Pharmacological inhibition of TrxR enhances cell surface TF activity in THP-1 cells. (A) THP-1 cells were treated with curcumin (25 µM) for varying time periods, and TrxR activity was measured. (B-C) THP-1 cells were treated with curcumin (25 µM) for varying time periods. Cell surface TF (B) and prothrombinase (C) activity was analyzed. (D) THP-1 cells were treated with curcumin (25 µM) for 2 hours and then fixed with 4% paraformaldehyde. After fixation, cells were washed and stained with AF488-annexin V and analyzed using confocal fluorescence microscopy, and the intensity of fluorescence was quantified. Images were obtained at 63× magnification (oil immersion). (E) THP-1 cells were treated with curcumin (25 µM) for 2 hours. After removing curcumin and washing cells once, cells were incubated with annexin V (400 nM) for 30 minutes to block cell surface PS before measuring TF activity. (F) THP-1 cells were treated with specific inhibitors of TrxR (auranofin or PMX-464) at varying concentrations for 2 hours, and cell surface TF activity was measured. (G) THP-1 cells were treated with 10 µM auranofin or 250 µM PMX-464 for 2 hours. Thereafter, a control vehicle or annexin V (400 nM) was added to cells for 30 minutes before measuring cell surface TF activity. THP-1 cells were treated with 25 µM curcumin (H), 10 µM auranofin (I), or 3 different doses of PMX-464 (50, 100, or 250 µM) (J) for a time period between 0 and 60 minutes, and the activation of MKK3/6 and p38 MAPK was analyzed by western blot analysis. (K) THP-1 cells were pretreated with the p38 MAPK activation inhibitor SB203580 (20 µM) for 1 hour, followed by PMX-464 (250 µM) or auranofin (10 µM) for 2 hours, and then cell surface TF activity was measured. *P < .05; **P < .01; ***P < .001; ns, not significant (compared with the values obtained in their respective controls or as indicated in the figures). DAPI, 4′-6-diamidino-2-phenylindole.