Apical endothelial CCL21 promotes comparable TEM of wt and DOCK2^–/–T lymphocytes under physiologic shear flow. (A) Wt and DOCK2^–/– T lymphocytes accumulated at low shear flow (0.25 dyne/cm²) for 2 minutes on TNFα-stimulated bEnd.3 overlaid with CCL21 (2 μg/mL) were subjected to physiologic shear stress (2 dyne/cm²) for 15 minutes. The migratory phenotype of each population was determined as described in “Materials and methods” and was expressed as a fraction of T cells accumulated on the endothelial monolayer at the first 2 minutes. Data are mean ± range values determined in 2 fields. (B) Phase-contrast images taken from time-lapse recordings of wt and DOCK2^–/– T cells in the process of TEM. Time zero was set at the beginning of each TEM. Micrographs are representative of about 100 transmigrating T cells. Bar = 10 μm. Arrows depict the lymphocyte-cell region that undergoes TEM. (C) Time course of TEM determined for wt and DOCK2^–/– lymphocyte populations. Time zero was set at the end of the accumulation phase. (D) Mean duration of the passage time required for individual apically adherent T cells to complete their TEM. At least 20 cells were analyzed for each experimental group. The experiments described in all panels are representative of 5 independent runs with different mice and bEnd.3 batches.