基于TLR4/NF-κB/NLRP3通路探讨二精丸减轻AD大鼠神经炎症的作用及机制  

Effect of Erjing Pills on alleviating neuroinflammation of AD rats based on TLR4/NF-κB/NLRP3 pathway and its mechanism

作　　者：黄丽萍[1,5] 卢龙辉 杨喜洋 谢永艳 徐子薇 朱徐东 王晶晶 吴志鑫 唐健富 吴毅 陈耀辉 HUANG Li-ping;LU Long-hui;YANG Xi-yang;XIE Yong-yan;XU Zi-wei;ZHU Xu-dong;WANG Jing-jing;WU Zhi-xin;TANG Jian-fu;WU Yi;CHEN Yao-hui(School of Pharmacy,Jiangxi University of Chinese Medicine,Nanchang 330004,China;Jiangxi Provincial People's Hospital,Nanchang 330029,China;the First Hospital of Putian City,Putian 351199,China;NMPA Key Laboratory of Quality Evaluation of Traditional Chinese Patent Medicine,Jiangxi Provincial Engineering Research Center for Drug and Medical Device Quality,Jiangxi Institute for Drug Control,Nanchang 330046,China;Key Laboratory of TCM Pharmacology of Jiangxi Province,Nanchang 330004,China)

机构地区：[1]江西中医药大学药学院,江西南昌330004 [2]江西省人民医院,江西南昌330029 [3]莆田市第一医院,福建莆田351199 [4]国家药品监督管理局中成药质量评价重点实验室,江西省药品与医疗器械质量工程研究中心,江西省药品检验检测研究院,江西南昌330046 [5]江西省中药药理重点实验室,江西南昌330004

出　　处：《中国中药杂志》2023年第3期770-777,共8页China Journal of Chinese Materia Medica

基　　金：江西省主要学科学术和技术带头人项目(20182BCB22005);江西省自然科学基金项目(20171BAB205083);江西省重点研发计划项目(20181BBC70046,20192BBG70071,20202BBGL73006);江西省中医药管理局课题(2021A346);江西中医药大学校级科技创新团队发展计划项目(CXTD22007)。

摘　　要：研究二精丸对D-半乳糖联合β淀粉样蛋白_(25-35)(Aβ_(25-35))致阿尔茨海默病(AD)大鼠神经炎症的改善作用及可能机制。SD大鼠随机分为假手术组、模型对照组、阳性药物组(多奈哌齐,1 mg·kg^(-1))、二精丸高剂量组(9 g·kg^(-1))、二精丸低剂量组(4.5 g·kg^(-1)),每组14只。D-半乳糖腹腔注射2周后,连续灌胃给药5周;D-半乳糖腹腔注射3周后,大鼠双侧海马注射Aβ25-35。灌胃给药4周后,新物体识别实验检测大鼠学习记忆能力。末次给药24 h后,取组织,免疫荧光法检测大鼠脑组织小胶质细胞活化情况;免疫组化检测大鼠海马CA1区Aβ1-42和磷酸化Tau404蛋白(phosphory protein Tau^(404),p-Tau404)阳性表达;酶联免疫吸附(enzyme linked immunosorbent assay,ELISA)法检测大鼠脑组织炎症因子白细胞介素1β(interleukin-1β,IL-1β)、肿瘤坏死因子α(tumor necrosis factor,TNF-α)、白细胞介素6(IL-6)水平;Western blot法检测大鼠脑组织Toll样受体4(Toll-like receptor,TLR4)/核因子κB(nuclear factor kappaB,NF-κB)/核苷酸结合寡聚化结构域样受体3(nucleotide-binding oligomerization domain-like receptors 3,NLRP3)通路相关蛋白表达。结果显示,与假手术组比较,模型对照组大鼠新物体识别指数明显下降,海马区Aβ_(1-42)沉积和p-Tau^(404)阳性蛋白显著增多,脑组织齿状回处小胶质细胞活化水平显著增高,海马组织IL-1β、TNF-α、IL-6水平明显升高;海马TLR4、p-NF-κB P65/NF-κB P65、p-IκBα/IκBα、NLRP3蛋白表达水平显著升高。与模型对照组比较,二精丸可提高大鼠新物体识别指数,减少海马区Aβ_(1-42)沉积和p-Tau^(404)阳性蛋白表达,抑制脑组织齿状回小胶质细胞活化;降低海马组织IL-1β、TNF-α、IL-6炎症因子水平;下调TLR4、p-NF-κB P65/NF-κB P65、p-IκBα/IκBα、NLRP3蛋白的表达。综上所述,二精丸可能通过改善小胶质细胞活化,降低神经炎症因子IL-1β、TNF-α、IL-6表达水平,抑制TLR4/NF-κB/NLRP3This paper aimed to study the effect of Erjing Pills on the improvement of neuroinflammation of rats with Alzheimer′s di-sease(AD) induced by the combination of D-galactose and Aβ25-35and its mechanism. SD rats were randomly divided into a sham group, a model control group, a positive drug group(donepezil, 1 mg·kg^(-1)), an Erjing Pills high-dose group(9.0 g·kg^(-1)), and an Erjing Pills low-dose group(4.5 g·kg^(-1)), with 14 rats each group. To establish the rat model of AD, Erjing Pills were intragastrically administrated to rats for 5 weeks after 2 weeks of D-galactose injection. D-galactose was intraperitoneally injected into rats for 3 weeks, and then Aβ25-35was injected into the bilateral hippocampus. The new object recognition test was used to evaluate the learning and memory ability of rats after 4 weeks of intragastric administration. Tissues were acquired 24 h after the last administration. The immunofluorescence method was used to detect the activation of microglia in the brain tissue of rats. The positive expressions of Aβ1-42and phosphory protein Tau404(p-Tau404) in the CA1 area of the hippocampus were detected by immunohistochemistry. The levels of inflammatory factors interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), and interleukin-6(IL-6) in the brain tissue were determined by enzyme-linked immunosorbent assay(ELISA). Toll-like receptor 4(TLR4)/nuclear factor kappa B(NF-κB)/nucleotide-binding oligomerization domain-like receptors 3(NLRP3) pathway-associated proteins in the brain tissue were determined by Western blot. The results showed that as compared with the sham group, the new object recognition index of rats in the model control group decreased significantly, the deposition of Aβ_(1-42)and p-Tau^(404) positive protein in the hippocampus increased significantly, and the levels of microglia activation increased significantly in the dentate gyrus. The levels of IL-1β, TNF-α, and IL-6 in the hippocampus of the model control group increased significantly, and the expression

关 键 词：二精丸 神经炎症 阿尔茨海默病 TLR4/NF-κB/NLRP3通路 小胶质细胞 

分 类 号：R285.5[医药卫生—中药学]