超顺磁性聚乙烯亚胺纳米粒的制备及其骨肉瘤细胞、组织靶向转染效果观察  

Preparation of PEI-SPION-NPs and observation their targeted magnetofection effects in osteosarcoma cells and tissues in vivo

作　　者：岑超德 张永 罗聪 陈韬 杨晓兰[4] 曹永飞 刘承伟 CEN Chaode;ZHANG Yong;LUO Cong;CHEN Tao;YANG Xiaolan;CAO Yongfei;LIU Chengwei(Guizhou Provincial Orthopedics Hospital,Guiyang 550000,China;不详)

机构地区：[1]贵州省骨科医院,贵阳550000 [2]贵阳市第一人民医院 [3]重庆医科大学附属儿童医院 [4]重庆医科大学

出　　处：《山东医药》2021年第8期15-19,共5页Shandong Medical Journal

基　　金：重庆市技术创新与应用发展专项面上项目(cstc2019jscx-msxmX0121)。

摘　　要：目的制备超顺磁性聚乙烯亚胺纳米粒(PEI-SPION-NPs),观察其作为靶向基因输送载体转染骨肉瘤细胞和组织的可行性。方法通过脱水缩合反应将低分子量聚乙烯亚胺(PEI)修饰超顺磁性氧化铁纳米粒(SPION),制备PEI-SPION-NPs,对其分子结构、形态、粒径、饱和磁化强度、Zeta电位进行检测。将PEI-SPION-NPs与pDNA分别以不同比例混合,琼脂糖凝胶电泳观察PEI-SPION-NPs对pDNA的结合能力和保护作用。培养骨肉瘤MG-63细胞,分为实验1组(加入PEI-SPION-NPs/pDNA+磁场作用)、实验2组(加入PolyMag200/pDNA+磁场作用)、实验3组(加入PEI/pDNA)及对照组(加入pDNA),采用CCK-8法检测细胞存活率,倒置荧光显微镜观察转染情况,流式细胞仪测算转染率。制备裸鼠皮下骨肉瘤模型,分为Control组(皮下注射生理盐水)、PEI-SPION-NPs组(皮下注射PEI-SPI⁃ON-NPs/pGL3-Luc+磁场作用)、PEI-NPs组(皮下注射PEI/pGL3-Luc复合物+磁场作用),检测注射部位荧光强度,分别于造模第5、6、7、8、9周测量各组裸鼠体质量。结果PEI-SPION-NPs的粒径低于PEI-SPION-NPs/pDNA;PEI-SPI⁃ON-NPs的饱和磁化强度为(21.5±1.6)emu/g,Zeta电位为(31.2±1.5)mV;PEI-SPION-NPs可结合并保护pDNA,并可成功转染MG-63细胞。实验1组和对照组细胞存活率高于实验2组和实验3组,实验1组转染效率高于实验3组和对照组(P均<0.05)。PEI-SPION-NPs/pGL3-Luc与PEI-NPs/pGL3-Luc均可在裸鼠骨肉瘤组织中成功转染,PEI-SPI⁃ON-NPs组较PEI-NPs组荧光更为集中。造模第7周后PEI-SPION-NPs组裸鼠体质量高于PEI-NPs组(P均<0.01)。结论PEI-SPION-NPs是有效的非病毒基因载体,在磁场作用下可成功转染骨肉瘤细胞和组织。Objective To prepare the polyethylenimine-superparamagnetic iron oxide nanoparticles-nanoparticles(PEI-SPION-NPs)and to observe the feasibility of PEI-SPION-NPs as targeted gene delivery vector to transefect into osteosarcoma cells and tissues in vivo.Methods We used the polyethylenimine(PEI)-modified superparamagnetic iron oxide nanoparticles(SPION)to prepare the PEI-SPION-NPs by the dehydration condensation reaction.Their molecular structure,morphology,particle size,saturation magnetization and Zeta potential were detected.PEI-SPION-NPs were mixed with pDNA in different proportions,and agarose gel electrophoresis was performed to observe the binding ability and protective effect of PEI-SPION-NPs on pDNA.MG-63 cells were selected as target cells for transfection,which were cultured and then were divided into the group 1(PEI-SPION-NPs/pDNA+the static magnetic field),group 2(PolyMag200/pDNA+the static magnetic field),group 3(PEI-NPs/pDNA),and the control group(naked pDNA);the cell survival rate was determined by CCK-8 assay,the transfection effect was detected by inverted fluorescence microscope,and the transfection efficiency was measured by flow cytometry.The hypodermal osteosarcoma models in nude mice were prepared,which were then divided into the control group(subcutaneous injection of normal saline),PEI-SPION-NPs group(subcutaneous injection of PEI-SPION-NPs/pGL3-Luc+the static magnetic field),and PEI-NPs group(subcutaneous injection of PEI/pGL3-Luc complexes+the static magnetic field).Fluorescence intensity of injection site was detected and body weight of nude mice in each group was measured at 5,6,7,8 and 9 weeks after osteosarcoma modeling,respectively.Results The hydrodynamic diameters of PEI-SPION-NPs were shorter than those of PEI-SPION-NP/pDNA complexes,the saturation magnetization of PEI-SPION-NPs was(21.5±1.6)emu/g and the zeta potential was(31.2±1.5)mV;PEI-SPION-NPs could bind to and protect pDNA,and can be successfully transfected into MG-63 cells.The cell survival rates of the group 1 and contr

关 键 词：超顺磁性聚乙烯亚胺纳米粒 骨肉瘤 磁辅助转染技术 靶向基因治疗 

分 类 号：R738.1[医药卫生—肿瘤]