Fig. 4

SARS-CoV-2 Spike 1 protein directly enhances human platelet activation. a SARS-CoV-2 Spike protein and Spike subunit 1 (S1) potentiated platelet aggregation and ATP release in response to collagen, thrombin, and ADP in vitro, whereas Spike subunit 2 (S2) did not. Washed platelets from healthy donors were preincubated with Spike protein, S1 or S2 at 2 μg/mL for 5 min, then stimulated with collagen (0.6 μg/mL), thrombin (0.025 U/mL), or ADP (5 μM). Aggregation and ATP release (with luciferase) were assessed under stirring at 1200 rpm. Representative results and summary data of 4 experiments are presented. b Spike protein stimulated platelets for PAC-1 binding and CD62P expression in the absence of agonist. In addition, Spike protein and S1 increased PAC-1 binding and CD62P expression induced by thrombin in platelets. Platelets were incubated with Spike protein (2 μg/mL, 60 min) in the absence of agonist, or preincubated with Spike protein, S1 or S2 at 2 μg/mL for 5 min and stimulated with thrombin (0.025 U/mL) for 10 min, and then analyzed using a flow cytometer. Representative results and summary data of 4 experiments are presented. c Spike protein (2 μg/mL) and S1 (2 μg/mL) potentiated platelet spreading on immobilized fibrinogen. After preincubation with Spike protein (2 μg/mL) for 5 min, platelets were allowed to spread on the fibrinogen-coated surfaces at 37 °C for the indicated times. Representative results and summary data of 3 experiments are presented. d Spike protein and S1 potentiated clot retraction induced by thrombin. Platelets from healthy donors were normalized at a concentration of 4 × 10⁸/mL and preincubated with Spike protein (2 μg/mL) or S1 (2 μg/mL) for 5 min, then stimulated with thrombin (1 U/mL). Representative images and summary data are presented from 3 experiments using platelets from different donors. Statistical analyses were performed using one-way ANOVA, followed by Tukey’s post hoc analysis in (a), (b) and (c). NS no significance; *P < 0.05; **P < 0.01. Two-way ANOVA and Tukey’s post hoc test was performed in (d); ## significant difference (P < 0.01) between Spike and control group; ** significant difference (P < 0.01) between S1 and control group