基于磷脂酰肌醇3⁃激酶/蛋白激酶B信号通路探讨针刺对超负荷运动致骨骼肌损伤大鼠氧化应激损伤及骨骼肌细胞凋亡的影响  

Effects of acupuncture on oxidative stress injury and apoptosis of skeletal muscle cells in rats with skeletal muscle injury caused by overload exercise based on phosphatidylinosital 3⁃kinase/protein kinase B signaling pathway

作　　者：刘祥华[1] 罗湘筠[1] 李文倩 LIU Xianghua;LUO Xiangjun;LI Wenqian(Department of Acupuncture,The First Affiliated Hospital of Hunan TCM Junior College(Hunan Provincial TCM Hospital),Zhuzhou,Hunan 412000,China)

机构地区：[1]湖南中医药高等专科学校附属第一医院(湖南省直中医医院)针灸科,湖南株洲412000

出　　处：《安徽医药》2020年第12期2443-2448,I0004,共7页Anhui Medical and Pharmaceutical Journal

摘　　要：目的探究针刺对超负荷运动致骨骼肌损伤大鼠氧化应激损伤及骨骼肌细胞凋亡的影响,初步了解针刺对超负荷运动致骨骼肌损伤的治疗机制。方法于2018年12月至2019年6月选取60只6周龄SPF级SD大鼠,采用随机数字表法将SD大鼠分为四组:空白组(Control,C)、超负荷运动组(Overload,O)、针刺组(Acupuncture,A)和超负荷运动+针刺组(Overload and Acupuncture,OA),各15只;O组及OA组实行超负荷运动,运动结束立即针刺A组及OA组足三里、阳陵泉、内关和肾俞穴,留针20 min,持续14 d;检测各组SD大鼠骨骼肌中超氧化物歧化酶(SOD)、丙二醛、琥珀酸脱氢酶(SDH)和谷胱甘肽过氧化物酶(GSH⁃Px)活性;使用流式细胞仪检测SD大鼠骨骼肌细胞凋亡情况,使用蛋白质印迹法检测细胞凋亡相关蛋白B细胞淋巴瘤/白血病⁃2(Bcl⁃2)、Bcl⁃2相关X蛋白(Bax)、半胱氨酸天冬氨酸蛋白酶3(Caspase⁃3)活性;检测SD大鼠血清中肌酸激酶水平并使用苏木精⁃伊红(HE)染色观察SD大鼠骨骼肌情况;使用蛋白质印迹法检测磷脂酰肌醇3⁃激酶/蛋白激酶B(PI3K⁃Akt)信号通路相关蛋白表达情况。结果相比C组丙二醛水平(4.75±0.29)mol/L、骨骼肌细胞凋亡水平(9.89±1.02)%和肌酸激酶(457.54±45.02)kU/L,O组丙二醛水平(13.97±0.55)mol/L、骨骼肌细胞凋亡水平(32.58±3.50%)、Bax、Caspase⁃3蛋白表达水、血清肌酸激酶水平(1985.50±88.87)kU/L均显著升高(P<0.05);相比C组GSH⁃Px水平(142.25±8.39)kU/L、SOD水平(2.50±0.25)kU/L、SDH水平(35.75±8.78)kU/L,O组GSH⁃Px水平(55.68±6.97)kU/L、SOD水平(1.02±0.17)kU/L、SDH水平(10.58±6.50)kU/L、Bcl⁃2、PI3K、磷酸化蛋白激酶B(p⁃AKT)、哺乳动物雷帕霉素靶蛋白(m⁃TOR)水平均显著降低(P<0.05),Akt蛋白表达差异无统计学意义(P>0.05);相比O组,A组丙二醛水平(4.99±0.32)mol/L、骨骼肌细胞凋亡水平(11.52±1.02)%、Bax、Caspase⁃3蛋白表达水、血清肌酸激酶水平均(557.56±54.25)kUObjective To explore the effects of acupuncture on oxidative stress injury and apoptosis of skeletal muscle cells in rats with skeletal muscle injury caused by overload exercise,and to preliminarily understand treatment mechanism of acupuncture for skeletal muscle injury caused by overload exercise.Methods Sixty 6⁃week⁃old SPF⁃level SD rats were enrolled from December2018 and June 2019,and were assigned into four groups by random number table method:blank group(Control,C),overload exer⁃cise group(Overload,O),acupuncture group(Acupuncture,A)and overload exercise+acupuncture group(Overload and Acu⁃puncture,OA),15 cases in each group.O group and OA group were given overload exercise.Immediately after end of exercise,acu⁃puncture was performed at Zusanli,Yanglingquan,Neiguan and Shenshu acupoints in group A and OA group,retaining needle for 20 min,lasting for 14 d.The activities of superoxide dismutase(SOD),malondialdehyde(MDA),succinate dehydrogenase(SDH)and glutathione peroxidase(GSH⁃Px)in skeletal muscle of each group were detected.Flow cytometry was applied to detect apopto⁃sis of skeletal muscle cells.Western blot was applied to detect activities of apoptosis⁃related proteins[B⁃cell lymphoma/leukemia⁃2(Bcl⁃2),Bcl⁃2 related X protein(Bax),cysteinyl aspartate specific protease3(Caspase⁃3)].The level of serum creatine kinase was detected.The skeletal muscle conditions of rats were observed by hematoxylin⁃eosin(HE)staining.The expression of PI3K⁃Akt sig⁃naling pathway⁃related protein was detected by Western blot.Results Compared with MDA level[(4.75±0.29)mol/L],apoptosis level of skeletal muscle cells[(9.89±1.02)%]and creatine kinase[(457.54±45.02)kU/L]in group C,levels of MDA[(13.97±0.55)mol/L],apoptosis level of skeletal muscle cells[(32.58±3.50%)],expressions levels of Bax and Caspase⁃3 proteins,and lev⁃el of serum creatine kinase[(1985.50±88.87)kU/L]in group O were significantly increased.Compared with GSH⁃Px level[(142.25±8.39)kU/L],SOD level[(2.50±0.25)kU/L]and

关 键 词：运动损伤 肌 骨骼 针刺疗法 骨骼肌损伤 PI3K⁃Akt信号通路 氧化应激 磷脂酰肌醇3⁃激酶 超氧化物歧化酶 琥珀酸脱氢酶 谷胱甘肽过氧化物酶 大鼠 Sprague⁃Dawley 

分 类 号：R245[医药卫生—针灸推拿学]