Direct observations of the kinetics of migrating T-cells suggest active retention by endothelial cells with continual bidirectional migration



models. In comparing results here to previous work with activated T-cells, it may be worth
noting that there is considerable inter-donor variation in migration of T-cells (e.g., compared to
neutrophils) in our experience. Gathering data from several studies in our laboratory, we have
observed transendothelial migration through EC treated with TNF and IFN in direct microscopic
flow assays between 2 and 50% of adherent PBL (mean =17%; n=26). When we did study
activated lymphocytes, they migrated more efficiently through cytokine-stimulated endothelial
monolayers than resting lymphocytes, in agreement with previous reports [7-10]. Despite this, the
activated lymphocytes did not migrate efficiently into an underlying collagen gel, indicating that
activation per se was insufficient to generate the necessary signals for migration away from the
subendothelial space. Nevertheless, the possibility that antigen-presentation by EC specifically
facilitates migration of cognate T-cells cannot be discounted [29].

The data presented here reveal some problems relating to interpretation of widely-used,
filter-based migration assays. Using prolonged incubations under static conditions, there is a
high level of lymphocyte adhesion and significant transmigration through filters, even without
cytokine stimulation of EC. With cytokine treatment, lymphocytes still take far longer to cross
filters than to cross endothelial monolayers, and far longer than is required to enter tissue in vivo.
When we reduced the initial contact time between PBL and EC before wash-off of non-adherent
cells, fewer PBL adhered, but the proportion of the adherent cells that migrated through the filter
was still small after many hours. Practically, this suggests that mechanisms found to support or
regulate migration in such assays, might not be specific to the transendothelial (as opposed to the
trans-filter) stage.

In our experience, neutrophils migrate through endothelial monolayers and migrate



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