Parmodulin 2 and vorapaxar inhibit proinflammatory signaling in endothelial cells. HUVECs were incubated with the indicated concentrations of (A) parmodulins or (B) orthosteric inhibitors for 30 minutes before exposure to SFLLRN. Increase in [Ca²⁺]_i flux after stimulation with 5 μM SFLLRN was evaluated and compared with vehicle controls that were not exposed to PAR1 antagonists. Data are presented as means ± SEM (n = 3-6). (C) HUVECs were incubated with either 3 μM parmodulin 2 (PM2) or 0.3 μM vorapaxar before exposure to vehicle (black) or SFLLRN (red). Release of von Willebrand factor (VWF) into supernatants was quantified by ELISA. Data are presented as means ± SEM (n = 4). ***P < .001. (D) Immunofluorescence microscopy of HUVECs incubated with vehicle, parmodulin 2, or vorapaxar, exposed to buffer or 1 U/ml thrombin, and subsequently stained with PE-phalloidin and DAPI. (E) Representative tracings of real-time monitoring of transendothelial electric resistance (TER) of HMVECs incubated with vehicle, 10 μM parmodulin 2, or 0.3 μM vorapaxar and subsequently exposed to SFLLRN (arrow). (F) Quantification of inhibition of SFLLRN- and thrombin-induced barrier dysfunction in HMVECs by parmodulin 2 and vorapaxar. *P < .05, **P < .01, ***P < .001.