摘要
目的构建小鼠巨噬细胞炎性蛋白-1α(MIP-1α)的真核表达质粒,观察其作为分子佐剂对单纯疱疹病毒II型(HSV-II)DNA疫苗免疫效果的影响。方法以LPS刺激RAW264.7细胞提取总RNA。采用RT-PCR扩增出MIP-1α基因的全部编码序列,利用克隆载体pUCm-T,将其亚克隆入pcDNA3中,构建出小鼠MIP-1α的真核表达质粒Pm;将其转染COS-7细胞,并用Boyden趋化小室法检测MIP-1α的生物学活性。然后用其与HSV-IIgD的DNA疫苗一起免疫BALB/c小鼠,检测免疫小鼠的特异性抗体、脾T细胞增殖反应及病毒攻击小鼠后对小鼠的保护率,观察MIP-1α对HSV-IIDNA疫苗免疫效果的影响。结果成功构建了小鼠MIP-1α的重组真核表达质粒;免疫BALB/c小鼠发现,其作为分子佐剂可加强HSV-IIgDDNA疫苗的免疫效果。结论小鼠MIP-1α可作为HSV-IIgDDNA疫苗的分子佐剂,为研制新型有效的HSV-IIDNA疫苗提供一定的依据。
AIM： To construct the recombinant eukaryotic expression plasmid of murine macrophage inflammatory protein-1α（MIP-1α） and investigate the effect of MIP-1α as an adjuvant on immune response induced by herpes simplex virus type Ⅱ glycoprotein D （HSV-Ⅱ gD） DNA vaccine. METHODS： Using total RNA from RAW204.7 cells stimulated with LPS, the whole code sequence of murine MIP-1α was amplified by reverse transcription polymerase chain reaction（RT-PCR） and inserted into pcDNA3 at Hind Ⅲ/Xba Ⅰ restriction sites. The recombinant eukaryotic expression plasmid Pm was transiently expressed in COS-7 cells and its specificity was demonstrated by RT-PCR and boyden chemotaxis chamber. BALB/c mice were immunized with gD DNA vaccine and/or MIP-1α, and the effect of MIP-1α on gD DNA vaccine was evaluated by detecting anti-HSV-Ⅱ antibody, antigen-specific lymphoproliferative responses, and examining survival rates after mice were challenged intravaginally with HSV-Ⅱ. RESULTS： The recombinant eukaryotic expression plasmid of murine MIP-1α was constructed, and it was revealed that immune responses of HSV-Ⅱ gD DNA vaccine were enhanced by coimmunization with MIP-1α. CONCLUSION： The murine MIP-1α can be used as an adjuvant of HSV-Ⅱ gD DNA vaccine.
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2006年第3期286-289,共4页
Chinese Journal of Cellular and Molecular Immunology
