Liu Yue, Lu Yun, Tian Xuezhong, Cui Geng, Zhao Yanmei, Yang Qiang, Yu Shunlu, Xing Guosheng, Zhang Boxun
Department of Orthopedics, Tianjin Hospital, No. 406 Jiefang South Road, Hexi District, Tianjin 300211, China.
Biomaterials. 2009 Oct;30(31):6276-85. doi: 10.1016/j.biomaterials.2009.08.003. Epub 2009 Aug 15.
We aimed to develop a hybrid scaffold with a porous structure and similar composition as natural bone for the controlled release of bone morphogenetic protein-2 (BMP-2) to enhance bone regeneration. We fabricated a gelatin/nanohydroxypatite (nHAP) scaffold by glutaraldehyde chemical cross-linking a gelatin aqueous solution with nHAP granules at a 5:1 ratio (v/w). Then, fibrin glue (FG) mixed with recombinant human BMP-2 (rhBMP-2) was infused into the gelatin/nHAP scaffold and lyophilized to develop an rhBMP-2-loaded gelatin/nHAP/FG scaffold. On scanning electron microscopy, the composite had a 3-D porous structure. The rhBMP-2 release kinetics from the hybrid scaffold was sustained and slow, and release of rhBMP-2 was complete at 40 days. Immunohistochemistry, azo-coupling and alizarin S-red staining were used to study in vitro differentiation of human bone-marrow mesenchymal cells (hBMSCs). Strong positive staining results confirmed that rhBMP-2 released from the scaffold could improve osteocalcin (OCN) and alkaline phosphatase (ALP) expression and calcium deposition formation. RT-PCR results showed significantly high mRNA expression of ALP and OCN in hBM-MSCs cultured on the gelatin/nHAP/FG scaffold with rhBMP-2. DNA assay demonstrated that the scaffold was noncytotoxic and could promote hBMSC proliferation from the components of the hybrid scaffold, not released rhBMP-2. The hybrid scaffolds were then used to repair critical-size segmental bone defects of rabbit radius. Gross specimen, X-ray, bone histomorphology and bone mineral density assay demonstrated that the rhBMP-2-loaded gelatin/nHAP/FG scaffold had good osteogenic capability and could repair the segmental bone defect completely in 12 weeks.
我们旨在开发一种具有多孔结构且成分与天然骨相似的混合支架，用于骨形态发生蛋白-2（BMP-2）的控释，以促进骨再生。我们通过戊二醛将明胶水溶液与纳米羟基磷灰石（nHAP）颗粒以5:1的比例（v/w）进行化学交联，制备了明胶/nHAP支架。然后，将与重组人BMP-2（rhBMP-2）混合的纤维蛋白胶（FG）注入明胶/nHAP支架中并冻干，以制备负载rhBMP-2的明胶/nHAP/FG支架。扫描电子显微镜观察显示，该复合材料具有三维多孔结构。混合支架中rhBMP-2的释放动力学持续且缓慢，rhBMP-2在40天时完全释放。采用免疫组织化学、偶氮偶联和茜素S-红染色法研究人骨髓间充质细胞（hBMSCs）的体外分化。强阳性染色结果证实，从支架释放的rhBMP-2可改善骨钙素（OCN）和碱性磷酸酶（ALP）的表达以及钙沉积形成。RT-PCR结果显示，在含有rhBMP-2的明胶/nHAP/FG支架上培养的hBM-MSCs中，ALP和OCN的mRNA表达显著升高。DNA检测表明，该支架无细胞毒性，且可通过混合支架的成分而非释放的rhBMP-2促进hBMSC增殖。然后，将这些混合支架用于修复兔桡骨的临界尺寸节段性骨缺损。大体标本、X射线、骨组织形态学和骨密度检测表明，负载rhBMP-2的明胶/nHAP/FG支架具有良好的成骨能力，可在12周内完全修复节段性骨缺损。
