Requirement for migration through confined spaces but not for adhesion in eliciting loss of shape integrity. (A) Proportions of T cells (nine controls, four patients, from four experiments) that were Annexin V⁺ or PI⁺, after migration in increasing collagen concentrations for 24 h in collagen gels of increasing matrix density. (B) Confocal and diffusion interference contrast (DIC) microscopy of control (green) and patient (red) T cells while migrating through transwell pores (orange arrowheads) toward CXCL12. Hoechst, blue. Representative of three patients and three controls from three experiments. (C) Percentage of time T cells (six controls, three patients, from three experiments) spent elongated in 0.2% agarose gel matrices. (D) Proportions of T cells (eight controls, four patients, from three experiments) elongated during migration on 2D ICAM-coated plates or in 3D collagen matrices. (E) Proportions of DOCK8-deficient T cells elongated after migration on collagen-coated slides (2D) or in collagen gels (3D), untreated or with anti–integrin β1 blocking antibodies. Means are shown for three patients and three controls tested under each condition from three experiments. Two-way ANOVA was performed to compare treatment with or without antibody. (F) Similar to E except that one patient and one control were tested after transfection of nonspecific or ITGB1 siRNA. Median fluorescence intensities of CD29 for control cells were 6,386 (NS siRNA) and 2,019 (ITGB1 siRNA), and for patient cells were 6,363 (NS siRNA) and 1,396 (ITGB1 siRNA). Bars, 10 µm. A, C, and D show means ± SD. Linear mixed effects modeling was performed for A, unpaired two-tailed Student’s t test for C and D, and two-way ANOVA to compare treatment with or without antibody for E. Statistical significance indicated by ***, P < 0.001; ****, P < 0.0001; ns, nonsignificant.