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目的:探讨巨噬细胞炎性蛋白1α(Macrophage inflammatory protein-1α,MIP-1α)和酪氨酸激酶受体3配体(Flt3ligand,Flt3l)基因佐剂联合应用增强人乳头瘤病毒16型(Human papillomavirus16,HPV16)E7DNA疫苗免疫效果的可能性。方法:构建真核表达质粒pcDNA3/MIP-1α、pcDNA3/Flt3l和pcDNA3/HPV16E7。C57BL/6小鼠随机分为8组,分别为pcDNA3组、pcDNA3/MIP-1α组、pcDNA3/Flt3l组、pcDNA3/MIP-1α和pcDNA3/Flt3l混合组、pcDNA3/HPV16E7组、pcDNA3/MIP-1α和pcDNA3/HPV16E7混合组、pcDNA3/Flt3l和pcDNA3/HPV16E7混合组以及pcDNA3/MIP-1α、pcDNA3/Flt3l和pcDNA3/HPV16E7混合组。每组14只,肌肉注射质粒免疫小鼠,每3周免疫1次,共3次;末次免疫后2周每组随机抽取6只小鼠,取脾脏制备淋巴细胞悬液,用CCK-8试剂盒检测其对TC-1靶细胞的特异性杀伤能力。其余小鼠在腹股沟处皮下注射5×104个TC-1细胞,观察肿瘤生长情况。结果:成功构建了真核表达质粒pcDNA3/MIP-1α、pcDNA3/Flt3l和pcDNA3/HPV16E7;pcDNA3/MIP-1α、pcDNA3/Flt3l和pcDNA3/HPV16E7混合组的CTL杀伤能力明显高于其它各组(P<0.01);肿瘤细胞攻击后16天,pcDNA3组小鼠全部长出肿瘤,而pcDNA3/MIP-1α、pcDNA3/Flt3l和pcDNA3/HPV16E7混合组的成瘤率仅为12.5%,明显低于其他组,统计学分析示8组之间成瘤率差异有显著性(P=0.007)。60天后各组成瘤率之间的差异无统计学意义(P=0.229)。结论:联合应用MIP-1α和Flt3l基因佐剂增强了HPV16E7DNA疫苗的免疫效果,一定程度上延缓了肿瘤的发生。
Objective: To investigate the effects of MIP-1α (MIP-1α) and Flt3ligand (Flt3l) gene adjuvant on the expression of human papillomavirus type 16 papillomavirus16, HPV16) E7 DNA vaccines. Methods: Eukaryotic expression plasmids pcDNA3 / MIP-1α, pcDNA3 / Flt3l and pcDNA3 / HPV16E7 were constructed. C57BL / 6 mice were randomly divided into 8 groups: pcDNA3 / MIP-1α group, pcDNA3 / Flt3l group, pcDNA3 / MIP-1α and pcDNA3 / Flt3l mixed group, pcDNA3 / HPV16E7 group, pcDNA3 / MIP-1α And pcDNA3 / HPV16E7 mixed group, pcDNA3 / Flt3l and pcDNA3 / HPV16E7 mixed group and pcDNA3 / MIP-1α, pcDNA3 / Flt3l and pcDNA3 / HPV16E7 mixed group. The mice in each group were immunized with intramuscular plasmid and immunized once every 3 weeks for 3 times. Six mice were randomly selected from each group 2 weeks after the last immunization. Spleen lymphocytes were prepared and treated with CCK-8 reagent The cassette was tested for its ability to specifically kill TC-1 target cells. The remaining mice were injected subcutaneously with 5 × 10 4 TC-1 cells in the groin to observe tumor growth. Results: The eukaryotic expression plasmids pcDNA3 / MIP-1α, pcDNA3 / Flt3l and pcDNA3 / HPV16E7 were successfully constructed. The CTLs killing ability of pcDNA3 / MIP-1α, pcDNA3 / Flt3l and pcDNA3 / HPV16E7 mixed groups was significantly higher than that of other groups <0.01). On the 16th day after the tumor cells were challenged, all the mice in the pcDNA3 group grew tumors. However, the tumorigenic rate of the pcDNA3 / MIP-1α, pcDNA3 / Flt3l and pcDNA3 / HPV16E7 mixed groups was only 12.5% Statistical analysis showed that there was a significant difference in tumor formation rate between the 8 groups (P = 0.007). After 60 days there was no significant difference between the constituent tumor rates (P = 0.229). Conclusion: The combination of MIP-1α and Flt3l gene adjuvant enhances the immunogenicity of HPV16E7 DNA vaccine and delays the tumor to a certain extent.