插入IL-2基因优化HBV DNA疫苗的研究

Study of optimization HBV DNA vaccine by insertion of IL-2 gene

目的：观察IL-2基因的插入对DNA疫苗免疫效应的影响，探讨优化DNA疫苗设计、提高DNA疫苗兔疫效应的途径。方法：采用PCR产物直接克隆和重组DNA技术构建了人IL-2和HBsAg融合基因的真核表达载体pcDNA3．1+-S/IL-2，通过脂质体基因转移技术导入Cos-7细胞中检测其瞬间表达，并经肌肉注射免疫C57Bl/6小鼠，以检测和比较它们的细胞和体液免疫应答。结果：通过酶切、PCR及测序证实已正确完整地插入 IL-2基因，成功构建了 pcDNA3．1+-S/IL-2重组质粒。体外转染Cos-7后可见基因的表达和分泌，pcDNA3．1+-S/IL-2可以提高免疫小鼠的抗 HBs抗体滴度和脾淋巴细胞诱生的IL-2的生物活性水平，增加 HBsAg特异性的脾淋巴细胞增殖指数。结论：IL-2和 HBsAg基因的融合表达对 DNA疫苗的免疫反应起协同和增强作用，提示IL-2基因插入可能是增强DNA疫苗的免疫效应的可行途径。

Abstract Objective:To observe and compare the effects of immune responses of HBV DNA caccine by insertion IL-2 gene.Try to ex-Plore the efficient way to enhance the immune effect of DNA vaccines. Methods:The recombinant expression plasmid vector peDNA3.1-S/IL-2with fusion gene of HBsAg and IL-2 cloned was constructed by PCR method and DNA recombinant technique.Then, it was transfered into Cos-7 cells by cation lyposomes and was detected its transient expressing product in vitro.At last,observed and compared the cellular and humoralimmune responses to HBsAg in C57BL/6 mice by intramuscular injection.Results:The pcDNA3. 1-S/IL-2 was constructed successfully by themethod of restriction endonucleases digestion, RCR and DNA sequencing. It can increase the titers of anti-HBs antibody. The bioactivity of IL-2and lymphocyte proliferation responses to HBsAg induced by the mouse spleen immunized with psDNA3.1-S/IL-2 were improved also. Conclu-sion:These results showed the expression of HBsAg-IL-2 fused gene can enhance the immune responses and suggested the insertion of IL-2 maybe a practicable way to enhance the DNA vaccine efficacy.