咔唑类生物碱衍生物CZ-7促进少突胶质前体细胞分化的研究  

Carbazole Alkaloid Derivative CZ-7 Contributes to the Differentiation of Oligodendrocyte Progenitor Cells

作　　者：王莎莎 张钊 毕皓植 楚世峰 贺文彬 臧应达 张东明 陈乃宏 WANG Shasha;ZHANG Zhao;BI Haozhi;CHU Shifeng;HE Wenbin;ZANG Yingda;ZHANG Dongming;CHEN Naihong(School of Basic Medicine,Shanxi University of Traditional Chinese Medicine,Taiyuan 030619 Shanxi,China;State Key Laboratory of Bioactive Substances and Functions of Natural Medicines,Institute of Materia Medica&Neuroscience Center,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100050,China)

机构地区：[1]山西中医药大学中药与食品工程学院,山西太原030619 [2]中国医学科学院药物研究所神经科学中心天然药物活性物质与功能国家重点实验室,北京100050

出　　处：《中药新药与临床药理》2020年第7期781-787,共7页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：中国医学科学院医学与健康科技创新工程经费资助项目(2016-I2M-1-004);中医脑病学山西省重点实验室开放课题基金资助项目(CME-OP-2017001);山西省重点研发计划(重大区域创新合作)项目(201803D421006)。

摘　　要：目的研究黄皮茎枝提取物咔唑类生物碱衍生物CZ-7对少突胶质前体细胞(OPCs)分化的影响。方法通过体外分离培养新生SD大鼠少突胶质前体细胞,并对分离的少突胶质前体细胞进行纯化;纯化后加入不同浓度CZ-7(0.1、1、10μmol·L^-1)进行干预。在光学显微镜下连续观察细胞形态变化;采用免疫荧光染色法鉴定细胞;CCK-8法检测CZ-7对少突胶质前体细胞细胞活力的影响;免疫荧光染色法检测CZ-7对少突胶质前体细胞分化成熟的影响;Western Blot法检测CZ-7对髓鞘碱性蛋白(MBP)表达的影响;qPCR法检测CZ-7对髓鞘相关基因MBP、PLP1表达的影响。结果分离纯化后获得了纯度较高的少突胶质前体细胞,继续培养至第7天时可自然分化成熟为少突胶质细胞。与对照组比较,CZ-7(0.1、1、10μmol·L^-1)不影响少突胶质前体细胞细胞活力(P>0.05)。少突胶质前体细胞纯化后加入CZ-7共孵育48 h,与对照组比较,CZ-7不同浓度组的少突胶质前体细胞(NG2阳性表达细胞)数明显减少(P<0.01),成熟少突胶质细胞(CC-1阳性表达细胞)数明显增多(P<0.01),成熟少突胶质细胞占总少突胶质细胞的百分比也明显升高(P<0.01);MBP蛋白表达量明显上调(P<0.05,P<0.01);髓鞘相关基因MBP、PLP1 mRNA表达水平明显上调(P<0.05,P<0.01)。结论CZ-7能够明显促进SD大鼠少突胶质前体细胞分化成熟,从而有利于髓鞘再生。Objective To study the effect of carbazole alkaloid derivative CZ-7 on the differentiation of rat oligodendrocyte progenitor cells(OPCs).Methods Primary SD rat OPCs were cultured and purified,and treated by different concentrations of CZ-7(0.1μmol·L^-1,1μmol·L^-1 and 10μmol·L^-1)three days after.Morphology of cells were continuously observed by optical microscope and identified by immunofluorescence.The cell viability of CZ-7-treated-OPCs was detected by CCK-8 assay.The effect of CZ-7 on the differentiation of OPCs was detected by immunofluorescence.The expression of MBP protein in OPCs after CZ-7 treatment was detected by Western Blot.The effect of CZ-7 on mRNA expression of genes related to oligodendrocyte maturity and myelination was detected by qPCR.Results High purity OPCs were obtained after isolation and culture.After 7 days of culture,the cells could differentiate naturally.CZ-7 treatment at doses of 0.1,1,and 10μmol·L^-1 did not affect the cell viability of OPCs(P>0.05).After 48 hours of CZ-7 treatment,the proportions of OPCs were significantly decreased(P<0.01),the proportions of OLs were significantly augmented(P<0.01)and the proportions of mature OLs/total OLs were significantly increased in CZ-7 groups(P<0.01).The expression levels of MBP protein were significantly increased(P<0.05,P<0.01).The expression levels of genes related to oligodendrocyte maturity and myelination were significantly increased(P<0.05,P<0.01).Conclusion CZ-7 facilitates the differentiation and maturation of OPCs,which may promote remyelination.

关 键 词：黄皮咔唑类生物碱衍生物CZ-7 少突胶质前体细胞 少突胶质细胞 分化 髓鞘再生 大鼠 

分 类 号：R285.5[医药卫生—中药学]