DO11.10小鼠OVA免疫耐受模型及耐受机制研究

发布时间：2018-01-08 02:14

本文关键词：DO11.10小鼠OVA免疫耐受模型及耐受机制研究　出处：《西北农林科技大学》2011年硕士论文　论文类型：学位论文

更多相关文章： DO11.10小鼠 卵清蛋白 外周免疫耐受

【摘要】：免疫耐受是机体免疫系统对特定抗原的免疫无应答或低应答现象,依据形成时间、器官不同,可分为中枢免疫耐受和外周免疫耐受。外周免疫耐受由成熟T、B淋巴细胞在外周免疫器官经克隆清除、克隆无能、免疫忽视及多种调节性T细胞作用形成。外周免疫耐受与人类、动物多种疾病密切相关。生理性免疫耐受打破后,易引起多种自身免疫病,如系统性红斑狼疮、多发性硬化症、Ⅰ型糖尿病等;避免移植物排斥反应也需建立抗原特异性免疫耐受;对病毒及肿瘤抗原产生免疫耐受,会引起机体发生肿瘤病及病毒病,临床治疗这些疾病则需要打破机体对肿瘤或病毒抗原的免疫耐受。上述问题的解决,必须以阐明机体外周免疫耐受形成维持或打破的细胞和分子机制为前提。因此,建立小鼠针对特定非己抗原的外周免疫耐受模型,并以这种模型为基础研究其形成的机理有着重要的理论和实践意义。 DO11.10小鼠经分子生物学手段以BALB/c小鼠为背景,特异性转入识别OVA323-339肽段的TCR基因而建立的常用的免疫耐受研究小鼠。国内外目前以该小鼠为模型,在黏膜免疫耐受方面做了很多工作,但该品系小鼠整体外周免疫耐受方面的研究报道不多。为此我们开展了本研究,获得的研究结果如下: 1.经尾静脉途经注射OVA抗原3次,每次间隔5d,再用OVA皮下免疫。结果显示,试验组DO11.10小鼠脾淋巴细胞对OVA抗原刺激后的增殖能力远低于对照小鼠,但其淋巴细胞的SI值要高于同样处理的BALB/c小鼠;流式细胞术测定结果显示,试验组DO11.10小鼠脾脏CD4~+CD25~+T比例和数量显著高于对照组小鼠,同样处理的BALB/c小鼠CD4~+CD25~+T比例和数量变化不如DO11.10小鼠高。结果表明,本研究成功建立了DO11.10小鼠的OVA免疫耐受模型,这种模型小鼠对OVA抗原的高敏感性与人工导入的TCR基因片段紧密相关。 2.分别用CD4~+T细胞表位肽(OVA323-339,KISQAVHAAHAEINEAG)、CTL表位肽(OVA257-264,SIINFEKL)刺激DO11.10小鼠脾淋巴细-胞,结果显示,CD4~+T细胞表位肽刺激的试验组淋巴细胞增殖能力远低于对照,CTL表位肽的相应结果在试验组和对照组之间差异不大;CD4~+T细胞表位肽刺激的淋巴细胞分泌TGF-β量远高于对照组,但试验组和对照组小鼠脾淋巴细胞分泌IL-10的水平没有显著差异。结果表明,OVA免疫耐受的DO11.10小鼠外周免疫耐受模型建立是通过抑制/杀伤OVA特异性T细胞这一淋巴细胞克隆而实现的,其中TGF-β而非IL-10发挥了重要作用。
[Abstract]:Immune tolerance is a phenomenon that the immune system has no or low response to a specific antigen. According to the time of formation, different organs can be divided into central immune tolerance and peripheral immune tolerance. Peripheral immune tolerance by mature T. B lymphocytes were cloned and removed from peripheral immune organs, clonal incompetence, immune neglect and the formation of various regulatory T cells, peripheral immune tolerance and human. When physiological immune tolerance is broken, it is easy to cause many autoimmune diseases, such as systemic lupus erythematosus, multiple sclerosis, type I diabetes, etc. Antigen-specific immune tolerance is also needed to avoid graft rejection. Immune tolerance to viruses and tumor antigens will lead to oncology and viral diseases. Clinical treatment of these diseases needs to break the immune tolerance of tumor or virus antigen. It is necessary to elucidate the cellular and molecular mechanisms of maintenance or breakage of peripheral immune tolerance. Therefore, a model of peripheral immune tolerance for specific non-hexane antigens in mice should be established. On the basis of this model, it has important theoretical and practical significance to study the mechanism of its formation. The background of DO11.10 mice was BALB/c mice by molecular biological methods. The commonly used immune tolerance study mice established by specific transfer of the TCR gene to recognize the OVA323-339 peptide have done a lot of work on mucosal immune tolerance with this mouse as a model at home and abroad. However, there are few reports on the whole peripheral immune tolerance of this strain of mice. Therefore, we have carried out this study, and the results are as follows: 1. OVA antigen was injected through caudal vein three times every 5 days, then immunized with OVA subcutaneously. The proliferation ability of spleen lymphocytes stimulated by OVA antigen in DO11.10 mice was much lower than that in control mice, but the SI value of lymphocytes was higher than that of BALB/c mice treated with the same treatment. The results of flow cytometry showed that the ratio and quantity of CD4 ~ CD25 ~ T in spleen of DO11.10 mice in the test group was significantly higher than that in the control group. The ratio and quantity of CD4 ~ CD25 ~ T in BALB/c mice treated with the same treatment were lower than those in DO11.10 mice. In this study, the OVA immune tolerance model of DO11.10 mice was successfully established. The high sensitivity to OVA antigen in the model mice was closely related to the TCR gene fragment introduced artificially. 2. OVA257-264 was used as CTL epitope peptide of CD4 ~ T cell epitope (OVA323-339). SI-INFEKL stimulated splenic lymphatic fine cell in DO11.10 mice. The results showed that the lymphocyte proliferation ability of the experimental group stimulated by CD4 ~ T cell epitope peptide was much lower than that of the control group. The corresponding results of CTL epitope peptide were not significantly different between the experimental group and the control group. The level of TGF- 尾 secreted by CD4- T cell epitope peptide was much higher than that of control group, but there was no significant difference between the experimental group and the control group. The peripheral immune tolerance model of DO11.10 mice with OVA tolerance was established by inhibiting / killing the lymphocyte clone of OVA specific T cells. TGF- 尾 and not IL-10 play an important role.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R392.9

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