TAK-442, a Direct Factor Xa Inhibitor, Inhibits Monocyte Chemoattractant Protein 1 Production in Endothelial Cells via Involvement of Protease-Activated Receptor 1

Figures

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  FIGURE 1. FXa- and thrombin-induced MCP-1 production in HUVECs. MCP-1 concentration was determined in cell supernatant collected 20 h after the addition of the agonist, FXa (A) or thrombin (B). In inhibition studies (C,D), each inhibitor was added 1 h prior to the addition of the agonist and effects of TAK-442 (TAK), melagatran (Mel), and vorapaxar (Vor) on MCP-1 productions induced by 1 U/mL FXa (C) and 0.3 U/mL thrombin (D) were measured. Data are shown as mean ± SEM (n = 3). ∗P ≤ 0.025 compared with the control value of cells treated without FXa or thrombin (one-tailed Williams’ test) (A,B). ∗P ≤ 0.025 and †P ≤ 0.05 compared with the control value of cells treated with FXa or thrombin and no inhibitor (one-tailed Williams’ test and Student’s t-test, respectively, following ANOVA) (C,D).

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  FIGURE 2. Effects of TAK-442, melagatran, and vorapaxar on the intracellular calcium ion concentration ([Ca2+]i) induced by FXa, thrombin, and SFLLRN-NH2 in human PAR1-transfected Chinese hamster ovary (hPAR1/CHO-K1) cells. Calcium signal was recorded after the addition of FXa (0.03 U/mL) (A), thrombin (0.003 U/mL) (B), or PAR1 agonist peptide SFLLRN-NH2 (3 nM) (C) using FRIPR. Each inhibitor, TAK-442, melagatran, or vorapaxar, was pre-incubated with the cells for 10 min before the treatment with each agonist. Data are expressed as the percentage inhibition of calcium signal obtained after the addition of agonist in inhibitor-treated wells (n = 4) compared with control wells (no inhibitor added). The drug concentration need to suppress the [Ca2+]i by 50% (IC50) was determined.