离子掺杂纳米羟基磷灰石对类人胶原骨材料结构与功能的影响

Bones of human beings and animals are composed of nano-sized hydroxyapatite (HAp) and collagen. The perfect matching of the inorganic and organic components in nanoscale contributes greatly to the wonderful combination of hardness and toughness of bones. So how to realize the careful control over the size and surface properties of HA is regarded as the key to the construction of artificial bones and other inorganic-organic composites, based on which the collagens may be effectively co-assembled. We simulate natural biological minerals in the structure characteristics and introduce abundant foreign cationic and/or anionic ions (Mg2+, Zn2+, Sr2+, F-, CO32-, etc.) into the reaction system in the preparation of HAp nanomaterials, and successfully synthesize series of monodisperse-uniform hybrid nanoHAps with different morphologies and sizes by facile hydrothermal routes. We utilize novel hybrid nanoHAps as the inorganic phase and human-like collagen (HLC) with superior biocompatibility as the main organic phase, and prepare series of degradable artificial bones with interconnected pores in the present of the cross linking agents. While the as-obtained porous scaffold should fulfill the requirements of the bone tissue engineering, including: a) biocompatibility, b) inducing strong bone bonding, c) better mechanical constancy in load bearing sites prior to regeneration of new tissue, d) porous structure and pore size of more than 100 μm for cell penetration, tissue in growth and vascularisation, e) the rate of their biodegradation matching with one of new tissue formation. Moreover, we may explore the relationship among the repaired efficiency of porous HAp scaffold for bone defect, the sizes of nanoHAps, and the types and amounts of foreign ions.

人类及动物的骨骼已经被证实由纳米尺度的羟基磷灰石（HAp）及胶原蛋白构成，有机无机组分在纳米尺度的完美复合是骨骼兼具韧性与硬度的优异机械性能的根源。如何在纳米尺度实现对HAp尺寸及表面性质的控制，并以此为基础与胶原蛋白进行高效复合是获得性能优异的人工骨材料乃至其它有机-无机复合材料的关键。本实验模拟天然生物矿物质的结构组成，在HAp制备体系中引入不同种类和不同量的杂质离子，获得系列不同形貌尺寸的单分散nanoHAp；利用杂化的nanoHAp与类人胶原蛋白交联，构筑可生物降解的多孔性骨修复框架材料。该框架材料应具有良好的生物相容性和骨诱导活性，能有效充当新骨形成的支架，在体内的降解速度能与新骨生成的速率完美匹配。该框架材料内部孔与孔相互贯通且孔径大于100μm，适合于细胞渗入、增殖和新血管的形成。我们将通过具体实验，揭示人工骨的性能与nanoHAp粒子形貌尺寸及掺杂离子之间的关系。

本项目通过反应体系的优化和多种离子掺杂的调控，合成出了系列形貌尺寸可控的离子掺杂羟基磷灰石纳米晶棒、纳米梭、纳米线、纳米片、纳米带、纳米柱等，并初步探讨了相关材料的理化性质。以用稀土铕和铽为掺杂离子，采用两种模式掺杂合成羟基磷灰石纳米晶：1）采用一步水热合成法，2）采用两步水热法。通过氟离子辅助镁离子共同掺杂，克服了钙镁离子半径相差很大而不易进行晶胞内离子替换的问题，获得了尺寸约为 20nm×100nm 的富镁的麦穗状单晶纳米梭。采用了氟离子或稀土离子协同锌离子掺杂模式，获得了单分散的锌离子共掺杂单晶纳米棒、纳米片和超细纳米线等。通过稀土铕离子和铽离子协助铜离子实现了有效地共掺杂，首次制备出铜离子掺杂羟基磷灰石超薄单晶纳米带和纳米棒。以稀土铕离子和氟离子为模型离子，对羟基磷灰石进行离子掺杂矿化合成，成功获得了直径约 1 纳米的超细纳米线、柔性超细超长纳米线、纳米梭和纳米柱。.本项目利用制备的离子掺杂的纳米羟基磷灰石与类人胶原蛋白复合，并利用京尼平、橄榄多酚、BDDE、EDC、DEO、多酚氧化酶、TG酶等为交联剂，自然陈化成型法制备出了系列人工骨。通过对复合材料的相貌分析、成分分析、力学实验和生物学实验，验证所得材料的内部结构、力学性能、细胞毒性和对骨缺损修复能力。结果表明所有材料均达到了松质骨的强度，有机相与无机相比值约为1:3~1:4。将小鼠成骨细胞前体细胞(MC3T3-E1 cells)种植于吸附法制备的支架上，进行细胞增殖分析、CFDA荧光染色观察、扫描电子显微镜(SEM)观察、碱性磷酸酶（ALP）测定和骨钙素（OCN）测定，结果表明：两种不同的方法构建HLC/TBC支架都有较好的细胞相容性，不仅可以使细胞较好的粘附、还可使细胞生长良好、增殖旺盛，可引导种植的细胞向支架孔隙内生长，并且能维持细胞天然的形态，促进细胞分泌细胞外基质。骨缺损修复实验表明人工骨的骨缺损修复效效果优异。