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目的比较截短、全长、全长融合等3种不同形式的乙型肝炎病毒核心抗原(HBcAg)核酸疫苗在小鼠体内的体液免疫和细胞免疫效果,获得最佳的免疫原设计方案。方法构建经密码子优化的不同形式HBcAg核酸疫苗质粒:pRec2.0-Ct(表达截短至144 aa的Core蛋白)、pRec2.0-C(表达全长183 aa的Core蛋白)和pRec2.0-C-preS1(表达全长Core和PreS1融合蛋白),转染293T细胞,Western blot检测目的抗原基因的表达。将3种核酸疫苗分别免疫BALB/c小鼠,ELISA法检测小鼠血清抗-HBc抗体水平,IFNγELISPOT法检测小鼠体内HBcAg特异性T细胞免疫应答。结果 3种核酸疫苗质粒经酶切及测序证实构建正确;3种核酸疫苗质粒在293T细胞中均可表达相应蛋白;经3种核酸疫苗免疫后的小鼠均产生了HBcAg特异性抗体和T细胞应答,其中pRec2.0-C组抗-HBc抗体水平和T细胞免疫反应水平均显著高于pRec2.0-C-preS1组和pRec2.0-Ct组(P<0.05)。结论将HBcAg截短至144个氨基酸或与preS1融合均可降低其体液免疫及细胞免疫应答。
OBJECTIVE: To compare the humoral and cellular immunity effects of three different forms of hepatitis B virus core antigen (HBcAg) nucleic acid vaccine, such as truncated, full-length and full-length fusion in mice, and to obtain the best immunogen design. Methods Construction of different codon-optimized HBcAg nucleic acid vaccine plasmids: pRec2.0-Ct (Core protein truncated to 144 aa), pRec2.0-C (Core protein with full-length 183 aa expression) and pRec2.0 C-preS1 (full-length Core and PreS1 fusion protein) was transfected into 293T cells, and the expression of the target antigen gene was detected by Western blot. BALB / c mice were immunized with three kinds of DNA vaccines respectively. Serum levels of anti-HBc antibodies were detected by ELISA, and HBcAg-specific T cell immune responses were detected by IFNγ ELISA. Results The three DNA vaccine plasmids were confirmed by restriction enzyme digestion and sequencing. The three kinds of DNA vaccine plasmids could express the corresponding proteins in 293T cells. The HBcAg specific antibodies and T cells The levels of anti-HBc antibody and T cell immune response in pRec2.0-C group were significantly higher than those in pRec2.0-C-preS1 group and pRec2.0-Ct group (P <0.05). Conclusion The truncation of HBcAg to 144 amino acids or the fusion with preS1 can reduce humoral and cellular immune responses.