EV71灭活疫苗交叉保护性及其与甲肝联合疫苗免疫效果的初步评价

发布时间：2018-01-01 23:16

本文关键词：EV71灭活疫苗交叉保护性及其与甲肝联合疫苗免疫效果的初步评价　出处：《北京协和医学院》2012年硕士论文　论文类型：学位论文

【摘要】：肠道病毒71型(Enterovirus71, EV71)是手足口病(hand, foot and mouth disease, HFMD)的主要病原体,病毒感染多发于婴幼儿,表现为手足口病、无菌性脑膜炎、急性驰缓性麻痹、脑干脑炎和其他一些严重的神经系统疾病,包括肺水肿和心肺功能障碍。近年来,我国EV71感染出现流行趋势,2008年国家已将手足口病纳入丙类传染病管理,预防EV71感染、降低发病率成为重要的公共卫生问题。疫苗接种是预防控制最有效的方法,中国医学科学院医学生物学研究所筛选出EV71FY-23K-B株作为毒种研制成人用灭活疫苗。由于EV71存在不同的亚型,不同株、不同亚型间是否有交叉保护作用关系到疫苗的预防效果。为充分了解我所疫苗对于其他不同EV71毒株的交叉保护性,本研究应用FY-23K-B株免疫血清对收集的K9、S1、R20-9J-3、S19、合49、BrCr等20个EV71不同毒株,同时对同属的脊髓灰质炎病毒(polio Ⅰ/Ⅱ/Ⅲ)、柯萨奇A组16型(CoxA16)、埃可病毒2/6型(ECHO2/6)等8株非EV71肠道病毒株进行中和试验。结果表明FY-23K-B株制备的疫苗对于A型株和不同的C4亚型株表现出广谱抗原性,其中4U(中和抗体)的特异性免疫血清已具有显著的中和效应,而2U血清中和效力相对较低,只能中和约35%的毒株；没有发现该株血清对其它8个非EV71的肠道病毒株的交叉保护作用。开发联合疫苗是疫苗发展的方向,为此我们就EV71与甲型肝炎病毒(Hepatitis A virus,HAV)灭活疫苗联合免疫的可行性进行了研究。我们将EV71(EU/ml)/HAV (EU/ml)进行不同的组合成为640/320、640/160、640/80、160/1280、160/640、320/640、640/640共7个联苗组,免疫ICR小鼠,并与相同剂量的单价疫苗组进行对比。初免后28天和加强后7天检测机体的特异性体液免疫和细胞免疫应答水平,结果发现初免后28天各组小鼠100%产生了保护性HAV抗体,160/640(EV71/HAV)组最高(4676.6mIU/ml),最低组为HAV80EU/ml单价组(131.5mIU/ml),加强后抗体平均增长12.2倍,其中160/640(EV71/HAV)组高达28101.8mIU/ml,但该组与320/640、640/640组间差异无统计学意义(P0.05)。初免后血清中EV71抗体阳转率即达到100%,各组平均GMT值为14.5,并表现出剂量效应,其中640/80(EV71/HAV)组抗体水平最高,但其与640/640、640/320、640/160组间差别无统计学意义(P0.05)。流式细胞术检测发现初免后各组小鼠脾脏中CD8+细胞比例高于CD4+细胞(P0.001),加强后则相反(P0.001),而HAV全病毒和EV71多肽刺激脾细胞产生的阳性反应弱,加强后脾细胞分泌的IL-4高于IFN-γ(P0.05)。说明一定剂量的HAV和EV71联合免疫可诱导机体产生良好的体液免疫应答,但细胞免疫应答较弱,就体液免疫应答而言,我们认为640/640(EV71/HAV)组的剂量配比是比较理想的；未发现两种抗原间存在相互干扰或协同作用。
[Abstract]:Enterovirus 71 (Enterovirus71, EV71) is HFMD (hand foot, and mouth disease, HFMD) of the main pathogens, virus infection in infants, manifested as HFMD, aseptic meningitis, acute flaccid paralysis, brain stem encephalitis and other serious neurological diseases, including pulmonary edema and heart and lung dysfunction. In recent years, China's EV71 infection epidemic trend, 2008 has been included HFMD class C infectious disease management and prevention of EV71 infection, reduce the incidence become an important public health problem.
Vaccination is the most effective method of prevention and control, medical and Biological Research Academy of Medical Sciences Chinese selected EV71FY-23K-B strain of virus inactivated vaccine development for adult. Due to the existence of EV71 subtype, different strains, whether there is cross protection in relation to the vaccine prevention effect in different subtypes. In order to fully understand what I vaccine for cross protection of different EV71 strains, FY-23K-B strains used in this study to collect the serum K9, S1, R20-9J-3, S19, 49 BrCr, 20 EV71 strains, also belong to the polio virus (polio I / II / III), coxsackievirus group A type 16 (CoxA16), ECHO virus type 2/6 (ECHO2/6) and 8 strains of non EV71 strains of intestinal virus neutralization test. The results showed that FY-23K-B strain vaccine prepared for type A strains and different C4 subtype strains showed broad-spectrum antigenicity, 4U (neutralizing antibody) specific immune Serum has a significant neutralization effect, while the neutralizing potency of 2U is relatively low, which only neutralizes about 35% of the strains. No cross protection effect of the serum on other 8 non EV71 enterovirus strains was found.
The development of the combined vaccine is a vaccine development direction, therefore we EV71 and hepatitis A virus (Hepatitis A virus, HAV) inactivated vaccine combined with immune feasibility is studied. We will EV71 (EU/ml) /HAV (EU/ml) by different combinations of 640/320640/160640/ 80160/1280160/640320/640640/640 a total of 7 vaccine group, ICR mice were immunized at and with the same dose of vaccine group were compared. After immunization 28 days and 7 days after the body to strengthen detection of specific humoral and cellular immune response level, results showed that 28 days after immunization of mice produced 100% protective HAV antibody, 160/640 (EV71/HAV) was the highest (4676.6mIU/ml), HAV80EU/ml group was the lowest the unit group (131.5mIU/ml), anti average growth of 12.2 times, including 160/640 (EV71/HAV) group up to 28101.8mIU/ml, but there was no significant difference between the group and the 320/640640/640 group (P0.05) . after immunization of serum EV71 antibody positive rate was 100%, the average GMT value of 14.5, and showed a dose effect of 640/80 (EV71/HAV) group, the highest antibody level, but the difference between the 640/640640/320640/160 group had no statistical significance (P0.05). Flow cytometry found after immunization of CD8+ cells the proportion of mice spleen cells was higher than that of CD4+ (P0.001), strengthen the contrary (P0.001), and the positive reaction of HAV virus and EV71 polypeptide stimulated spleen cells produced weak spleen cells secrete IL-4 after strengthening is higher than that of IFN- gamma (P0.05). A dose of HAV and EV71 combined immunization can induce good humoral immune responses, but the cellular immune response is weak, humoral immune response, we believe that the 640/640 (EV71/HAV) dose ratio group is ideal; not found two antigens exist between mutual interference and synergistic effect.

【学位授予单位】：北京协和医学院
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R392

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