Skip to main content
. 2017 Aug 31;113(14):1776–1788. doi: 10.1093/cvr/cvx175

Figure 4.

Figure 4

Cgnl1 regulates Rac1 and RhoA signalling and actin cytoskeleton dynamics. (A) Chemo-luminescence measurement of the GTP-bound small G-proteins in cell lysates from siCgnl1 or sisham-transfected HUVECs after 20 and 40 min of cell seeding, showing the levels of GTP-Rac1, GTP-Rho-A, and GTP-cdc42. Values represent means ± SD. *P < 0.05 versus sisham in corresponding time group (n > 4). Student’s t-test. (B) Micrographs of typical results of a cell adhesion assay in which sisham and siCgnl1 treated HUVECs are seeded on a gelatine/collagen coated surface and analyzed for cell-spreading after 10 min adhesion. Scale bar represents 10 µm. (C) Actin visualized by phalloidine staining enables distinction between adherent cells (flat cell morphology) and non-adherent cells (round cell morphology). Scale bar represents 100 µm. Bargraph shows round/flat cell ratio in the siCgnl1 and sisham transfected groups. Values represent means ± SD. *P < 0.05 versus sisham in corresponding group (n = 6). Student’s t-test. (D) Serial images of time-lapse imaging of HUVECs GFP cells seeded in 3D collagen coculture with pericytes in siCgnl1 and sisham group. Different time points (T) are shown. 1 time point represents 1-h post-seeding. Scale bar represents 50 μm. (E) Quantification of AR and roundness per HUVEC-GFP+ structure (from T = 0 to 50 post-seeding). Each symbol represents average ± SD of five time points. Each time point is composed of 5 individual measurements. P < 0.0001 for AR and roundness, siCngl1 versus sisham group, linear regression analysis, overall comparison.