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

描述（由申请人提供）：树突状细胞（dc）是外周免疫系统的哨兵，通过液相摄取和内吞作用不断地探测周围的病原体。当检测到病原体来源的碎片时，dc吞噬外来物质并在此过程中被激活。吞噬蛋白被加工并以肽/MHC复合物的形式展示给II类MHC限制性T细胞，而脂质被合并到CD1/(32m/)脂质复合物中，呈递给CD1限制性T细胞。关于II类MHC分子的内体抗原加工、装载和呈递的大部分已知信息都是通过对B细胞系的生化和细胞生物学分析，以及在更有限的程度上对培养的dc进行的分析来了解的。现在，新的方法使得在表达绿色荧光蛋白(GFP)- II类MHC融合蛋白的小鼠表皮中，活体II类MHC阳性dc原位摄取红色荧光抗原成为可能。经过抗原处理的树突状细胞向皮肤引流淋巴结的转移可以在活细胞中追踪。首先，我们将研究皮肤源性dc和淋巴结驻留dc的蛋白和脂质抗原递呈途径。我们将验证我们的假设，即在炎症期间，皮肤免疫反应的启动需要细胞介导的蛋白质和脂质抗原的运输，由表皮驻留dc（朗格汉斯细胞）到皮肤引流淋巴结。存在于淋巴结T细胞区域的树突状细胞的抗原呈递是T细胞启动的必要条件。我们假设抗原从LC转移到非迁移性淋巴结驻留dc是成功启动T细胞的必要条件。因此，LCs表达正常抗原内吞作用但有缺陷的内体加工的突变小鼠应该具有有限的表型。相反，表达内体加工缺陷的非迁移性dc的小鼠应表现出T细胞启动能力降低。我们将研究表皮来源的朗格汉斯细胞和淋巴结驻留DC亚型的抗原加工和II类MHC递呈途径。最后，我们成功地建立了一种新的小鼠模型来研究cd1介导的抗原在体内的递呈。CD1d参与皮肤免疫反应的可能性将在CD1d-黄色荧光蛋白敲入小鼠中进行研究。

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.