重组人BMP-2/壳聚糖/硫酸葡聚糖复合微球诱导异位成骨的micro-CT评价  

ECTOPIC OSTEOGENESIS EVALUATION OF RECOMBINANT HUMAN BONE MORPHOGENETIC PROTEIN 2 LOADED CHITOSAN/DEXTRAN SULFATE BY MICRO-CT

作　　者：夏远军[1] 余翔[2] 章莹[1] 尹庆水[1] 夏虹[1] 郑晓辉[3] 谢会斌[1] 黄显华[1] 

机构地区：[1]广州军区广州总医院创伤骨科,广州510010 [2]广州中医药大学第一临床医学院 [3]广州中医药大学第一附属医院骨科

出　　处：《中国修复重建外科杂志》2016年第3期286-291,共6页Chinese Journal of Reparative and Reconstructive Surgery

基　　金：国家自然科学基金青年基金资助项目(51303031);广东省自然科学基金资助项目(S2012010009743);广东省应用基础研究计划项目(2013J4100120)~~

摘　　要：目的采用micro-CT评价重组人BMP-2/壳聚糖/硫酸葡聚糖（recombinant human BMP-2/chitosan/dextran sulfate,rhBMP-2/CS/DS）复合微球诱导异位成骨情况。方法采用离子交联法制作rhBMP-2/CS/DS复合微球,扫描电镜观察其形态,ELISA法检测微球体外缓释效应。取6-8周龄雄性SD大鼠48只,随机分为4组（n=12）;制备股四头肌肌袋模型后,分别将相同体积明胶海绵（A组）、CS/DS微球（B组）、rhBMP-2（C组）及rhBMP-2/CS/DS复合微球（D组）植入肌袋肌间隙中。术后观察大鼠一般情况;于4、8、12、16周处死动物,取异位骨化组织块行micro-CT扫描及三维重建,检测组织骨密度（tissue mineral density,TMD）、骨体积分数（bone volume fraction,BVF）、骨小梁厚度（trabecular thickness,Tb.Th）、骨小梁数量（trabecular number,Tb.N）、皮质骨骨密度（bone mineral density,BMD）及组织矿含量（tissue mineral content,TMC）。结果扫描电镜观察示,制备的rhBMP-2/CS/DS复合微球球形较规整,分散较均匀,无团聚,微球表面较光滑;微球体外释放rhBMP-2于2 h后出现1个突释放期,2 d时达峰值,释放周期约20 d。各组大鼠均存活至实验完成,术后各时间点,A、B组CT扫描均未见放射性显影,三维重建未发现成骨;而C、D组放射性显影强度及三维重建骨组织逐渐增加,且D组强于C组。各时间点A、B组TMD、BVF、Tb.Th、Tb.N、BMD及TMC均为0。随时间延长,C、D组除Tb.N外,其余骨组织参数均呈总体增加趋势,各时间点与A、B组比较差异均有统计学意义（P〈0.05）;C、D组间4周时比较差异无统计学意义（P〉0.05）,8-16周时D组显著高于C组,比较差异有统计学意义（P〈0.05）。结论 rhBMP-2/CS/DS复合微球异位诱导成骨能力显著强于单独rhBMP-2。Objective To evaluate the ectopic osteogenesis of recombinant human bone morphogenetic protein 2（rhBMP-2） loaded chitosan（CS）/dextran sulfate（DS） by micro-CT. Methods rhBMP-2/CS/DS microspheres were prepared by the ionic crosslinking and its shape was observed under the scanning electron microscope. The release of rhBMP-2 was determined from resultant microspheres by ELISA assay. Forty-eight Sprague Dawley male rats were randomly divided into 4 groups（n=12）, quadriceps muscle bag model was made, gelatin sponge（group A）, CS/DS microspheres（group B）, rhBMP-2（group C）, and CS/DS/rhBMP-2 microspheres（group D） were implanted into the bags respectively. The tissue samples with heterotopic ossification were harvested for micro-CT scanning at 4, 8, 12, and 16 weeks. The tissue mineral density（TMD）, bone volume fraction（BVF）, trabecular thickness（Tb.Th）, trabecular number（Tb.N）, bone mineral density（BMD）, and tissue mineral content（TMC） were measured. Results The prepared rhBMP-2/CS/DS microspheres with smooth surfaces were spherical and evenly disperses without obvious agglomeration. At 2 hours, microsphere started a sudden release period in vitro; the release reached a peak at 2 days; and the release cycle lasted about 20 days. The rats survived to the end of the experiment. At each time point after operation, no radiation developed and no osteogenesis was observed by three dimensional reconstruction in groups A and B. However, radioactive strength and reconstructed bone tissue gradually increased in groups C and D, and group D had more radioautography and more bone tissues than group C. At each time point, TMD, BVF, Tb.Th, Tb.N, BMD, and TMC of groups A and B were zero. Ectopic bone formed with time, the other parameters showed an increasing trend except Tb.N in groups C and D, showing significant difference when compared with groups A and B at each time point（P〈0.05）. There was no significant difference between groups C and D at 4 weeks（P〉0.05）; the p

关 键 词：BMP-2 壳聚糖 硫酸葡聚糖 异位成骨 MICRO-CT 三维重建 大鼠 

分 类 号：R318.08[医药卫生—生物医学工程]