【摘要】：日本血吸蟲病(Schistosomiasis japonicium)是一種嚴(yán)重危害人、畜健康,影響社會和經(jīng)濟(jì)發(fā)展的人畜共患寄生蟲病。近年來我國血吸蟲病防治工作取得了舉世矚目的成就,但由于血吸蟲病流行的生態(tài)環(huán)境沒有得到根本改變,傳染源控制困難,血吸蟲病在江湖川漢地區(qū)流行仍較嚴(yán)重。作為滅螺、藥物防治和其他綜合防治措施的重要補(bǔ)充,血吸蟲病疫苗的研究就顯得十分必要。迄今研制的一些日本血吸蟲疫苗保護(hù)效果還不夠理想,免疫佐劑是探索提高疫苗保護(hù)效果的有效途徑之一。 本論文應(yīng)用已經(jīng)在動物實(shí)驗中證明可誘導(dǎo)較高免疫保護(hù)作用的重組蛋白SjGCP和Sj28GST與3種佐劑(弗式佐劑、ISA206佐劑、ISA70M佐劑)配伍免疫BALB/c小鼠,再攻擊感染日本血吸蟲(尾蚴40±2條),收集成蟲和檢測每克肝臟蟲卵數(shù),計算減蟲率和肝臟減卵率。結(jié)果在動物實(shí)驗中,兩種重組抗原均誘導(dǎo)了部分的抗日本血吸蟲感染的免疫保護(hù)效果。 本論文進(jìn)一步利用流式細(xì)胞儀技術(shù)(FCM)和免疫酶聯(lián)吸附法(ELISA)檢測抗原免疫的小鼠機(jī)體細(xì)胞和體液免疫情況,對這兩種抗原結(jié)合三種佐劑誘導(dǎo)的免疫保護(hù)機(jī)制進(jìn)行初步的研究。通過流式細(xì)胞術(shù)檢測了第3次免疫1周后的小鼠的CD4+、CD8+細(xì)胞,細(xì)胞內(nèi)細(xì)胞因子IL2、IL4、IL10、IFNγ及IL12。通過ELISA方法檢測免疫前、第3次免疫小鼠1周后以及尾蚴攻擊6周后血清中的抗體IgG及其亞型IgG、IgG2a、IgG2b、IgG3,抗體IgE、IgA、IgM的水平。與空白組相比,免疫組CD4+、CD8+的數(shù)值以及CD4+/CD8+比值,細(xì)胞因子IL2、IL4、IL10、IFNγ的數(shù)值的變化,說明了免疫組的小鼠機(jī)體誘導(dǎo)產(chǎn)生了Thl/Th2型混合的細(xì)胞免疫反應(yīng),且Thl型占優(yōu)勢地位。ELISA結(jié)果顯示,重組蛋白SjGCP-GST和Sj28GST引起了小鼠機(jī)體較為強(qiáng)烈的體液免疫應(yīng)答,抗體類型主要是IgG。本實(shí)驗說明了重組蛋白SjGCP和Sj28GST免疫保護(hù)效果是細(xì)胞免疫和體液免疫共同作用的結(jié)果。本文數(shù)據(jù)為抗日本血吸蟲病疫苗的成功研制,提供了具有一定意義的參考。
[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 鹵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 緯 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 緯 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.
【學(xué)位授予單位】：上海師范大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R392

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