Figure 7. Myosin II contractility regulates 3D migration independent of pore size.

(a) Fibroblast 3D migration rates for control (white) 5 μM (light grey) and 25 μM (dark grey) blebbistatin for each ECM architecture. N=3, n=48. (b) MIPs comparing activated β1 integrin (9EG7) in fibroblasts between control and overnight exposure to 25 μM blebbistatin in FB4 collagen. Inset illustrates that activated β1 integrin remains associated with collagen fibrils surrounding the cell body region in the absence of myosin II contractility. (c) Reducing integrin binding with an inhibitory antibody against β1 integrin (mAb13; 1 and 10 μg ml⁻¹) partially rescues contractility-deficient migration, but in a pore size-dependent manner. Red dashed lines indicate control levels. N=3, n≥70.(d) Timelapse series of a fibroblast expressing EYFP-paxillin in FB16 ECM treated with 25 μM blebbistatin (added 30 min before first frame). Numerous protrusions (red asterisks) form but do not aid cell body movement (yellow circle), while an elongated tail (cyan arrowheads) hinders migration. (e) HFF treated with blebbistatin and mAb13 (1 μg ml⁻¹) show fewer protrusions while showing no elongated tails and slipping through the matrix. *P<0.05 (ANOVA). + Significantly different from control conditions; P<0.05 (ANOVA). Errors bars: s.e.m. Scale bars: (b) 10 μm; (d,e) 30 μm.