【摘要】 背景:研究表明,重组人骨形态发生蛋白2/壳聚糖-磷酸钙支架复合体具有良好的细胞相容性,且材料内部的多孔结构和孔径也能满足于细胞长入和骨的再生,但要应用于临床还需证明能否体内成骨。目的:构建壳聚糖-磷酸钙与重组人骨形态发生蛋白2的复合材料,并观察其植入兔肌袋模型后的异位成骨能力。设计、时间及地点:观察性实验,于2007-06/08在暨南大学附属第一医院中心实验室、外科实验室及暨南大学理工学院材料科学与工程系实验室完成。材料:将固相的磷酸三钙粉末及液相的壳聚糖溶液在室温下混合,制备壳聚糖-磷酸钙支架。再将壳聚糖-磷酸钙支架放入重组人骨形态发生蛋白2溶液中浸泡,真空抽吸后冷冻干燥,制备壳聚糖-磷酸钙材料/重组人骨形态发生蛋白2复合材料。方法:测量改良后支架的孔隙率及抗压强度,扫描电镜观察其超微结构;20只新西兰大白兔局部麻醉后在左后肢大腿外侧切口,暴露大腿肌肉,分离形成肌袋。随机分为3组,空白对照组5只不植入材料,单纯壳聚糖-磷酸钙材料组5只、壳聚糖-磷酸钙/重组人骨形态发生蛋白2复合材料组10只分别植入壳聚糖-磷酸钙材料、壳聚糖-磷酸钙/重组人骨形态发生蛋白2复合材料。主要观察指标:在植入后4周,通过大体观察、X射线摄片、组织学检测其体内异位成骨能力及生物反应性。结果:制备的复合重组人骨形态发生蛋白2后的壳聚糖-磷酸钙支架的孔隙率为87%,压缩弹性模量为20MPa。扫描电镜结果显示支架材料内部为多孔结构,孔径超过100μm。空白对照组苏木精-伊红染色可见肌肉纤维间少量淋巴细胞浸润。单纯壳聚糖-磷酸钙材料组大体观察可见材料周围有薄层的结缔样组织包裹,材料与周围肌肉连接松散,苏木精-伊红染色可见材料植入区为圆形空腔。壳聚糖-磷酸钙/重组人骨形态发生蛋白2复合材料组大体观察可见材料周围有薄层的结缔样组织包裹,材料中间可见空洞,有降解现象及少量淡红色纤维样结构长入,X射线示左大腿内材料高密度影无明显改变,苏木精-伊红染色可见少量血管样组织长入,Masson三色染色可见编制骨岛中有胶原纤维形成。结论:壳聚糖-磷酸钙/重组人骨形态发生蛋白2复合材料具有良好的孔隙率、抗压强度及超微三维结构,复合材料在兔体内具有异位成骨能力。更多还原

【Abstract】 BACKGROUND:It has been demonstrated that chitosan and calcium phosphate cement with human bone morphogenetic protein-2(rhBMP-2) has good cell compatibility, which more suitable for the cell growth and bone regeneration for its porous structure and pore diameter.However, whether the composite materials can osteogenesis in vivo needs further study.OBJECTIVE:To construct the chitosan-calcium phosphate/rhBMP-2 composite materials, further more, to observe the ectopic osteogenesis of the composite materials after implanting it into rabbit muscular tissue.DESIGN, TIME AND SETTING:The observational experiment was performed at the Central Laboratory and Surgical Laboratory of the First Affiliated Hospital of Jinan University and the Laboratory of Materials Science and Engineering, Jinan University Technology College from June to August in 2007.MATERIALS:The scaffold of chitosan-calcium phosphate cement was produced and taken into rhBMP-2 solution, followed by vacuum aspiration and freeze-drying to harvest the chitosan-calcium phosphate/rhBMP-2 composite materials.METHODS:The porosity and compressive strength of chitosan-calcium phosphate/rhBMP-2 composite materials were measured, and its microstructure was observed via scanning electron microscopy.A total of 20 New Zealand rabbits were divided into the blank control(n =5), pure chitosan-calcium phosphate(n=5) and composite materials(n =10) groups.The scaffolds were implanted intramuscularly.There was no treatment in the blank control group except exposed the muscle of upper leg.MAIN OUTCOME MEASURES:The gross observation, X-ray films, as well as histological examination were used to measure the ectopic ossification and biological response after 4 weeks of implantation.RESULTS:The porosity of rhBMP-2/chitosan-calcium phosphate composite scaffold was 87%, with compressive elastic modulus of 20 MPa.The scanning electron microscope showed that there was interconnected pore structure, which over 100 μm in diameter in the material.In the blank control group, hematoxylin-eosin staining showed that there was sparse lymphocytic infiltration around muscle fibers.In the pure chitosan-calcium phosphate group, the gross observation demonstrated that the materials were encapsulated by connective tissues, the connection between material and muscle was loose, and there was circular cavity in the implanted area.In the composite materials group, the scaffold was encapsulated by connective tissues, with cavity in the middle, and fibriform structure decomposed into the scaffold.The X-ray revealed that the high density shadow was not obvious changes.Hematoxylin-eosin staining and Masson displayed that connective tissue involving blood vessels and collagen fibers formed in the woven bone islands.CONCLUSION:The chitosan-calcium phosphate/rhBMP-2 composite materials is three-dimensional, porous with good compression strength, which has ability of ectopic osteogenesis in rabbits.更多还原