载阿霉素新型改性壳聚糖纳米微粒控释载体对骨肉瘤细胞的影响  

Effect of a doxorubicin-loaded modified chitosan sustain-release nanoparticle carrier on osteosarcoma cells

作　　者：金文意 徐瑞[2] 高明勇[1] 刘子林 常春雨[2] Jin Wenyi;Xu Rui;Gao Mingyong;Liu Zilin;Chang Chunyu(Department of Orthopeadics,Wuhan University Renmin Hospital,Wuhan,Hubei 430030,China;College of Chemistry and Molecular Sciences,Wuhan University,Wuhan,Hubei 430072,China)

机构地区：[1]武汉大学人民医院骨科,430060 [2]武汉大学化学与分子科学学院,430072

出　　处：《中华生物医学工程杂志》2020年第1期1-8,共8页Chinese Journal of Biomedical Engineering

基　　金：国家自然科学基金(81870980,81371339)。

摘　　要：目的构建一种新型阿霉素-壳聚糖乳酸盐纳米缓释载体用于骨肉瘤治疗研究。方法2%乳酸水溶液制备水溶性的壳聚糖乳酸盐,pH 5.0的反应体系中,壳聚糖乳酸盐与TPP以10:1的投料比制备载阿霉素的壳聚糖乳酸盐纳米微粒(ChLa-Dox NPs),动态光散射(DLS)分析其粒径、多分散系数、Zeta电位,扫描电镜进行微观形貌学表征,ChLa-Dox NPs的生物学效应检测则通过缓释载体体外缓释动力学、MG63骨肉瘤细胞系细胞毒性实验、细胞迁徙能力及流式细胞术检测。结果在pH 5.0及壳聚糖乳酸盐:TPP的投料比为10:1的反应条件下制备的ChLa-DoxNPs平均粒径为142.4±1.21μm,多分散系数(PDI)为0.14±0.01,Zeta电位为+29.2 mV。ChLa-DoxNPs在体外10h时阿霉素达到缓释平台期,缓释12 h内阿霉素累积释放率为(69.4±2.6)%;MG63细胞系骨肉瘤细胞模型实验中,空载纳米微粒组细胞存活率为92.36±3.17%,而不同载药量的ChLa-Dox NPs组具备显著的骨肉瘤细胞杀伤作用。相比空载体组,ChLa-Dox NPs对骨肉瘤细胞迁徙能力有显著的抑制作用,明显促进骨肉瘤凋亡的发生[(凋亡率为(54.08±2.53%)]。结论经改良制备的载阿霉素壳聚糖乳酸盐纳米微粒符合抗肿瘤纳米载体的理化特性要求,生物相容性好,体外缓释动力学及抗骨肉瘤相关分子-细胞生物学检测表明其具备较好的阿霉素缓释特性,体外能有效的抑制骨肉瘤细胞增殖、迁徙并促进其凋亡,具有较明确的临床转化前景。Objective To develop a novel doxorubicin(Dox)-loaded chitosan lactate nanoparticles(ChLa-Dox NPs)for experimental treatment of osteosarcoma.Methods The hydrosoluble chitosan lactate(ChLa)was prepared with 2%aqueous lactic acid solution.In a reaction system with pH=5 and a charge ration of ChLa:TPP=10:1,the ChLa-Dox NPs were generated.Subsequently,dynamic light scattering(DLS)was employed to investigate the particle size distribution,polydispersity(PDI)and Zeta potential.The microstructure of nanoparticles was identified with transmission electron microscopy(TEM).The biological characterization of ChLa-Dox NPs were explored with multiple assays including in-vitro sustain-release,osteosarcoma cell-line MG63 cytotoxicity,cell migration and flow cytometry.Results Under pH=5 and a charge ratio of ChLa:TPP=10:1,the genrated ChLa-Dox NPs were 142.4±1.21μm in size,0.140±0.01 by PDI,and 29.2 mV by Zeta potential.The controlled release assay in vitro showed that the sustain release of ChLa-Dox Np reached a plateau at 10 h and the accumulative release amount of Dox at 12 h was(69.4±2.6)%.In cell model tests using osteosarcoma MG63 cell-line,the cell viability was 92.36±3.17%in the group treated with blank NPs;in contrast,treatment of osteosarcoma cells with ChLa NPs loaded with different doses of Dox showed significantly cytotoxicity.Compared with blank NPs,ChLa-Dox NPs showed significant inhibition of migration ability and remarkably promotion of apoptosis in osteosarcoma cells[apoptosis rate:(54.08±2.53)%].Conclusion ChLa-Dox NPs generated with optimized formulation demonstrated efficient release profile and good biocompatibility in vitro.The in-vitro sustain-release kinetics and molecular/cell biological tests on anti-osteosarcoma activity showed that ChLa-Dox NPs exhibited satisfactory properties of doxorubicin sustain-release,effective inhibition of cell proliferation and migration,and promotion of apoptosis of osteosarcoma cells in vitro.These point to a promising prospect of clinical translational attempts.

关 键 词：骨肉瘤 多柔比星 壳聚糖乳酸盐 纳米微粒 缓释载体 

分 类 号：R73[医药卫生—肿瘤]