骺板细胞外基质源性仿生支架的制备及性能评估

Preparation and performance evaluation of a biomimetic scaffold derived from the extracellular matrix of a native growth plate

背景:骺板细胞外基质具有丰富的胶原、蛋白多糖及信号分子,其成分和特征最接近天然骺板组织,是构建织工程骺板的最理想原料之一。目的:采用骺板细胞外基质构建取向柱状仿生支架材料,并对其进行性能评估。方法:取胎牛膝关节标本,采用超湿法粉碎、差速离心及有机溶剂分离等技术联合处理获得细胞外基质微丝,利用定向结晶、冷冻干燥技术及物理/化学交联方法制备出骺板细胞外基质源性仿生柱状支架材料,扫描电镜观察支架的内部结构及孔径,检测支架的孔隙率及吸水膨胀率,同时对支架进行组织学观察。结果与结论:(1)苏木精-伊红染色显示,细胞外基质微丝呈纤维状,形状规则,均匀一致;甲苯胺蓝、番红O及Ⅱ型胶原免疫组织化学染色呈阳性,说明保留了天然骺板细胞外基质的有效成分;(2)显微镜显示支架横截面呈多孔蜂窝状结构,孔径均匀一致,形状规则,纵切面显示平行排列的柱状结构,均匀一致,孔径分布均匀,相互连通好,支架外壁纤维呈纵向取向排列,类似于天然骺板软骨的柱状分布结构。扫描电镜显示支架横切面呈多孔蜂窝状,均匀分布,形状规则,支架的横向孔径为(117.5±15.37)μm。支架孔隙率为(92.05±1.54)%,吸水膨胀率为(95.95±1.07)%;(3)结果表明,骺板细胞外基质源性仿生柱状组织工程支架材料在成分来源和结构特征方面均达到了模仿天然骺板软骨的特点。

BACKGROUND： The extracellular matrix from the growth plate contains abundant sources of collagens,proteoglycan and signaling molecules. These components and characteristic are very similar to the natural growth plate tissue, and therefore, it is an ideal material to construct tissue-engineered growth plate. OBJECTIVE： To prepare the columnar biomimetic scaffold using native growth plate extracellular matrix and to evaluate the scaffold performance. METHODS： The knee joint of fetal bovine was separated and purified by pulverization technology in moist environment, differential centrifugation and organic solvent to obtain extracellular matrix microfilaments. Then the extracellular matrix microfilaments were treated by directed crystallization technology, freeze drying and physicochemistry crosslinking method in order to prepare the extracellular matrix biomimetic scaffold. The scaffold was evaluated by scanning electron microscope for internal structure and pore size, porosity and swelling rate of water absorption as well as by histological observation. RESULTS AND CONCLUSION： The extracellular matrix microfilaments were fibriform with the even and regular shape shown by hematoxylin-eosin staining, and were also positive for toluidine blue, Sarranine O and collagen II staining, indicating the existence of active components of the growth plate in the scaffold.（2） The scaffold under the scanning electron microscope showed porous honeycomb structure on the transverse section plane with uniform pore size and regular shape, while there was a parallel arrangement of columnar structure on the longitudinal section plane of the scaffold with well-distributed and mutual communication, and scaffold fibers on the outer wall were longitudinally arranged, which imitated the columnar structure from the native growth plate. The pore diameter of scaffold was（117.5±15.37） μm, the porosity was（92.05±1.54）% and the water absorption was（95.95±1.07）%. To conclude, the columnar scaffold from the extracellular matrix of the growth plate imitates the native growth plate in terms of components and structures.