Promyelocytic extracellular chromatin converts HUVECs into a procoagulant phenotype. (A) Thrombin and intrinsic Xa and extrinsic Xa production were measured on HUVECs treated with NB4 extracellular chromatin from the different treatment groups. (B) Inhibition assays of protein production were performed using DNase I or APC to degrade extracellular chromatin before incubation with ECs. Coagulation time (C) and fibrin formation (D) of ECs stimulated with ATRA- or PBS-derived NB4 chromatin for 24 hours in the presence of DNase I or APC were measured. Data shown from 3 independent experiments and presented as mean ± SD. (E) FXa (red) and FVa (green) costaining (yellow) was observed on filopods near the retracted edges of ECs and on newly formed thin filaments (arrow), similar to the binding sites for lactadherin. (F) ECs pretreated with ATRA-derived chromatin and incubated with healthy plasma showed considerable fibrin strand formation arranged radially along with filopodia (arrow), which formed a fibrin network. The inset bar represents 5 μm in panels E-F. One of 6 independent experiments is shown. *P < .01, **P < .001 vs no inhibitor group and ^#P < .05 vs DNase I (+) APC (+) in panel B; **P < .001 vs PBS-treated NB4 cells in panels A, C, D; ^#P < .05, ^##P < .01, ^###P < .001 vs DNase I (-) APC (-) in panels C-D; ^&P < .05 vs DNase I (+) APC (+) in panel D.