Annexin Peptide Ac2-26 Suppresses TNFα-Induced Inflammatory Responses via Inhibition of Rac1-Dependent NADPH Oxidase in Human Endothelial Cells
Figure 4
The stimulatory effect of tumour necrosis factor (TNFα) on adhesion molecule gene upregulation requires Rac1 in HMECs.
(A) TNFα (2–50 ng/ml) reduced Nox4 gene expression (B) and pretreatment of Ac2-26 (0.5 µM) did not affect the TNFα mediated mRNA downregulation. TNFα (2–50 ng/ml) alone (C) and in combination of Ac2-26 (0.5 µM) (D) did not affect Nox2 mRNA expression. TNFα (20 ng/ml) stimulated ICAM-1 mRNA (E) and protein (G) and VCAM-1 mRNA (F) and protein (G) upregulation was blunted by Ac2-26 (0.5 µM) and diphenyleneiodonium (DPI, 1 µM) in cells trasnfected with control GFP plasmid. Dominant negative Rac1 (N17Rac1) reduced TNFα-mediated ICAM-1 and VCAM-1 upregulation and this inhibition was not potentiated by pretreatment of Ac2-26 and DPI. mRNA expression was normalized to control with TNFα stimulation in cells transfected with GFP. GAPDH was used to confirm equal loading. Data are mean ± SEM, n = 5 to 8. * P<0.05 vs control without TNFα stimulation; †P<0.05 vs control with TNFα stimulation. There was no difference (not significant, NS) between control Ac2-26 and DPI in the presence of TNFα in cells transfected with N17Rac1.