OC 22.1 - Proteome and phosphoproteome analyses of SARS-CoV-2 infected humanized ACE2 mice platelets revealed reprogramming of platelets from early hyperreactive to late antiviral phenotypes

Sunday, June 25, 2023

14:45 – 15:00 ET

Room: 515

Session Available As: On-Demand

Presenting Author(s)

Saravanan Subramaniam, PhD (he/him/his)

Research Assistant Professor
Boston University
Boston, Massachusetts, United States

Co-Author(s)

RH

Ryan Matthew Hekman

Research Assistant
Center for Network Systems Biology, Boston University, Boston, MA, USA
Boston, Massachusetts, United States
AJ

Archana Jayaraman

Postdoc
Pulmonary Center, Department of Medicine, School of Medicine, Boston University MA, USA
Boston, Massachusetts, United States
DK

Devin Kenney

Graduate student
National Emerging Infectious Diseases Laboratories (NEIDL), Boston University, Boston, MA, USA
Boston, Massachusetts, United States
AK

Aoife Kateri O'Connell

Research Assistant
National Emerging Infectious Diseases Laboratories (NEIDL), Boston University, Boston, MA, USA
Boston, Massachusetts, United States
PM

Paige Montanaro

Research Assistant
National Emerging Infectious Diseases Laboratories (NEIDL), Boston University, Boston, MA, USA
Boston, Massachusetts, United States
BB

Benjamin Blum

Graduate student
Center for Network Systems Biology, Boston University, Boston, MA, USA
Boston, Massachusetts, United States
SW

Sarah Walachowski

Postdoc
Pulmonary Center, Department of Medicine, School of Medicine, Boston University MA, USA
Boston, Massachusetts, United States
ME

Maria Ericsson

Core specialist
Electron Microscopy Core Facility, Harvard Medical School, Boston, MA, USA
Boston, Massachusetts, United States
KR

Katya Ravid

DSc, PhD, Barbara E. Corkey Professor of Medicine
Department of Medicine and Whitaker Cardiovascular Institute, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA
Boston, Massachusetts, United States
NA

Nicholas A Crossland

Assistant Professor
National Emerging Infectious Diseases Laboratories (NEIDL), Boston University, Boston, MA, USA
Boston, Massachusetts, United States
AE

Andrew Emili

Professor
Center for Network Systems Biology, Boston University, Boston, MA, USA
Boston, Massachusetts, United States
FD

Florian Douam

Assistant Professor
National Emerging Infectious Diseases Laboratories (NEIDL), Boston University, Boston, MA, USA
Boston, Massachusetts, United States
MB

Markus Bosmann

Associate Professor
Pulmonary Center, Department of Medicine, School of Medicine, Boston University MA, USA
Boston, Massachusetts, United States

Background: Platelets are effectors of hemostasis and play a major role in coordinating immune and inflammatory activities. Since lung has resident megakaryocytes/platelets we hypothesized that lung resident megakaryocytes/platelets might have significant contribution to COVID-19 pathogenesis and CAC.

Aims: We aimed to characterize early phase alterations of platelets isolated from humanized ACE2 mice infected with SARS-CoV-2.

Methods: Heterozygous K18-hACE2 and C57BL/6J mice were used to study SARS-CoV-2 infectivity. Platelet response to SARS-CoV-2 infection was quantified by mass spectrometry analysis of proteomics and phosphoproteomics. Western blotting, ELISA, and multiplex plasma profiling were performed to validate the proteomics and phosphoproteomics data.

Results: SARS-CoV-2 inoculated (1x106 PFU, i.n.) K18-hACE2 mice started to lose weight at 4-dpi and showed 90% lethality at 7-dpi in association with viral neuroinvasion. Lungs and kidneys of infected K18-hACE2 mice showed mild increase in CD61+ aggregates compared to mock. Gene ontology and pathway analyses of platelet proteomics and phosphoproteomics revealed that SARS-CoV-2 infection significantly upregulates the complement-coagulation cascades (F2/F12/Tfpi/C1ra/Cd55/C4bp) and platelet activation-adhesion-degranulation proteins (Vwf/Itgb3/Itgb5/Selp/Pecam1) and chemokine (Pf4/Cxcl5/Ccl8) signaling at 2-dpi. However, interferon (Ddx58/Trim25/Mapk3) signaling was dominant at 4-dpi. Activation of proteomics and phosphoproteomics protein markers were highly correlated with platelet activation and interferon signaling at 2- and 4-dpi, respectively. Plasma chemokines (Ccl8/Pf4) and cytokines (IL6) were significantly expressed at 2-/ 4-dpi. Quantitatively higher amounts of prothrombin after 2-/4-dpi in platelet lysates suggested the activated platelets during SARS-CoV-2 provided a positive environment for prothrombinase complex assembly and cleaved prothrombin to active thrombin and might promote procoagulant activity. Our platelet proteome data compared with platelet transcriptional data from COVID-19 patients revealed that interferon signaling factors in human platelet transcriptomic data highly correlated with 4-dpi mouse platelets.

Conclusion(s): SARS-CoV-2-infected humanized ACE2 mice recapitulate thrombin-mediated early (before the appearance of clinical signs) platelet reprogramming towards activation-degranulation-aggregation.

Learning Objectives:

Upon completion, participant will be able to understand the phenotypic changes of platelets during SARS-CoV-2 infection.
Upon completion, participant will be able to get the concept of platelet-mediated COVID-19-associated coagulopathy (CAC).
Upon completion, participant will be able learn, for the first time, a dual-phased (procoagulant and antiviral) landscape of mouse platelet proteomes in experimental SARS-CoV-2 infection.