细胞因子IL-27和CpG ODN佐剂对呼吸道合胞病毒重组蛋白疫苗免疫效果的影响

Effects of IL-27 and CpG ODN adjuvant on immune efficacy of respiratory syncytial virus recombinant vaccine

目的研究细胞因子佐剂IL-27单独或与CpG2216佐剂联合使用对呼吸道合胞病毒(RSV)重组蛋白疫苗免疫效果的影响。方法将编码IL-27的质粒(以下简称IL-27)单独或与CpG2216佐剂作为联合佐剂与G1F/M2共免疫Balb/c小鼠3次,每次免疫10 d后取血,分离血清,用间接ELISA法测定3次免疫后的血清IgG抗体效价以及末次IgG1、IgG2a抗体效价。第3次免疫后2周处死小鼠,取脾细胞,用LDH法检测CTL活性、ELISPOT法检测分泌IL-4和IFN-γ的淋巴细胞水平、流式细胞仪检测CD4^+、CD8^+的效应记忆和中央记忆T细胞。结果 ELISA结果表明:除PBS组外,各组均诱导了高水平的IgG抗体无显著性差异(P<0.05),IgG2a、IgG1分别为Th1和Th2型抗体,IL-27+G1F/M2组、IL-27+CpG2216+G1F/M2组IgG2a和IgG1的比值明显高于其他组,说明IL-27作为佐剂对G1F/M2所诱导的体液免疫应答无增强作用但可以调节免疫应答的类型,使免疫应答更平衡;CTL杀伤实验显示:IL-27^+CpG2216+G1F/M2组的CTL杀伤活性显著高于其他各组(P<0.05);ELISPOT结果显示:IL-27^+CpG2216+G1F/M2组中分泌IFN-γ的淋巴细胞数量显著高于其他组分泌IFN-γ的淋巴细胞数量,且各组分泌IFN-γ的淋巴细胞数量显著高于同组分泌IL-4的淋巴细胞数量(P<0.05),即均诱导了Th1型优势应答;流式细胞仪检测结果说明IL-27+G1F/M2组和IL-27^+CpG2216+G1F/M2组均能诱导产生大量的CD44^+CD62L^+双阳性细胞。结论细胞因子佐剂IL-27和CpG2216佐剂联合使用可以增强吸道合胞病毒重组疫苗诱导的细胞免疫应答而且IL-27对Th1型优势应答具有调节作用。

This study performed to investigate the effects of IL-27 and CpG ODN adjuvant on immune efficacy of respiratory syncytial virus（RSV） recombinant vaccine.Balb/c mice were immunized with RSV vaccines G1F／M2 three times at an interval of 10 days,with pcDNA3.1/IL-27 alone or pcDNA3.1/IL-27 plus CpG2216 as adjuvant.Mice were phlebotomized at day 10 after every immunization and the serum was separated for measuring specific antibody IgG titers and specific antibodies IgG1,IgG2 a by indirect ELISA.Two weeks after the third immunization,the mice were sacrificed and splenocytes were prepared for assessing specific lysis activity using a LDH Cytotoxicity Assay Kit.Furthermore,IFN-γ and IL-4 secreting cells were quantified using an ELISPOT kit,while the percentage of CD4~＋/CD8~＋ effector cells or memory cells in the spleen cells was analyzed with flow cytometer.We found that specific IgG antibodies were induced and showed no significant difference in mice of all groups,except PBS group,after the third immunization;ELISA indicated that IgGl is Th1 type antibody and IgG2 a is Th2 type antibody.The ratio of IgG1/IgG2 a in IL-27＋G1F/M2 group was obviously higher than that of the groups without adjuvant.These results show that IL-27 as the adjuvant of G1F/M2 can not enhance humoral immune response,but can make Th1 and Th2 type response reduced.The CTL activity of IL-27＋ CpG2216＋GlF/M2 groups（P〈0.05） was significant higher than that of the other groups.IL-27＋ CpG2216＋ G1F/M2 groups induced significantly more INF-γ secreting cells than other groups did（P〈0.05）.In addition,the level of INF-γ secreting cells was remarkably higher than that of IL-4secreting cells in each group,suggesting a Th1 biased response had been induced.These results of flow cytometer indicatd that IL-27＋CpG2216 elicited more CD44~＋CD62L~＋ cells.In conclusion,IL-27＋CpG2216,as an adjuvant,can enhance significantly cellular immune responses,and IL-27 can regulate Th1 type responses of RSV recombinant vaccine G1F/M2.