丙型肝炎病毒JFH1在不同三维培养体系中复制水平的比较

目的：检测中空纤维丝、热可逆凝胶和低吸附培养板3种三维培养体系中丙型肝炎病毒（HCV）增殖水平，寻找效率较高的HCV体外培养系统。方法：利用中空纤维丝、热可逆凝胶及低吸附培养板培养永生化人肝细胞HuS-E/2，通过XTT法检测细胞增殖情况。以基因型为2a型的HCV病毒株JFH1进行感染，在感染后不同时间点收集细胞标本，利用实时PCR法检测细胞中HCV的滴度。结果：在中空纤维丝和热可逆凝胶中培养的HuS-E/2细胞增殖曲线为持续递增，至培养结束时细胞数为初始培养数的2～3倍，JFH1感染后中空纤维丝方法培养的细胞中HCV滴度为2.00×10³ copies/1 μg total RNA，热可逆凝胶培养的细胞中HCV滴度为2.95×103 copies/1 μg total RNA,低吸附培养板方法培养的细胞中HCV滴度为1.18×10³ copies/1μg total RNA。结论：与中空纤维丝及低吸附培养板比较，热可逆凝胶法为基础的JFH1体外培养系统细胞生长稳定，HCV复制效率较高。

Comparison of HCV replication levels in different threedimensional culture systems

Objective To compare the replication efficiency of HCV in different three-dimensional(3-D) culture systems for a better choice of HCV culture system in vitro.Methods The human immortalized hepatocytes HuS-E/2 cells were cultivated in hollow fiber,mebiol gel and hydrocell plate.The cells were incubated with the medium containing genotype 2a HCV JFH1 for infection.The cell proliferation was monitored with XTT assay.HCV amplification in 3-D cultured cells was measured with real time PCR.Results The cell proliferation were continuously increased in hollow fiber system and mebiol gel system.The cell number was 2-3 times increased at the end of culture.After genotype 2a HCV-JFH1 infection,the titers of HCV were 2.95×103 copies/1 μg total RNA,2×103 copies/1 μg total RNA,and 1.18×103 copies/1 μg total RNA in mebiol gel,hollow fiber and hydrocell plate respectively.Conclusion Compared with hollow fiber culture system and hydrocell plate culture system,the cell proliferation is stable and the HCV replication is more efficient in the 3-D culture system for HCV based on mebiol gel system.