肠道病毒71型与脊髓灰质炎病毒抗体交叉反应性研究

Cross-reactivity between anti-enterovirus 71 and anti-pliovirus antibodies

通过临床血清样本研究及实验动物验证,探讨肠道病毒71型(EV71)与脊髓灰质炎病毒(PV)之间的抗体交叉反应性。以酶联免疫吸附试验(ELISA)分别检测健康儿童血清中IgG型抗体与EV71及PV的反应性;用原核表达与亲和纯化等方法分别获得EV71和PV的衣壳蛋白VP1及非结构蛋白3C;将PV-VP1、EV71-VP1、EV71灭活疫苗及PV灭活疫苗(IPV)分别经皮下免疫BALB/c小鼠制得免疫血清,以蛋白免疫印迹实验分别研究PV-VP1或EV71-VP1小鼠阳性血清与EV71-VP1和PV-VP1的反应情况,进一步以竞争ELISA试验分别研究EV71-VP1对PV-VP1与其特异性免疫血清特异结合的影响。微量中和试验研究PV阳性抗血清对EV71的体外中和效应。结果:已接种PV疫苗但未曾感染EV71的健康儿童的IgG抗体与EV71有较高的反应性,且针对EV71及PV两种病毒的IgG抗体的相对含量成正相关;蛋白免疫印迹实验显示PV-VP1与EV71-VP1免疫的小鼠抗血清中存在交叉抗体,竞争ELISA试验进一步表明EV71-VP1蛋白能明显抑制PV-VP1与其特异性免疫血清抗体的结合,反之亦然。但中和实验揭示PV阳性血清在体外不能中和EV71病毒。PV之间存在大量交叉反应性抗体,但PV免疫后产生的与EV71交叉反应的抗体是非中和性质抗体。非中和抗体可能通过免疫调理效应在EV71病毒的致病的过程中发挥重要作用。

The aim of this study was to investigate the cross-reactivity between the antibodies against enterovirus 71（EV71） and poliovirus（PV）.Reactivity of the two viruses to IgG antibodies specific to EV71 or PV in serum samples from children were tested respectively using ELISA assay;neutralizating effect of PV-seropositive human serum against EV71 was detected with microtiter neutralization assay;VP1 and 3C proteins were expressed in E.coli and purified with affinity purification.Antisera were prepared from the BALB/c mice immunized subcutaneously with PV-VP1,formalin-inactivated EV71 and PV vaccine.Reactivity of anti-serum of PV-VP1（or EV71-VP1） to EV71-VP1（or PV-VP1） and inhibitory effect of EV71-VP1 on the binding of PV-VP1 to its specific IgG were investigated by Western blot and ELISA assay,respectively.The sera of healthy children who has immunized by OPV but not infected by EV71 display high reactivity to EV71 and the levels of EV71-specific and PV-specific antibodies in serum samples of EV71-seronegative children correlate well with each other.In addition,ELISA assay shows EV71 VP1 protein has a marked inhibition effect on the binding of PV VP1 to specific IgG antibodies,and vice versa.The cross-reactivity was also demonstrated in a Western blot assay with anti-sera made from PV-VP1-immunized mice.However,the antibodies against PV do not have crossreactive neutralizing activity against EV71.In conclusion,PV immunization elicits cross-reactive,non-neutralizing antibodies against EV71.These cross-reactive antibodies may play an important role in pathogenesis of EV71 infection through opsonization.