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目的:研究Flt3L与CCL5作为联合佐剂在prime/boost免疫策略中对HBc抗原特异性免疫应答的增强及抗肿瘤作用。方法:将两种细胞因子质粒与携带HBc抗原的DNA疫苗经肌内注射法共免疫小鼠,免疫3次后再用原核表达的HBc颗粒蛋白或HBcDNA疫苗加强,观察对稳定表达HBcAg的小鼠黑色素瘤细胞(B16-HBc)的生长抑制作用;并分别采用MTT法检测荷瘤小鼠脾淋巴细胞增殖、流式细胞术检测脾CD8+T淋巴细胞中IFN-γ表达、ELISA法检测脾淋巴细胞培养上清IL-2、IL-4含量及乳酸脱氢酶(LDH)释放法检测特异性CTL杀伤活性。结果:与对照组相比,佐剂联合DNA疫苗免疫经蛋白加强组(DDP/Adj)显著抑制肿瘤生长;佐剂联合DNA疫苗免疫组(DDD/Adj)及DDP/Adj组均可促进特异性淋巴细胞增殖反应(P<0.05),且DDP/Adj高于DDD/Adj组(P<0.05);DDD/Adj及DDP/Adj组小鼠脾脏CD8+T淋巴细胞中IFN-γ表达、IL-2表达水平及CTL杀靶活性均高于对照组(P<0.01或P<0.05),IL-4表达水平在各组无显著区别(P>0.05)。结论:在prime/boost免疫策略中,采用Flt3L与CCL5两种细胞因子联合应用可显著促进荷瘤小鼠产生抗原特异性免疫应答及抗肿瘤作用。
OBJECTIVE: To investigate the anti-tumor effect of Flt3L and CCL5 as a combination adjuvant on specific HBc antigen-specific immune response in prime / boost immunization strategy. Methods: The two kinds of cytokines plasmids were immunized with the DNA vaccine carrying HBc antigen by intramuscular injection. After 3 times of immunization, the mice were boosted with prokaryotic HBc particle protein or HBcDNA vaccine. The mice stably expressing HBcAg Melanoma cells (B16-HBc). The proliferation of tumor-bearing mice spleen lymphocytes was detected by MTT assay, the expression of IFN-γ in spleen CD8 + T lymphocytes was detected by flow cytometry, the splenic lymphocytes The specific cytotoxic activity of CTL was detected by the cell culture supernatant IL-2, IL-4 content and lactate dehydrogenase (LDH) release assay. Results: Compared with the control group, adjuvant combined with DNA vaccine significantly inhibited tumor growth by protein-enhanced group (DDP / Adj); adjuvant combined with DNA vaccine (DDD / Adj) and DDP / Adj group (P <0.05), and the ratio of DDP / Adj was higher than DDD / Adj group (P <0.05). The expression of IFN-γ in spleen CD8 + T lymphocytes of DDD / Adj and DDP / 2 expression and target killing activity of CTL were all higher than that of the control group (P <0.01 or P <0.05). There was no significant difference in the expression level of IL-4 between the two groups (P> 0.05). Conclusion: The combination of Flt3L and CCL5 cytokines can significantly promote antigen-specific immune response and anti-tumor effect in prime / boost immunization strategy.