肌肉免疫DC-EBV-LMP2诱导免疫应答的研究

摘要

目的探讨肌肉免疫DC-EBV-LMP2诱导的免疫效果.方法从BALB/C小鼠分离骨髓细胞,诱导培养获得小鼠DC,以100 MOI的rAd-LMP2感染获得DC-EBV-LMP2,继续培养48h,诱导DC-EBV-LMP2成熟.用免疫酶法确定LMP2在DC中表达.0,2,4周,以2×105个DC-EBV-LMP2/只免疫BALB/C小鼠,于第5,8周分别LMP2特异性水平和T细胞亚群的变化.结果结果显示,肌肉免疫DC-EBV-LMP2可诱导BALB/C鼠LMP2特异性免疫应答,且第5周强于第8周.CD8+T/CD3+T细胞亚群的比值在第5周高于对照组,而第8周低于对照组.结论DC-EBV-LMP2肌肉免疫BALB/C鼠可诱导LMP2特异性CTL.%　　Objective To explore immunity response induced by DC-EBV-LMP2 with intramuscle immunity. Methods Bone marrow cells were separated from BALB/C mice, and cultured in RPMI1640 medium supplemented with 10%fetal calf serum and recombinant murine cytokines in vitro. Murine dendritic cells (mDCs) were harvested, and infected by recombinant adenovirus including latent membrane proteinⅡ(LMP2) gene (rAd-LMP2) of Epstein-Barr virus(EBV)at a dose of 100 multiplicity of infection (MOI). DC-EBV-LMP2 was induced and matured at 48 hours later. The LMP2 expression in mDCs was detected by immunoenzyme. BALB/C mice were immunized with 2×105 DC-EBV-LMP2 per mouse by intramuscle immunity at 0,2,4 weeks. LMP2 specific cytotoxic lymphocytes (CTLs) and T cell subset in BALB/c mice were detected at 5th week and 8th week with enzyme-linked immunosorbent spot (ELISPOT) and flow cytometry, respectively. Results LMP2 specific CTLs in BALB/c mice were detected both at 5th week and 8th week after intramuscle immunity with DC-EBV-LMP2. LMP2 specific CTL in BALB/c mice is more powerful at 5th week(308±167 per 106cells,negative control 18± 6 per 106cells) than at 8th(196±81 per 106cells,negative control 7±2 per 106cells). The ratio of CD8+T to CD3+T subset is higher at DC-EBV-LMP2 group than injected PBS and DC groups at 5th week, but lower than in PBS and DC groups at 8th week. The antibody to LMP2 was detected in mice after DC-EBV-LMP2 vaccinated. Conclusion LMP2 specific CTLs can be induced by the means of intramuscle immunity DC-EBV-LMP2 in BALB/c mice.