一种仿生修饰分子rFN／CAD的设计、制备及初步应用

Design,preparation and preliminary use of a novel biomimetic agent rFN/CAD

目的制备一种具有黏附和成骨双重功能的仿生分子,为种子细胞在支架材料界面的高效黏附和靶向分化提供新途径。方法筛选纤维连接蛋白和钙黏附蛋白前体分子,通过分子模拟、基因克隆、蛋白表达纯化等技术设计并进行制备。通过离心促黏附实验和成骨诱导分化实验检测其体外活性。结果成功获得长1954bp的FNⅢ7-10/CAD11 EC1-2重组基因和由641个氨基酸组成的融合蛋白。体外功能研究发现聚苯乙烯表面的融合蛋白涂层可显著提高人骨髓间质干细胞的黏附和成骨分化能力。结论 rFN/CAD具有促黏附和促成骨的双重生物活性,可作为一种理想的仿生修饰分子用于生物界面的构建。

Objective To prepare a novel biomimetic agent with adhesive and osteoinduction capabilities, and provide a new approach for improving seeding cell adhesion and osteoblastic differentiation on a scaffold surface. Methods A novel bi- functional recombinant protein was engineered and prepared by bioinformatics, standard molecular cloning, protein expression and purification techniques. A centrifugal cell adhesive assay and an osteoinduction assay were employed to determine the biofunction ex vivo. Results A 1954bp long DNA chimera of FN Ⅲ7-10/CADll EC1-2 and recombinant FN Ⅲ7-10/CADll EC1-2 of 641 amino acids were obtained. Polystyrene surface coated with this protein significantly increased the adhesive property and osteodifferentiation of human bone marrow mesenchymal stem cells. Conclusion rFN/CAD possesses dual biofunctions of osteoblastic adhesion and differentiation, which can be further applied as an ideal biomimetic molecule for surface modification.