In the mouse, much work has been done to characterize the function of Ly6c- “patrolling” monocytes by utilising intravital microscopy. However, this technique cannot be used outside of animal models, and therefore relatively little is known about the behaviour of their human counterpart, CD14dimCD16+ monocytes. To address this, we developed a re-circulating pump system to grow human endothelial cells under shear flow for 72h before adding human CD14dimCD16+ monocytes to the circulating media and performing live cell time-lapse imaging with a Nikon TiE fluorescence microscope. Using this approach we observed CD14dimCD16+ monocytes adhere and crawl on both macro-vascular HUVEC and micro-vascular HDMEC. The tracks of CD14dimCD16+ monocytes were twice as long when crawling on HDMEC compared with HUVEC under the same shear flow rate and 3 times fewer cells detached and re-entered the flow, indicating a preference for crawling on micro-vascular endothelium. HDMEC show higher levels of CD31, ICAM-1, E-selectin and VEGFR2, and lower levels of VCAM-1 and NRP-1 in a resting state, suggesting that the former represent ligands preferentially used for CD14dimCD16+ monocyte migration on micro-vascular cells, a role that will require further elucidation by studying migration under blocking conditions.
- Copyright © 2013 by The American Association of Immunologists, Inc.