可控缓释硒自固化骨修复材料的制备及其活性机理研究

The application of the special biochemistry function of Selenium in bone repair is a frontier problem in biomaterials research field. According to this problem, the applicants propose controlled synthesis self-harden calcium phosphate bone repair material with Selenium sustained-release ability. The selenium will be sealed in microspheres before introduced to the self-harden α-calcium phosphate cement. The controlled release of selenium will be realized by regulate the doping amount, the diameter and the capsule wall material of the microspheres. Moreover, the selenium releasing mechanics from the compound system and the effect of the loaded selenium on the network forming and chemical bonding of the α-calcium phosphate cement will be researched systematically. Furthermore, the novel materials will be co-cultured with cells and bacteria to investigate their in vitro biological activity, such as osteogenic properties, anti-tumor properties and anti-bacteria properties. Deep analysis will be carried out on the experiment data to explore the relationship between these properties with selenium in the material, and make clear the main factors those effect the biological activity and safety of the composite material, and screen out the optimally controllable material and the safe and effective dose of selenium for further study. This research can provide a novel idea not only for bone repair but also for bone tumor preventive and treatment. Meanwhile the experiments can provide data and theory support for further study of rational and new use of the biochemical characteristics of selenium.

本项目针对将硒元素独特的生物化学功能应用于骨修复领域的前沿问题，提出制备可控缓释活性硒的α-磷酸钙自固化骨修复材料，将硒以微球封装后与可自固化的α-磷酸钙骨水泥复合，通过调控封装微球的量、直径与囊壁材质等方式，实现硒释放的可控。系统研究负载不同形态的硒对磷酸钙骨水泥网络结构形成和化学键合作用的影响机理及硒药物缓释动力学规律。并将材料与细胞、细菌共同培养，探明材料体外成骨性能、抑瘤性能、抗菌性能等生物活性及相互作用机理，明确影响复合硒的磷酸钙自固化骨修材料生物活性和生物安全性的内在要素，筛选获得性能理想的优控材料及硒的安全有效剂量，为进一步实验研究奠定基础。本研究可为临床上骨修复及骨肿瘤的预防和治疗提供一种新思路，同时也可为合理利用硒的生化特性，开辟硒新用途，提供实验依据及理论支持。

在本项目资助下，通过课题组成员的共同努力，进行了载不同形态硒的CS微球材料和可控缓释硒自固化磷酸钙骨修复材料的制备，对材料结构、性能进行测试、表征；以SBF模拟体液为环境，研究了材料的体外缓释硒性能，并以乳腺癌细胞MCF-7、人骨肉瘤细胞MG-63等肿瘤细胞为细胞模型，对这两种材料的体外抗肿瘤活性进行了研究。得到如下结论：.（1）载无机/有机形态硒CS微球的制备，单凝聚法制备的载硒CS微球载药量、包封率偏低、粒径偏大不适合。乳化交联是较理想的制备方法，微球形貌完整光滑球形，Na2SeO3/CS微球平均粒径20μm 、包封率和载药量可控在75.82%-27.54%，和1.90%-9.01% 范围，SeMet/CS微球平均粒径10μm，包封率28.82%-19%、载药量0.49%-2.19%。结构表征微球中包裹了预期硒目标药物。.（2）制得的自固化缓释硒磷酸钙骨修复的复合材料，结构性能表征显示：相对于纯TCP，复合材料的凝固时间符合临床手术要求，固化强度提高，注射性良好，TCP均匀的包裹在载硒CS微球表面并形成了针片状 HA 晶体，微球的添加对复合TCP材料的晶相组分无显著影响。 .（3）载硒CS微球在SBF介质中缓释效应显著，有效缓释时间可达32d,在释药过程中对释放介质的pH影响较小。缓释后，微球的形貌由规则的球形变成了不规则的椭球形，光滑表面变成皱褶粗糙，且微球间的相互粘连增加，证明微球的缓释效应显著。载硒CS微球对人乳腺癌细胞MCF-7具有明显抑制作用，抑制率与缓释硒的浓度呈正比，且在48h-72h时间范围的抑制作用最强。. 载硒自固化磷酸钙骨修复复合材料缓释硒的效应良好，有效缓释达22d，缓释后形成了蜂窝状完善的网状三维立体多孔结构，利于组织细胞和血管及神经的黏附长入。体外细胞实验表明，复合TCP对人乳腺癌细胞MCF-7及MG-63人骨肉瘤细胞增生的抑制作用显著，且对两株细胞增生的抑制性差异不显著。本研究结果通过模型药物缓释硒，避免硒化物在应用中易过量而引发的中毒难题。结果发表在相关杂志上和申请专利。