During an inflammatory response, circulating neutrophils must cross the blood vessel endothelium in a luminal-to-abluminal direction to gain access to injured tissues. A recent study in Nature Immunology now shows that transendothelial migration does not always proceed down a one-way street — neutrophils can undergo 'reverse transendothelial migration' and potentially spread the inflammatory response to other tissues.

Much of the mechanics of transendothelial migration is well appreciated, but many previous microscopy studies have relied on in vitro systems or in vivo models with insufficient resolution for studying the dynamics of this response. Woodfin et al. developed a technique to efficiently label endothelial cell junctions in mice by local injection of a fluorescently labelled monoclonal antibody specific for platelet endothelial cell adhesion molecule (PECAM). By using this method in lys-EGFP-ki mice (in which all endogenous neutrophils and monocytes express green fluorescent protein), they were thus able to monitor leukocyte transendothelial migration with high spatiotemporal resolution in real time in vivo.

Confocal microscopy imaging of transendothelial migration during acute inflammation (induced by interleukin-1β (IL-1β), N-formyl-methionyl-leucyl-phenylalanine (fMLP) or ischaemia–reperfusion injury) showed that most leukocytes cross the endothelial barrier via a paracellular route. However, in ∼10% of cases, transient pores appeared in the endothelial cell body and leukocytes exited the blood by a transcellular route.

Interestingly, several distinct types of paracellular transendothelial migration were observed. Although most leukocytes crossed the endothelium without pausing, a small proportion of cells either showed a 'hesitant' form of transendothelial migration (moving back and forth in the endothelial cell junction before finally entering the sub-endothelial tissue space) or underwent reverse transendothelial migration, crossing the endothelium in an abluminal-to-luminal direction. The type of transendothelial migration undertaken was influenced by the nature of the inflammatory stimulus; few leukocytes showed hesitant or reverse transendothelial migration in response to IL-1β or fMLP, but ∼15% of leukocytes underwent such disrupted polarized transendothelial migration following ischaemia–reperfusion injury. Further studies indicated that only neutrophils, and not monocytes, showed hesitant or reverse transendothelial migration.

Addressing the mechanisms behind disrupted polarized transendothelial migration, the authors found that following ischaemia–reperfusion injury there was decreased expression of junctional adhesion molecule C (JAM-C) at endothelial cell junctions. This loss of JAM-C expression was not seen during IL-1β-induced inflammation and appeared to be at least partly induced by reactive oxygen intermediates (ROI) released during ischaemia–reperfusion injury. An increased frequency of disrupted polarized transendothelial migration was seen when mice were treated with an antibody that blocked JAM-C function and in genetically modified mice deficient in endothelial cell JAM-C. These results suggested that JAM-C is involved in controlling the directionality of transendothelial migration.

The authors uncovered a potentially sinister outcome to neutrophil reverse transendothelial migration. In models of cremaster muscle or lower limb ischaemia–reperfusion injury, neutrophils that had undergone reverse transendothelial migration showed an increased activatory state (characterized by high expression levels of intercellular adhesion molecule 1 (ICAM1) and ROI) and accumulated in the pulmonary vasculature. Furthermore, there was a significant correlation between the degree of lung inflammation observed following ischaemia–reperfusion injury and the frequency of ICAM1^hi neutrophils in the pulmonary vascular bed.

Taken together, these data show that endothelial cell-expressed JAM-C is important for directing the one-way trafficking of neutrophils from the vascular lumen into inflamed tissues. Disruption of JAM-C function promotes reverse transendothelial migration of neutrophils, and this process may be involved in disseminating a local inflammatory response to other distal organs.

Author: Yvonne Bordon - Copyright © 2011 Nature Publishing Group, a division of MacMillan Publishers Limited; used with permission