Nallely García-Larragoiti1,2,3, Young Chan Kim4, César López-Camacho4, Alan Cano-Méndez1,2, Sandra López-Castaneda1,2, Darinel Hernández-Hernández5, Ángel G Vargas-Ruiz5, Ma Soledad Vázquez-Garcidueñas1, Arturo Reyes-Sandoval3,4,6, Martha E Viveros-Sandoval1,2,3. 1. División de Estudios de Posgrado, Facultad de Ciencias Médicas y Biológicas "Dr. Ignacio Chávez,", Universidad Michoacana de San Nicolás de Hidalgo, Morelia, México. 2. Centro Multidisciplinario de Estudios en Biotecnología, Facultad de Medicina Veterinaria y Zootecnia, Universidad Michoacana de San Nicolás de Hidalgo, Morelia, México. 3. Universidad Michoacana de San Nicolás de Hidalgo, Morelia, México. 4. Nuffield Department of Medicine, The Jenner Institute, University of Oxford, Oxford, UK. 5. Departamento de Hematología y Oncología, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Tlalpan, México. 6. Unidad Adolfo López Mateos, Instituto Politécnico Nacional, Ciudad de México, México.
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.
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 DENVinfection and promotes a more efficient infection. OBJECTIVE: The present study aimed to investigate the ability of platelet response to stimulation with full-length DENVNS1 protein and its domains. METHODS:DENVNS1 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 DENVNS1 proteins. Platelet activation was confirmed by expression of αIIbβIII and P-selectin by flow cytometry. Platelet aggregation was also assessed using DENVNS1 protein and its individual domains as agonists. RESULTS:DENVNS1 protein and its domains induce P-selectin and αIIbβ3 complex expression on platelet surfaces. DENVNS1 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 DENVNS1 protein and also its domains promote platelet recognition, activation, and aggregation.
Authors: Baranca Buijsers; Fadel Muhammad Garishah; Silvita Fitri Riswari; Rosalie M van Ast; Setyo Gundi Pramudo; Rahajeng N Tunjungputri; Gijs J Overheul; Ronald P van Rij; André van der Ven; Bachti Alisjahbana; Muhammad Hussein Gasem; Quirijn de Mast; Johan van der Vlag Journal: Front Immunol Date: 2021-12-20 Impact factor: 7.561