负载重组人骨形态发生蛋白-2壳聚糖纳米微球的制备及异位成骨活性研究  

Preparation and ectopic bone formation activity of recombinant human bone morphogenetic protein-2 loaded chitosan nanospheres in vivo

作　　者：王玮[1] 尹庆水[1] 张余[1] 

机构地区：[1]广州军区广州总医院骨科医院, 510010

出　　处：《中华创伤骨科杂志》2012年第2期147-152,共6页Chinese Journal of Orthopaedic Trauma

基　　金：基金项目：全军医学科学技术研究“十一五”计划（08Z010）

摘　　要：目的 制备负载重组人骨形态发生蛋白-2（rhBMP-2）的壳聚糖纳米微球,并考察其成骨活性. 方法 应用离子交联法制备空白壳聚糖纳米微球和rhBMP-2壳聚糖纳米微球,应用透射电镜观察微球的形态,激光粒径分析仪测定其粒径的分布,检测其载药量、包封率及累积释药率.取24只SD大鼠,随机分为4组,每组6只.以无菌手术分别在大鼠左侧股部建立肌袋.A组大鼠肌袋内植入rhBMP-2壳聚糖纳米微球（含rhBMP-2 1 mg）,B组大鼠肌袋内植入rhBMP-2 1 mg,C组大鼠肌袋内植入空白壳聚糖纳米微球,D组大鼠肌袋内不做任何处理.评估rhBMP-2壳聚糖纳米微球的成骨活性.结果离子交联法制备的壳聚糖纳米微球球形规整、分散均匀,微球平均粒径为230.0 nm,分布较集中,包封率为66.87％±4.58％,载药率为（33.44±2.29） μg/mg.A、B、C、D组的ALP活性平均分别为（1.94±0 35）、（1.48±0.56）、（0.20±0.07）及（0.18±0.06） kat/g,差异有统计学意义（F=42.959,P=0.000）,A、B组明显高于C、D组,且A组高于B组.4组钙含量平均分别为（5.20±1.42）、（3.80±1.40）、（0.19±0.08）、（0.20±0.08）μg/mg,差异有统计学意义（F=39.242,P=0.000）,A、B组明 显高于C、D组,且A组高于B组. 结论 离子交联法可成功制备出均一的rhBMP-2壳聚糖纳米微球,该微球具有良好的载药性能和缓释性能,且其骨诱导活性优于单纯rhBMP-2.Objective To prepare recombinant human bone morphogenetic protein-2（rhBMP-2） loaded chitosan nanospheres and to evaluate their ectopic bone formation activity in vivo. Methods The rhBMP-2 loaded chitosan nanospheres were prepaid using the ionic crosslinking method with TPP as a crosslinking agent.The morphological properties and particle size distribution were evaluated by transmission electron microcopy （TEM） and particle size analyzer.An intramuscular ectopic bone formation test was carried out to compare the ectopic bone formation activity in 24 SD rabbits that were randomized into 4 even groups.rhBMP-2 loaded chitosan nanospheres （1 mg）,rhBMP-2 （1 mg）,chitosan nanospheres and nothing were implanted into the muscle pouch at the left thigh,respectively,in groups A,B,C and D. Results The rhBMP-2 loaded chitosan nanospheres were well distributed and spherical in shape.The average particle diameter was 230.0 nm,the entrapment efficiency was 66.87% ± 4.58%,and the loading rate was 33.44 ± 2.29 μg/mg.The mean ALP activity was respectively 1.94 ± 0.35 kat/g,1.48 ± 0.56 kat/g,0.20 ±0.07 kat/g and 0.18 ±0.06 kat/g in groups A,B,C and D,with a significant difference between the 4 groups （ F =42.959,P =0.000）.The mean Ca2 ＋ content was respectively 5.20 ± 1.42 μg/mg,3.80 ± 1.40 μg/mg,0.19 ± 0.08 μg/mgand 0.20 ± 0.08 μg/mg in groups A,B,C and D,with a significant difference between the 4 groups （ F =39.242,P =0.000）.Groups A and B were obviously higher than groups C and D and group A was obviously higher than group B in terms of ALP activity and Ca2＋ content. Conclusion The rhBMP-2 loaded chitosan nanospheres prepared by ionic crosslinking method show a good property and obvious ectopic bone formation activity.

关 键 词：骨形态发生蛋白质类 壳聚糖 纳米复合物 骨生成 

分 类 号：R318.08[医药卫生—生物医学工程]