含乙型肝炎病毒C基因的重组腺相关病毒对树突状细胞的作用  

Recombinant adeno-associated virus carrying hepatitis B virus C gene (rAAV-HBV-C) effectively pulses dendritic cells (DC) without affecting their maturity

作　　者：丛敏[1] 王萍[1] 阎钟钰[1] 唐淑珍[1] 王宝恩[1] 贾继东[1] 刘勇[2] 尤红[1] 

机构地区：[1]首都医科大学附属北京友谊医院肝病中心,北京100050 [2]美国阿肯色州立医科大学基因治疗中心

出　　处：《中华肝脏病杂志》2005年第1期17-19,共3页Chinese Journal of Hepatology

基　　金：北京市科技新星计划(2004B32)

摘　　要：目的 研究腺相关病毒(AAV)为载体的含有乙型肝炎病毒(HBV)C基因重组病毒(rAAV-HBV-C)感染树突状细胞(DC)的效率及对DC生长和成熟的影响。 方法 从正常人外周血中分离单个核细胞收取单核细胞进行体外培养,分别用rAAV-HBV-C和293细胞裂解物感染刺激单核细胞,感染后的单核细胞在粒细胞-巨噬细胞集落刺激因子(GM-CSF)、白细胞介素4(IL-4)和肿瘤坏死因子α(TNF α)存在的条件下继续培养获得成熟的DC。用逆转录聚合酶链反应及流式细胞仪细胞内染色检测目的基因(HBV-C)的转录及表达;通过观察不同时间DC的形态及检测收获DC时其表面分化抗原(CD)的表达来评价DC的生长和成熟状态。 结果 rAAV-HBV-c感染后的DC可转录HBV-C基因并表达HBV-C抗原,病毒感染组与对照组收获的DC在细胞形态与CD表达方面差异无统计学意义。 结论 rAAV-HBV-C可有效感染DC,感染的病毒对DC的生长及成熟没有显著影响。Objective Recombinant virus pulsated dendritic cells (DCs) may affect their survival, growth and maturity. This study is to test the infection efficiency of recombinant adeno-associated virus carrying hepatitis B core antigen (rAAV-HBV-c) to DCs and the growth and maturity of them. Methods Peripheral blood mononuclear cells (PBMCs) were isolated from healthy blood donors. Adherent monocytes were pulsed by rAAV-HBV-c and 293 lysate as controls on the first day of isolation. DCs were cultivated in AIM-V media with 1000 u/ml granulocyte macrophage stimulating factor (GM-CSF), 1000 u/ml interleukin-4 (IL-4) and 50 ng/ml tumor necrosis factor- α(TNF α) separately in vitro. DCs were examined at different times and the expressions of several clusters of differentiations (HLADR, CD14, CD80, CD83, CD86) were studied using FACS after being cultured for 7 days. The transcription and expression of HBV-C gene were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and intracellular staining fluorescence activated cell sorter (FACS), respectively. Results The rAAV-HBV-c infected and uninfected monocytes gradually matured and their morphology had no significant differences. The CDs expressed on the surfaces of the two groups of DCs were also similar (HLADR: 96.1% vs 94.5%; CD86: 87.7% vs 89.8%; CD83: 75.6% vs 78%; CD80: 52% vs 54.3%; CD14: 6.4% vs 4.5%). HBV-C gene mRNA expression was measured using RT-PCR and 89.5% of the rAAV-HBV-c infected DCs showed their protein expression using FACS. Conclusion rAAV-HBV-c can effectively pulse DCs without affecting the growth and maturity of them.

关 键 词：HBV DC 感染 乙型肝炎病毒 单核细胞 表达 C基因 结论 培养 目的 

分 类 号：R346[医药卫生—基础医学]