Figure 3

WGA mediated TNTs are dependent upon the junctional protein CD31. (A) Example images of HUVECs stained with a human anti-CD31 antibody (green) in control and cells treated with 10 μg/ml WGA for 30 mins. White arrows indicate the redistribution of CD31 from cell-cell contacts in control to clusters in WGA treated cells. Red arrows depict aggregates of CD31 at the plasma membrane Scale bar is 50 μm. (B) Mean data validating CD31 knock-down by RNA interference. In 3 independent experiments the CD31 siRNA reduced the protein abundance by ~87%. (C) Mean data and analysis (n/N = 3/29 each point) from images shown in D showing the effect of WGA is significantly inhibited when CD31 is reduced. Red diamonds indicate cells transfected with CD31 siRNA. Data are represented as mean ± SD. *P < 0.05. (D) Example images of cell mask stained cells depicting WGA-induced TNTs in HUVECS transfected with control siRNA (arrows) but not in cells transfected with siRNA targeted against CD31. Scale bar 30 μm. (E) Phalloidin staining (red) suggest that WGA does not induce F-actin redistribution in HUVECs transfected with CD31 siRNA as compared to control. Scale bar 50 μm.