cKO P2 neutrophils display enhanced motility in highly confined 3D matrices and microconstrictions. (A) Cell trajectories of neutrophils migrating in a dense collagen gel (4 mg/mL). t indicates the period of the displayed tracking, and r is the mean displacement ratio in this period. (B) Representation of the mean speed of neutrophils migrating in a 3D matrix at different collagen concentrations (2 and 4 mg/mL). At least 157 cells for the WT group and 277 cells for the cKO P2 group were measured. Data are represented as box plots (+median), and whiskers range from the 10th percentile to the 90th percentile. Graphs are representative of ≥3 independent experiments. (C) MSD of neutrophils migrating in a 3D collagen matrix (4 mg/mL). The plot corresponds to analysis of trajectories shown in panel A. (D) Directionality of neutrophils migrating in a 4-mg/mL collagen gel prior to (random) or after CXCL2 (20 ng/mL) stimulation. At least 274 cells for the WT group and 409 cells for the cKO P2 group were traced. (E) Speed over time for cells migrating in a 4-mg/mL collagen gel. Temporal evolution of the speed before and after CXCL2 stimulation is shown. Graph corresponds to plots shown in panel D. (F) Schematic representation of the migration assay through microconstrictions. The lower panel shows an example of a cell migrating through a 1 µm × 15 μm section constriction. (G,I) Proportion of neutrophils that were able to pass through a constriction. *P < .05 in 4 of 5 independent experiments (G) and in all 3 independent experiments (I). (H,J) Average time needed by neutrophils to pass through the constriction. The statistical significance of the different experiments was defined as described in "Materials and methods." *P < .05.