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. 2018 Jul 15;29(14):1732–1742. doi: 10.1091/mbc.E17-06-0422

FIGURE 1:

FIGURE 1:

B-cells on nanotopographic surfaces exhibit enhanced actin polymerization. (A) Interference reflection micrographs of surfaces with parallel nanoridges. The single arrow indicates a ridge and the double arrow represents the spacing between ridges. Left panel: 3-µm ridge spacing; right panel: 5-µm ridge spacing. Scale bars are 3 and 5 µm, respectively. (B) Representative EGFP-actin–expressing A20 cell spread on an activating, flat glass surface. Scale bar: 5 μm. (C) Representative EGFP-actin A20 cell spread on a surface with a 3-µm ridge spacing. (D) Representative EGFP-actin A20 cell spread on a surface with 5.0-µm ridge spacing. Heat maps showing the MNA of actin fluorescence from representative EGFP-actin–expressing A20 cells on (E) a flat substrate, (F) a surface with a 3-μm ridge spacing, and (G) a surface with a 5-µm ridge spacing. Scale bars: 5 μm. (H) Peak-to-mean ratio of actin fluorescence intensity for all time points over entire cells (N = 9 cells for flat, N = 16 cells for 5 µm, and N = 15 cells for 3 µm) (p < 0.001 KS test). (I) A representative EGFP-actin–expressing A20 B-cell on a surface with a 3-µm ridge spacing. Scale bar: 3 µm. (J) Actin fluorescence intensity profile along a line perpendicular to the ridges (see representative white line in I). Note the enrichment of actin adjacent to ridges (thick gray lines). (K) Histogram of the widths of actin-enriched regions as a function of distance from the center of the nearest ridge (N = 14 cells). (L) A representative Lifeact-GFP–expressing primary B-cell on a surface with a 5-µm ridge spacing. Scale bar: 5 µm. (M) Heat map showing the MNA of actin fluorescence from a representative Lifeact-GFP–expressing primary B-cell on 5-µm spaced ridges. Scale bar: 5 µm. (N) Peak-to-mean ratio of actin fluorescence intensity for all time points in primary cells (N = 9 cells both on flat and 5-μm ridges, p < 0.001 KS test). All box-whisker plots are as follows: central marks in the box denote median values, boxes denote the 25th and 75th percentile values, and whiskers denote extreme values of the distributions. Outliers are shown in red.