部分作用机制。方法:40只雄性SD大鼠,适应性饲养后随机分为正常对照组、脾虚组、足三里组、非经非穴组,每组10只。除正常对照组外,其余3组采用三因素复合法复制脾虚证模型,造模持续14 d。足三里组电针大鼠双侧"足三里"穴,非经非穴组电针双侧髂嵴上非经非穴点,疏密波,频率2 Hz/100 Hz,每日1次,每次20 min,干预10 d。正常对照组和脾虚组大鼠在每天同一时间固定20 min,不予电针干预。采用高效液相色谱法检测大鼠骨骼肌三磷酸腺苷(ATP)、磷酸腺苷(AMP)的含量;Western blotting法检测大鼠骨骼肌腺苷酸活化蛋白激酶(AMPK)、磷酸化AMPK(p-AMPK)、Atg1/失调51样激酶-1(ULK1)、磷酸化ULK1(p-ULK1)、微管相关蛋白1轻链3-Ⅰ(LC3-Ⅰ)以及LC3-Ⅱ蛋白的表达。结果:脾虚组大鼠骨骼肌ATP含量明显低于正常对照组(P<0.01),足三里组ATP含量高于脾虚组(P<0.05)但低于正常对照组(P<0.05),非经非穴组与脾虚组差异无统计学意义(P>0.05);脾虚组、足三里组AMP/ATP较正常对照组上调(P<0.01,P<0.05);脾虚组p-AMPK/AMPK较正常对照组差异无统计学意义(P>0.05),足三里组p-AMPK/AMPK较正常对照组和脾虚组均上调(均P<0.05);足三里组p-ULK1/ULK1和LC3-Ⅱ/LC3-Ⅰ均明显高于正常对照组和脾虚组(均P<0.01)。结论:电针"足三里"可能是通过激活AMPK,AMPK直接磷酸化ULK1,形成稳定的ULK1/AMPK复合物,提高线粒体自噬能力,从而起到调理脾胃、补中益气治疗脾虚证的作用。Objective To observe the expression change of mitophagy-related proteins in skeletal muscle in rats with spleen deficiency syndrome and to explain the partial action mechanism of acupuncture at Zusanli（ST 36） for spleen deficiency syndrome. Methods Forty male SD rats, after normal feeding, were randomly divided into a normal group, a spleen deficiency group, a Zusanli group and a non-acupoint group, ten rats in each group. Except the normal group, the three factors modeling method was used for 14 days to establish the model of spleen deficiency syndrome on the other 3 groups. The rats in the Zusanli group were treated with EA at bilateral ＂Zusanli＂（ST 36）, while the rats in the non-acupoint group were treated with EA at bilateral non acupoint（dense-sparse wave, frequency of 2 Hz/100 Hz, 20 min per treatment, once a day for 10 days）. The rats in the normal group and spleen deficiency group were treated with immobilization for 20 min per day, and no EA was given. The HPLC method was applied to measure the content of adenosine triphosphate（ATP） and adenosine monophosphate（AMP） in skeletal muscle. The Western blotting method was applied to measure the expression of adenosine monophosphate activated protein kinase（AMPK）, p-AMPK, ULK1, p-ULK1,LC3-Ⅰand LC3-Ⅱ in skeletal muscle. Results The ATP content in the spleen deficiency group was significantly lower than that in the normal group（P〈0.01）; the ATP content in the Zusanli group was significantly higher than that in the spleen deficiency group（P〈0.05） but lower than that in the normal group（P〈0.05）, there was no significant difference between the non-acupoint group and the spleen deficiency group（P〉0.05）. Compared with the normal group, the AMP/ATP in the spleen deficiency group and the Zusanli group were significantly up-regulated（P〈0.01, P〈0.05）. The differences of p-AMPK/AMPK between the spleen deficiency group and the normal group was not significant（P〉0.05）. Compared with the normal group and sp

关 键 词：脾虚 穴 足三里 电针 线粒体自噬 腺苷酸活化蛋白激酶(AMPK) 三磷酸腺苷(ATP) 

分 类 号：R245[医药卫生—针灸推拿学]