rhBMP-2／无定形磷酸钙纳米缓释微粒成骨活性的实验研究

目的探讨无定形磷酸钙（ACP）对rhBMP-2的缓释作用，检测rhBMP-2／ACP纳米缓释微粒的成骨活性。方法用扫描电镜观察已制备rhBMP-2／ACP缓释微粒的大小、形态：测定rhBMP-2的体外释放情况并描绘曲线；用MTT法检测缓释微粒对兔骨髓间充质干细胞（MSC）增殖情况的影响：用碱性磷酸酶（ALP）试剂盒检测细胞ALP活性以反映缓释微粒对其分化的影响．并与ACP的作用进行比较；将缓释微粒植入大鼠股部肌袋．通过X线、组织形态学观察评价缓释微粒的异位成骨能力。结果缓释微粒大小为100nm左右．具有典型的无定形球形面貌。开始时为快速释放期．随后呈缓慢持续释放。缓释微粒能显著促进MSC的增殖和分化，植入大鼠股部肌袋12周．材料大部分降解．有明显的骨形成。结论ACP可作为rhBMP-2合适的缓释载体材料．生成的纳米缓释微粒具有良好的降解性能和成骨活性。

Experimental study on the osteogenic activity of rhBMP-2 loaded amorphous calcium phosphate slow release microsphere

Objective To study the effect of amorphous calcium phosphate （ACP） on the slow release activity of rhBMP-2 microspheres and evaluate its osteogepic activity. Methods The size and appearance of the microspheres were observed by scanning electron microscope. The release profile of rhBMP-2/ACP was described with curve figure. The effect of the slow release microsphere on the proliferation of rabbit bone marrow stem cells （MSC） was measured by cell culture technique and MTT colorimetric assay. The ALP kit was used to evaluate the ALP activity of the cells and show the effect of the microsphere on differentiation of the cells compared with that of ACP only. The rhBMP-2/ACP or ACP microspheres were implanted bilaterally into leg muscle pouches of rats to observe ectopic bone formation activity with histological and X-ray measure. Results The diameter of rhBMP-2/ACP microspheres were about 100 nm with amorphous sphere appearance. The rhBMP-2/ACP microspheres released rapidly at the beginning , then released lasting slowly. The microspheres could significantly promote proliferation and differentiation of MSC. Conclusion It is demonstrated that ACP is an appropriate carrier material for rhBMP-2 to achieve slow release, the produced microspheres provides good slow release and osteogenic activity.