多孔磷酸钙人工骨与重组人骨形成蛋白2复合修复骨缺损的实验研究

EXPERIMENTAL STUDIES ON THE POROUS CALCIUM PHOSPHATE CEMENT COMBINED WITH RECOMBINANT HUMAN BONE MORPHOGENETIC PROTEIN 2 FOR BONE DEFECTS REPAIR

目的研究多孔磷酸钙人工骨(porouscalciumphosphatecement,PCPC)与重组人骨形成蛋白2(recombinanthumanbonemorphogeneticprotein2,rhBMP-2)复合后体外的缓释作用及其对兔骨缺损的修复作用。方法采用物理吸附法将rhBMP-2(0.4mg)溶液吸附至PCPC中,制备成PCPC/rhBMP-2复合材料。冻干后,扫描电镜观察复合材料内部形态。以包覆壳聚糖的PCPC/rhBMP-2为实验组,单纯PCPC/rhBMP-2为对照组,测试在模拟体液中的rhBMP-2缓释行为。取新西兰大白兔12只,股骨远端制成直径4.2mm,深5.0mm的骨缺损模型。将包覆壳聚糖的PCPC/rhBMP-2复合材料修复骨缺损作为实验组,以植入单纯PCPC作为对照组。术后观察动物一般情况,于4周和8周取材行X线片和组织学观察。结果扫描电镜显示PCPC/rhBMP-2复合材料孔隙中吸附了大量的rhBMP-2。rhBMP-2体外缓释对照组rhBMP-2于150h基本全部释放;实验组rhBMP-2于350h缓释量约达99%,较对照组慢。动物实验动物术后切口无感染,于4周行动自如。X线片示术后4周对照组骨缺损区材料清晰,实验组骨缺损区密度大部分接近宿主骨,材料模糊;8周对照组材料边缘较术后4周模糊,实验组骨缺损区密度已基本接近宿主骨。组织学观察,术后4周对照组可见少量成骨细胞和破骨细胞,实验组可见成熟骨组织和骨髓腔,新生骨逐渐取代材料;8周对照组可见大量成骨细胞和破骨细胞,少量新生骨并向材料内长入,实验组可见成熟骨小梁和骨髓组织。结论PCPC是rhBMP-2较理想的载体材料,复合后具有良好的诱导成骨作用,可作为一种新型复合人工骨修复骨缺损,具有良好的临床应用前景。

Objective To study in vitro sustained release behaviour of the recombinant human bone morphogenetic protein 2（rhBMP-2） from the sample which porous calcium phosphate cement （PCPC） was combined with rhBMP- 2, and to evaluate the effect of PCPC/rhBMP- 2 composite on repairing bone defect in the animal study. Methods rhBMP-2 was absorbed into PCPC by vacuum-adsorption and freeze dried at - 40 ℃ , the PCPC/rhBMP-2 enwrapped with chitosan as the experimental group, the pure PCPC/rhBMP -2 as the control group, then the sustained release of rhBMP- 2 from PCPC was determined in simulated body fluid （SBF） by UV-VIS spectrophotometer. At same time, the PCPC/rhBMP-2 composites with chitosan were implanted into the （4.2 mm× 5.0 mm femora defects of rabbits, which were considered as the experimental group, whereas in the control group only PCPC was implanted. The effect of repairing bone defect was evaluated in the 4th and 8th week postoperatively by radiograph and histomorphology. Results The PCPC have a high absorption efficiency to rhBMP-2, and the release of rhBMP-2 was sustained release system. The release of rhBMP-2 from PCPC in the experimental group （99% after 350 hours） was slower than that in the control group （100% after 150 hours）. In the experimental group, the radiological and histomorphological evaluations showed that the interfaces between the materials and host bones became blurred both at 4th and 8th week. The implanted materials were partially absorbed, and the implanted areas exhibited the formation of new bone. In the control group, a little amount of new bones was observed. Conclusion The PCPC shows great clinical potential as a carrier for rhBMP-2. The PCPC/rhBMP- 2 composite possesses much potentialities of osteoinductivity and the ability of repairing bone defect, so it can be used as a novel bone substitute clinically.