摘要
目的:电针可通过增强自噬流、促进神经元再生、重塑轴突和髓鞘等实现对脊髓损伤的保护作用,但其在神经源性尿潴留中的作用尚不明确。本研究探讨电针通过增强腺苷活化蛋白激酶(AMP activated protein kinase,AMPK)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)通路介导的自噬治疗神经源性尿潴留的作用机制。方法:建立骶髓损伤后神经源性尿潴留大鼠模型。将造模成功的大鼠随机分为模型组、电针组(采用电子刺激针对次髎、中极、三阴交行电针,每天1次,每次20 min,共7 d)、电针+AMPK抑制剂组(在电针组处理的基础上,在第1和4天,于L_(2~3)椎间隙周围肌内注射100μg的AMPK抑制剂compound C)。另设正常组不进行任何处理。采用多通道生理记录仪记录膀胱最大容量、膀胱基础压力、漏尿点压力、膀胱顺应性;HE染色观察膀胱组织形态;透射电镜观察自噬情况;免疫荧光染色观察微管相关蛋白1轻链3(LC3)II、Beclin1蛋白质的表达;蛋白质印迹法检测膀胱组织中AMPK、磷酸化的AMPK(p-AMPK)、mTOR、磷酸化的mTOR(p-mTOR)、LC3II、Beclin1的蛋白质表达水平。结果:与正常组相比,模型组膀胱最大容量、漏尿点压力、膀胱顺应性,膀胱组织中p-AMPK、LC3II、Beclin1蛋白质表达水平升高,p-mTOR蛋白质表达水平降低(均P<0.05)。与模型组相比,电针组膀胱最大容量、膀胱顺应性、膀胱组织中p-mTOR蛋白质表达水平降低,p-AMPK蛋白、LC3II、Beclin1蛋白质水平升高(均P<0.05);与电针组相比,电针+AMPK抑制剂组膀胱最大容量、膀胱顺应性,膀胱组织中p-mTOR蛋白水平升高,p-AMPK、LC3II、Beclin1蛋白质水平降低(P<0.05)。模型组膀胱体积变大,各层次不清,出现不同程度变性,组织损伤严重,出现自噬小体;电针组膀胱较模型组小,各层次清晰可见,出现自噬小体;电针+AMPK抑制剂组各层次稍凌乱,出现损伤。结论:电针通过AMPK/mTOR通路激活自噬,从而减轻脊髓损伤导致的神经源性尿潴留。
Objective:Electroacupuncture can enhance autophagic flow,promote neuronal regeneration,axonal and myelin remodeling to achieve the protection of spinal cord injury,but its role in neurogenic urine retention is not completely clear.This study aims to investigate whether the mechanism of electroacupuncture in the treatment of neurogenic urine retention is through autophagy mediated by adenosine monophosphate activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR)pathway.Methods:A rat model of neurogenic urine retention after sacral spinal cord injury was established.The rats with successful model were randomly divided into a model group,an electroacupuncture group(electro-acupuncture for Ciliao,Zhongji,and Sanyinjiao by electronic stimulation,once a day,20 min each time for 7 days),and an electroacupuncture+AMP-activated protein kinase(AMPK)inhibitor group(on the basis of the treatment of electroacupuncture group,100μg of AMPK inhibitor compound C was injected intramuscularly around the L_(2-3)intervertebral space on the 1st and 4th day).The normal group did not receive any treatment.The maximum bladder volume,bladder basal pressure,leak point pressure,and bladder compliance were recorded by multi-channel physiological recorder;the morphology of bladder tissue was observed by HE staining;autophagy was observed under transmission electron microscope;the expressions of LC3II and Beclin1protein were observed by immunofluorescence staining;the protein levels of AMPK,phosphorylated-AMPK(p-AMPK),m TOR,phosphorylated-mTOR(p-mTOR),microtubule associated protein 1 light chain 3(LC3)II and Beclin1 in bladder tissue were detected by Western blotting.Results:Compared with the normal group,the maximum bladder capacity,leak point pressure,bladder compliance,p-AMPK,LC3II,Beclin1 protein expressions in the bladder tissue of the model group increased,and the p-mTOR protein expressions were decreased(all P<0.05);compared with the model group,the maximum bladder capacity,bladder compliance,p-mTOR protein expression in the bladder tissue of the electroacupuncture group were decreased,and the p-AMPK,LC3II,and Beclin1 protein expressions were increased(all P<0.05);compared with the electroacupuncture group,the maximum bladder capacity,bladder compliance,p-mTOR protein expression in the bladder tissue of the electroacupuncture+AMPK inhibitor group were increased,the p-AMPK,LC3II,and Beclin1 protein expressions were decreased(all P<0.05).In the model group,the bladder became larger,with unclear and varying degrees of degeneration,severe tissue damage and autophagosome appeared;the bladder of the electroacupuncture group was smaller than that of the model group,and all levels were clearly visible with autophagy bodies;the layers were slightly disordered and damaged in the electroacupuncture+AMPK inhibitor group.Conclusion:Electroacupuncture can activate autophagy through AMPK/mTOR pathway,thereby reducing neurogenic urine retention caused by spinal cord injury.
作者
李正飞
张任
赵国瑞
匡尧
LI Zhengfei;ZHANG Ren;ZHAO Guorui;KUANG Yao(Department of Massage,First Teaching Hospital of Tianjin University of Traditional Chinese Medicine,Tianjin 300193;National Clinical Research Center of Chinese Medicine Acupuncture and Moxibustion,Tianjin 300193,China)
出处
《中南大学学报（医学版）》
CAS
CSCD
北大核心
2022年第4期488-496,共9页
Journal of Central South University ：Medical Science
基金
天津市教委科研计划项目(2018KJ028)。
关键词
电针
磷酸腺苷活化蛋白激酶
哺乳动物雷帕霉素靶蛋白
自噬
神经源性尿潴留
electroacupuncture
adenine monophosphate activated protein kinase
mammalian target of rapamycin
autophagy
neurogenic urine retention
