Figure 4
From: Pro-angiogenic Ginsenosides F1 and Rh1 Inhibit Vascular Leakage by Modulating NR4A1
F1 and Rh1 regulate NR4A1 expression and transcriptional activity. (A) Effect of F1 and Rh1 on VEGF-induced NR4A1 mRNA expression in HUVECs. The mRNA expression of NR4A1 after treatment with F1 or Rh1 in VEGF-stimulated cells was analyzed by Real-time RT PCR. β-actin was used as a reference gene in the analysis. Data are presented as mean ± SD (n = 3). (B) Effect of F1 and Rh1 on VEGF-induced NR4A1 protein expression in HUVECs. The protein expressions of NR4A1 after treatment with F1 or Rh1 in VEGF-stimulated cells were assessed with by Western blot analysis. Data are presented as mean ± SD (n = 3). Full-length blots are presented in Supplementary Fig. 9 (C) Effect of F1 and Rh1 on NR4A1 transcriptional activity. NurRE-Luc reporter plasmid-transfected HEK293T cells were treated with the indicated condition for 1 hr (0.1 μM of Csn-B and 10 μM of F1 or Rh1). NR4A1 transcriptional activities were analyzed using the dual luciferase reporter gene assay. Data are presented as mean ± SD (n = 3). (D) Effect of F1 and Rh1 on Csn-B-induced NR4A1 mRNA expression in HUVECs. NR4A1 mRNA expressions after co-treatment of F1 or Rh1 with Csn-B were analyzed by Real-time RT PCR. β-actin was used as a reference gene in the analysis. Data are presented as mean ± SD (n = 3). (E) Effect of F1 and Rh1 on the VEGF-induced junctional disruption in HUVECs. The cells were treated with F1 or Rh1 (10 μM, 1 hr) after VEGF stimulation (2.5 nM, 1 hr). VE-cadherin (green) was stained with immunofluorescence and the nucleus (blue) was stained by DAPI. Scale bars, 10 μm. (F) Effect of F1 and Rh1 on the Csn-B-induced junctional disruption in HUVECs. The cells were co-treated with 1 μM of Csn-B and 10 μM of F1 or Rh1 for an hour. VE cadherin (green) was stained by immunofluorescence and the nucleus (blue) was stained by DAPI. Scale bars, 10 μm. (G) Schematic representation of the inhibitory effect of F1 and Rh1 on VEGF-induced vascular leakage in ECs. Statistical significance was calculated based on three independent experiments (*P < 0.05; **P < 0.01; ***P < 0.001, P-values between depicted groups).
