黄芪糖蛋白对EAE小鼠的神经保护作用及其修复机制分析  

Neuroprotective Effect and Repair Mechanism of Huangqi Glycoprotein on Mice with Experimental Autoimmune Encephalomyelitis

作　　者：邢雁霞[1] 刘斌钰[1] 赵一锦 张丽红 张光先 薛慧清 马存根[1] 章培军[1] XING Yan-xia;LIU Bin-yu;ZHAO Yi-jin;ZHANG Li-hong;ZHANG Guang-xian;XUE Hui-qing;MA Cun-gen;ZHANG Pei-jun(Institute of Brain Science,Datong University,Datong 037009,China;Medical College,Thomas Jefferson University,Philadelphia 19107,USA;"2011"Collaborative Innovation Center,Research Center of Neurobiology,Shanxi University of Chinese Medicine,Taiyuan 030619,China)

机构地区：[1]大同大学脑科学研究所,山西大同037009 [2]托马斯·杰斐逊大学医学院,宾夕法尼亚州费城19107 [3]山西中医药大学“2011”协同创新中心,神经生物学研究中心,太原030619

出　　处：《中国实验方剂学杂志》2018年第24期7-13,共7页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家国际科技合作专项(2013DFA30700);国家自然科学基金项目(81473577);山西省自然科学基金项目(2013011052-4);大同市应用基础研究计划项目(2017135)

摘　　要：目的:观察黄芪糖蛋白(HQGP)对实验性自身免疫性脑脊髓炎(EAE)小鼠的保护作用,并探讨其神经修复机制。方法:复制髓鞘少突胶质细胞糖蛋白35-55(MOG35-55)多肽诱导的小鼠EAE模型,20只C57BL/6雌性小鼠随机分为EAE组和HQGP组。HQGP组于免疫后第3日起,按1 mg·kg-1·d-1腹腔注射HQGP,持续给药18 d。通过五级临床症状评分和小鼠体质量变化观察HQGP对EAE小鼠的干预效应;取小鼠脊髓进行苏木精-伊红(HE)和固蓝(LFB)染色观察HQGP的神经保护作用;免疫组织化学法检测小鼠脊髓白细胞分化抗原(CD)68+巨噬细胞的浸润以及诱导型一氧化氮合酶(i NOS),微管相关蛋白2(MAP-2)和神经元核抗原(Neu N)的表达;免疫荧光组织化学双染法检测小鼠脊髓CD11b+细胞和趋化因子配体-5(CCL-5)的表达;酶联免疫吸附测定法(ELISA)检测小鼠脾脏单个核细胞(MNC)培养上清液中相关细胞因子的变化。结果:HQGP有效缓解EAE小鼠的临床症状,推迟发病时间,减轻中枢神经系统的炎性反应和髓鞘脱失;HQGP能减少小鼠脊髓CD68+巨噬细胞和CD11b+细胞的数量,抑制脊髓i NOS和趋化因子CCL-5的表达,增加MAP-2和Neu N的表达水平;HQGP能明显降低EAE小鼠肿瘤坏死因子-α(TNF-α)和白细胞介素(IL)-6的分泌,促进IL-10和γ-干扰素(IFN-γ)的分泌。结论:HQGP具有治疗EAE的潜在作用,其神经保护机制与抑制炎症反应、调控免疫细胞表达及细胞因子的分泌、减轻髓鞘脱失并促进轴突修复和神经元发育有关。Objective:To observe the protection of Huangqi glycoprotein (HQGP)on mice with experimental autoimmune encephalomyelitis (EAE)and further explore its mechanism of nerve repair.Method:Twenty female C57BL/6mice were immunized subcutaneously with myelin oligodendrocyte glyeoprotein 35-55 (MOG35_55)polypeptide and randomly divided into EAE group and HQGP group.Mice in HQGP group were received intraperitoneal injection of HQGP (1mg·kg^-1·d^-1)on day 3post-immunization with continuous administration for 18days.To observe the effect of HQGP in EAE,clinical assessment of EAE scores was evaluated and the body weight of mice was recorded every day.Spinal cords of mice were obtained for hematoxylin- eosin (HE)and Luxol fast blue (LFB)staining to observe the neuroprotective effect of HQGP.The infiltration of cluster of differentiation 68^+(CD68^+)maerophages and the expression of inducible nitric oxide synthase (iNOS),mierotubule-assoeiated protein 2(MAP-2)and neuronal nuelei (NeuN)in spinal cord of mice were detected by immunohistoehemistry.Double immunofluoreseenee histochemistry was used to detect the expression of CDllb^+ cells and ehemokine ligand-5(CCL-5)in spinal cord of mice.The levels of eytokines in the supernatant of cultured splenic suspension of mononuclear cells (MNCs)were determined by enzyme-linked immunosorbent assay (ELISA).Result:HQGP delayed onset and improved the clinical symptoms of EAE,accompanied by inhibiting inflammation and alleviating demyelination in the spinal cord.Compared with EAE group,HQGP reduced the infiltration of CD68+maerophages and CD1lb^+ cells,inhibited the expression of iNOS and CCL-5 and increased the expression of MAP-2and NeuN in the spinal cord.Further studies showed that HQGP could reduce the production of tumor necrosis factor (TNF)-α and interleukin (IL)-6,increase the level of IL-10 and y- interferon (IFN-y).Conclusion:HQGP has the potential role for the treatment of EAE and the mechanism is related to anti-inflammation,regulation of immune cell expression and cytokine se

关 键 词：黄芪糖蛋白 实验性自身免疫性脑脊髓炎 免疫细胞 细胞因子 诱导型一氧化氮合酶 微管相关蛋白2 神经元核抗原 

分 类 号：R22[医药卫生—中医基础理论] R24[医药卫生—中医学] R28