弓形虫SAG1-ROP2复合基因疫苗与免疫佐剂大肠埃希菌肠毒素B亚基对小鼠的免疫保护性研究

发布时间：2018-06-21 06:39

本文选题：弓形虫 + 复合基因疫苗　；参考：《山东大学》2010年硕士论文

【摘要】： 弓形虫是一种广泛寄生于人和动物有核细胞内的寄生原虫,可以感染包括人类在内的所有哺乳动物,此虫呈世界性分布,人群感染率相当高,许多国家的平均感染率在25%-50%。弓形虫侵袭的细胞种类多,累及器官广泛,临床表现复杂。在免疫抑制或免疫缺陷患者中(如器官移植、恶性肿瘤、AIDS、长期使用免疫抑制剂等)可引起严重的器官损害(多见于脑和眼)。先天性感染是弓形虫通过母婴传播的,可引起胎儿流产、死胎或畸胎,是造成儿童先天畸形、弱智及智力发育迟缓的重要原因之一。此外,弓形虫病也给畜牧业生产造成严重的经济损失。迄今尚无治疗弓形虫病的特效药物,因此研制安全有效的疫苗极为迫切。大量的动物和人体实验表明,DNA疫苗不仅可以预防,还可以治疗病原生物的感染。由于弓形虫生活史较复杂,抗原成分具有发育阶段的特异性或差异性。因此在研究有关弓形虫疫苗的过程中,发展多种抗原组合、针对不同生活史发育阶段的复合多价疫苗是研究弓形虫疫苗过程中的一个发展方向与共识,这有助于克服单一抗原成分作为候选疫苗的不足。 SAG1抗原作为弓形虫速殖子表面的主要抗原之一,具有高度的免疫原性和免疫保护性,是迄今应用最多的弓形虫疫苗候选抗原。SAG1蛋白分布于弓形虫速殖子表膜、速殖子内以及纳虫泡的管状结构中,是诱导宿主免疫应答的主要靶抗原。能够诱导机体产生IgG, IgM, IgE, IgA及sIgA,并可诱导细胞因子产生。 ROP2蛋白是弓形虫棒状体分泌的一种蛋白,存在于弓形虫速殖子、缓殖子及包囊期,它可诱导机体产生以IgA, IgM, IgG为主的体液免疫,还可被人特异性T细胞克隆识别而产生高水平的IFN-γ。SAGl和ROP2在虫体入侵宿主的过程中相互促进,其相应的抗体可有效拮抗弓形虫感染。 DNA疫苗诱导体液免疫和细胞免疫的效率相对较低,尤其是在大型动物和人类中,影响了它的实际应用。我们选用大肠埃希菌肠毒素B亚基(LTB)真核表达质粒作为基因佐剂,以增强弓形虫复合抗原基因SAG1-ROP2的免疫效果。 LT是由产肠毒素E. coli分泌并存在于菌体周质中的一种外毒素,由A、B两种亚单位组成,其中B亚单位既有较强的免疫原性,又有较强的黏膜佐剂效应,且毒力小、安全性高,是最有潜力的黏膜免疫佐剂之一。口服和鼻内接种LT均可诱导黏膜和全身性的免疫应答,有效增强抗原蛋白的免疫原性。该研究选用弓形虫复合抗原基因SAG1-ROP2及基因佐剂LTB通过黏膜方式免疫小鼠,观察并研究其免疫保护性。结果表明复合基因疫苗的免疫效果明显优于单基因疫苗SAG1及ROP2, LTB可以对该复合基因疫苗起到很好的免疫增强作用。在弓形虫研究领域,国内外尚无此项研究报道。
[Abstract]:Toxoplasma gondii (Toxoplasma gondii) is a parasitic protozoa widely parasitic in the nucleated cells of humans and animals. It can infect all mammals including human beings. The infection rate of Toxoplasma gondii is very high in the world. The average infection rate in many countries ranges from 25 to 50. Toxoplasma gondii invades many types of cells, involving a wide range of organs, and complex clinical manifestations. In immunosuppressive or immunodeficient patients (e.g. organ transplantation, malignant tumor AIDS, long-term use of immunosuppressive agents, etc.), severe organ damage can be caused (more commonly seen in the brain and eyes). Congenital infection is transmitted from mother to child by Toxoplasma gondii, which can cause abortion, stillbirth or teratogenesis. It is one of the important causes of congenital malformation, mental retardation and mental retardation in children. In addition, toxoplasmosis also causes serious economic loss to animal husbandry production. There is no effective drug for Toxoplasma gondii so it is urgent to develop a safe and effective vaccine. A large number of animal and human experiments have shown that DNA vaccine can not only prevent, but also treat the infection of pathogenic organisms. Because of the complex life history of Toxoplasma gondii, the antigen components have the specificity or difference of development stage. Therefore, in the research of Toxoplasma gondii vaccine, the development of a variety of antigen combinations, for different stages of life cycle development of the complex multivalent vaccine is a development direction and consensus in the research process of Toxoplasma gondii vaccine. SAG1 antigen, one of the major antigens on the surface of Toxoplasma gondii tachyzoites, is highly immunogenicity and immunogenicity. Toxoplasma gondii vaccine candidate antigen. SAG1 protein is widely used in Toxoplasma gondii Tachyzoites epidermis, tachyzoites and the tubular structure of nalcidia vesicles, and is the main target antigen to induce host immune response. It can induce the production of IgG, IgM, IgE, IgA and Siga, and induce cytokine production. ROP2 protein is a kind of protein secreted by Toxoplasma gondii rodlike body, which exists in Toxoplasma gondii tachyzoites, bradyzoites and cysts. It can induce humoral immunity mainly composed of IgA, IgM and IgG, and can be recognized by human specific T cell clone to produce high levels of IFN- 纬. SAGl and ROP2, which promote each other in the process of invading the host. The efficiency of DNA vaccine in inducing humoral and cellular immunity was relatively low, especially in large animals and humans, which affected its practical application. The eukaryotic expression plasmid of Escherichia coli enterotoxin B subunit LTB was used as gene adjuvant. In order to enhance the immune effect of Toxoplasma gondii complex antigen gene SAG1-ROP2, LT is an exotoxin secreted by enterotoxin E. coli, which is composed of two subunits of Agna B, and exists in the peripheral cytoplasm of Toxoplasma gondii (Toxoplasma gondii). The B subunit has both strong immunogenicity and strong mucosal adjuvant effect. It is one of the most potential mucosal adjuvants with low virulence and high safety. Oral and nasal inoculation with LT could induce mucosal and systemic immune responses and enhance the immunogenicity of antigenic proteins. In this study mice were immunized with SAG1-ROP2 and LTB by mucosal method to observe and study the immune protection of Toxoplasma gondii complex antigen gene SAG1-ROP2 and gene adjuvant LTB. The results showed that the immune effect of the composite gene vaccine was significantly better than that of the single gene vaccine SAG1 and ROP2LTB could enhance the immune response of the vaccine. In the field of Toxoplasma gondii, there is no such research report at home and abroad.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R392

【引证文献】