Fig. 3
From: SARS-CoV-2 binds platelet ACE2 to enhance thrombosis in COVID-19
SARS-CoV-2 directly enhances 1 platelet activation in vitro. a SARS-CoV-2 dose-dependently potentiated platelets aggregation and ATP release in response to collagen, thrombin, and ADP in vitro. Washed platelets from healthy donors were preincubated with SARS-CoV-2 in the indicated concentration for 30 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 SARS8 CoV-2 induced PAC-1 binding and CD62P expression in the absence of agonist; and potentiated integrin PAC-1 binding and CD62P expression induced by thrombin in platelets. Platelets were preincubated with SARS-CoV-2 virus (1 × 105 PFU, 60 min) or with SARS-CoV-2 virus (1×105 PFU, 30 min), and treated with thrombin (0.025 U/mL, 10 min), and then analyzed using a flow cytometer. Representative flow cytometry histograms and summary data of 5 experiments are presented. c Representative confocal fluorescence images (phalloidin) showing that SARS-CoV-2 potentiated platelet spreading on immobilized fibrinogen (100 μg/mL). After preincubation with SARS-CoV-2 (1 × 105 PFU) for 30 min, platelets were allowed to spread on the fibrinogen-coated surfaces at 37 °C for indicated times. Representative results and summary data of 4 experiments are presented. d SARS-CoV-2 potentiated clot retraction induced by thrombin. Platelets from healthy donors were normalized at a concentration of 4 × 108/mL and preincubated with SARS-CoV-2 (1 × 105 PFU) for 30 min, then stimulated with thrombin (1 U/mL). Representative results and summary data of 4 experiments are presented. e Immunofluorescent staining of Nucleocapsid protein (NP, red) and CD41 (green) in human platelets incubated with SARS-CoV-2 virus (1 × 105 PFU) for 3 h. Representative images from 3 experiments using platelets from different healthy donors. f Scanning electron microscope (SEM) of SARS-CoV-2 particles on the surface of platelets. SEM of healthy human platelets (3 × 108 platelets/mL) incubated with SARS-CoV-2 (1 × 105 PFU) for 30 min. Platelets were washed for 3 times and fixed immediately after incubation and processed for SEM experiment. Representative images of single platelet from control group (platelet1) and SARS-CoV-2 treatment group (platelet2 and platelet3) are shown from three different experiments. Arrows point toward the SARS-CoV-2 virus. g. Transmission electron microscopy (TEM) of SARS-CoV-2 particles in platelets. TEM of healthy human platelets (3 × 108 platelets/mL) incubated with SARS-CoV-2 (1 × 105 PFU) for 3 h. Platelets were washed for 3 times and fixed immediately after incubation and processed for TEM experiment. Representative images from control group (platelet1) and SARS CoV-2 treatment group (platelet2 and platelet3) are shown from three different experiments. Arrows point toward the SARS-CoV-2 particles. Statistical analyses were performed using unpaired two-tailed Student’s t test 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); *P < 0.05 and **P < 0.01 compared with control group