HLA-E?????????????幹??????????HepG2????е???

目的 构建表达人类白细胞抗原-E(HLA-E)基因慢病毒载体,探讨慢病毒介导HLA-E基因在肿瘤免疫中的意义.方法 2006年10月至2007年11月在中山大学附属第三医院肝移植中心应用pGC-El-GFP-HLA-E慢病毒载体转导肝癌HepG2细胞,检测慢病毒载体的转导效率、细胞内HLA-E信使核糖核酸(mRNA)及蛋白质水平的表达.结果 pGC-El-GFP-HLA-E慢病毒载体转导肝癌HepG2细胞72h的转导效率为(95.0±1.3)%;通过RT-PCR检测24h后细胞内HLA-E mRNA水平是肝癌HepG2细胞的(8.73±1.05)倍,72h后为(293.48±42.01)倍,差异具有统计学意义(P<0.01).细胞免疫组化显示转导HLA-E后细胞内的蛋白水平明显增强.结论 慢病毒栽体系统能够使转导的基因在靶细胞中得到稳定表达,是基因治疗中的理想载体.

Construction of recombinant lentivirol expression vector carrying human HLA-E gene and Its expression in HepG2 cells

??Objective:To construct the lentiviral expression vector of HLA-E gene and investigate its significance for further study on tumor immunity. Methods:After the lentiviral expression vector of HLA-E gene (pGC-E1-GFP-HLA-E) was transduced HepG2 cell lines,the transduction efficiency??the HLA-E mRNA level by RT-PCR and the expression of HLA-E by immunohistochemistry were detected between October 2006 and November 2007 in the Liver Transplautation Center of Third Affiliated Hospital of Sun Yat-sen University. Results:The transduction efficiency of HepG2 cell lines using the lentiviral vectors was 95.0%±1.3% which allowing the stable expression of HLA-E after 72 hr.The HLA-E-mRNA level in vitro was detected more 8.73±1.05 times and 293.48±42.01 times than normal level by RT-PCR (P??0.01).The expression of HLA-E by immunohistochemistry was displayed significant enhancement. Conclusion:The lentiviral expression vector is ideal vetor of gene therapy and made genes transducted stable expression in target cell.