基于NLRP3/ASC/Caspase-1细胞焦亡通路探讨补肾启智方改善缺血性脑卒中大鼠认知功能的作用机制  

Action Mechanism of Bushen Qizhi Formula in Improving Cognitive Function in Ischemic Stroke Rats Based on NLRP3/ASC/Caspase-1 Pyroptosis Pathway

作　　者：巩俐 刘雪梅[2] 罗超琴 王翎沣 马承平 张艺馨 卢佳慧 傅晨[2] GONG Li;LIU Xuemei;LUO Chaoqin;WANG Lingfeng;MA Chengping;ZHANG Yixin;LU Jiahui;FU Chen(Beijing University of Chinese Medicine,Beijing China 100029;Dongfang Hospital Affiliated to Beijing University of Chinese Medicine,Beijing China 100078)

机构地区：[1]北京中医药大学,北京100029 [2]北京中医药大学东方医院,北京100078

出　　处：《中医学报》2025年第3期608-616,共9页Acta Chinese Medicine

基　　金：国家自然科学基金青年科学基金项目(82104808);2024年度北京中医药大学研究生自主科研课题项目(ZJKT2024054)。

摘　　要：目的:探讨补肾启智方通过调控核苷酸结合寡聚结构域样受体蛋白3(NOD-like receptor protein3,NLRP3)/凋亡相关斑点样蛋白(apoptosis-associated speck-like protein containing CARD,ASC)/半胱天冬酶-1(Caspase-1)信号通路,介导海马神经元焦亡,改善大脑中动脉闭塞(middle cerebral artery occlusion,MCAO)大鼠认知障碍的作用机制。方法:将40只Wistar大鼠随机分为sham组、MCAO组、补肾启智方组(BSQZ,14.4 g·kg^(-1))、焦亡抑制剂组(Ac-YVAD-CMK,5 mg·kg^(-1)),每组10只。除sham组外,其余大鼠采用改良Longa线栓法建立MCAO模型。采用Morris水迷宫实验和新物体识别(novel object recognition,NOR)实验评估大鼠的学习与记忆功能;ELISA法检测大鼠海马组织中白细胞介素-1β(interleukin-1β,IL-1β)和白细胞介素-18(interleukin-18,IL-18)含量;免疫荧光法检测大鼠海马组织中消皮素D-N(gasdermin D-N,GSDMD-N)阳性细胞数目;Western blot法检测大鼠海马组织中GSDMD-N、NLRP3、ASC、活化半胱天冬酶-1(c-Caspase-1)表达水平。结果:与sham组比较,MCAO组、BSQZ组、Ac-YVAD-CMK组大鼠Zea Longa评分升高(P<0.01),Garcia JH评分降低(P<0.01)。Morris水迷宫实验显示,在定位航行实验的第3、4、5天,与sham组比较,MCAO组大鼠的逃逸潜伏期延长(P<0.01);与MCAO组比较,BSQZ组、Ac-YVAD-CMK组大鼠的逃逸潜伏期缩短(P<0.05)。与sham组比较,MCAO组大鼠在目标象限的停留时间、穿越平台次数减少(P<0.01);与MCAO组比较,BSQZ组、Ac-YVAD-CMK组大鼠在目标象限的停留时间、穿越平台次数增加(P<0.05)。NOR实验显示,与sham组比较,MCAO组大鼠的相对识别指数下降(P<0.01);与MCAO组比较,BSQZ组、Ac-YVAD-CMK组大鼠的相对识别指数升高(P<0.01)。与sham组比较,MCAO组大鼠海马组织中IL-1β、IL-18含量及GSDMD-N阳性细胞数目增加(P<0.01);与MCAO组比较,BSQZ组、Ac-YVAD-CMK组大鼠海马组织中IL-1β、IL-18含量及GSDMD-N阳性细胞数目减少(P<0.01)。Western blot结果显�Objective:To investigate the effects of Bushen Qizhi Formula in mediating hippocampal neuron pyroptosis and ameliorating the mechanism of action of cognitive impairment in middle cerebral artery occlusion(MCAO)rats by modulating nucleotide-binding oligomerization structural domain-like receptor protein 3(NLRP3)/apoptosis-associated speck-like protein containing CARD(ASC)/cysteine aspartate protein(CARD)/Caspase-1 signaling pathway.Methods:Forty Wistar rats were randomly divided into sham group,MCAO group,Bushen Qizhi Formula group(BSQZ,14.4 g·kg^(-1)),and focal death inhibitor group(Ac-YVAD-CMK,5 mg·kg^(-1)),with 10 rats in each group.Except for the sham group,the MCAO model was established in the remaining rats by using the modified Longa wire bolus method.The learning and memory functions of rats were assessed by Morris water maze experiment and novel object recognition(NOR)experiment;interleukin-1β(IL-1β)and interleukin-18(IL-18)levels were detected in the hippocampal tissue of rats by ELISA.18(IL-18)in rat hippocampal tissues;the number of gasdermin D-N(GSDMD-N)positive cells in rat hippocampal tissues was detected by immunofluorescence;and the expression levels of GSDMD-N,NLRP3,ASC,and activated cysteine protease-1(c-Caspase-1)were detected in rat hippocampal tissues by Western blot.Results:Compared with the sham group,Zea Longa scores of rats in the MCAO group,BSQZ group,and Ac-YVAD-CMK group were elevated(P<0.01),and Garcia JH scores were decreased(P<0.01).The Morris water maze experiments showed that,on the 3rd,4th,and 5th days of the localization and navigation experiments,compared with the sham group,the escape of rats in the MCAO group latency was prolonged(P<0.01);compared with the MCAO group,the escape latency of rats in the BSQZ group and Ac-YVAD-CMK group was shortened(P<0.05).Compared with the sham group,the residence time in the target quadrant and the number of traversing platforms of rats in the MCAO group decreased(P<0.01);compared with the MCAO group,the residence time in the target quadr

关 键 词：补肾启智方 缺血性脑卒中 NLRP3/ASC/Caspase-1信号通路 细胞焦亡 卒中后认知障碍 

分 类 号：R285.5[医药卫生—中药学]