壳聚糖-丝素支架材料的细胞毒性检测

Cytotoxicity test of chitosan-silk fibroin scaffold material to BMSCs in vitro

目的:对壳聚糖-丝素(chitosan-silk fibroin)共混多孔支架体外进行细胞毒性实验,探讨壳聚糖-丝素支架对骨髓间充质干细胞的细胞毒性作用情况,为该材料临床应用提供依据。方法:制备壳聚糖-丝素共混多孔支架,并制备该支架材料的浸提液。通过MTT法检测25%、50%、100%浓度的壳聚糖-丝素浸提液对兔骨髓间充质干细胞(BMSCs)的细胞毒性作用,以第1,3,5,7天为检测时间点,观察细胞生长状况,计算细胞相对增殖率,并对材料的细胞毒性进行分级。以正常培养基为阴性对照组,含0.64%苯酚的培养液为阳性对照组。结果:BMSCs在不同浓度实验组及阴性对照组中生长均好,细胞呈长梭形等形态,细胞形态饱满有光泽,MTT法结果表明同一时间点各浓度实验组与阴性对照组间之间差异均无统计学意义(P>0.05),各时间点各浓度浸提液对BMSCs细胞的相对增殖率均在95%以上,毒性分级为0级或1级,基本上无毒性。结论:壳聚糖-丝素多孔支架材料细胞相容性良好,基本上无细胞毒性,符合生物材料细胞毒性试验的安全标准,是一种很有潜力的组织工程支架材料。

Objective：To investigate cytotoxicity of chitosan-silk fibroin blend porous scaffold material to BMSCs in vitro.Methods： Chitosan was mixed with silk fibroin at ratio of 6∶4,then the mixture was prepared for chitosan-silk fibroin porous scaffold by freeze drying.Cell observation method and MTT assay were used to detect the cytotoxicity of chitosan-silk fibroin porous scaffold extract fluid to bone marrow-derived mesenchymal stem cells（BMSCs）.The cytotoxicity of the scaffold extract fluid at concentrations of 25%,50%,and 100% to BMSCs was detect by MTT assay.The relative growth rate was calculated and the cytotoxicity was graded at day 1,3,5,and 7.Normal culture media was considered as negative control group,and 0.64% phenol solution was considered as positive control group.Results： BMSCs grew well in the experimental groups and negative control group and there was no significant difference in cell morphology between these groups.The relative growth rates of the experimental groups were all over 95%,and the cytotoxicity grade was 0 or 1 level at day 1,3,5,and 7.The chitosan-silk fibroin porous scaffold is basically non-toxic to BMSCs.Conclusion： The chitosan-silk fibroin scaffold material has a good cytocompatibility.The cytotoxicity of the scaffold material satisfies the safety standard.It is a potential scaffold material for tissue engineering.