Autophagy-mediated antigen presentation for CD1d-restricted NKT cell recognition

在拟议的研究中，我们的目标是研究自噬是否通过将抗原（Ags）递送到CD1d负载区室来调节自然杀伤T（NKT）细胞反应。自噬是一种高度保守的溶酶体分解代谢途径，在此过程中，细胞器和长寿命蛋白质被降解，以提供细胞生存所需的营养。除了参与稳态、细胞死亡和细胞清除外，自噬似乎在促进抗原呈递细胞（APC）中通过自噬体的抗原呈递中起关键作用。NKT细胞代表与常规T细胞以及自然杀伤细胞共享功能特征的T淋巴细胞亚群。NKT细胞被认为通过识别经由CD1d分子呈递的脂质Ag而枢转地参与先天性和适应性免疫过程之间的介导。我们假设自噬通过将脂质抗原转运到CD1d装载区室来促进NKT细胞反应。为了解决上述假设，我们将分析自噬体在稳态和免疫激活后是否与专业APC中的CD1d负载区室共定位，以及APC中自噬的破坏是否会降低体外和体内的CD1d呈递和NKT细胞活化。用于进行所需分析的实验设置将包括流式细胞术、人和小鼠骨髓细胞系、ELISA、siRNA介导的敲减、涉及条件性敲除小鼠的小鼠实验以破坏自噬以及通过共聚焦显微镜进行的免疫细胞化学共定位研究。 这些结果有望进一步加深我们对自噬介导的Ag通过CD1d分子呈递和连续NKT细胞应答之间的分子相互作用的理解。调节CD1d限制性NKT细胞活性可能为治疗和预防（自身）免疫介导的疾病提供新的治疗选择

In the proposed research, we aim to investigate whether autophagy regulates natural killer T (NKT) cell responses through delivery of antigens (Ags) to CD1d-loading compartments. Autophagy is a highly conserved lysosomal catabolic pathway during which cell organelles and long-lived proteins are degraded in order to provide nutrients for cell survival. Aside from its involvement in homeostasis, cell death and cellular clearance, autophagy appears to play a crucial role in facilitating antigen presentation via autophagosomes in antigen-presenting cells (APCs). NKT cells represent a T lymphocyte subpopulation sharing functional characteristics with both conventional T cells as well as natural killer cells. NKT cells are believed to be pivotally involved in the mediation between innate and adaptive immune processes by recognizing lipid Ags presented via CD1d molecules. We hypothesize that autophagy facilitates NKT cell responses through transport of lipid Ags to CD1d-loading compartments. To address the aforementioned hypothesis, we will analyze whether autophagosomes colocalize with CD1d-loading compartments in professional APCs during the steady state and following immune activation as well as whether disruption of autophagy in APCs reduces CD1d presentation and NKT cell activation in vitro and in vivo. The experimental set up used to carry out the required analyses will include flow cytometry, human and mouse myeloid cell lines, ELISA, siRNA-mediated knockdown, mice experiments involving conditional knockout mice for disruption of autophagy as well as immunocytochemical co-localization studies via confocal microscopy. The results are expected to further our understanding of the molecular interactions between autophagy-mediated Ag presentation via CD1d molecules and consecutive NKT cell responses. Modulation of CD1d-restricted NKT cell activity might provide new therapeutic options for the treatment and prevention of (auto-) immune-mediated conditions