硼酸盐生物玻璃人工骨诱导人骨髓间充质干细胞成骨的分子调控机制研究

The difficulty of osteomyelitis treatment is to cure the local infection and repair the bone defect at the same time. However,the current releasing system used in clinic cannot meet the demands of osteomyelitis treatment.In our previous studies,we found that bioactive borate glass is an ideal candidate for the antibiotic carrier which can simutiously stop the local infection and repair the bone defect. However, its mechanism of bone induction is still unknown. In our feasibility experiments on mesenchymal stem cells (MSC) derived from human bone marrow, the results indicated that the bioactive borate glass signficiantly increased MSC propagation and osteoblast differentiation. In addition, the expressions of BMP-2 and key genes (Wnt3a, Wnt5a) in Wnt signal pathway were significantly upregulated after the bioactive borate glass treatment. Our hypothesis is that the mechanism of bioactive borate glass on the osteoblast differentiation of MSC may be associated with the BMP and Wnt signaling pathways. In this proposal, we will perform in vivo and in vitro studies to further investigate the signaling mechanism of bioactive borate glass on inducing bone formation in human marrow MSC. Moreover, we will determine the expessions and interactions of key bone inductive factors in the BMP-2 and Wnt signaling pathways. This proposed study will establish a fundament for the material improvement and the clinic transformation of bioactive borate glass.

骨髓炎治疗的难点是抗感染和骨修复。本项目组长期致力于硼酸盐生物玻璃人工骨的研究，发现该材料具有良好的生物相容性、成骨活性、降解可调控及良好的抗生素缓释效果等优点，在治愈骨髓炎的同时促进骨缺损的良好修复。为探讨该材料促进骨修复的机制，我们在预实验中观察了其对人骨髓间充质干细胞（MSC）成骨基因及相关信号分子的影响，结果显示其可促进人骨髓MSC快速增殖并向成骨细胞分化，且BMP-2及Wnt信号通路的成骨关键基因(BMP-2和 Wnt5a)表达明显上调。本课题拟在此研究基础上，通过人骨髓MSC体外及裸鼠体内实验研究硼酸盐生物玻璃诱导人骨髓MSC成骨的分子调控机制。采用重组慢病毒、qRT-PCR、免疫组化等技术，明确该材料通过BMP-2及 Wnt信号通路调控人骨髓MSC成骨的分子靶点及作用途径，以期阐明硼酸盐生物玻璃诱导骨再生的分子调控机制，为材料的改进及临床转化提供理论依据.

大段骨缺损修复一直是临床研究的重点难点。本课题组长期研究发现硼酸盐生物玻璃具有良好的成骨活性，但其对种子细胞成骨分化的分子调控机制尚有待进一步明确。本项目利用新西兰大白兔大段骨缺损模型及硼酸盐浸提液对BMSCs刺激模型，通过生物信息学分析、荧光定量PCR、western blot和抑制剂阻断通路等研究手段系统的探讨硼酸盐生物玻璃对大段骨缺损的修复功能及可能的调控机制。研究结果表明BMP/Smad通路在硼酸盐生物玻璃修复骨缺损中发挥重要调控作用。但在研究中我们同时发现硼酸盐生物玻璃生物毒性问题一直存在争议而难于实施转化应用，我们经过不断探索设计出了新的修复材料即原位光交联水凝胶HA-NB/Gelatin用于骨缺损修复，体外实验发现其具有良好的光成胶性能和生物相容性，BMSCs在其中活性良好，移植体内28天后大部分降解。将HA-NB/Gelatin负载羟基磷灰石后修复大鼠颅骨缺损，通过micro-CT、三色荧光、VG染色证实其修复效果优于单纯HA-NB/Gelatin水凝胶，表明其可作为良好的生物材料用于骨缺损修复，同时具有良好的临床转化前景。