1. NOD.H-2～（h4）鼠T细胞克隆的研究 2. HPV，HSV基因工程预防疫苗的构建

发布时间：2018-02-28 17:44

本文关键词： 树突状细胞 T细胞克隆 NOD.H-2~(h4)鼠　出处：《山西医科大学》2006年博士论文　论文类型：学位论文

【摘要】： 目的实验性免疫性甲状腺炎是自身免疫性疾病研究的一个典型实例。为了开创一个研究的新途经,阐明T、B细胞在实验性免疫性甲状腺炎中的免疫作用及DC递呈Tg产生的T细胞克隆的免疫作用,阐明实验性免疫性甲状腺炎的诱发机制,我们以甲状腺球蛋白(Thyroglobulin,Tg)为抗原,树突状细胞为抗原递呈细胞建立了抗原特异性的T细胞克隆,并进行了被动转移实验。材料及方法 NOD.H-2h4鼠饲养于SPF无菌环境中,喂养0.05%碘化钠8周后,每两周采血,用ELISA测Tg水平,然后每只小鼠用100ug Tg的剂量混匀于福氏完全佐剂中免疫小鼠两次,用淋巴细胞增殖实验优化Tg的含量,寻找树突状细胞摄取递呈Tg的最适抗原量,然后,将免疫后NOD.H-2h4小鼠的脾细胞作为T细胞与同基因鼠的并与抗原孵育过的树突状细胞共培养。同时选用这六个T细胞克隆中的四个T细胞克隆---F11,2D11,H8,G8,将NOD.H-2h4鼠分为X射线照射组和正常NOD.H-2h4鼠组,进行了T细胞克隆的被动转移实验。结果建立了六个Tg特异性的T细胞克隆,这些细胞克隆在体外培养时间大于两年以上,流式细胞仪检测其表型均为CD4+,而且分泌IFN-γ,不分泌IL-4。T细胞克隆被动转移实验表明:所有的四个T细胞克隆均不能引起正常NOD.H-2h4鼠的甲状腺炎。四个克隆中三个T细胞克隆可引起X线照射组NOD.H-2h4鼠的甲状腺炎,而小鼠的照射剂量(475拉德)足够去除受鼠的抑制性细胞的功能。2D11克隆被动转移的结果亦表明:T细胞克隆在体内有辅助B细胞产生抗体的能力。结论抗原特异性的T细胞克隆的建立产生及维持,开创了一个研究自身免疫性疾病的新途经。树突状细胞起抗原递呈细胞的作用,在T细胞系及T细胞克隆的建立中是极为重要的,否则,T细胞即使在Tg及IL-2存在的培养环境中也仅能存活数天。T细胞克隆被动转移的实验,除了可以了解病理产生过程,转移的T、B细胞存活的时间长短外,也有助于了解T、B细胞的免疫调节作用及自身免疫性甲状腺炎的诱发机制,为研究实验性免疫性甲状腺炎的病理产生过程提供了理论依据和实验手段。
[Abstract]:Purpose. Experimental immune thyroiditis is a typical example of autoimmune disease. To elucidate the immune function of ThyroglobulinTg in experimental thyroiditis and the immune effect of T cell clone produced by DC, and to elucidate the inducing mechanism of ThyroglobulinTg, we take ThyroglobulinTg as antigen. Antigen-specific T cell clones were established from dendritic cells as antigen presenting cells and passive transfer experiments were carried out. Materials and methods. NOD.H-2h4 mice were fed in SPF aseptic environment. After feeding 0.05% sodium iodide for 8 weeks, blood samples were collected every two weeks, TG levels were measured with ELISA, and then each mouse was mixed with 100ug TG for twice in Freund's complete adjuvant. Lymphocyte proliferation test was used to optimize the content of TG, to find the optimal antigen quantity for dendritic cells to absorb and present TG, and then, The spleen cells of immunized NOD.H-2h4 mice were co-cultured with dendritic cells of allogeneic mice and incubated with antigens. At the same time, four of the six T cell clones, F112D11D11H8G8, were used to divide NOD.H-2h4 mice into X-rays. Shooting group and normal NOD.H-2h4 group, The passive transfer test of T cell clone was carried out. Results. Six TG-specific T cell clones were established, which were cultured for more than two years in vitro. The phenotypes of these clones were detected as CD4 by flow cytometry. Moreover, the passive transfer assay of secreting IFN- 纬 and non-secreting IL-4.T cell clone showed that none of the four T cell clones could induce thyroiditis in normal NOD.H-2h4 mice, and three of the four clones could induce thyroiditis in NOD.H-2h4 mice exposed to X-rays. The results of passive transfer of mouse inhibitory cells. 2D11 clones showed that the clone of T cells had the ability to assist B cells to produce antibodies in vivo. Conclusion. The establishment and maintenance of antigen-specific T cell clones has opened up a new way to study autoimmune diseases. Dendritic cells act as antigen-presenting cells. It is very important in the establishment of T cell line and T cell clone, otherwise T cell can survive for only a few days in the culture environment of TG and IL-2. In addition to the survival time of metastatic TG-B cells, it is helpful to understand the immunomodulatory effect of TG-B cells and the inducing mechanism of autoimmune thyroiditis. It provides theoretical basis and experimental means for studying the pathological process of experimental immune thyroiditis.
【学位授予单位】：山西医科大学
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R392;R581.4

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