【作者】 魏述永； 童杰； 薛强； 尚芳红； 李燕君； 刘杨； 李许； 杨盛； 徐晓玉；

【Author】 WEI Shu-Yong;TONG Jie;XUE Qiang;SHANG Fang-Hong;LI Yan-Jun;LIU Yang;LI Xu;YANG Sheng;XU Xiao-Yu;Pharmaceutical College of Southwest University & College of Chinese Medicine,Chongqing Engineering Research Center for Pharmacodynamics Evaluation;Rongchang Campus of Southwest University;

【机构】 西南大学药学院中医药学院、重庆市药效评价工程技术中心； 西南大学荣昌校区；

【摘要】 目的:分析葛根素在共培养SD大鼠皮层神经元和星形胶质细胞中的分布差异。方法:原代分离培养神经元和星形胶质细胞,分别以MAP2、GFAP抗体结合DAPI染色进行免疫荧光鉴定并以Transwell膜为骨架建立两种细胞的共培养体系,建立受试细胞内葛根素含量的荧光分光光度分析方法,分别在单独培养及共培养的神经元与星形胶质细胞培养液内加入葛根素溶液至终浓度为1000 ng/mL,并进行药物加入后不同时间两种细胞内葛根素含量的差异分析。结果:免疫荧光鉴定两种细胞的纯度分别为神经元96.50±0.87、星形胶质细胞97.67±0.10,荧光分光光度法的专属性、精密度、回收率及线性关系良好,神经元内葛根素的含量高于星形胶质细胞,药物加入后20min,细胞内药物浓度达到峰值,加入药物后60min-90min,星形胶质细胞内未见葛根素检出,神经元样品内药物维持在12.8 ng/mL。结论:葛根素在神经元内比星形胶质细胞内具有更高的浓度及更长的保留时间。更多还原

【Abstract】 AIM:To investigate the distribution and retention time of puerarin in co-cultured cortical neurons and astrocytes in Sprague-Dawley（SD） rats.METHODS:Cortical neurons and astrocytes of new-born SD rats were separated and identified by immunofluorescence,and the cells were co-cultured in a system based on Transwell membrane for three days,then the distribution and retention time of puerarin in these two kinds of co-cultured and separately cultured cells were detected by fluorescence spectrophotometry and fluorescence microscope.RESULTS:Immunofluorescence identification indicated that the purity of cultured neurons and astrocytes were 96.50±0.87 and 97.67±0.10,respectively.The specificity,linearity,precision,and percentage recovery of the fluorescence spectrophotometric method were all feasible.The concentration of puerarin in both co-cultured and separately cultured neurons was higher than that of astrocytes,the peak time of puerarin in the tested cells was about 20 min,and after puerarin was added for 60 min,little drug was detected in astrocytes,but in both separately cultured and co-cultured neurons,the concentration of puerarin achieved a stable concentration of about 12.8ng·mL^-1.CONCLUTION:Higher concentration and longer retention time of puerarin were observed in neurons than in astrocytes.更多还原