分层交错螺旋型人工骨的构建及其取向成骨效能研究

A general issue for conventional artificial materials in repair of large bone defects, is the limitation of bone ingrowth and bone filling, which results in non-osseointegration with host bone and regenerative failure. Our previous study found that a spiral-cylindrical scaffold can guide oriented migration of bone-forming cells, which eventually grow throughout the material, achieving functional osseointegration. In the proposed project, we will break the macroscale design concept to topologically optimize of the scaffold microstructure. A layered crisscross-spiral scaffold will be prepared using electro-spun nanofibers to simulate bone microenvironment. By regulating the composition and arrangement of nanofibers and incorporating growth factors in the multi-layer scaffold, the seed cells (BMSCs) will be placed in differential microenvironment, and a biomimetic artificial bone will be built with tubular vascularized tissue and cylinder bone tissue from the centre to the periphery. Moreover, 3D microstructure of the scaffold affecting cell adhesion, migration, proliferation, differentiation and its possible mechanism will be studied, and a rabbit radius defect model will be established to investigate the oriented osteogenic effect. The newly designed bone scaffold is expected to achieve structural and functional biomimetics at a microscopic level, and a systematic study is proposed to investigate spatial configuration and arrangement of the scaffold regulating oriented cell growth, which provides theoretical and experimental basis for repairing large bone defects.

常规的人工骨支架在修复大段骨缺损时，普遍存在骨长入深度有限和新生骨充盈不足的问题，故难以与宿主骨完全融合而导致治疗失败。课题组前期研究发现，具有螺旋取向结构的柱体支架能引导骨形成细胞定向迁移，并最终贯穿材料内部生长，实现功能性骨整合。本项目将突破宏观尺度的设计理念，对取向支架的微结构进一步拓扑优化，采用静电纺丝技术，分层构建模拟骨微环境的交错螺旋型纳米纤维支架。通过改变纳米纤维的组成和空间排列方式，以及分层添加生长因子为种子细胞BMSCs提供差异化诱导环境，构建中心通道血管化组织、外围柱体骨组织的仿生人工骨，探索支架的三维微观结构对细胞粘附、迁移、增殖及分化等的作用机制，并用兔桡骨缺损模型评价其体内取向成骨效能。本项目拟构建的新型人工骨支架，从微观水平实现结构和功能的仿生，并系统研究支架三维构型及空间排列方式调控细胞取向生长的规律，有望为大范围骨缺损的修复治疗提供理论和实验基础。

常规的人工骨支架在修复骨缺损时，普遍存在骨长入深度有限和新生骨充盈不足的问题，故难以与宿主骨完全融合而导致治疗失败。采用静电纺丝技术，改变纳米纤维的组成和空间排列方式，分层构建模拟骨微观结构的分层交错螺旋型纳米纤维支架，并调控细胞取向生长是本项目的立项初衷。项目启动后，课题组通过调控静电纺丝参数和结合辅助电极的方式对构建分层交错螺旋型纳米纤维支架进行了探索，先后尝试了改变电压连接方式、辅助电极高度、辅助电极间距、收杆接收距离、接收杆转速、接收杆直径、平移装置移动速率和辅助电极倾斜方向，最终成功实现了以小直径接收杆在低转速下构建分层交错取向排列的纳米纤维支架。为了使人工骨支架实现更好的骨整合效果，除了从支架结构上实现仿生模拟以外，还应使材料的组分更接近自然骨的化学成分并具有一定的功能特征。因此，课题组选取了自然骨的主要无机成分——羟基磷灰石作为研究对象，并用阿仑膦酸盐和四氧化三铁对其进行表面修饰，制备了多功能化羟基磷灰石。随后，将多功能化羟基磷灰石负载于聚合物支架上，进一步评价了其理化性能和生物学性能。实验结果显示，阿仑膦酸盐和四氧化三铁的化学修饰没有让羟基磷灰石晶体发生晶格形变，但使其具有了亚铁磁性和可控释药的性能。体外细胞培养和体内动物实验表明，功能化羟基磷灰石能够抑制破骨细胞活性、促进成骨细胞的增殖和分化，提升了植入物的骨整合能力并加速了骨重建进程。在本项目支持下，目前课题组已发表SCI论文一篇，申请国家发明专利一项。