GEF-H1 is required for shear stress–induced neutrophil spreading. (A) GEF-H1+/+ and GEF-H1−/− neutrophils were stimulated with fMLP, plated on ICAM-1–coated coverslips, left untreated or exposed to 4 dynes/cm2 constant shear stress for 5 min, and then fixed and stained. DAPI images were used to identify primary objects and phalloidin images were subsequently used to determine cell spread areas. Representative images of phalloidin staining are shown with cell outlines overlaid in white. Bar, 20 µm. (B) Frequency distributions showing cell spread areas were derived from observations pooled from three independent experiments. At least 180 cells were measured for each condition. Outliers that were 1.5 times the interquartile range above the third quartile or below the first quartile were removed. Student's t test was used to determine p-values by comparing mean spread areas (n = 3). (C) Representative high-magnification time course images of individual cell responses to shear stress. Time after induction of shear stress is indicated. Bar, 10 µm. Arrows indicate the direction of shear.
