Figure 3.

Silencing CBS affects proliferation, migration, and vessel formation. A) Effect of various doses of Hcy for 24 h on the rate of proliferation of HUVECs ascertained through [³H]thymidine incorporation assay. Data represent means ± sd (n = 3), 1-way ANOVA. ^#P > 0.05 (not significant). B) Immunoblot data demonstrating the effect of various doses of Hcy for 24 h on eNOS (Ser1177) phosphorylation in HUVECs grown in serum-supplemented EBM medium. C) Effect of silencing CBS using CBS siRNA-1 and CBS siRNA-2 on the relative rate of proliferation of HUVECs compared with scrambled siRNA-treated cells ascertained through [³H]thymidine incorporation assay after 72 h transfection. Data represent means ± sd (n = 3), 2-tailed Student’s t test. *P ≤ 0.05. D) Effect of various doses of AOAA for 24 h on the rate of proliferation of HUVECs ascertained through ³H-thymidine incorporation assay. Data represent means ± sd (n = 3), 1-way ANOVA. *P ≤ 0.05, **P ≤ 0.01, ^#P > 0.05. E) Wound healing assay with scrambled control siRNA or CBS siRNA-treated HUVECs after 48 h transfection in serum-supplemented EBM medium to determine the effect of CBS on migration and wound healing. Data represent means ± sd (n = 3), 2-tailed Student’s t test. **P ≤ 0.01. F) Tube formation assay of scrambled control siRNA- or CBS siRNA-treated HUVECs after 72 h transfection on 2 mg/ml growth-factor reduced Matrigel. The images were acquired 4 h after plating serum-starved HUVECs on matrigel in serum-free EBM medium. Data represent means ± sd (n = 3), 2-tailed Student’s t test. **P ≤ 0.01.