Analysis of Class II MHC and CD1d antigen presentation pathways in skin-derived d

描述(申请人提供)：树突状细胞(DC)是外周免疫系统的哨兵，通过液体相摄取和内吞作用持续检测其周围的病原体。当检测到病原体衍生的片段时，DC会吞噬外来物质，并在此过程中被激活。被吞噬的蛋白质被加工并以肽/MHC复合体的形式展示给第二类MHC限制性T细胞，而脂质被结合到CD1/(32M/脂质复合体)中，呈现给CD1限制性T细胞。关于内体抗原的处理、装载和呈递II类MHC分子的大部分知识是通过对B细胞系的生化和细胞生物学分析来了解的，在更有限的范围内，还可以对培养的DC进行分析。现在，新的方法使得在表达绿色荧光蛋白(GFP)-II类MHC融合蛋白的小鼠的表皮中原位显示II类MHC阳性的活树突状细胞摄取红色荧光抗原成为可能。在活细胞中可以追踪到抗原经验丰富的树突状细胞向皮肤引流淋巴结的运输。首先，我们将研究皮肤来源的树突状细胞和常驻淋巴结的树突状细胞中的蛋白质和脂类抗原呈递途径。我们将检验我们的假设，即在炎症过程中，皮肤免疫反应的启动需要驻留在表皮的树突状细胞(朗格汉斯细胞)通过细胞介导的蛋白质和脂肪抗原的运输到皮肤引流淋巴结。位于淋巴结T细胞区的DC的抗原提呈对于T细胞的启动是必不可少的。我们假设抗原从LC转移到非迁移性淋巴结驻留的DC是成功的T细胞启动所必需的。因此，LCS表达正常抗原内吞作用但内体加工缺陷的突变小鼠应该具有有限的表型。相反，表达非迁移性DC的小鼠在内体处理中存在缺陷，应该表现出降低的T细胞免疫能力。我们将研究表皮来源的朗格汉斯细胞和淋巴结居留DC亚型中的抗原处理和II类MHC呈递通路。最后，我们成功地建立了一种新的小鼠模型来研究CD1介导的体内抗原递呈。将在CD1d黄色荧光蛋白敲入小鼠中研究CD1d对源自皮肤的免疫反应的可能参与。

DESCRIPTION (provided by applicant): Dendritic cells (DCs) are sentinels of the peripheral immune system that continuously survey their surroundings for pathogens, by fluid phase uptake and endocytosis. When pathogen-derived fragments are detected, DCs engulf the foreign material and in the process become activated. Engulfed proteins are processed and displayed as peptide/MHC complexes to Class II MHC-restricted T cells, while lipids are incorporated into CD1/(32m/lipid complexes for presentation to CD1-restricted T cells. Much of what is known about endosomal antigen processing, loading and presentation of Class II MHC molecules was learned through biochemical and cell biological analysis of B cell lines, and to a more limited extent, of cultured DCs. New methodology now makes it possible to visualize the uptake of red fluorescent antigen by live Class II MHC-positive DCs in situ in the epidermis of mice that express green fluorescent protein (GFP)-Class II MHC fusion proteins. Trafficking of antigen-experienced DCs to skin-draining lymph nodes can be tracked in live cells. First, we will study the protein and lipid antigen-presentation pathways in skin-derived DCs and lymph node-resident DCs. We will test our hypothesis that during inflammation, priming of immune responses in the skin requires cell-mediated transport of both protein and lipid antigens by epidermis- resident DCs (Langerhans cells) to skin-draining lymph nodes. Antigen presentation by DCs that reside in the T cell area of lymph nodes is essential for T cell priming. We hypothesize that antigen transfer from LC to a non-migratory lymph node-resident DCs is necessary for successful T cell priming to occur. Accordingly, mutant mice in which LCs express normal antigen endocytosis but defective endosomal processing should have limited phenotype. In contrast, mice expressing non-migratory DCs defective in endosomal processing should exhibit reduced T cell priming capacities. We will study antigen processing and Class II MHC presentation pathways in epidermis-derived Langerhans cells and lymph node-resident DC subtypes. Finally, we have succeeded to generate a new mouse model to study CD1-mediated antigen presentation in vivo. The possible involvement of CD1d to immune responses originating in the skin will be investigated in CD1d- yellow fluorescent protein knock-in mice.