摘要
目的制备hIL-18和TK双基因修饰的人脑胶质瘤活疫苗,检测HSV-TK/GCV系统对疫苗的体外调控作用。方法采用表达HSV-TK基因的慢病毒液感染U87-hIL18细胞,经G418和BSD抗性基因筛选获得U87-hIL18-TK单克隆细胞株,扩增传代获得双基因修饰疫苗,Elisa检测疫苗上清IL-18浓度,RT-PCR检测疫苗TK基因表达,MTT法检测更昔洛韦(GCV)对体外培养疫苗的杀伤效应。结果 TK基因成功转导入U87-hIL18细胞中,获得稳定表达IL-18、TK基因的U87-hIL18-TK胶质瘤活疫苗,体外杀伤效应显示10μg/mlGCV对活疫苗的杀伤率达90.3%。结论成功构建免疫基因和自杀基因共同修饰的胶质瘤活细胞疫苗,其在体内的安全性及免疫效能有待进一步研究。
Objective To establish hIL-18 and TK genes co-modified human glioma live vaccine,and detect the in vitro modulatory effect of HSV-TK/GCV system on the vaccine.Methods The lentivirus containing HSV-TK gene was added to U87-hIL18 cells,and U87-hIL18-TK monoclonal cell strains were obtained by G418 and BSD resistance gene screening,and amplified and passaged for producing double gene-modified vaccine,then the specific expression of IL-18 and HSV-TK gene in live the vaccine were evaluated by Elisa assay and RT-PCR respectively.The sensitivity of the live vaccine to ganciclovir （GCV） was evaluated by MTT assay.Results TK gene was successfully transduced into U87-hIL18 cells.Elisa and RT-PCR test showed that IL-18 and TK gene were stably expressed in the tumor vaccine respectively.MTT proved that HSV-TK/GCV system was able to completely control the proliferation of the live vaccine in vitro.The 90.3% of the live vaccines were killed by 10 μg/ml GCV.Conclusions The glioma live vaccine co-modified with immunogene and suicide gene has been constructed successfully and further investigations are required to elucidate its safety in vivo and immunopotency.
出处
《中国微侵袭神经外科杂志》
CAS
北大核心
2010年第6期271-274,共4页
Chinese Journal of Minimally Invasive Neurosurgery
基金
广东省自然科学基金(编号:001122)
全军医学科研"十五"计划项目基金(编号:01MA038)
中国博士后科学基金(编号:20070420766)
