ROS overproduction is involved in IS-induced platelet hyperactivation. (A) Flow cytometric analysis of ROS levels in platelets of CKD mice after AST-120 treatment by DCFH-DA staining (n = 6). Data are mean ± SD. **P < .01 vs sham group; ^##P < .01 vs CKD group, ANOVA. (B) Flow cytometric analysis of ROS levels in platelets of IS-treated mice (n = 6). Data are mean ± SD. **P < .01 vs vehicle group, Student t test. (C) Flow cytometric analysis of ROS levels in platelets after being treated with indicated concentrations of IS (n = 6). Data are mean ± SD. **P < .01 vs control group, ANOVA. (D-G) Platelets were pretreated with 5 mM NAC for 15 minutes and then stimulated with 500 μM IS for another 15 minutes. (D) Flow cytometric analysis of ROS levels in platelets (n = 6). (E) Flow cytometric analysis of P-selectin expression in platelets in response to 2 μg/mL collagen (n = 6). (F) Flow cytometric analysis of the percentage of platelet-monocyte aggregates in response to 2 μg/mL collagen in vitro (n = 6). (G) Representative images of thrombus formation on a collagen-coated flow chamber under shear-flow conditions (i). Scale bar, 50 μm. Quantification of the area of platelet adhesion for each group (n = 6) (ii). (D-G) Data are mean ± SD. **P < .01 vs control or collagen group; ^##P < .01 vs IS 500 μM group, ANOVA. DCFH-DA, dichlorodihy-drofluorescein diacetate.