TCRζ^dim T cells exhibit enhanced migratory capacity in vitro. (A) Healthy donor PBLs (5 × 10⁶) were applied to the gelatin-coated Transwell containing a monolayer of TNF-α–stimulated HUVECs and cells in upper and lower chambers harvested at 24 hours prior to staining for expression of CD3ϵ and TCRζ and analysis by flow cytometry. Representative dot plots are shown. (B) Subset analysis of migrating cells. After 24 hours cells were harvested and stained for TCRζ and CD3, CD4, or CD8, as in panel A. Data are expressed as the percentage of cells migrating relative to the total number of each cell subset added to the Transwell. The significance of differences between migration of TCRζ^bright and TCRζ^dim cells is shown. (C) The chemokines CXCL10, CCL5, or CXCL12 were added to each lower chamber of the Transwell at 50 ng/mL and the total number of PBLs migrating determined after 24 hours; *P < .013; **P = .063; ***P < .004 compared to cells migrating in the absence of exogenous chemokine. (D) The effects of each chemokine on the migration of TCRζ^bright and TCRζ^dim cells were determined by flow cytometry, as described. Differences in migration between subsets was highly significant (P < .002), with the exception of migration in response to CXCL12. Within subset differences between “medium” and CXCL12-stimulated cells were also significant (P < .023).