益元灸通过AMPK-Cx43途径改善脊髓损伤后神经源性膀胱大鼠储尿功能的机制研究  

Mechanism of Yiyuan moxibustion on improving urinary storage function in rats with neurogenic bladder after spinal cord injury through the AMPK-Cx43 pathway

作　　者：张芝兰 黄晓萌 韦慧麟 黄靖 尚文雅 李冰 任亚锋 ZHANG Zhilan;HUANG Xiaomeng;WEI Huilin;HUANG Jing;SHANG Wenya;LI Bing;REN Yafeng(Rehabilitation Medicine School of Henan University of Chinese Medicine,Zhengzhou 450046 China;The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450003 China)

机构地区：[1]河南中医药大学康复医学院,郑州450046 [2]河南中医药大学第一附属医院

出　　处：《北京中医药大学学报》2024年第6期835-844,共10页Journal of Beijing University of Traditional Chinese Medicine

基　　金：河南省中医药科学研究专项(No.2022JDZX015,No.2021JDZY022)。

摘　　要：目的基于腺苷酸活化蛋白激酶(AMPK)-缝隙连接蛋白43(Cx43)途径探讨益元灸对骶上脊髓损伤后神经源性膀胱大鼠储尿功能的影响及其机制。方法将80只雌性SD大鼠按随机数字表法选取12只作为假手术组,其余68只大鼠按照改良Hassan Shaker脊髓横断法制备骶上脊髓损伤模型,待脊髓损伤模型稳定后,筛选出符合标准的脊髓损伤后神经源性膀胱模型。将造模成功的大鼠按照随机数字表法分为模型组、益元灸组、抑制剂组、益元灸+抑制剂组,每组14只。益元灸组大鼠进行益元灸治疗,抑制剂组大鼠尾静脉注射AMPK抑制剂多索吗啡(0.2 mg/kg),益元灸+抑制剂组大鼠予益元灸联合尾静脉注射多索吗啡(0.2 mg/kg),假手术组和模型组不进行任何干预,连续14 d。干预结束后采用尿流动力学评价大鼠膀胱功能;HE染色观察膀胱组织形态学变化;比色法检测膀胱组织中三磷酸腺苷(ATP)含量;免疫组化法检测膀胱组织中磷酸化AMPK(p-AMPK)、Cx43、酪氨酸蛋白激酶受体(C-kit)阳性表达;蛋白质印迹法检测膀胱组织中AMPK、p-AMPK、Cx43、C-kit蛋白表达;实时荧光定量PCR法检测膀胱组织中Cx43、C-kit mRNA的表达。结果与模型组比较,益元灸组大鼠膀胱最大容量、膀胱顺应性增高,漏尿点压力降低;细胞相对规则有序,空泡细胞减少,组织水肿减轻;膀胱组织中ATP含量减少,p-AMPK阳性表达率升高,Cx43、C-kit阳性表达率下降,p-AMPK蛋白表达升高,Cx43、C-kit蛋白表达下降,Cx43、C-kit mRNA表达下降(均P<0.05)。与益元灸+抑制剂组比较,益元灸组大鼠膀胱最大容量、膀胱顺应性增高,漏尿点压力降低;细胞排列较紧密,组织水肿减轻;膀胱组织中ATP含量减少,p-AMPK阳性表达率升高,Cx43、C-kit阳性表达率降低,p-AMPK蛋白表达升高,Cx43、C-kit蛋白表达下降,Cx43、C-kit mRNA表达下降(均P<0.05)。结论益元灸可改善骶上脊髓损伤后神经源性膀胱大鼠的储尿功Objective To investigate the effect and mechanism of Yiyuan moxibustion on urinary storage function in rats with neurogenic bladder(NB)after suprasacral spinal cord injury(SCI)based on adenosine activated protein kinase(AMPK)-connexin 43(Cx43)pathway.Methods Twelve female Sprague-Dawley rats were selected as the sham operation group using the random number table method,and 68 rats were used to prepare the suprasacral SCI model according to the modified Hassan Shaker spinal cord transection method.After the SCI model was stable,the standard NB model after SCI was screened out.The rats with successful modeling were divided into the model group(n=14),the Yiyuan moxibustion group(n=14),the inhibitor group(n=14),and the Yiyuan moxibustion+inhibitor group(n=14)according to the random number table method.The rats in the Yiyuan moxibustion group were treated with Yiyuan moxibustion.The rats in the inhibitor group were injected with AMPK inhibitor dorsomorphin(0.2 mg/kg)by tail vein.The rats in the Yiyuan moxibustion+inhibitor group were treated with Yiyuan moxibustion combined with tail vein injection of dorsomorphin(0.2 mg/kg).The sham operation group and the model group did not receive any intervention for 14 days.After the intervention,the bladder function of rats was evaluated by urodynamics.HE staining was used to observe morphological changes of the bladder tissue;the content of adenosine triphosphate(ATP)in bladder tissue was detected by colorimetry.The positive expressions of phosphorylated AMPK(p-AMPK),Cx43 and tyrosine protein kinase receptor(C-kit)in bladder tissue were detected by immunohistochemistry.The protein expressions of AMPK,p-AMPK,Cx43 and C-kit in bladder tissue were detected by Western blotting.Results Compared with the model group,the maximum bladder capacity and bladder compliance of the rats in the Yiyuan moxibustion group were increased,the leak point pressure was decreased,the cells in the Yiyuan moxibustion group were relatively regular and orderly,the vacuolar cells were reduced,and the tiss

关 键 词：脊髓损伤 神经源性膀胱 腺苷酸活化蛋白激酶-缝隙连接蛋白43途径 三磷酸腺苷 酪氨酸蛋白激酶受体 

分 类 号：R245.8[医药卫生—针灸推拿学]