摘要
目的探讨心跳骤停心肺复苏后脑损伤(BIC)模型中,治疗性低温(TH)对神经元氧化应激损伤的作用及相关机制。方法雄性SD大鼠60只,7~10周,体重280~320 g。采用随机数字表法分为五组:假手术组(S组)、BIC组、低温治疗组(TH组)、沉默冷诱导RNA结合蛋白(CIRP)后低温治疗组(THC组)和空载后低温治疗组(THN组),每组12只。采用经食管连续快速起搏诱发室颤,心跳骤停4 min后,行心肺复苏术(CPR),建立BIC模型。于BIC模型制备后1 d,采用免疫荧光法检测海马活性氧簇(ROS)荧光强度,Western blot法检测海马CIRP、硫氧还蛋白1(Trx1)、磷酸化凋亡信号调节激酶1(ASK1)蛋白含量及其下游p38和c-Jun N-末端激酶(JNK)磷酸化含量。于BIC模型制备后3 d,采用原位末端标记(TUNEL)法检测海马神经元凋亡细胞百分比。结果与S组比较,BIC模型制备后1 d BIC组ROS荧光强度明显升高,CIRP含量明显降低,p-ASK1、p-p38含量明显升高;TH组和THN组ROS荧光强度均明显升高,CIRP含量明显升高,p-JNK含量明显降低;THC组ROS荧光强度明显升高,CIRP、Trx1含量明显降低,p-ASK1、p-p38含量明显升高(P<0.05)。与BIC组比较,BIC模型制备后1 d TH组和THN组ROS荧光强度明显降低,CIRP、Trx1含量明显升高,p-ASK1和p-JNK含量明显降低;TH组p-p38含量明显降低;THC组ROS荧光强度明显降低,p-ASK1和p-p38含量明显升高(P<0.05)。与THC组比较,BIC模型制备后1 d TH组和THN组ROS荧光强度明显降低,CIRP、Trx1含量明显升高,p-ASK1、p-p38和p-JNK含量明显降低(P<0.05)。与S组比较,BIC模型制备后3 d BIC组、TH组、THC组和THN组神经元凋亡细胞百分比明显升高(P<0.05)。与BIC组比较,BIC模型制备后3 d TH组、THC组和THN组凋亡细胞百分比明显降低(P<0.05)。与THC组比较,BIC模型制备后3 d TH组和THN组凋亡细胞百分比明显降低(P<0.05)。结论TH可以减轻BIC模型制备后神经元氧化应激反应以及减少神经元凋亡,沉默CIRP后该作用减弱,显示TH复苏神经元的机制可能与激活CIRP/Trx1/ASK1通路有关。
Objective To explore the effect of therapeutic hypothermia(TH)on neuronal oxidative injury,and its possible mechanism in a model of brain injury after cardiac arrest(BIC).Methods According to the random number table,60 male Spragne-Dawley rats,aged 7-10 weeks,weighing 280-320 g,were divided into shame group(group S),group BIC,group TH,TH after cold-inducible RNA-binding protein(CIRP)silent group(group THC),and TH after negative control group(group THN),12 in each group.The BIC model was established by cardiopulmonary resuscitation after 4 minutes of cardiac arrest induced by transesophageal electrical stimulation.The reactive oxygen species(ROS)by immunofluorescence staining,the expression of CIRP,thioredoxin1(Trx1),apoptosis-regulated kinase 1(ASK1)and its downstream p38 and c-Jun N-terminal kinase(JNK)phosphorylation levels by Western blot of 5 groups were tested 1 day after BIC.The apoptosis rate of hippocampal neurons was observed by TdT-nediated dUTP nick end labeling(TUNEL)staining 3 days after BIC.Results Compared with group S,ROS in group BIC increased,the expression of CIRP decreased,the expression of p-ASK1 and p-p38 increased;ROS in groups TH and THN increased,the expression of CIRP increased,the expression of p-JNK decreased;ROS of group THC increased,the expression of CIRP and Trx1 decreased,and the expression of p-ASK1 and p-p38 increased(P<0.05)1 day after BIC.Compared with group BIC,ROS in groups TH and THN decreased,the expression of CIRP and Trx1 increased,the expression of p-ASK1 and p-JNK decreased;the expression of p-p38 in group TH decreased;ROS of group THC decreased,and the expression of p-ASK1 and p-p38 increased(P<0.05)1 day after BIC.Compared with group THC,ROS in groups TH and THN decreased,the expression of CIRP and Trx1 increased,the expression of p-ASK1,p-p38 and p-JNK decreased(P<0.05)1 day after BIC.Compared with group S,the apoptosis rate of neurons in groups BIC,TH,THC and THN increased(P<0.05)3 days after BIC.Compared with group BIC,the apoptosis rate of neurons in groups TH,THC and THN decreased(P<0.05)3 days after BIC.Compared with group THC,the apoptosis rate of neurons in groups TH and THN decreased(P<0.05)3 days after BIC.Conclusion TH can alleviate neuronal oxidative injury and reduce neuronal apoptosis,while after silencing CIRP,the roles of TH are weakened.Therefore,the potential mechanism of TH resuscitating neurons may be related to the activation of CIRP/Trx1/ASK1 pathway.
作者
李娟
周洁洁
张洁
惠康丽
徐苗苗
段满林
徐建国
LI Juan;ZHOU Jiejie;ZHANG Jie;HUI Kangli;XU Miaomiao;DUAN Manlin;XU Jianguo(Department of Anesthesiology,Jinling Hospital,School of Medicine,Nanjing University,Nanjing 210002,China)
出处
《临床麻醉学杂志》
CAS
CSCD
北大核心
2021年第1期72-77,共6页
Journal of Clinical Anesthesiology
基金
国家自然科学基金(81671884)。
