可溶性HLA-G二聚体在体外抑制树突状细胞成熟和M1型巨噬细胞极化

发布时间：2018-05-14 22:15

本文选题：可溶性HLA-G二聚体 + 树突状细胞　；参考：《华中科技大学》2013年硕士论文

【摘要】：HLA-G是一种非经典的HLAⅠ类分子，可以通过与抑制性受体结合诱导广泛的免疫耐受。大量研究证实可溶性HLA-G二聚体与其抑制性受体的结合力更强，可以诱导更强的免疫耐受。这些抑制性受体包括ILT-4和ILT-2，主要表达在T细胞，B细胞，巨噬细胞，NK，DC等表面。DC是重要的抗原提呈细胞，连接了固有免疫与适应性免疫，它们可以发现、捕获，加工处理外源性抗原并提呈给T细胞。它们与原始T细胞的相互作用在诱导T细胞分化成Th1/Th2细胞、调节性T细胞中起重要作用。近年来有不少文献报道可溶性HLA-G二聚体能够通过与抑制性受体ILT-4结合抑制DC的成熟，但也有人报道可溶性HLA-G二聚体对DC的成熟没有影响。本研究拟探讨可溶性HLA-G二聚体对DC成熟的作用，以期这种二聚体可以抑制DC的成熟，，进一步诱导T细胞、B细胞的免疫耐受，降低移植排斥反应的程度。巨噬细胞参与和促进炎症反应，根据其微环境的不同可分为经典的M1与选择性激活的M2。前者与炎症反应和组织损伤有关；后者可促进血管生成和组织修复，并且参与寄生虫感染与过敏反应，在一定条件下两者可以相互转化。由于巨噬细胞表面也存在可溶性HLA-G二聚体的受体，所以本实验将探讨可溶性HLA-G二聚体对这两种细胞极化的影响，研究可溶性HLA-G二聚体在炎症反应中的作用。论文分为2个部分，主要内容与结果如下： 1.可溶性HLA-G二聚体在体外可抑制树突状细胞成熟由人外周血分离得到PBMC，贴壁单核细胞在GM-CSF、IL-4的作用下分化成未成熟DC，而TNF-或IFN-γ可促进DC成熟。同时在此过程中加入可溶性HLA-G二聚体，并设置721.221上清作为对照。用流式抗体CD1a，CD83，CD40染色，流式细胞仪检测各组细胞表面这三种分子的表达水平。检测结果显示：可溶性HLA-G二聚体组DC表面CD83及CD40表达水平比721.221上清对照组明显下降。 2.可溶性HLA-G二聚体在体外可抑制M1型巨噬细胞极化由人外周血分离得到PBMC，贴壁单核细胞在GM-CSF与LPS刺激下得到M1；用M-CSF与IL-4刺激得到M2。并在实验组加入可溶性HLA-G二聚体，并设置721.221上清作为对照。用流式抗体CD14，CD80，CD206染色，流式细胞仪检测M1组细胞表面CD14+CD80+的表达水平，M2组细胞表面CD14+CD206+的表达水平。流式细胞仪检测结果显示：可溶性HLA-G二聚体可以降低M1表面CD80的表达水平，而对M2表面的CD206表达没有影响。
[Abstract]:HLA-G is a nonclassical HLA class I molecule that can induce extensive immune tolerance by binding to inhibitory receptors. A large number of studies have confirmed that soluble HLA-G dimer has stronger binding power with its inhibitory receptor and can induce stronger immune tolerance. These inhibitory receptors, including ILT-4 and ILT-2, are mainly expressed in T cell B cells, and macrophages, such as NKDC, are important antigen presenting cells linking innate and adaptive immunity, which can be found, captured, and captured. Exogenous antigen was processed and presented to T cells. Their interaction with primordial T cells plays an important role in inducing T cells to differentiate into Th1/Th2 cells and regulate T cells. In recent years, it has been reported that soluble HLA-G dimer can inhibit DC maturation by binding with inhibitory receptor ILT-4, but it is also reported that soluble HLA-G dimer has no effect on DC maturation. The aim of this study was to investigate the effect of soluble HLA-G dimer on DC maturation in order to inhibit DC maturation, induce T cell B cell immune tolerance and reduce the degree of graft rejection. Macrophages participate in and promote inflammatory response, according to their microenvironment can be divided into classic M1 and selectively activated M 2. The former is related to inflammation and tissue damage, while the latter can promote angiogenesis and tissue repair, and participate in parasitic infection and allergic reaction, which can transform each other under certain conditions. Because there are soluble HLA-G dimer receptors on the surface of macrophages, the effect of soluble HLA-G dimer on these two cell polarization and the role of soluble HLA-G dimer in inflammatory response were studied. The paper is divided into two parts. The main contents and results are as follows: 1. Soluble HLA-G dimer inhibits dendritic cell maturation in vitro PBMCs were isolated from human peripheral blood. Adherent monocytes differentiated into immature DCs under the action of GM-CSF- IL-4, while TNF- or IFN- 纬 promoted DC maturation. At the same time, soluble HLA-G dimer was added in the process, and 721.221 supernatant was set as control. Flow cytometry was used to detect the expression of these three molecules on the surface of the cells. The results showed that the expression of CD83 and CD40 on DC surface in soluble HLA-G dimer group was significantly lower than that in 721.221 supernatant group. 2. Soluble HLA-G dimer inhibits the polarization of M1 macrophages in vitro PBMCs were isolated from human peripheral blood. Monocytes were stimulated by GM-CSF and LPS to obtain M1 and M2 stimulated by M-CSF and IL-4. The soluble HLA-G dimer was added to the experimental group, and the supernatant of 721.221 was set as the control. Flow cytometry was used to detect the expression level of CD14 CD80 on the surface of M1 group and M2 group. The results of flow cytometry showed that soluble HLA-G dimer could decrease the expression of CD80 on M1 surface, but had no effect on CD206 expression on M2 surface.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R392

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