Department of Anatomy, Yonsei University College of Medicine, Seoul, Republic of Korea.
Brain Korea 21 Plus Project for Medical Science, Yonsei University College of Medicine, Seoul, Republic of Korea.
Exp Mol Med. 2019 Apr 9;51(4):1-13. doi: 10.1038/s12276-019-0227-1.
Precise spatiotemporal regulation of leukocyte extravasation is key for generating an efficient immune response to injury or infection. The integrins LFA-1(CD11a/CD18) and Mac-1(CD11b/CD18) play overlapping roles in neutrophil migration because they bind the same as well as different ligands in response to extracellular signaling. Using two-photon intravital imaging and transmission electron microscopy, we observed the existence of preferred sites for neutrophil entrance into the endothelial cell monolayer and exit from the basement membrane and pericyte sheath during neutrophil extravasation, namely, hotspots I and II, by elucidating distinctive roles of LFA-1 and Mac-1. To penetrate the vascular endothelium, neutrophils must first penetrate the endothelial cell layer through hotspot I (i.e., the point of entry into the endothelium). Neutrophils frequently remain in the space between the endothelial cell layer and the basement membrane for a prolonged period (>20 min). Subsequently, neutrophils penetrate the basement membrane and pericyte sheath at hotspot II, which is the final stage of exiting the vascular endothelium. To further investigate the roles of LFA-1 and Mac-1, we newly generated LFA-1 FRET (CD11a-YFP/CD18-CFP) mice and Mac-1 FRET (CD11b-YFP/CD18-CFP) mice. Using both FRET mice, we were able to determine that LFA-1 and Mac-1 distinctly regulate the neutrophil extravasation cascade. Our data suggest that the vascular endothelium functions as a double-layered barrier in the steps of neutrophil extravasation. We propose that the harmonized regulation of neutrophil penetration through the endothelium via hotspots I and II may be critical for vascular homeostasis during inflammation.
白细胞渗出的精确时空调节对于产生针对损伤或感染的有效免疫反应至关重要。整合素 LFA-1（CD11a/CD18）和 Mac-1（CD11b/CD18）在中性粒细胞迁移中发挥重叠作用，因为它们在响应细胞外信号时结合相同和不同的配体。通过双光子活体成像和透射电子显微镜，我们观察到中性粒细胞渗出过程中存在进入内皮细胞单层和从基底膜和周细胞鞘退出的首选部位，即热点 I 和 II，这阐明了 LFA-1 和 Mac-1 的独特作用。为了穿透血管内皮细胞，中性粒细胞必须首先通过热点 I（即进入内皮细胞的点）穿透内皮细胞层。中性粒细胞经常在内皮细胞层和基底膜之间的空间中停留很长时间（>20 分钟）。随后，中性粒细胞在热点 II 处穿透基底膜和周细胞鞘，这是离开血管内皮细胞的最后阶段。为了进一步研究 LFA-1 和 Mac-1 的作用，我们新生成了 LFA-1 FRET（CD11a-YFP/CD18-CFP）小鼠和 Mac-1 FRET（CD11b-YFP/CD18-CFP）小鼠。使用这两种 FRET 小鼠，我们能够确定 LFA-1 和 Mac-1 分别调节中性粒细胞渗出级联反应。我们的数据表明，血管内皮细胞在中性粒细胞渗出的步骤中充当双层屏障。我们提出，通过热点 I 和 II 协调调节中性粒细胞穿透内皮可能对炎症期间的血管稳态至关重要。
