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. 2017 Sep 26;8:504. doi: 10.3389/fneur.2017.00504

Figure 5.

Figure 5

Repulsive guidance molecule a (RGMa) inhibited cytoskeleton reassembly, filopodia, and lamellipodia formation in human umbilical artery endothelial cells (HUAECs) via neogenin and Unc5b. Before the immunofluorescence experiment, HUAECs were treated with vehicle, RGMa, VEGF, or VEGF plus RGMa for 40 min. F-actin was stained with phalloidin conjugated with FITC and phosphorylated focal adhesion kinase (p-FAK) connected with primary antibody was labeled with Alexa Fluor 555 donkey anti-rabbit (H + L) secondary antibody. (A) Immunofluorescence showed the cytoskeleton (green) change and p-FAK (red) distribution. (B) Immunofluorescence showing the cytoskeleton change of HUAECs transfected with SgRNA or neogenin gRNA. (C) Immunofluorescence showed the cytoskeleton change of HUAECs transfected with SgRNA or Unc5b gRNA. The filopodia are indicated as sharp spikes (arrowhead), and lamellipodia (arrow) are indicated as flat intensive staining. The merged images are shown in the upper panels, and the amplified indicated areas are shown in the lower panel for different groups. Photographs were obtained with laser scanning confocal microscopy (Nikon, A1 + R, magnification 400×). Results shown are representative images of ≥4 independent experiments.