脑泰方对脑小血管病模型大鼠脑白质病变及胼胝体Wnt/β-catenin通路的影响  

Effects of Naotai Formula(脑泰方)on Cerebral White Matter Lesions and Wnt/β-Catenin Pathway in Corpus Callosum of Rats with Cerebral Small Vessel Disease Model

作　　者：王香元 梅志刚 葛金文[1,3] WANG Xiangyuan;MEI Zhigang;Ge Jinwen(College of Integrated Chinese and Western Medicine,Hunan University of Chinese Medicine,Changsha,410208;College of Medical and Health Science,China Three Gorges University;Hunan Scientific Academy of Chinese Medicine)

机构地区：[1]湖南中医药大学中西医结合学院,湖南省长沙市410208 [2]三峡大学健康医学院 [3]湖南省中医药研究院

出　　处：《中医杂志》2024年第20期2135-2144,共10页Journal of Traditional Chinese Medicine

基　　金：国家重点研发计划中医药现代化研究重点专项(2018YFC1704904);国家中医药管理局青年岐黄学者培养项目。

摘　　要：目的从无翅型MMTV整合位点家族成员(Wnt)/β连环蛋白(β-catenin)通路探讨脑泰方对脑小血管病白质病变的作用及其机制。方法将10只WKY大鼠作为假手术组,30只自发性高血压大鼠(SHR)随机分为模型组、脑泰方低剂量组、脑泰方高剂量组,每组10只。除假手术组外,其余各组大鼠采用D-半乳糖背部皮下注射56天造模,并在造模第29天进行双侧颈总动脉狭窄术低灌注28天构建复合危险因素脑小血管病大鼠模型。脑泰方低、高剂量组大鼠手术当天分别给予脑泰方9、27 g/(kg·d)灌胃,假手术组和模型组给予10 ml/(kg·d)纯水灌胃,各组大鼠灌胃4周。给药前及给药后每周监测大鼠的血压;造模第50天开始连续6天进行水迷宫实验;术前及术后、给药后激光散斑检测脑血流量变化。最后一次灌胃次日取材,卢卡斯快蓝(LFB)和透射电镜观察大鼠胼胝体髓鞘损伤程度,ELISA法检测大鼠胼胝体白细胞介素1β(IL-1β)、肿瘤坏死因子α(TNF-α)、白细胞介素10(IL-10)、肿瘤坏死因子β(TNF-β)水平,免疫荧光法检测大鼠胼胝体中髓鞘碱性蛋白(MBP)表达水平和离子化钙结合适配分子1(IBA1)、神经胶质抗原2(NG2)、细胞增殖标志物Ki67抗原(Ki67)/NG2、2',3'-环核苷酸3'-磷酸二酯酶(CNPase)表达;Western Blot法检测胼胝体中MBP、无翅型MMTV整合位点家族成员3a(Wnt3a)、磷酸化糖原合成酶激酶3β(p-GSK-3β)、糖原合成酶激酶3β(GSK-3β)、磷酸化β连环蛋白(p-β-catenin)、β-catenin表达,RT-qPCR法检测胼胝体中GSK-3β、β-catenin mRNA表达。结果与假手术组比较,模型组大鼠各时间点收缩压升高,水迷宫实验中穿越平台次数、目标象限停留时间百分比减少,第4天和第5天逃避潜伏期明显延长,术后及给药后大鼠脑血流量下降(P<0.05或P<0.01);LFB染色及透射电镜观察可见胼胝体髓鞘损伤明显;胼胝体中TNF-α和IL-1β含量增加,TNF-β含量降低,MBP荧光强度和蛋白表�Objective To explore the effects and mechanisms of Naotai Formula(脑泰方)against white matter lesions associated with cerebrovascular disease via Wnt/β-catenin pathway.Methods Ten WKY rats were used as sham surgery group,and 30 spontaneously hypertensive rats(SHRs)were randomly divided into model group,lowand high-dose Naotai Formula group,with 10 rats in each group.In addition to the sham surgery group,rats in each group were modelled by dorsum subcutaneous injection of D-galactose for 56 days,by performing bilateral common carotid artery stenosis on the 29th day of modelling with low perfusion for 28 days to construct the rat models of cere⁃bral small-vessel disease with composite risk factors.The rats in the low-and high-dose Naotai Formula groups were given 9 and 27 g(/kg·d)of Naotai Formula by gavage on the day after surgery,and 10 ml(/kg·d)of pure water by ga⁃vage in the sham surgery group and the model group,with all rats in each group gavaged for 4 weeks.Blood pressure of rats was monitored weekly before and after medication administration;water maze experiments were performed for 6 consecutive days from the 50th day of modelling;and changes in cerebral blood flow were detected by laser scattering preoperatively,postoperatively,and after medication administration.The samples were taken on the next day after the last gavage,and the extent of myelin damage in the rat corpus callosum was observed by Lucas fast blue(LFB)and transmission electron microscopy,and the levels of interleukin 1β(IL-1β),tumour necrosis factorα(TNF-α),interleukin 10(IL-10),and tumour necrosis factorβ(TNF-β)were detected by ELISA,and the levels of expression of myelin basic protein(MBP)and ionized calcium binding adaptor molecule 1(IBA1),neuroglial antigen 2(NG2),cell proliferation marker Ki67 antigen(MBP),and cell proliferation marker Ki67 antigen(Ki67A)/NG2,2',3'-cyclic-nucleotide 3'-phosphodiesterase(CNPase)expression were measured by immunofluorescence.MBP in the corpus callosum,wingless MMTV integration site family

关 键 词：脑小血管病 脑白质病变 髓鞘损伤 脑泰方 少突胶质细胞 WNT/Β-CATENIN通路 

分 类 号：R285.5[医药卫生—中药学]