Figure 4.

Shear flow promotes actin-dependent ICAM-1 nanoclustering on TNFα-treated ECs. Representative STED images of ICAM-1 taken in (A) static ECs; (B) after 4 h of shear-flow stimulation, upstream of flow; (C) after 4 h of shear-flow stimulation, downstream of flow; and (D) after 4 h shear-flow-stimulated ECs and 20 min of CytoD treatment, upstream of flow. White arrows indicate the flow direction. Scale bars, 4 μm. (Insets) Magnified views of ICAM-1 fluorescent spots (1.5 × 1.5 μm regions). (E) Normalized ICAM-1 spot intensity for different conditions, as extracted from the analysis of STED images. The intensity of ICAM-1 has been normalized to the mean intensity of individual Abs nonspecifically attached to the glass substrate (i.e., glass condition). Box and whiskers plots (bars show median values, quartile distribution in boxes; 2.5–97.5 percentile in whiskers, individual symbols are outliers) show spot intensity data for one representative experiment per condition on at least n = 15 cells per condition. Significance has been calculated within the experiment shown. N = 2 independent experiments per condition. ^∗∗∗p < 0.001; ns, not significant. To see this figure in color, go online.