钙磷陶瓷对巨噬细胞与间充质干细胞相互作用的调控及其在骨诱导中的意义

Osteoinductive biomaterial shows great potential in regenerative medicine, but is still difficult to achieve a good clinical result similar to autogenous bone graft. Hence there is a need for further in-depth understanding of the mechanism of material’s osteoinduction in order to guide the design of novel bone-repairing material, which can regulate body’s response and reach self-repair. This proposal, which stands in the forefront of clinical study, with a unique perspective based on the critical role of inflammatory reaction in tissue repair, tries to comprehensively understand the relationship between material’s osteoinduction and inflammatory response caused by calcium phosphate (CaP) ceramics with different phase composition. It applies in vivo optical imaging technology to build the novel in vitro and in vivo models for non-invasive, real-time, quantitative and qualitative monitoring of temporal-spatial dynamic changes of macrophages and mesenchymal stem cells (MSCs) under the material’s effect. Moreover, it also provides a quick and efficient cell sorting method to investigate the effect of material’s phase composition on the secretion of signaling molecules of macrophages and on the response of MSCs to inflammation, as well study the influence of a local inflammatory microenvironment on homing and differentiation of MSCs and its underlying mechanism. This proposal provides a new idea for the design of novel osteoinductive biomaterial, by regulating the interaction between macrophages and MSCs to improve inflammatory environment and induce bone formation.

骨诱导性生物材料在再生医学领域具有巨大的潜力，然而目前仍难以达到与自体骨移植相同的临床功效。因此需要进一步深入认识材料的骨诱导机理，用于指导新型的“可调控机体应答、达成自身修复”骨修复材料的设计。本项目立足于临床骨修复材料研究的前沿，以独特的视角，基于炎性反应在组织修复中的重要作用，全面考察不同相组成的钙磷生物陶瓷所激发的炎性反应与材料骨诱导性之间的联系。采用活体光学成像技术构建新颖的体内外活体示踪模型，用于无创、实时、定性定量地监控在材料作用下巨噬细胞和间充质干细胞（MSCs）的竞争性动态变化；并快速、有效地分离细胞，考察材料相组成对巨噬细胞旁分泌的作用及对MSCs调节炎性反应过程的影响，并探讨其相互作用所导致的局部炎性微环境对MSCs所介导的骨发生的调控及分子机理。本项目提出材料可以通过调控机体内细胞间相互作用来改善炎性环境并诱导骨发生，为新型骨诱导性生物材料的设计提供新的思路。

钙磷活性生物陶瓷的骨诱导性已得到广泛认可，然而其诱导机理仍为完全了解。近年研究发现固有免疫反应在组织修复中具有重要作用，因此本项目考察钙磷陶瓷对免疫细胞（特别是巨噬细胞）行为的调控，以及材料-巨噬细胞-间充质干细胞（MSCs）相互作用在材料诱导成骨中的意义。通过构建小鼠大腿肌内植入模型发现钙磷陶瓷可诱导异位骨的生成，并且植入体的体内诱导成骨能力与材料所调控的宿主固有免疫反应（如巨噬细胞的浸润和极化）密切相关。体外细胞培养表明钙磷陶瓷影响了巨噬细胞的形貌、增殖和分泌功能。而巨噬细胞响应钙磷陶瓷所分泌的细胞因子可调控MSCs的细胞学行为，包括：钙磷陶瓷增加巨噬细胞趋化因子的分泌从而促进MSCs的迁移，并且陶瓷可通过显著下调促炎症因子的表达和上调抑炎症因子及多种生长因子的表达，从而形成一个有利于成骨的局部微环境，促进MSCs的成骨分化。同时，利用转基因和荧光活体示踪技术标记细胞，成功构建材料/巨噬细胞/MSCs接触式共培养体系，模拟真实生理环境，发现三者相互作用可抑制巨噬细胞的炎症功能并影响MSCs参与的骨发生。本研究加深了对材料骨诱导性机理的认识和了解，可用于指导具有免疫调控能力的骨诱导材料的设计与制备。