适合鸭瘟病毒弱毒增殖的MHC单倍型鸭胚肝间充质干细胞的建立

为了建立适于鸭瘟病毒(DPV)弱毒增殖的细胞系,利用4个主要组织相容性复合体(MHC)单倍型鸭品系,体外分离鸭胚肝细胞,在干细胞因子、白细胞抑制因子和成纤维细胞生长因子诱导下,获得鸭胚肝间充质干细胞(DuLMSC)。接种DPV疫苗株C-KCE后,结果显示:F1~F10代均能稳定出现CPE;将F10病毒接种细胞24 h后,通过扫描电镜在胞质、胞核及细胞间隙均能够观察到典型的疱疹病毒粒子;F1至F10代均能扩增出DPV UL2基因片段,序列测定结果表明F1、F5、F10代的扩增产物与NCBI中公布的DPV株(EU082088)的核苷酸同源性为100%;组织培养半数感染量和一步生长曲线结果表明,源自B4系单倍型鸭的DuLMSC细胞系增殖DPV的病毒含量明显高于B1、B2和B3系。本研究为稳定培养鸭瘟病毒提供了优良稳定的实验材料。

Establishment of MHC haplotype duck embryo liver mesenchymal stem cells for weak duck plague virus proliferation

In order to establish a proper cell line for producing weak duck plague virus(DPV),duck embryonic liver mesenchymal stem cells(Du LMSC)were isolated from 4 major histocompatibility complex duck haplotypes,respectively under SCF,LIF and b FGF.The Du LMSCs were infected with DPV vaccine strain of C-KCE.The results showed that,obvious cytopathic effects were stably observed from F1 through F10;typical herpesvirus particles were discerned at cytoplasm,nucleus and intercellular space at 24 h post infection using F10 cells;DPV UL2 genomic fragment could be PCR amplified from F1 through F10,and the nucleotide sequences of F1,F5 and F10 were identical to NCBI reference strain of EU082088;content of virus in B4 haplotype Du LMSC was higher than the other B1,B2 and B3 cell lines conformed by TCID50 from F1 to F10 and one-step growth curve detection.The study provided a good support cell line for propagating DPV vaccine in vitro.