重组人骨形态发生蛋白2壳聚糖纳米微球的制备及体外细胞毒性

背景：通过各种微球负载骨生长因子使骨形态发生蛋白达到缓释效果逐渐成为研究热点，但关于载药壳聚糖纳米微球的生物相容性特别是细胞毒性的报道较少。目的：对重组人骨形态发生蛋白2壳聚糖纳米微球进行细胞毒性检测，评估应用壳聚糖纳米微球作为重组人骨形态发生蛋白2缓释载体的生物安全性。方法：通过离子交联法制备空白壳聚糖纳米微球，应用透视电镜观察微球的形态，激光粒径分析其粒径分布；通过重组人骨形态发生蛋白2壳聚糖纳米微球体外细胞毒性试验评估微球的生物安全性。结果与结论：离子交联法制备的壳聚糖微球，球形规整，分散均匀，微球平均粒径为230 nm，分布较集中。载药及空白微球的反应分级为0或1级，均为合格。提示，离子交联法制备可成功制备出负载重组人骨形态发生蛋2的纳米微球，且微球细胞毒性检测合格，为进一步的骨组织工程研究提供理论实验基础。

Preparation of recombinant human bone morphogenetic protein 2 loaded chitosan nanosphere and its cytotoxicity in vitro

BACKGROUND:The study on bone morphogenetic protein (BMP) release approach has more important significance, but in recent years studies focus on the release effect of microspheres loaded various bone growth factors. However, the biocompatibility, especially cytotoxicity of drug-loaded chitosan nanoparticles, has not been reported. OBJECTIVE:To prepare and characterize a controlled release system for recombinant human bone morphogenetic protein 2 (rhBMP-2) and to evaluate the cytotoxicity and biological safety of chitosan nanospheres as a carrier for controlled release of rhBMP-2.METHODS:The rhBMP-2 loaded chitosan nanospheres were prepared using an ionic crosslinking method with ripolyphosphate  as the crosslinking agent. Transmission electron microscope was used to evaluate the morphological properties and particle size analyzer was used to analysis particle size distribution. A cytotoxic test in vitro was adopted to determine the biological safety of rhBMP-2 loaded chitosan nanospheres. RESULTS AND CONCLUSION:The rhBMP-2 loaded chitosan nanospheres were spherical in shape, and showed a smooth surface. The mean diameter of nanospheres was 230 nm. The cell relative growth rates (RGR) of rhBMP-2 loaded chitosan nanospheres and blank chitosan nanospheres groups were 0 or 1, and all were qualified. The rhBMP-2 loaded chitosan nanospheres prepared by ionic crosslinking method show a good property and biocompatibility. Such a novel type of controlled release system has the potential of being applied for the bone tissue engineering.