Bini Elisabetta, Foo Cheryl Wong Po, Huang Jia, Karageorgiou Vassilis, Kitchel Brandon, Kaplan David L
Department of Biomedical Engineering, Department of Chemistry, Bioengineering and Biotechnology Center, Tufts University, 4 Colby Street, Medford, Massachusetts 02155, USA.
Biomacromolecules. 2006 Nov;7(11):3139-45. doi: 10.1021/bm0607877.
Spider silk fibers have remarkable mechanical properties that suggest the component proteins could be useful biopolymers for fabricating biomaterial scaffolds for tissue formation. Two bioengineered protein variants from the consensus sequence of the major component of dragline silk from Nephila clavipes were cloned and expressed to include RGD cell-binding domains. The engineered silks were characterized by CD and FTIR and showed structural transitions from random coil to insoluble beta-sheet upon treatment with methanol. The recombinant proteins were processed into films and fibers and successfully used as biomaterial matrixes to culture human bone marrow stromal cells induced to differentiate into bone-like tissue upon addition of osteogenic stimulants. The recombinant spider silk and the recombinant spider silk with RGD encoded into the protein both supported enhanced the differentiation of human bone marrow derived mesenchymal stem cells (hMSCs) to osteogenic outcomes when compared to tissue culture plastic. The recombinant spider silk protein without the RGD displayed enhanced bone related outcomes, measured by calcium deposition, when compared to the same protein with RGD. Based on comparisons to our prior studies with silkworm silks and RGD modifications, the current results illustrate the potential to bioengineer spider silk proteins into new biomaterial matrixes, while also highlighting the importance of subtle differences in silk sources and modes of presentation of RGD to cells in terms of tissue-specific outcomes.
蜘蛛丝纤维具有卓越的机械性能，这表明其组成蛋白可能是用于制造组织形成生物材料支架的有用生物聚合物。从金蛛（Nephila clavipes）拖牵丝主要成分的共有序列中克隆并表达了两种具有RGD细胞结合域的生物工程蛋白变体。通过圆二色光谱（CD）和傅里叶变换红外光谱（FTIR）对工程化蜘蛛丝进行了表征，结果显示在用甲醇处理后，其结构从无规卷曲转变为不溶性β-折叠。重组蛋白被加工成薄膜和纤维，并成功用作生物材料基质来培养人骨髓基质细胞，在添加成骨刺激剂后，这些细胞被诱导分化为类骨组织。与组织培养塑料相比，重组蜘蛛丝和编码有RGD的重组蜘蛛丝均能促进人骨髓间充质干细胞（hMSCs）向成骨方向分化。与含有RGD的相同蛋白相比，不含RGD的重组蜘蛛丝蛋白在钙沉积方面显示出增强的骨相关结果。基于与我们之前对蚕丝和RGD修饰的研究比较，当前结果说明了将蜘蛛丝蛋白生物工程改造为新型生物材料基质的潜力，同时也突出了丝源的细微差异以及RGD向细胞呈现方式在组织特异性结果方面的重要性。
