细粒棘球蚴重组抗原Eg10和mMDH致树突状细胞免疫耐受的机制研究

发布时间：2018-04-11 22:12

  本文选题：细粒棘球蚴 + Eg10　； 参考：《宁夏医科大学》2015年硕士论文

【摘要】：目的初步明确细粒棘球蚴重组抗原Eg10和m MDH免疫小鼠产生的免疫耐受机制。方法1.体外培养小鼠骨髓来源的树突状细胞(Dendritic cell,DC),用Eg10和m MDH体外刺激DCs,通过扫描电镜观察DCs的形态变化,细胞免疫荧光观察DCs表达吲哚胺2,3双加氧酶(Indoleamine 2,3-dioxygenase,IDO)的水平;2.利用IDO抑制剂1-甲基色氨酸(1-Methyl-tryptophan,1-MT)处理DCs 3h,用来抑制IDO的表达及酶活性,然后用重组抗原Eg10和m MDH体外刺激DCs,通过混合淋巴细胞反应检测DCs刺激CD4+T细胞增殖及诱导CD4+CD25+FOXP3+调节性T细胞(Regulatory T cell,Treg)生成的能力;Q-PCR检测DCs中多种细胞因子的表达;流式细胞术(FCM)检测DC表面分子CD80/CD86、MHCII、CD40的表达情况。结果1.重组抗原Eg10和m MDH不能刺激DCs表面树突的生成,DCs呈现不成熟状态;2.重组抗原Eg10和m MDH刺激DCs后,IDO的表达增强,上清中犬尿氨酸的含量也升高,且1-MT能特异性降低IDO的表达水平和酶活性;3.与1-MT未处理的m MDH组相比,1-MT处理的m MDH组中DCs刺激CD4+T细胞增殖的能力升高,诱导CD4+CD25+FOXP3+Tregs生成的能力降低,差异有统计学意义;而Eg10组对1-MT的反应不明显;4.重组抗原Eg10和m MDH刺激DCs后,TNF-α、IL-6、IL-10 m RNA水平的表达呈不同程度的升高,1-MT处理的抗原组TNF-αm RNA表达升高,IL-6、IL-10m RNA表达降低;5.重组抗原Eg10和m MDH刺激DCs后,细胞表面分子的表达水平较低,1-MT处理并不能提高表面分子的表达水平。结论1.细粒棘球蚴重组抗原Eg10和m MDH体外刺激小鼠骨髓来源的DCs,不能使其发育成熟,DCs刺激CD4+T细胞增殖的能力较弱;2.细粒棘球蚴重组抗原m MDH通过刺激DCs表达高水平的IDO,引起免疫耐受。3.细粒棘球蚴重组抗原Eg10不是通过刺激DCs表达IDO引起免疫耐受的,可能存在其他机制。
[Abstract]:Objective to investigate the mechanism of immune tolerance induced by recombinant antigens Eg10 and m MDH of echinococcus granulosus in mice.Method 1.Dendritic cells derived from mouse bone marrow were cultured in vitro. DCS were stimulated with Eg10 and m MDH in vitro. The morphological changes of DCs were observed by scanning electron microscope. The expression of Indoleamine 23-dioxygenase (IDO) in DCs was observed by immunofluorescence.DCs was treated with 1-Methyl-tryptophane 1-MT-1, a IDO inhibitor, for 3 h to inhibit the expression and enzyme activity of IDO.Then the recombinant antigen Eg10 and m MDH were used to stimulate DCS in vitro, and the ability of DCs to stimulate the proliferation of CD4 T cells and to induce the production of CD4 CD25 FOXP3 regulatory T cell Treg-T cells was detected by mixed lymphocyte reaction. Q-PCR was used to detect the expression of various cytokines in DCs.Flow cytometry (FCM) was used to detect the expression of CD80 / CD86 and MHCII- 鈪，

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