骨形态发生蛋白2基因复合纤维蛋白凝胶/聚乳酸-聚己内酯双相支架修复节段性骨缺损(英文)

Bone morphogenetic protein-2 gene combined with fibrin gel and polylactic acid/polycaprolactone for repairing segmental bone defects

背景：骨形态发生蛋白2基因转染骨髓基质干细胞后,可促进其增殖分化并诱导异位骨形成。但这种方法操作复杂,体外细胞扩增需时较长,不便于临床应用。目的：将人工骨经骨形态发生蛋白2腺病毒载体与纤维蛋白凝胶和聚乳酸/聚己内酯处理,移植修复骨缺损。设计：随机对照动物实验。单位：实验于2003-09/2004-12在吉林大学中日联谊医院中心实验室完成。材料：聚乳酸/聚己内酯生物可降解材料块由中国科学院长春应用化学研究所提供,孔隙直径150-250μm,孔隙率90%以上;实验动物为3月龄新西兰大耳白兔60只。方法：于新西兰大耳白兔双侧桡骨中段造成1.5cm骨缺损,分别植入4种经不同方法处理的人工骨：①Ad-BMP-2组：骨形态发生蛋白2腺病毒载体＋纤维蛋白凝胶＋聚乳酸/聚己内酯。②重组BMP-2组：重组骨形态发生蛋白2＋纤维蛋白凝胶＋聚乳酸/聚己内酯;③对照基因组：β-半乳糖酐酶基因腺病毒载体＋纤维蛋白凝胶＋聚乳酸/聚己内酯。④PLA/PCL组：纤维蛋白凝胶＋聚乳酸/聚己内酯。主要观察指标：术后4,8,12组周摄双侧前肢正位X射线片测定骨痂灰度（代表新骨密度）;苏木精-伊红染色观察骨缺损修复情况,爱辛蓝和Vankossa染色观察软骨形成及矿化;电镜观察骨细胞成熟、支架降解吸收情况;术后12周行生物力学检测评价新骨抗弯强度。结果：术后12周Ad-BMP-2组缺损区在成骨活跃程度、骨再生量和再生髓腔结构等方面均显著优于重组BMP-2组,其骨缺损得到了较彻底的修复。对照基因组和PLA/PCL组均不能产生骨性愈合。结论：聚乳酸/聚己内酯协同纤维蛋白凝胶运载骨形态发生蛋白2基因修复节段性骨缺损可达到较好的效果。

BACKGROUND： Bone morphogenetic protein-2 （BMP-2） gene transfection can promote the proliferation and differentiation of bone marrow stromal stem cells and induce heterotopic bone formation. However, this method is complicated to operate and in vitro amplification needs a long time, so it is inconvenient in clinic. OBJECTIVE： After treated with adenovirus vector carrying BMP-2 （Ad-BMP-2）, fibrin gel and polylactic acid/polycaprolactone （PLA/PCL）, artificial bone was used to repair bone defects. DESIGN： A randomized control animal study. SETTING： Experiments were performed at the Central Laboratory of China-Japan Friendship Hospital Affiliated to Jilin University from September 2003 to December 2004. MATERIALS： PLA/PCL biodegradable materials were provided by Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, with the pore diameter of 150 250 V m and porosity of above 90%. Sixty New Zealand flap-eared white rabbits aged 3 months were selected. METHODS： Artificial bone treated by four different methods was implanted into 1.5 cm bone defect region on the midpiece of the bilateral radial bone of rabbits. Artificial bone treated with Ad-BMP-2＋ fibrin gel＋ PLA/PCL was implanted into rabbits in the Ad-BMP-2 group; treated with recombinant BMP-2＋ fibrin gel＋ PLA/PCL was implanted in the recombinant BMP-2 group; treated with adenovirus vector carrying β -galactosidase （Ad-Lacz）＋ fibrin gel＋ PLA/PCL was implanted in the control gene group; treated with fibrin gel＋ PLA/PCL was implanted in the PLA/PCL group. MAIN OUTCOME MEASURES： Four, eight, twelve weeks after surgery, X-ray was used to detect callus gray （representing new bone density） on the bilateral anterior limbs. Hematoxylin-eosin （HE） staining was employed to observe the repair of bone defects. Alcian Blue staining and Vankossa staining were used to examine chondrogenesis and mineralization. Electron microscope was applied to investigate maturity of bone cells, degradation and absorption of scaffolds. Twelve weeks after surgery, biodynanlic test was performed to evaluate bending strength of new bone. RESULTS： Twelve weeks after surgery, osteogenic activity, amount of bone regeneration and structure of medullary cavity were significantly better in the defect regions in the Ad-BMP-2 group than in the recombinant BMP-2 group. Bone defects were completely repaired. Bone union was not detected in the control gene and PLA/PCL groups. CONCLUSION： PLA/PCL combined with fibrin gel carrying BMP-2 gene can obtain good effects on repairing segmental bone defects.