基于Co离子和BMP-2诱导成骨的三维多孔骨修复系统的构建研究

The defective angiogenesis of bone defect and insufficient osteoinduction of bone graft materials are main problems in the treatment of bone repair. Both of cobalt ion (Co) and bone morphogenetic protein-2 (BMP-2) could enhance osteogenesis. Co can up-regulate the activity of hypoxia-induced factor-1 (HIF-1) signal, while it produces reactive oxygen species (ROS) which can destroy body cell. The BMP-2 possesses weak stability and it is easy to lose activity. In this study, body environment and ROS-responsive degradable poly(amino acid) (PAA) will be synthesized for reducing the damage of ROS produced by Co and keeping the stability and activity of BMP-2. The Co incorporated into hydroxyapatite (HA-Co) and BMP-2 encapsulated into PAA will be loaded into porous chitosan with controlled degradation. Following the degradation of chitosan, HA-Co and PAA will be released next. The ROS and body environment together promote the degradation of PAA, followed by the release of BMP-2. HA-Co and BMP-2 entitle porous chitosan with performance of angiogenesis and osteogenesis. This study will explore the influence of above composites on HIF-1 signal in the area of calvarial defect and research its bone repairing ability. This study possesses practically applicable value in clinic.

骨缺损修复存在局部新生血管能力不足和植骨材料诱导成骨能力有限等诸多不足。钴离子（Co）和骨形态发生蛋白-2（BMP-2）均可促进成骨；Co可激活缺氧诱导因子-1（HIF-1）发挥成血管和成骨作用，但产生的活性氧自由基（ROS）损伤机体细胞；BMP-2稳定性差，且易失活。为减少Co产生的ROS对机体细胞的损伤，保持BMP-2的稳定和活性，本项目设计了机体内环境联合ROS响应降解的氨基酸聚合物（PAA）；通过将掺Co的羟基磷灰石（HA-Co）与包埋BMP-2的PAA联合构建后载入可控降解壳聚糖多孔支架。壳聚糖降解释放HA-Co和PAA，Co产生的ROS和机体内环境共同促进PAA降解，PAA的降解促使BMP-2释放，HA-Co和BMP-2双释放因子赋予壳聚糖多孔支架诱导成血管和成骨的功能。本项目将深入探索该复合材料对颅骨缺损区HIF-1信号通路的调控作用，明确其骨修复功能，研究具有临床实用价值。

本项目设计了机体内环境联合ROS响应降解的氨基酸聚合物（PAA）；通过将掺Co的羟基磷灰石（Co-HA）与包埋BMP-2的PAA联合构建后载入可控降解壳聚糖多孔支架。基于以上目的，研究构建了Co-HA诱导骨髓间充质干细胞（BMSCs）成骨分化后蛋白和基因的表达研究，证实Co-HA诱导成骨和成血管的能力；研究同时构建了基于活性氧自由基响应降解的PAA聚合物，为包埋具有诱导成骨能力的BMP-2提供有效的载体；而且，通过构建孔隙均匀的三维多孔壳聚糖载体，有效将Co-HA和PAA/BMP-2均匀分散于其中，并将其植于新西兰大白兔颅骨及股骨髁骨骨缺损部位8w对其成骨性能进行体内研究，证实了其有效的引导成骨能力。研究结果显示，Co-HA能够诱导BMSCs成骨分化，促进成骨和成血管相关基因的表达；PAA在活性氧自由基环境下能够有效降解；多孔壳聚糖包埋Co-HA和PAA/BMP-2载体在新西兰大白兔体内8周可有效引导成骨，新生骨紧密贴附载体，体现了较好的生物相容性和引导成骨性能。该研究结果在一定程度上揭示了Co-HA可利用于临床骨修复的可能，且通过HIF-1通路促进成骨基因的表达；而且，三维多孔载体有效负载Co-HA和PAA/BMP-2骨缺损填充物表现出良好的引导成骨性能，具有一定的应用价值。