材料骨组织诱导中的蛋白质组学研究

The approach, in which bone defects are repaired with autologous bone generated in ectopic sites (i.e under skin or in muscle) using osteoinductive materials, is promising near future. Optimization of osteoinductive materials and understanding of the mechanism of material-induced bone formation are crucial in such an approach for bone regeneration. The critical role of surface microstructures in material-induced bone formation has been shown in previous studies. It is assumed that the surface microstructure increases surface area to concentrate higher amount of proteins (i.e growth factors)　as such to differentiate mesenchymal stem cells to osteogenic cells to form bone. While which proteins adsorbed onto the material surface responsible for osteoinduction and which specific proteins are expressed in material-induced bone formation are not know as yet. In this project, for the first time, the comparative proteomics research strategy is used to analyze 1) protein adsorption onto osteoinductive materials with various chemistry (i.e calciul phosphate ceramics and non-calcium phosphate ceramics such as titanium oxide ceramic and aluminum ceramic) from body fluid in vivo and simulated body fluid in vitro, 2) the protein expression of bone marrow stromal cells cultured on the osteoinductive materials and 3) protein expression in ectopic bone formation induced by osteoinductive materials as compared to BMP_induced bone formation. Two-dimensional electrophoresis and multidimensional chromatography are the techniques to separation proteins, while MALDI-TOF-MS is applied to identify the proteins. The potential difference in protein adsorption and protein expression will outline us the specific protein adsorption responsible for material-induced bone formation and specific protein expressions in material-induced bone formation. The results will finally explain the possible mechanisms of material-induced bone formation and pave a way for the clinical application of osteoinductive materials for bone regeneration.

应用骨组织诱导材料不外加细胞或生长因子在体内构建类似自体骨的移植物修复颌面骨缺损并支持牙种植体重建咬合具有切实可行的应用前景，骨诱导材料的优化及骨诱导相关机制的深入研究是其广泛应用的基础。前期研究表明材料微结构是启动骨组织诱导发生的关键因素，而材料微结构是直接诱导干细胞骨向分化，还是先聚集骨形成相关蛋白再诱导干细胞骨向分化尚不清楚；材料骨诱导过程中是否存在起关键作用的特异性蛋白质亦不明确。本项目应用比较蛋白质组学的研究策略，分别分析骨诱导材料和非骨诱导材料在体内外对蛋白质吸附的差异，及对干细胞骨向分化的相关蛋白表达的差异，以双向电泳及多维色谱分离蛋白质后用MALDI-TOF-MS质谱检测，结合生物信息学数据库对实验结果进行解析，并对有意义的差异蛋白进一步验证。本项目有望筛选出材料骨组织诱导过程中的特异性蛋白分子，回答材料诱导干细胞骨向分化的机制，为进一步骨诱导材料优化及临床应用提供依据。

应用骨组织诱导材料不外加细胞或生长因子在体内构建类似自体骨的移植物修复颌面骨缺损并支持牙种植体重建咬合具有切实可行的应用前景，骨诱导材料的优化及骨诱导相关机制的深入研究是其广泛应用的基础。前期研究表明材料微结构是启动骨组织诱导发生的关键因素，而材料微结构是直接诱导干细胞骨向分化，还是先聚集骨形成相关蛋白再诱导干细胞骨向分化尚不清楚；材料骨诱导过程中是否存在起关键作用的特异性蛋白质亦不明确。本项目应用比较蛋白质组学的研究策略，选用基于质谱的蛋白质非标记定量技术（Label-free quantitation）分别分析骨诱导材料和非骨诱导材料在体外对蛋白质吸附的差异，结合生物信息学数据库对实验结果进行解析，并对有意义的差异蛋白进行定性和定量分析。探究材料骨组织诱导过程中的特异性蛋白分子，回答材料诱导干细胞骨向分化的机制，为进一步骨诱导材料优化及临床应用提供依据。