摘要/Abstract
众多研究表明，含有三肽基序即精氨酸-甘氨酸-天冬氨酸（RGD）的材料有助于细胞在其上的粘附和生长.尽管作为天然蛋白质的柞蚕丝蛋白同样具有RGD序列，但其从柞蚕丝纤维中的再生过程复杂且不易加工成型，因此对此类蛋白质材料的研究较少.我们以水溶液共混的方式，在再生柞蚕丝蛋白中引入了更为常用的桑蚕丝蛋白，通过对桑蚕丝蛋白水溶液特殊有效的醇诱导和冻融方法，成功制备了非水溶性的柞蚕丝蛋白/桑蚕丝蛋白复合支架材料.研究结果显示，随着柞蚕丝蛋白含量增加，共混支架的孔径和强度均有所下降，但其热稳定性上升；FTIR测试表明，尽管柞蚕丝蛋白和桑蚕丝蛋白的一级结构完全不同，但其二级结构在以此方法制备的支架材料中均以β-折叠构象为主.体外细胞培养实验证实，小鼠成纤维细胞和成骨细胞都能在支架上增殖生长，并且共混支架较纯桑蚕丝蛋白支架具有更高的细胞增殖速率，表明其不仅具有良好的生物相容性，而且可能更适用于组织工程领域.
关键词: 动物丝蛋白, 氨基酸序列, 组织工程, 相容性, 力学性能
Lots of research has indicated that materials contain Arg-Gly-Asp (RGD) sequence can promote cell attachment and proliferation on them. Although Antheraea pernyi silk fibroin is a natural structural protein which contains RGD sequence, there are few studies on this kind of protein materials, for the regeneration of Antheraea pernyi silk fibroin from silk fibers is complicated and it is hard to be processed. In this paper, we present a water-insoluble Antheraea pernyi/Bombyx mori silk fibroin blending scaffold. The regenerated Antheraea pernyi silk fibroin (RASF) solution was prepared by dissolving degummed silk fibers at 100℃ and dialyzing at 4℃. The regenerated Bombyx mori silk fibroin (RBSF) solution was prepared by dissolving degummed silk fibers at 60℃ and dialyzing at 20℃. Regenerated silk fibroin solution was concentrated to 6 wt% solution in 10 wt% PEG solution. Based on RASF and RBSF solution, RBSF porous scaffold and RASF/RBSF blending scaffolds with different ratios were prepared through treating 1-butanol/SF solution under freezing at -20℃. The volume ratio of 1-butanol to solution was 1:2. RASF porous scaffold was not hard enough to hold itself, therefore the maximum content of RASF in blending scaffold was 70 wt%. With increasing of RASF content, pore sizes of scaffolds decreased from 250 μm to 150 μm and compressive strengths decreased from 280 kPa to 108 kPa, while the thermal stabilities increased. FTIR results demonstrated that the molecular conformation of silk fibroin was proven to be β-sheet, β-turn and α-helix. The biocompatibilities of scaffolds were demonstrated with in vitro cell culture. The results showed that L929 fibroblast and MC3t3-E1 osteoblast adhered, proliferated and migrated well into the scaffolds. The speed of cell proliferation accelerated with the increase of RASF content. Obviously, these regenerated silk fibroin scaffolds with good bio-compatibility could be used in tissue engineering field further.
Key words: animal silk protein, sequence, tissue engineering, compatibility, mechanical properties
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