基于软骨细胞外基质的取向支架的制备及评价

Development and characterization of oriented scaffolds derived from cartilage extracellular matrix

目的制备软骨细胞外基质(CECM)取向多孔支架,评价其理化特性及对脂肪干细胞(ADSCs)的相容性。方法切取新鲜猪关节软骨片,匀浆后离心筛选出直径50~500 nm左右的软骨纤维,经Triton X-100脱细胞后制备成浓度6%的CECM浆料,通过定向结晶和冷冻干燥后交联即得CECM取向支架。对CECM取向支架的理化性能和细胞相容性进行评价。结果 CECM取向支架横断面为均匀的多孔网状结构,孔壁上密布软骨纤维,纵断面为垂直管状结构;支架呈苏木精-伊红红染,甲苯胺蓝、番红O、天狼星红染色均呈阳性;支架孔隙率、吸水率和纵向压缩弹性模量分别为95.455%±0.910%、95.889%±1.071%和(40.208±5.097)k Pa;ADSCs在支架上黏附和增殖,并均匀长入支架孔隙内。结论 CECM取向支架的成分与天然软骨相似,生物相容性良好,孔隙结构、孔径大小适于种子细胞黏附、增殖和长入,是一种比较理想的软骨组织工程支架。

Objective This study aimed to prepare oriented scaffolds derived from a cartilage extracellular matrix （CECM） and to investigate their physicochemical property and compatibility with adipose-derived stem cells （ADSCs）. Methods A fresh porcine articular cartilage was cut into pieces. Cartilage nanofibers with diameters of 50-500 nm were collected through homogenization and centrifugation. These nanofibers were then decellularized by using Triton X-100 to produce 6% CECM. The oriented scaffolds derived from the nanoscale CECM were fabricated through unidirectional solidification and lyophili- zation. Afterward, these scaffolds were crosslinked. The physical and chemical performances and cell compatibility of CECM- oriented scaffolds were evaluated. Results The cross-sections of the scaffolds contained homogeneous reticular porous structures with nanofibers on the walls of the pores, and the longitudinal sections revealed vertical tubular structures. Hematoxylineosin staining revealed that the scaffolds were red without blue. Toluidine blue, safranin O, and Sirius red staining showed positive results. The porosity, water absorption rate, and vertical compressive elastic modulus of the scaffolds were 95.455%± 0.910%, 95.889%±1.071%, and （40.208±5.097） kPa, respectively. Conclusion The components of the oriented scaffolds derived from CECM are similar to those of native cartilage with favorable biocompatibility. The porous structures and sizes of the scaffolds are suitable for the adhesion, proliferation, and infiltration of ADSCs. The oriented scaffolds derived from CECM are relatively optimal for cartilage tissue engineering .