SAG1和ROP1混合基因真核表达质粒接种小鼠诱导产生拮抗致死性弓形虫感染的保护性免疫

A Cocktail Candidate DNA Vaccine Against Toxoplasma gondii with Genes Encoding SAGl and ROPI Induces Protective Immunity Against Lethal Challenge in Mice

目的检测混合SAG1和ROP1编码基因真核表达质粒疫苗诱导小鼠的免疫应答,评价其抗弓形虫感染的保护性免疫效果。方法将SAG1和ROP1编码基因片段克隆入pEGFP-N3表达载体,构建重组质粒;RT-PCR体外验证重组质粒在NIH3T3细胞中的表达;通过检测抗体、抗体分型及细胞因子来评价体液免疫和细胞免疫;流式细胞仪测定脾脏淋巴细胞亚群;腹腔内注射毒性株弓性虫速殖子攻击免疫小鼠。结果 RT-PCR结果显示重组质粒能在哺乳动物细胞内表达;SAG1和ROP1混合重组质粒疫苗诱导小鼠产生很强体液免疫和细胞免疫,对于毒性株弓形虫感染攻击具有免疫保护作用。结论不同候选抗原编码基因混合重组质粒能够诱导小鼠产生抗弓形虫感染保护性免疫,提示含有多种成份的混合DNA疫苗的研制可作为核酸免疫研究的策略之一。

Objective To examine the immune response elicited by SAG1 and ROP1 encoding plasmids and assess the protective effect of the DNA vaccination against toxoplasmosis. Methods DNA fragments encoding SAG1 and ROP1 were cloned into pEGFP-N3 expression vector to construct serial recombinant plasmids. In vitro expression of the recombinant plasmids in the NIH3T3 cells were verified by RT-PCR. Humoral and cellular responses were assayed using ELISA for detection of Ab,Ab isotype and cytokines. FCM was also used to sort the lymphocyte subsets of spleen cells. All mice were challenged with higly virulent tachyzoites via intraperitoneal injection. Results RT-PCR indicated that the SAG1 and ROP1 could be expressed in the expression recorrbinmant plasmid transfected NIH3T3 cell line. Mice genetically immunized with the recombinant plasmids elicited strong humoral and cellular immune responses and showed partial protective effect to mice challenged with highly virulent isolate to Toxoplasma gondii. Conclusions A strategy of vaccination with recombinant plasmids encoding different antigens might be useful to offer protection against toxoplasma and this could have an implication in the design of future multicomponent DNA vaccines.