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Mitochondria-induced platelet procoagulant activity
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Figure 5. Extracellular mitochondria induced procoagulant activity of platelets and enhanced clot retraction. (A-C) Platelets incubated with extracellular mitochon- dria (exMT) at a ratio of 1:1 or 1:5 for 30 min at 37°C expressed increased amounts of fibrinogen (A) (n=24, one-way analysis of variance, ANOVA) and coagulation factor Va (B) (n=24, one-way ANOVA), and had shortened clotting time (C) (n=24, one-way ANOVA). Platelets stimulated with 5 mg/mL of collagen or 50 nM of thrombin were again used as controls to mimic “coated platelets.” (D) Clot retraction in platelet-rich plasma was induced by 1 U/mL of thrombin in the presence of phosphate- buffered saline (PBS, left panel), exMT (1:1 ratio to platelets, middle panel), or 5 μg/mL of collagen (right panel). The graph in the right panel summarizes the results from multiple experiments (n=27, one-way ANOVA, *P<0.05 vs. PBS-treated). FVa: activated factor V; PS: phosphatidylserine.
Figure 6. CD36 antibody ABC reduced extracellular mito- chondria-induced platelet acti- vation. Non-injured mice were infused with 2 x 108/mouse of MitoTracker-Green-labeled extracellular mitochondria (exMT), together with CD36 antibody or isotype IgG. (A-F) Blood samples were collected 30 min after infusion to meas- ure levels of exMT-bound platelets (A), CD62p+ platelets (B), platelet-leukocyte com- plexes (C), platelet counts (D), and platelet-leukocyte-exMT tri-complexes (E) by flow cytometry (n=16, one-way analysis of variance). (F) Schematic summary of the study. An injured brain releas- es metabolically active exMT that interact with platelets. These exMT induce α-granule secretion by platelets, but do not induce platelet aggrega- tion. As a result, the exMT- bound platelets and platelet- derived microvesicles promote the systemic consumptive coagulopathy and inflamma- tion that are consistently found during the acute phase of traumatic brain injury. WBC: white blood cell; ROS: reactive oxygen species; VWF: von Willebrand factor; FVa: activat- ed factor V; MV: microvesicles.
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