Platelet activation and aggregation response to dengue virus nonstructural protein 1 and domains

Abstract

Background Platelets are now recognized as immunological sentries in the first line of defense that participate in the detection and response to pathogens. This frequently results in a decrease in the number of circulating platelets. Different mechanisms have been hypothesized to explain the thrombocytopenia in patients with severe dengue, one of them is the participation of the non‐structural protein 1 (NS1) of dengue virus (DENV), which can be secreted into circulation during DENV infection and promotes a more efficient infection. Objective The present study aimed to investigate the ability of platelet response to stimulation with full‐length DENV NS1 protein and its domains. Methods DENV NS1 plasmid was transfected into HEK‐293T. Proteins were purified by Niquel Sepharose affinity chromatography. Secreted proteins were assessed by sodium dodecylsulfate polyacrylamide gel electrophoresis, Coomassie staining and western blot. Platelet‐rich plasma was directly incubated with DENV NS1 proteins. Platelet activation was confirmed by expression of αIIbβIII and P‐selectin by flow cytometry. Platelet aggregation was also assessed using DENV NS1 protein and its individual domains as agonists. Results DENV NS1 protein and its domains induce P‐selectin and αIIbβ3 complex expression on platelet surfaces. DENV NS1 induce a stable platelet aggregation after the addition of a minimal dose of adenosine diphosphate (ADP), epinephrine (EPI), or collagen. Interestingly, only EPI could induce the formation of platelet aggregates after incubation with the protein domains of NS1. Conclusion Our results suggest that the full DENV NS1 protein and also its domains promote platelet recognition, activation, and aggregation.

Platelet Reactivity and Inflammatory Phenotype Induced by Full-Length Spike SARS-CoV-2 Protein and Its RBD Domain

“…Our results demonstrate that neither the complete spike protein nor the RBD domain induce platelet aggregation on their own when compared to known aggregation agonists, which achieve aggregation percentages of more than 90% ( p < 0.0001) ( Figure 2 A–H). Once we observed that proteins induced activation but not aggregation, based on previous results reported in our lab by García-Larragoiti et al [ 22 ] and in a previous work performed by Chiao-Hsuan et al [ 23 ], we decided to assess if platelet hyperreactivity induced by viral protein stimulation, in addition to subthreshold doses of known aggregation agonists, could restore platelets’ aggregation capacity.…”

Rotational thromboelastometry in critical phase of dengue infection: Association with bleeding

“…21 Protein disulfide isomerase on the surface of platelets has been found to cross react with antibodies against dengue virus nonstructural protein 1 (NS1) in patients with dengue hemorrhagic fever. 22 , 23 These interactions ultimately result in inhibition of platelet adhesion and aggregation, coagulation defects, and endothelial cell damage. 24 , 25 Moreover, platelet destruction due to the entry of virus into platelets and absorption of immune complexes to the membrane of the platelet may contribute to reduction in platelet aggregation.…”

“…Proposed mechanisms for reduced platelet aggregation in dengue hemorrhagic fever are the presence of dengue‐specific immune complexes and platelet destruction 21 . Protein disulfide isomerase on the surface of platelets has been found to cross react with antibodies against dengue virus nonstructural protein 1 (NS1) in patients with dengue hemorrhagic fever 22,23 . These interactions ultimately result in inhibition of platelet adhesion and aggregation, coagulation defects, and endothelial cell damage 24,25 .…”

“…and/or range. All data were statistically analyzed using SPSS23.0.0.0 (IBM Statistics for Windows version 23; IBM Corp., Armonk, NY, USA) under 95% confidence interval. Association of ROTEM parameters between patients with severe and mild to moderate bleeding manifestations was analyzed using chi-square testing.…”

Recent Findings on Platelet Activation, vWF Multimers and Other Thrombotic Biomarkers Associated with Critical COVID-19

“…As positive controls of platelet activation we used known activation agonists ADP, collagen and epinephrine and the acquisition was performed by flow cytometry as reported before by our group. 10 Results were analyzed using FlowJo v 10.8.0.…”