High-resolution imaging of the immunological synapse and T-cell receptor microclustering through microfabricated substrates
High-resolution imaging of the immunological synapse and T-cell receptor microclustering through microfabricated substrates
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DOI：
10.1098/rsif.2011.0025
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发表时间：
2011-10-07
影响因子：
3.9
通讯作者：
Wind, S. J.
中科院分区：
文献类型：
--
作者：
Biggs, M. J. P.;Milone, M. C.;Wind, S. J.
通过抗原呈递的T细胞活化与称为免疫突触（IS）的大分子膜组装的形成相关。IS的发生和阿曲他克林信号传导的开始的特征在于细胞膜微簇的形成和这些微簇组织成分离的微域。中心区富含T细胞受体（TCR）-主要组织相容性复合物微簇，称为中央超分子活化簇（cSMAC），形成该结构的靶心，而cSMAC周围的细胞界面的特征在于富含粘附和共刺激分子的区域。在体外，研究动态TCR微簇聚结和IS在T细胞群的发生是阻碍了细胞迁移的培养系统和分辨率的限制，导致横向细胞-细胞接触。在这里，我们详细介绍了一种新的系统，描述了微坑阵列的制造，旨在隔离单个T细胞抗原呈递细胞（APC）的共轭物，并促进IS形成在水平成像平面的高分辨率的微簇动力学研究。我们随后使用该系统来描述在T细胞群体中cSMAC的形成，并研究界面APC膜的形态。
T-cell activation via antigen presentation is associated with the formation of a macromolecular membrane assembly termed the immunological synapse (IS). The genesis of the IS and the onset of juxtacrine signalling is characterized by the formation of cell membrane microclusters and the organization of such into segregated microdomains. A central zone rich in T-cell receptor (TCR)-major histocompatibility complex microclusters termed the central supramolecular activation cluster (cSMAC) forms the bullseye of this structure, while the cellular interface surrounding the cSMAC is characterized by regions enriched in adhesion and co-stimulatory molecules. In vitro, the study of dynamic TCR microcluster coalescence and IS genesis in T-cell populations is hampered by cell migration within the culture system and resolution constraints resulting from lateral cell-cell contact. Here, we detail a novel system describing the fabrication of micropit arrays designed to sequester single T-cell-antigen presenting cell (APC) conjugates and promote IS formation in the horizontal imaging plane for high-resolution studies of microcluster dynamics. We subsequently use this system to describe the formation of the cSMAC in T-cell populations and to investigate the morphology of the interfacial APC membrane.