本文關鍵詞：DO11.10小鼠OVA免疫耐受模型及耐受機制研究　出處：《西北農林科技大學》2011年碩士論文　論文類型：學位論文

  更多相關文章： DO11.10小鼠 卵清蛋白 外周免疫耐受

【摘要】：免疫耐受是機體免疫系統對特定抗原的免疫無應答或低應答現象,依據形成時間、器官不同,可分為中樞免疫耐受和外周免疫耐受。外周免疫耐受由成熟T、B淋巴細胞在外周免疫器官經克隆清除、克隆無能、免疫忽視及多種調節(jié)性T細胞作用形成。外周免疫耐受與人類、動物多種疾病密切相關。生理性免疫耐受打破后,易引起多種自身免疫病,如系統性紅斑狼瘡、多發(fā)性硬化癥、Ⅰ型糖尿病等;避免移植物排斥反應也需建立抗原特異性免疫耐受;對病毒及腫瘤抗原產生免疫耐受,會引起機體發(fā)生腫瘤病及病毒病,臨床治療這些疾病則需要打破機體對腫瘤或病毒抗原的免疫耐受。上述問題的解決,必須以闡明機體外周免疫耐受形成維持或打破的細胞和分子機制為前提。因此,建立小鼠針對特定非己抗原的外周免疫耐受模型,并以這種模型為基礎研究其形成的機理有著重要的理論和實踐意義。 DO11.10小鼠經分子生物學手段以BALB/c小鼠為背景,特異性轉入識別OVA323-339肽段的TCR基因而建立的常用的免疫耐受研究小鼠。國內外目前以該小鼠為模型,在黏膜免疫耐受方面做了很多工作,但該品系小鼠整體外周免疫耐受方面的研究報道不多。為此我們開展了本研究,獲得的研究結果如下: 1.經尾靜脈途經注射OVA抗原3次,每次間隔5d,再用OVA皮下免疫。結果顯示,試驗組DO11.10小鼠脾淋巴細胞對OVA抗原刺激后的增殖能力遠低于對照小鼠,但其淋巴細胞的SI值要高于同樣處理的BALB/c小鼠;流式細胞術測定結果顯示,試驗組DO11.10小鼠脾臟CD4~+CD25~+T比例和數量顯著高于對照組小鼠,同樣處理的BALB/c小鼠CD4~+CD25~+T比例和數量變化不如DO11.10小鼠高。結果表明,本研究成功建立了DO11.10小鼠的OVA免疫耐受模型,這種模型小鼠對OVA抗原的高敏感性與人工導入的TCR基因片段緊密相關。 2.分別用CD4~+T細胞表位肽(OVA323-339,KISQAVHAAHAEINEAG)、CTL表位肽(OVA257-264,SIINFEKL)刺激DO11.10小鼠脾淋巴細-胞,結果顯示,CD4~+T細胞表位肽刺激的試驗組淋巴細胞增殖能力遠低于對照,CTL表位肽的相應結果在試驗組和對照組之間差異不大;CD4~+T細胞表位肽刺激的淋巴細胞分泌TGF-β量遠高于對照組,但試驗組和對照組小鼠脾淋巴細胞分泌IL-10的水平沒有顯著差異。結果表明,OVA免疫耐受的DO11.10小鼠外周免疫耐受模型建立是通過抑制/殺傷OVA特異性T細胞這一淋巴細胞克隆而實現的,其中TGF-β而非IL-10發(fā)揮了重要作用。
[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.
【學位授予單位】：西北農林科技大學
【學位級別】：碩士
【學位授予年份】：2011
【分類號】：R392.9

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本文編號：1395203