New types of chitosan-gelatin-hyaluronate-heparan sulfate (C-G-Ha-HS) hybrid three-dimensional scaffolds were fabricated by lyophilization method and their physical properties including the pore size, porosity, water absorption and degradation of the scaffolds were determined. Meanwhile, cell adhesion behavior, the cell status and viability as well as the biocompatibility in the fabricated scaffolds were detected by inoculating the neural stem/progenitor cells (NS/PCs) into the scaffold, and the results were compared with those of the cells inoculating into the chitosan-gelatin (C-G) scaffold. By the SEM observation and CCK-8 assay, it was found that the pore size of C-G-Ha-Hs scaffolds mainly range from 90 μm to 130nm, porosity increases significantly, water absorption rate are above 95% and the scaffolds can degrade in vitro. C-G-Ha-HS scaffolds significantly improve the adhesion ratio and proliferation of NS/PCs on them and the cells in the scaffold can connect into neural networks. Compared with C-G scaffold, C-G-Ha-HS scaffolds (5:5) obviously enhance the NS/PCs adhesion ratio by (200.8±8.0)% and effectively increase the cell viability by (357.7±8.9)% after cell culture of 6 days. The results show that the C-G-Ha-Hs scaffolds are the suitable material used to construct the three-dimensional environment for the culture of NS/PCs in vitro, and can be used for tissue engineering as well as drug screening.%采用冷冻干燥法制备一种新型三维壳聚糖-明胶-透明质酸-硫酸肝素(chitosan-gelatin-hyaluronate-heparan sulfate,C-G-Ha-HS)复合支架,分别测定支架的孔径、孔隙率、吸水率和降解率等物理性质,同时以壳聚糖-明胶(chitosan-gelatin,C-G)支架作为对照组,考察了神经干/祖细胞(neural stem/progenitor cells,NS/PCs)在该支架上的粘附率、生长状态以及细胞生存力等生物相容性指标.扫描电镜观察及CCK-8分析表明,C-G-Ha-HS支架孔径主要分布于90～130μm,孔隙率明显增大,吸水率大于95％且能够体外降解； C-G-Ha-HS支架明显改善了NS/PCs的粘附性及生存力,细胞在支架内相互交织成网状,C-G-Ha-HS支架(5:5)上的细胞粘附率提高至对照的(200.8±8.0)％,培养6天以后,细胞的存活率增加至对照的(357.7±8.9)％.这些结果初步显示,C-G-Ha-HS支架可以用于构建体外NS/PCs的三维培养,为进一步应用于神经组织工程和药物筛选提供了实验依据.
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