Skip to main content
. 2005 Oct 13;115(11):2992–3006. doi: 10.1172/JCI24586

Figure 4.

Figure 4

CA4P blocks VE-cadherin/β-catenin/Akt signaling pathway. (A) CA4P disengages VE-cadherin and β-catenin. The localization of VE-cadherin and β-catenin in FGF-2–stimulated HUVECs was monitored during 18 hours by microscopy upon CA4P treatment (10 nM). Notice the redistribution of VE-cadherin and β-catenin assuming a zigzag pattern at cell-cell contacts (arrowheads) before the apparition of gaps between the cells (asterisks). Magnification: ×40. Scale bar, 10 μm. (B) Quantification of the organization pattern of VE-cadherin and β-catenin. The organization pattern, i.e., linear or zigzag, was evaluated by microscopic counting of 10 fields at ×10 magnification and presented as the percentage of linear or zigzag pattern per microscopic field ± SEM (#P < 0.001, †P < 0.001 compared with linear and zigzag organization pattern observed at time 0, respectively; n = 10). (C) CA4P synergizes with human mAb against VE-cadherin to decrease cell viability. AdNullE4–- or AdNullE4+-infected HUVECs were incubated with either CA4P or human mAb against VE-cadherin (BV9) or in combination, and the number of viable cells was determined after a 48-hour incubation. Results of 4 experiments in duplicate are expressed as the mean number of viable cells ± SEM (*P < 0.05, **P < 0.01, #P < 0.001 compared with untreated cells; n = 4). (D) CA4P synergizes with neutralizing mAb against VE-cadherin to increase endothelial cell permeability. Modification of endothelial cell permeability of AdNullE4–- or AdNullE4+-infected HUVEC monolayers was assessed as described in Methods at designated time points. Results of 3 experiments in triplicate are expressed as the permeability ratio (experimental/control) ± SEM (*P < 0.05, **P < 0.01, #P < 0.001 compared with control; n = 3). (E) CA4P inhibits tyrosine phosphorylation of VE-cadherin and β-catenin. Levels of tyrosine phosphorylation of immunoprecipitated VE-cadherin and β-catenin were determined by Western blotting. (F) CA4P induces serine/threonine phosphorylation of β-catenin. Phosphorylation levels (Ser45 and Ser33/37/Thr41) of β-catenin were determined by Western blotting. (G) CA4P decreases Akt phosphorylation. Phosphorylation level (Ser 473) of Akt was determined by Western blotting. The condition where the serum-free medium (X-VIVO) was used corresponds to basal protein expression in the absence of stimulation.