针刺对惊厥性脑损伤大鼠海马神经元Caspase-12蛋白及内质网Ca^(2+)表达的影响  

Effect of acupuncture on expression of endoplasmic reticulum Ca2+and Caspase-12 protein in hippocampal neurons of rats with convulsive brain injury

作　　者：张慧[1] 马允 吴旭[3] 刘振邦 吴生兵 昂文平[1] 杨帆[1] ZHANG Hui;MA Yun;WU Xu;LIU Zhen-bang;WU Sheng-bing;ANG Wen-ping;YANG Fan(College of Acupuncture-moxibustion and Massage,Anhui University of Traditional Chinese Medicine,Hefei 230038,China;Wuhu Hospital of Traditional Chinese Medicine,Wuhu 241000,Anhui Province;College of Life Sciences,University of Science and Technology of China,Hefei 230012;Scientific Research and Experiment Center,Anhui University of Traditional Chinese Medicine,Hefei 230038,China)

机构地区：[1]安徽中医药大学针灸推拿学院,合肥230038 [2]芜湖市中医院,安徽芜湖241000 [3]中国科技大学生命科学学院,合肥230012 [4]安徽中医药大学科研实验中心,合肥230038

出　　处：《针刺研究》2022年第8期673-677,683,共6页Acupuncture Research

基　　金：国家自然科学基金项目(No.81072872);安徽高校自然科学研究项目(No.KJ2016A858)。

摘　　要：目的:观察针刺对惊厥性脑损伤大鼠不同时点海马神经元内质网Ca^(2+)表达、细胞凋亡和Caspase-12蛋白表达的影响,探讨针刺维持内质网钙稳态及抑制细胞凋亡级联反应的可能作用机制。方法:SD大鼠随机分为正常组、模型组、针刺组,每组36只。采用腹腔一次性注射惊厥剂量(50 mg/kg)戊四唑造成大鼠惊厥。针刺组于惊厥后即刻针刺“百会”“大椎”30 min。各组分别于惊厥发作后2、12、48 h采用TUNEL法检测海马神经元细胞凋亡情况;采用内质网Ca^(2+)荧光探针技术检测大鼠海马神经元内质网Ca^(2+)表达;采用免疫组织化学法观察各组大鼠海马组织中Caspase-12蛋白表达。结果:与正常组比较,模型组2、12、48 h海马神经元凋亡细胞数明显增加(P<0.01),海马神经元内质网Ca^(2+)表达降低(P<0.01),Caspase-12蛋白表达升高(P<0.01);与模型组比较,针刺组在惊厥发作2、12、48 h海马神经元凋亡减轻,凋亡细胞数明显减少(P<0.05,P<0.01),海马神经元内质网Ca^(2+)表达升高(P<0.05,P<0.01),Caspase-12蛋白表达降低(P<0.05,P<0.01)。结论:针刺有利于海马神经元内质网Ca^(2+)表达的恢复,从而维持内质网内钙稳态,避免诱发内质网应激介导的Caspase-12凋亡级联反应,减少细胞凋亡,从而治疗惊厥性脑损伤。Objective To observe the effect of acupuncture on endoplasmic reticulum calcium,apoptosis number and Caspase-12 protein expression in hippocampal neurons of convulsive rats,so as to explore its mechanisms underlying improvement of convulsion.Methods SD rats were randomly divided into normal control,model and acupuncture groups,with 36 rats in each group.Rats in the normal control group received intraperitoneal injection(i.p.)of normal saline(2 mL),and those of the other 2 groups received i.p.of pentylenetetrazole(50 mg/kg)for establishing convulsion model.Manual acupuncture stimulation was applied to“Baihui”(GV20)and“Dazhui”(GV14)for 30 min after modeling.The hippocampal tissues were taken at 2,12 and 48 h after modeling.The endoplasmic reticulum Ca^(2+) concentration(optical density,OD)was detected by using fluorescence probe technique and laser confocal microscopy,and the number of apoptosis of hippocampal neurons at the 3 time points detected by using terminal-deoxynucleoitidyl transferase mediated nick end labeling(TUNEL)stain.The expression of Caspase-12 protein in hippocampus at 3 time points was observed by immunohistochemistry.Results In comparison with the normal group,the number of apoptotic cells of hippocampal neurons and the expression levels of Caspase-12 protein in hippocampus at 2,12 and 48 h after seizures were obviously increased(P<0.01),and the OD value of Ca^(2+) at 3 time points significantly decreased(P<0.01)in the model group.Following acupuncture intervention,the increased levels of the number of apoptotic cells of hippocampal neurons and the expression of Caspase-12 protein in hippocampus at 3 time points and the decreased levels of OD value of Ca^(2+) at 3 time points were reversed in the acupuncture group(P<0.05,P<0.01).Conclusion Acupuncture intervention is effective in reducing the apoptosis of hippocampal neurons in convulsion rats,which may be related to its functions in down-regulating Caspase-12 expression and promoting influx of Ca^(2+) in the hippocampal neurons.

关 键 词：针刺 惊厥 CA^(2+) 神经元凋亡 CASPASE-12 内质网应激 

分 类 号：R245.311[医药卫生—针灸推拿学]