日本血吸虫重组抗原疫苗SjGCP-GST和Sj28 GST免疫保护力和免疫机制初步研究

发布时间：2018-11-12 20:58

【摘要】：日本血吸虫病(Schistosomiasis japonicium)是一种严重危害人、畜健康,影响社会和经济发展的人畜共患寄生虫病。近年来我国血吸虫病防治工作取得了举世瞩目的成就,但由于血吸虫病流行的生态环境没有得到根本改变,传染源控制困难,血吸虫病在江湖川汉地区流行仍较严重。作为灭螺、药物防治和其他综合防治措施的重要补充,血吸虫病疫苗的研究就显得十分必要。迄今研制的一些日本血吸虫疫苗保护效果还不够理想,免疫佐剂是探索提高疫苗保护效果的有效途径之一。本论文应用已经在动物实验中证明可诱导较高免疫保护作用的重组蛋白SjGCP和Sj28GST与3种佐剂(弗式佐剂、ISA206佐剂、ISA70M佐剂)配伍免疫BALB/c小鼠,再攻击感染日本血吸虫(尾蚴40±2条),收集成虫和检测每克肝脏虫卵数,计算减虫率和肝脏减卵率。结果在动物实验中,两种重组抗原均诱导了部分的抗日本血吸虫感染的免疫保护效果。本论文进一步利用流式细胞仪技术(FCM)和免疫酶联吸附法(ELISA)检测抗原免疫的小鼠机体细胞和体液免疫情况,对这两种抗原结合三种佐剂诱导的免疫保护机制进行初步的研究。通过流式细胞术检测了第3次免疫1周后的小鼠的CD4+、CD8+细胞,细胞内细胞因子IL2、IL4、IL10、IFNγ及IL12。通过ELISA方法检测免疫前、第3次免疫小鼠1周后以及尾蚴攻击6周后血清中的抗体IgG及其亚型IgG、IgG2a、IgG2b、IgG3,抗体IgE、IgA、IgM的水平。与空白组相比,免疫组CD4+、CD8+的数值以及CD4+/CD8+比值,细胞因子IL2、IL4、IL10、IFNγ的数值的变化,说明了免疫组的小鼠机体诱导产生了Thl/Th2型混合的细胞免疫反应,且Thl型占优势地位。ELISA结果显示,重组蛋白SjGCP-GST和Sj28GST引起了小鼠机体较为强烈的体液免疫应答,抗体类型主要是IgG。本实验说明了重组蛋白SjGCP和Sj28GST免疫保护效果是细胞免疫和体液免疫共同作用的结果。本文数据为抗日本血吸虫病疫苗的成功研制,提供了具有一定意义的参考。
[Abstract]:Schistosomiasis japonicum (Schistosomiasis japonicium) is a zoonotic parasitic disease that seriously harms human and animal health and affects social and economic development. In recent years, the control of schistosomiasis in China has made great achievements. However, because the ecological environment of schistosomiasis has not been fundamentally changed and the source of infection is difficult to control, schistosomiasis is still prevalent in Jianghu, Sichuan and Han areas. As an important supplement of snail control, drug control and other comprehensive control measures, the research of schistosomiasis vaccine is very necessary. The protective effect of some Schistosoma japonicum vaccines developed up to now is not ideal, and the immune adjuvant is one of the effective ways to improve the protective effect of the vaccine. In this paper, the recombinant proteins SjGCP and Sj28GST, which have been proved in animal experiments to induce higher immune protection, were used to immunize BALB/c mice with three adjuvants (Freund adjuvant, ISA206 adjuvant, ISA70M adjuvant). Then infected Schistosoma japonicum (40 卤2 cercariae), collected the adult worm and detected the egg number per gram liver, calculated the worm reduction rate and liver egg reduction rate. Results in animal experiments, two recombinant antigens induced partial immune protection against Schistosoma japonicum infection. The cellular and humoral immunity of antigen-immunized mice was detected by flow cytometry (FCM) and immunosorbent enzyme linked adsorption (ELISA). The mechanism of immune protection induced by these two antigens combined with three adjuvants was preliminarily studied. CD4, CD8 cells, intracellular cytokines IL2,IL4,IL10,IFN 纬 and IL12. were detected by flow cytometry in mice one week after the third immunization. The levels of antibody IgG and its subtype IgG,IgG2a,IgG2b,IgG3, antibody IgE,IgA,IgM were detected by ELISA method before immunization, 1 week after the third immunization and 6 weeks after cercariae attack. Compared with the control group, the changes of CD4, CD8, CD4 / CD8 ratio and cytokine IL2,IL4,IL10,IFN 纬 in the immunized group indicated that the immunized mice induced a mixed cellular immune response of Thl/Th2 type. The results of ELISA showed that the recombinant proteins SjGCP-GST and Sj28GST elicited a stronger humoral immune response in mice, and the main antibody type was IgG.. The results showed that the protective effect of recombinant protein SjGCP and Sj28GST was the result of both cellular and humoral immunity. The data provided a valuable reference for the successful development of Schistosomiasis japonicum vaccine.
【学位授予单位】：上海师范大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R392

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