全酿酒酵母HPV16-E7疫苗的构建及免疫原性研究

Construction of the whole yeast HPV16-E7 vaccine and study on its immunogenicity

目的构建人乳头状瘤病毒16型(HPV16)E7重组全酿酒酵母疫苗,评价疫苗的免疫效应。方法将HPV16 E7 cDNA亚克隆入酵母表达载体pYES2/NT,重组质粒转化酿酒酵母,经诱导表达、热灭活制备全酵母HPV16-E7疫苗,免疫C57BL/6小鼠。ELISA、MTT、LDH法分别检测疫苗诱导的细胞因子、脾淋巴细胞增殖和细胞毒性T淋巴细胞(CTL)应答水平。结果全酵母疫苗能够有效表达HPV16-E7蛋白;疫苗小鼠免疫,能够刺激脾淋巴细胞增殖,并诱导出Th1型细胞因子和特异性CTL杀伤效应;免疫效果优于单纯HPV16-E7蛋白免疫(P<0.01)。结论全酵母疫苗既能表达HPV16-E7蛋白,又能有效诱导特异性CTL免疫应答。研究结果为进一步进行疫苗抗肿瘤疗效研究,提供了实验基础。

Objective To construct a whole recombinant yeast vaccine containing human papillomovirus type 16 （HPV16） E7 gene, and study on the immune response induced by the HPV16 - E7 vaccine. Methods The HPV16 E7 coding cDNA was subcloned into yeast expression vector and positive transformant stains were obtained after transform the recombinant plasmids into the competent saccharomyces cerevisiae. Then the whole yeast HPV16 - E7 vaccine was prepared by heat- inactivation. C57BL/6 mice were immunized with the vaccine, and the serum cytokines, the proliferation of spleen lymphocyte and the CTL function induced with the vaccine were detected by ELISA, MMT and LDH, respectively. Results HPV16 E7 gene could be efficiently expressed by the whole yeast vaccine. The vaccine could stimulate the proliferation of spleen lymphocyte, moreover, activate the Thl cytokines and the specific CTL function of mice. The immune effect of the whole yeast vaccine was more efficient than that of HPV16 - E7 protein （P〈0. 01）. Conclusions The whole yeast vaccine could express HPV16 -E7 protein, and induce the specific CTL immune response efficiently. Further exploration of anti- tumor effect of the yeast vaccine can be based on these results.