The SARS-CoV-2 Spike protein disrupts human cardiac pericytes function through CD147 receptor-mediated signalling: a potential non-infective mechanism of COVID-19 microvascular disease

  1. Elisa Avolio
  2. Michele Carrabba
  3. Rachel Milligan
  4. Maia Kavanagh Williamson
  5. Antonio P. Beltrami
  6. Kapil Gupta
  7. Karen T. Elvers
  8. Monica Gamez
  9. Rebecca R. Foster
  10. Kathleen Gillespie
  11. Fergus Hamilton
  12. David Arnold
  13. Imre Berger
  14. Andrew D. Davidson
  15. Darryl Hill
  16. Massimo Caputo
  17. Paolo Madeddu

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Abstract

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a broad range of clinical responses including prominent microvascular damage. The capacity of SARS-CoV-2 to infect vascular cells is still debated. Additionally, the SARS-CoV-2 Spike (S) protein may act as a ligand to induce non-infective cellular stress. We tested this hypothesis in pericytes (PCs), which are reportedly reduced in the heart of patients with severe coronavirus disease-2019 (COVID-19). Here we newly show that the in vitro exposure of primary human cardiac PCs to the SARS-CoV-2 wildtype strain or the α and δ variants caused rare infection events. Exposure to the recombinant S protein alone elicited signalling and functional alterations, including: (1) increased migration, (2) reduced ability to support endothelial cell (EC) network formation on Matrigel, (3) secretion of pro-inflammatory molecules typically involved in the cytokine storm, and (4) production of pro-apoptotic factors causing EC death. Next, adopting a blocking strategy against the S protein receptors angiotensin-converting enzyme 2 (ACE2) and CD147, we discovered that the S protein stimulates the phosphorylation/activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) through the CD147 receptor, but not ACE2, in PCs. The neutralisation of CD147, either using a blocking antibody or mRNA silencing, reduced ERK1/2 activation, and rescued PC function in the presence of the S protein. Immunoreactive S protein was detected in the peripheral blood of infected patients. In conclusion, our findings suggest that the S protein may prompt PC dysfunction, potentially contributing to microvascular injury. This mechanism may have clinical and therapeutic implications.

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  1. Version published to 10.1042/cs20210735
  2. Version published to 10.1101/2020.12.21.423721v2 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2020.12.21.423721: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board StatementIRB: Human myocardial samples (right ventricle or atrium) were discarded material from surgical repair of congenital heart defects (ethical approval number 15/LO/1064 from the North Somerset and South Bristol Research Ethics Committee).
    Consent: Adult patients and paediatric patients’ custodians gave informed written consent.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Primary antibodies (ACE2, dilution 1:100; CD147, 1:500, 6x-HIS-tag (Invitrogen MA1-21315), 1:1000) were incubated for 16 hrs at 4°C. β-Actin was used as a loading control (Sigma, A5441, 1:10000)
    6x-HIS-tag
    suggested: None
    β-Actin
    suggested: (Sigma-Aldrich Cat# A5441, RRID:AB_476744)
    A5441
    suggested: (Sigma-Aldrich Cat# A5441, RRID:AB_476744)
    Where required, cells were incubated with the anti-CD147 neutralizing antibody (20 μg/mL).
    anti-CD147
    suggested: None
    PCs were pre-incubated with anti-ACE2 (20 μg/mL, as described before5) or anti CD147 (20 μg/mL) antibodies for 1 hat 37 °C and then exposed to the S protein (500 ng/mL - 2·9 nM) for 1 h at 37°C.
    anti-ACE2
    suggested: None
    anti CD147
    suggested: None
    Software and Algorithms
    SentencesResources
    Western blot data were analyzed using the ImageJ software.
    ImageJ
    suggested: (ImageJ, RRID:SCR_003070)
    Statistics: Data were analyzed using Prism version 8.0 and expressed as individual values and as means ± standard error of the mean.
    Prism
    suggested: (PRISM, RRID:SCR_005375)

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

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  4. Version published to 10.1101/2020.12.21.423721v1 on bioRxiv