Effect of apical CXCL12 and shear on lymphocyte TEM toward subendothelial (basal) CCL5 and contribution of α4 or αL integrins. (A) T cells in contact with TNF-α–activated HUVECs under shear were observed during a shear-free (static) period in the presence or absence of specified chemokines (note that rate of detachment was not measured in this analysis because the objective was to assess TEM of adherent cells). (B) Lymphocytes contacting endothelium were analyzed for the indicated migratory categories under continuous shear flow. Concentrations of apical CXCL12 are indicated and were presented with or without basal CCL5 (100 ng/mL). For inhibition of Gi proteins, lymphocytes were incubated overnight in culture medium with 100 ng/mL PTX. (C) Effects of blocking α4 (VLA-4) or αL (LFA-1) integrins on the indicated adhesive and migratory phenotypes of T cells. Cells were pretreated with indicated blocking mAbs (10 μg/mL) for 10 minutes and were perfused into the chamber in medium containing 1 μg/mL of the respective mAbs and chemokine configurations identical to those used in panel B group 3. Migratory phenotypes developing under persistent shear stress were analyzed as in Figure 1. Data are mean ± SEM from 4 experiments that used different lymphocyte donors and HUVEC preparations. *P < .05; **P < .01.
