稳定表达人白细胞介素2肝干细胞系的建立  

Establishment of a hepatic stem cell line stably expressing human interleukin-2

作　　者：郑和鸣[1] 蔡秀军[1] 

机构地区：[1]浙江大学医学院附属邵逸夫医院普外科,杭州310016

出　　处：《中国免疫学杂志》2010年第4期341-344,共4页Chinese Journal of Immunology

基　　金：浙江省自然科学基金资助项目(No.Y207078);浙江省自然科学基金资助项目(No.R205075)

摘　　要：目的:通过逆转录病毒介导,建立能稳定高效表达白介素2的肝干细胞WB-F344。方法:构建逆转录病毒载体Plpcx-IL-2质粒,转染包装细胞PT67,收集细胞培养上清,检测病毒的滴度。将高滴度的PT67/Plpcx-IL-2病毒液感染肝干细胞WB-F344,用嘌呤霉素加压筛选出抗性克隆,并用RT-PCR、间接免疫荧光实验、免疫组化实验和Western blot实验证实抗性细胞在转录水平和蛋白水平均有白介素2的表达情况。结果:成功构建含白介素2的逆转录病毒载体,获得滴度为105CFU/ml的感染性病毒液。建立的细胞系在转录水平和蛋白水平均有白介素2的表达。结论:成功建立了稳定表达人白细胞介素2的肝干细胞系WB-F344/Plpcx-IL-2,为下一步开展研究以肝干细胞为载体肝癌免疫基因治疗奠定了基础。Objective：Establishing the hepatic stem cell（WB-F344） line that expresses human interleukin-2 stably and effectively.Methods：We constructed the retroviral vector Plpcx-IL-2 containing human interleukin2 gene,and transfected it into PT67 packaging cells.The infectious viruses were used to infect the hepatic stem cell WB-F344,then the puromycin resistant clones were acquired after puromycin selection.The expression of IL-2 was detected using RT-PCR,immunocytochemistry,Western blot and indirect immunofluorescence,respectively.Results：The retroviral vector containing human IL-2 gene was constructed.The titer of virus in the supernatant reached 105 CFU/ml after transfecting PT67 packeting cells.The puromycin resistant clones were acquired after infection of WB-F344 and the cell line WB-F344/Plpcx-IL-2 expressing human interleukin-2 stably was established.Conclusion：By retroviral-mediated transfecting human interleukin-2 gene,the hepatic stem cell stably and effectively expressing interleukin-2 are obtained.This provides a good basis for further research on immuno-gene therapy of liver cancer.

关 键 词：人白细胞介素2 WB-F344细胞 稳定表达 

分 类 号：R392[医药卫生—免疫学]