Ca-P生物陶瓷表面/界面的形成、表征及其对材料骨诱导性的影响

【作者】 范红松；

【导师】 张兴栋；

【作者基本信息】 四川大学 ， 生物医学工程， 2002， 博士

【摘要】 充分调动人体自我康复能力，达到病变或缺损组织或器官的永久性修复，是当代生物医学科学发展的目标。实现这一目标的关键是必须发展出具有仿生结构的替换材料并赋予其诱导组织再生的生物功能。磷酸钙是自然骨的无机组成，不仅具有良好的生物相容性，且能和新骨形成化学键性结合，特别是近年来发现的钙磷生物材料可具有骨诱导性，有可能诱导骨组织再生，重建病变或缺损的骨，并实现其永久修复。但Ca-P生物材料是否具有骨诱导性一直存在争议，其中一个重要原因是对骨诱导机理的解释不够完善。材料植入体内后与宿主的反应首先并通过表面／界面发生，材料的表面／界面及其行为应当与材料的骨诱导性直接相关。本文通过材料表面／界面及其形成、表征的影响因素和与材料生物学性能关系的研究，探讨了材料的表面／界面及其行为与材料骨诱导的关系。 材料与宿主间的相互作用，是涉及许多因素的一个十分复杂的过程，为简化研究模型，揭示各主要因素的作用，本文首先通过体外实验，逐个研究了各主要的材料学和生物环境因素对磷酸钙陶瓷表面／界面的形成和表征的影响；进一步通过具有不同表面／界面特征的磷酸钙陶瓷对蛋白质和骨组织细胞黏附和细胞分化、增殖的体外实验，对影响材料生物学性能的表面／界面特征进行了初步探讨；最后，通过体内实验，验证和修正体外实验结果，并通过综合分析得出结论。 体外模拟实验研究发现，在水溶液体系中，几乎所有Ca-P陶瓷通过溶解——沉积反应均可在其表面生成二次结晶的磷灰石层，但形成磷灰石表面层的能力和表征强烈受到材料组成、结构和溶液介质组成的影响。微环境中一定浓度的钙离子、磷酸根离子和碳酸根离子的存在是形成碳酸化羟基磷灰石表面层的必要条件。与此同时，溶液中的CO₃^2-离子及其他Na、Mg等离子亦可随Ca²⁺、PO₄^3-的沉积进入磷灰石晶格，导致含CO₃^2-等离子的结晶不完善的磷灰石层形成，这种磷灰石层正好与骨磷灰石的组成和结构相对应，通常称之为类骨磷灰石。影响Ca-P材料形成类骨磷灰石层能力的关键因素是材料的溶解性，并与其相组成、多孔结构及显微结构及结晶完整性密切相关。几种多孔磷酸钙陶瓷形成类骨磷灰石层的能力排序为：HA／TCP＞烧结不完善的HA＞高温烧结的HA，与CaP陶瓷骨诱导能力正好对应。体内外研究亦发现，溶液介质组成及材料组成和结构等因素强烈影响其表面类骨磷灰石层的特征。蛋白质等有机大分子可增进磷灰石晶体成核并抑制其过度生长，所形成的磷灰石层，以蛋白质连接HA微晶构成的网状层为特征，与之对比，在无蛋白质及有机大分子的溶液介质中形成的磷灰石层以片状磷灰石结晶为特点。前者更近于生物矿化的骨的特征。 体外蛋白吸附，骨组织细胞培养及体内实验表明，由结晶不完善的HA微晶和有机大分子参与和调制形成的网状结构的磷灰石层可有效地锚附蛋白、细胞并为细胞分化和增殖以及骨组织形成提供适宜的条件；与之对比，无蛋白质等有机大分子参与形成的片状磷灰石结晶形成的磷灰石层，即使其组成结构类似于骨磷灰石，仍可被蛋臼和细胞识别出为非骨基质。前者表征着与材料生物学性能有关，或有利于增进材料生物学性能，特别是骨诱导性的特征，在此种意义上，准确的类骨磷灰石层可定义为：由蛋白质等有机大分子参与和调制形成的碳酸化的非化学计量的类似网状结构的磷灰石层。 进一步的体内实验研究证明，具有上述特征的类骨磷灰石层的多孔磷酸钙陶瓷及可于体内形成类骨磷灰石层的多孔CaF陶瓷具有明显的诱导成骨作用，反之则不表现出或仅微弱地表现出骨诱导作用。结合以前研究结果，可以得出：具有一定特征的类骨磷灰石层的形成是诱发CaI生物材料骨诱导性的一个必要条件。更多还原

【Abstract】 The basic principle for the design of modern biomaterials is to take advantage of the body’s natural ability to heal, especially to endow biomaterials with the biological function to induce tissue or organ regeneration. As the main inorganic composition of body’s hard tissue, calcium phosphate biomaterials have excellent biocompatibility and the ability bonding with bone after implantation and are not only osteoconductivity, but also osteoinductivity.Weather the non-living biomaterials has the ability to induce bone formation has not be accepted as a general phnomena; one of the key reason is that the mechanisum of osteoinduction is still unclear. After implantation in vivo, the reaction between the materials and the doner is happenend fisrt at the surface/interface, so the properties of surce/interface are important to the biological property of the materials, including its osteoinductivity. In this thesis we focused on the surface/interface, by the research of the formation and the characterilization of the surface/interface of calcium phosphate biomaterials and the related affecting factors, including the micro-biological environment and the materials related factors, to discuss the relationship between the surface/interface characteristics and the osteoinduction of calcium phosphate ceramics.It’s a complex course with so much related factors to refer to the interaction between the materials and the doners. To simplify the study model and outstand the infuluences of key factors, this thesis is focuesed on the key materials and bio-micro-envirnment factors, which have effects to the formation and characteristics of calcium phosphate ceramics surface/interface. Secondly, by the study of the effects of calcium phosphate with different surface/interface characteristics to the adhension and difffirenration and prolifferation of bone tissue cells, to study the surface/interface characteristics which is related with biological factors; and lastly, by in vivo study, to correct the results in vitro and give the conclusions.By in vitro biomemetic study, it has been founded that in the aqueous system, almost all of the Ca-P ceramics can form a second nuclean apatite layer by disolution-reprecipitate. The forming ability and the characteristic of the surface apatite are effected by the composition of the materials, micro-structure and thesolutions be used. Calcium and phosphate ions and carbonate ions are necessary for the carbonate apatite formation. At the same time, carbonate, sodium and others ions were incoparated into the apatite to induce the formation of bone like apatite, this kind of apatite is corresponding with the bone apatite. The key factor is the solubility of the ceramics, the phase composition, microstructure and the sintering temperature of the porous ceramics also have strong effects to the ability of bone like apatite formation. The ability of bone-like apatite formation of calcium phosphate ceramics is: HA/TCP> HA sintered imperfectly >HA sintered on high temperature. The order of the ability of bone-like apatite formation is corresponding with the order of osteoinduction of Ca-P ceramics.By the studies in vitro and in vivo, it has been demonstrated that the composition of the media and the composition and the structure of the materials has strongly affect the characteristics of the bone-like apatite layer. The formation of the net-like bone like apatite with the coporation of protein molecules is suitable for the adsorption and attachment and proliferation of the protens and cells. Contraray to that, the plate-like bone like apatite without the coparation of protein molecules, although has almost the same constructure of bone apatite, could still be recoganized as the un-real biomimimal matrix, the ability to protein and cells attachment, proliferation is not so good. Based on the results, the bone like apatite could be denificated as: the nonstomichemical amophase apatite layer formed by incoparation of carbonate and the corporation and manipulation of proteins.By the study in vivo,更多还原