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目的比较IL-15在DNA疫苗初始和重组天坛痘苗病毒加强免疫阶段对免疫原性的影响。方法应用PCR方法构建重组片段I8R-pE/L-IL-15-GFP-G1L,通过同源重组将IL-15基因插入到表达HIV-1CN54Gag,Pol和Env基因的重组天坛艾滋病疫苗rTV的基因组中。HIV DNA疫苗在0、3、6周与质粒pIL-15共免疫BALB/c小鼠,或仅使用DNA疫苗,rTV和rTV-IL-15在12周分别进行加强免疫。末次免疫后1、4周应用ICS和ELISPOT方法检测小鼠脾淋巴细胞中HIV特异性细胞免疫。末次免疫后1、4周和12周ELISA方法检测小鼠血清中Gp120特异性抗体。结果 IL-15基因正确插入重组天坛艾滋病疫苗rTV基因组中,成功构建了重组病毒rTV-IL-15。各组免疫小鼠均诱导出HIV特异性细胞和体液免疫。尽管各组诱导的HIV特异性CD8+T细胞免疫反应差异无统计学意义,但加强免疫后4周,DNA-IL-15/rTV-IL-15组分泌IFN-γ、TNF-α以及IFN-γ/TNF-α双因子的CD8+T效应记忆细胞百分比均值显著高于DNA-IL15/rTV组(P<0.05),分别为0.86%,0.37%和0.31%。DNA/rTV-IL-15组的gp120抗体滴度在加强免疫后1周即达到1∶40 381,显著高于DNA-IL-15/rTV组和DNA-IL-15/rTV-IL-15组(P=0.000 1,P=0.020 5),且抗体滴度持续至12周未见明显下降。而DNA-IL-15/rTV组和DNA-IL-15/rTV-IL-15组的抗体滴度则从加强免疫1周后逐渐上升,在12周达到高峰(1∶11 143,1∶16 889),DNA-IL-15/rTV组4周和12周的抗体滴度显著高于1周(P=0.009,P=0.012)。结论在初始和加强免疫阶段使用IL-15佐剂可以提高CD8+T效应记忆细胞应答水平。
Objective To compare the effects of IL-15 on the immunogenicity during initial vaccination and recombinant vaccinia virus vaccination. Methods The recombinant fragment of I8R-pE / L-IL-15-GFP-G1L was constructed by PCR. The IL-15 gene was inserted into the genome of rTVTV rTV expressing HIV-1CN54Gag, Pol and Env by homologous recombination . The HIV DNA vaccine co-immunized BALB / c mice with plasmid pIL-15 at week 0, 3, 6, or boosted with DNA vaccine alone, rTV and rTV-IL-15 for 12 weeks, respectively. One and four weeks after the last immunization, HIV-specific cellular immunity in splenic lymphocytes of mice was detected by ICS and ELISPOT methods. Gp120 specific antibodies in serum of mice were detected by ELISA method at 1, 4 weeks and 12 weeks after the last immunization. Results The IL-15 gene was correctly inserted into rTV genome of recombinant adenovirus vaccine and recombinant virus rTV-IL-15 was successfully constructed. Each group of immunized mice induced HIV-specific cellular and humoral immunity. Although the immunological reaction of HIV-specific CD8 + T cells induced by each group was not statistically different, the expression of IFN-γ, TNF-α and IFN-γ in DNA-IL-15 / The mean percentage of CD8 + T effect memory cells of γ / TNF-α two factors was significantly higher than that of DNA-IL15 / rTV group (0.86%, 0.37% and 0.31% respectively). The titer of gp120 antibody in DNA / rTV-IL-15 group was 1:40 381 1 week after boosting, which was significantly higher than that in DNA-IL-15 / rTV group and DNA-IL-15 / rTV-IL-15 group (P = 0.000 1, P = 0.020 5), and the antibody titers did not decrease significantly for 12 weeks. The antibody titers in DNA-IL-15 / rTV group and DNA-IL-15 / rTV-IL-15 group increased gradually from 1 week after booster immunization and reached the peak at 12 weeks (1:11 143, 1:16 889). The titer of antibody in DNA-IL-15 / rTV group was significantly higher than that in 1 week (P = 0.009, P = 0.012) at 4 weeks and 12 weeks. Conclusions The use of IL-15 adjuvant during the initial and boost phases can increase the level of CD8 + T effector memory response.