Carman Christopher V
Center for Vascular Biology Research, Division of Molecular and Vascular Medicine, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA.
Methods Mol Biol. 2012;757:215-45. doi: 10.1007/978-1-61779-166-6_15.
Immune system functions rely heavily on the ability of immune cells (i.e., blood leukocyte) to traffic throughout the body as they conduct immune surveillance and respond to pathogens. A monolayer of vascular endothelial cells (i.e., the "endothelium") provides a critical, selectively permeable barrier between two principal compartments of the body: the blood circulation and the tissue. Thus, knowledge of the basic mechanisms by which leukocytes migrate across the endothelium (i.e., undergo "transendothelial migration"; TEM) is critical for understanding immune system function. Cultured endothelial cell monolayers, used in combination with isolated blood leukocytes, provide a basis for highly useful in vitro models for study of TEM. When used in conjunction with high spatial and temporal resolution imaging approaches, such models have begun to reveal complex and dynamic cell behaviors in leukocytes and endothelial cells that ultimately determine TEM efficiency. In this chapter, we provide protocols for setting up a basic in vitro TEM system and for conducting high-resolution dynamic live-cell and three-dimensional fixed-cell imaging of TEM.
免疫系统的功能在很大程度上依赖于免疫细胞（即血液白细胞）在进行免疫监视和对病原体作出反应时在全身游走的能力。单层血管内皮细胞（即“内皮”）在身体的两个主要部分之间提供了一个关键的、选择性渗透的屏障：血液循环和组织。因此，了解白细胞穿过内皮（即经历“跨内皮迁移”；TEM）的基本机制对于理解免疫系统功能至关重要。与分离的血液白细胞结合使用的培养内皮细胞单层为研究TEM的高度有用的体外模型提供了基础。当与高空间和时间分辨率成像方法结合使用时，此类模型已开始揭示白细胞和内皮细胞中最终决定TEM效率的复杂动态细胞行为。在本章中，我们提供了建立基本体外TEM系统以及对TEM进行高分辨率动态活细胞和三维固定细胞成像的方案。
