甲氨蝶呤对体外培养神经干细胞的毒性作用.pdfVIP

292 Tianjin Med J, Apr 2009 , Vol 37 No 4 甲氨蝶呤对体外培养神经干细胞的毒性作用* △ 张绪梅 黄国伟 田志红 任大林 观察甲氨蝶呤（MTX）对体外培养的神经干细胞（NSC）毒性作用的特点，并初步探讨其毒性机制。 摘要 目的： 方 体外培养胎鼠大脑皮质NSCs ，采用Nestin 免疫荧光法进行鉴定；光镜下观察MTX 干预后NSC 形态变化；采用5- 法： 溴脱氧尿苷（BrdU）掺入法和Nestin/BrdU 免疫组化双标记检测MTX 对NSCs 增殖的影响；流式细胞术分析细胞周期 变化；高压液相法检测细胞内同型半胱氨酸（Hcy）的浓度。 400 μg/L MTX 可引起明显的细胞形态改变；免疫荧光 结果： 双标记结果显示，与对照组相比，MTX 干预组可明显抑制NSCs 增殖（P 0.05）；MTX 干预48 h 后，细胞停留在G1 期 的比例比空白对照组明显增加；MTX 还可引起细胞内Hcy 浓度升高。 MTX 可能通过改变细胞周期和升高细胞 结论： 内Hcy 浓度而对体外培养NSCs 产生明显的毒性损伤作用。 关键词 甲氨蝶呤 干细胞 神经系统 细胞周期 半胱氨酸 Toxic Effects of Methotrexate on Primary Cultured Neural Stem Cells of Fetal Rats in Vitro ZHANG Xumei, HUANG Guowei, TIAN Zhihong, REN Dalin Department of Nutrition and Food Hygiene, School of Public Health,Tianjin Medical University, Tianjin 300070, China Abstract Objective: To study the toxic effects induced by methotrexate on primary cultured neural stem cells from fetal rats in vitro, and explore the possible mechanism for toxic effects. Methods: NSCs were identified by immunofluorescent stain- ing using antibody against Nestin. Morphological observation were used to evaluate the toxicity of methotrexate on NSCs by light microscope. BrdU incorporating method and Nestin/BrdU double-label immunofluorescence technique were used to de - tect NSCs proliferation under the treatment of methotrexate. The process of cell cycle was analyzed by flow cytometry. High per - formance