锶硼玻璃结构与形成层状HA的关联及其诱导骨再生过程

The strontium-contained borate glass scaffolds with the porosity of 60 % and the compressive strength of 30 M Pa were fabricated in our early work, which could repair the big volume of segmental defects in the tibia bone of rabbits. These scaffolds showed an excellent performance in this animal experiment. But the mechanism for the glass transformation into bone tissue is still unknown. In this project, the relationship between the glass structure and its biological properties will be explored through the four steps: 1) The regulation for the formation of multi-layered HA deposited from the borate glass will be characterized during the bio-mineralization process by testing the glass network structure, which is composed of 3-fold coordination of boron group unit; 2) The effect of pH value of the interface of the glass and HA at the micro-environment on the differentiation and proliferation of osteoblasts will be monitored in-real time by measuring the cells attached on the HA surface using micro-electrodes; 3) The forming process of the composite of multi-layer HA and multi-layer collagen secreted from these cells will be observed in real time by an active cell monitored electric dynamical microscope; 4) The gene expression level during cell differentiation, and the stimulation of B and Sr to bone tissue regeneration will be analyzed by gene group technology, quantitative polymerase chain reaction (PCR) and immunofluorescence staining chemistry. From the above study on the mechanism for transformation of the glass into bone tissue, not only the bone tissue regeneration process stimulated by B and Sr will be revealed molecular-biologically, but also the third-generation biomaterial products with self-intellectual property rights will be developed, based on these theoretical and practical achievements of the design and research on bioactive borate glass.

项目组前期制备了孔隙率为60％，抗压强度达30兆帕的含锶硼酸盐玻璃支架，在修复大面积节段性骨缺损的动物实验中，取得了良好的效果。但是对锶硼玻璃能转化为骨组织的机理不甚明嘹。为探索玻璃结构与其生物性能的关联：1）表征硼酸盐玻璃网络结构中三配位硼基团单元在生物矿化时形成层状羟基磷灰石（HA）规律；2）采用微pH计检测在粘附HA表面上的成骨细胞酸碱性，研究固体表面微环境的pH值对成骨细胞分化增殖的影响；3）采用实时活细胞电动显微镜，观察成骨细胞粘附HA表面后，形成层叠HA与层叠细胞复合的过程；4）采用基因组技术，定量PCR、免疫荧光染色，分析锶、硼在成骨细胞分化时基因表达水平变化，分析锶、硼在骨再生过程的刺激作用。通过对玻璃转化为骨组织的机理研究，不仅揭示含锶硼酸盐玻璃诱导骨再生的分子机制，而且为设计、开发新型的硼酸盐玻璃产品提供理论和实践依据，为研制具有自主知识产权的第三代骨修复产品指明方向。

大面积节段性骨缺损发生后很难实现有效的修复，是骨科临床亟需解决的关键技术之一。基于生物学和工程学原理和方法形成的组织工程学为解决类似的问题提供了科学依据和技术支撑，然而现有的一些骨组织工程支架材料不能完全满足临床需要。这是因为作为组织工程的支架材料，除应该具备良好的可降解性和相容性之外，还应该具有促进生物组织（包括骨髓间充质干细胞诱导成骨分化，成骨细胞增殖、粘附等）快速生长的特性。能够随着植入体内的逐步降解，形成新的与自身功能和形态相适应的组织或器官，从而达到再生或修复病损、缺损组织或器官的目的。相关的研究是当前生物医用材料工程领域的研究热点和难点之一。.本项目的前期研究表明，含锶硼酸盐玻璃不但具有良好的生物活性、相容性和生物降解性，而且还有良好的促成骨作用。若将其制备成具有一定形状和三维连通结构的多孔细胞支架,还具有良好的力学性能和理想的孔结构，在修复大面积节段性骨缺损的动物实验取得了良好的效果。但是对锶或其它无机离子对硼玻璃能够转化为骨组织的机理及作用不够明瞭，对其生物矿化为层状羟基磷灰石的规律、原理和有关材料表面状态、微环境的pH值等对成骨细胞分化增殖的影响也不够清晰。.本项目通过材料性能表征、细胞及动物实验、基因组技术等手段，探索了锶硼玻璃结构与形成层状HA的关联及其诱导骨再生过程机理。具体解释了1）硼酸盐玻璃在生物矿化时形成层状HA的材料化学问题及微环境问题。揭示了硼酸盐玻璃的组成、形成层状HA及固体表面与细胞应答反应和最终形成骨板组织等一系列理化性能和生化现象的内在联系。2）分析和理清了锶离子、铜离子等在促成骨细胞增殖和促血管形成方面的机理及基因表达水平变化等生物学上的科学问题。更为重要的是本项目的完成为硼酸盐玻璃在生物医学工程领域中的应用提供新思路，为新一代骨组织工程支架材料的开发提供了理论基础和实践依据，也为研制具有自主知识产权的新型骨组织工程材料提供了思路和方向。