Fig. 3. InsP6 increases platelet aggregate size in a fibrinogen-dependent manner.
(A, B) Whole blood was incubated with 16 μM InsP3 (light grey), InsP5 (grey), InsP6 (dark grey) or PBS (white) in absence (A) or presence (B) of 2.8 μg/ml of the GPIIb/IIIa blocking antibody abciximab for 1 min while rotating at 10 rpm. Then the samples were subjected to flow for 2 min at 720 rpm (= 1800 s-1) in the cone and plate analyzer Impact-R. After washing and staining with May-Grünwald solution the average size of surface-bound platelet aggregates was analyzed using the internal image analyzer. (C, D) Washed blood supplemented either only with 1.5 mg/ml fibrinogen (white) or with additional 1.5 mg/ml fibrinogen (light grey), 10 μg/ml VWF (dark grey), 3 μg/ml human collagen type III (CIII, black) ± 16 μM InsP6 (panel C, striped columns with InsP6) or ± 16 μM InsP5 (panel D, striped columns with InsP5) was incubated for 5 min and then mixed for 1 min while rotating at 10 rpm. Then the samples were subjected to flow for 2 min at 720 rpm (= 1800 s-1) in the cone and plate analyzer Impact-R. After washing and staining with May-Grünwald solution the average size of surface-bound platelet aggregates was analyzed using the internal image analyzer. Mean values ± SEM of three independent experiments (each performed in duplicates) are shown. Unpaired t-test: *p<0.05, **p<0.005 ***p<0.0001; n.s.: not statistically significant. (E) One representative image of each series of the experiments summarized in panel C are shown.