低分子质量壳聚糖修饰多壁碳纳米管的制备及细胞毒性研究

Preparation and cytotoxicity study of multi-wailed carbon nanotubes modified with low-molecular-weight chitosan

目的 利用低分子质量壳聚糖（CS）制备可在水溶液中稳定分散的壳聚糖/多壁碳纳米管（CS/MWCNTs）材料,并观察其与人脐静脉内皮细胞（HUVEC）的相互作用.方法 以物理吸附法对MWCNTs进行CS修饰,利用透射电镜、纳米粒度及Zeta电位分析仪对其进行表征.将CS/MWCNT进行荧光标记,以不同浓度与细胞作用24 h,通过激光共聚焦显微镜观察细胞摄入情况,并检测细胞毒性及细胞内活性氧自由基含量.结果 当低分子质量CS与MWCNTs的质量比大于10∶1时,可很好地将MWCNTs进行分散,CS/MWCNTs可在水相中稳定存在.细胞摄入实验显示,进入细胞内的碳纳米管主要位于胞浆内.毒性检测结果显示,在较高质量浓度（10、20 μg/ml）时,CS分散后的MWCNTs毒性较小.而与2种碳纳米管（MWCNTs与CS/MWCNTs）作用的细胞内的活性氧含量均随着浓度升高而显著提高,差别无统计学意义（P＞0.05）.结论 水溶性的CS/MWCNTs材料拥有极好的分散性,性状稳定,细胞毒性低,这对后期将其应用于以MWCNTs为载体的治疗研究具有重要意义.

Objective To prepare stable aqueous dispersions of chitosan/multi-walled carbon nanotubes （CS/MWCNTs） composites,and observe the effects of CS/MWCNTs on the growth of human umbilical vein endothelial cells （HUVEC）.Methods CS/MWCNTs composites were prepared by electrostatic interactions between negatively charged MWCNTs and positively charged low-molecular-weight CS.The prepared CS/MWCNTs were characterized by transmission electron microscopy and Zetasizer nano-analyser.The cellular uptake of the fluorescently labeled CS/MWCNTs was observed by laser confocal microscopy after incubating with HUVEC for 24 h at different concentrations.In vitro cytotoxicity and cellular reactive oxygen were also detected.Results When the mass ratio of low-molecular-weight CS to MWCNTs was equal or greater than 10∶1,the CS/MWCNTs can be stabilized in solution.Cellular uptake experiments showed that the CS/MWCNTs could enter into the cells and locate mainly in the cytoplasm.Cytotoxicity study showed that the CS/MWCNTs composites was less toxic than MWCNTs alone at high concentration （10 and 20 μg/ml）.However,there was no significant differencein the level of cellular reactive oxygen between the two groups （P〈0.05）.Conclusions CS/MWCNTs composites showed low cytotoxicity and high stability,which would be a promising carrier for drug delivery.