Fig. 8
Oroxylin inhibits SHP-2 oxidative inactivation to suppress GPVI signaling in platelets. A, OroA inhibited SHP-2 oxidation in washed human platelets induced by collagen in a concentration-dependent manner. Washed human platelets were pre-incubated with different concentrations of OroA or NAC for 5 min, and then stimulated with collagen (1 µg/mL). Platelets were lysed and immunoprecipitated with SHP-2 antibody. The oxidation of SHP-2 was examined by immunoblotting with anti-oxPTP antibody. Representative results and summary data of 3 experiments are presented. B, OroA inhibited tyrosine phosphorylation (pTyr) in washed human platelets (B1) and washed mouse platelets (B2) induced by collagen (1 µg/mL). Representative results and summary data of 3–6 experiments are presented. C, OroA inhibits the phosphorylation of Src, Syk, PLCγ2 and Integrin β3 in washed human platelets (C1) and washed mouse platelets induced by collagen (1 µg/mL) (C2). Representative results and summary data of 3–6 experiments are shown. D, OroA inhibited Ca2+ influx induced by collagen (1 µg/mL). Washed human platelets loaded with Fura-2AM were incubated with OroA (50 or 100 µM) for 5 min and measured for 300 s with collagen (1 µg/mL). Representative results and summary data of 3 experiments are presented. E, OroA inhibits the phosphorylation of PKC in washed human platelets (E1) and washed mouse platelets (E2). Representative results and summary data of 3–5 experiments are shown. All statistical analyses were performed using one-way ANOVA followed by Tukey’s multiple comparison test. NS, no significance; *P < 0.05; **P < 0.01; ***P < 0.001