Thrombin-induced ROS generation requires PAR4, but not PAR1. (A) H2DCFDA labelled human platelets were pre-treated with vehicle control (0.1% DMSO), PAR1 antagonist SCH79797 (1 µM) or PAR4 antagonist tcY-NH2 (400 µM), stimulated with thrombin (2 U/mL), and monitored for ROS production by flow cytometry. Data are mean±SEM (n=3), *P≤0.05, ***P≤0.001. (B) Washed platelets from WT and PAR4 KO mice (2.5×108/mL) were preloaded with 10 µM H2DCFDA and left unstimulated, or stimulated with thrombin (2 U/mL) and ROS production was measured by flow cytometry. Data are mean±SEM (n=3), *P<0.05 H2DCFDA labelled platelets from WT and PAR4 KO mice were left unstimulated, or stimulated with thrombin (2 U/mL) and ROS production was measured by flow cytometry. Data are mean±SEM (n=3), ***P≤0.001.