Monocytes and Macrophages, Targets of Severe Acute Respiratory Syndrome Coronavirus 2: The Clue for Coronavirus Disease 2019 Immunoparalysis

  1. Asma Boumaza
  2. Laetitia Gay
  3. Soraya Mezouar
  4. Eloïne Bestion
  5. Aïssatou Bailo Diallo
  6. Moise Michel
  7. Benoit Desnues
  8. Didier Raoult
  9. Bernard La Scola
  10. Philippe Halfon
  11. Joana Vitte
  12. Daniel Olive
  13. Jean-Louis Mege

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Abstract

Background

Coronavirus disease 2019 (COVID-19) clinical expression is pleiomorphic, severity is related to age and comorbidities such as diabetes and hypertension, and pathophysiology involves aberrant immune activation and lymphopenia. We wondered if the myeloid compartment was affected during COVID-19 and if monocytes and macrophages could be infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

Methods

Monocytes and monocyte-derived macrophages (MDMs) from COVID-19 patients and controls were infected with SARS-CoV-2 and extensively investigated with immunofluorescence, viral RNA extraction and quantification, and total RNA extraction followed by reverse-transcription quantitative polymerase chain reaction using specific primers, supernatant cytokines (interleukins 6, 10, and 1β; interferon-β; transforming growth factor–β1, and tumor necrosis factor–α), and flow cytometry. The effect of M1- vs M2-type or no polarization prior to infection was assessed.

Results

SARS-CoV-2 efficiently infected monocytes and MDMs, but their infection is abortive. Infection was associated with immunoregulatory cytokines secretion and the induction of a macrophagic specific transcriptional program characterized by the upregulation of M2-type molecules. In vitro polarization did not account for permissivity to SARS-CoV-2, since M1- and M2-type MDMs were similarly infected. In COVID-19 patients, monocytes exhibited lower counts affecting all subsets, decreased expression of HLA-DR, and increased expression of CD163, irrespective of severity.

Conclusions

SARS-CoV-2 drives monocytes and macrophages to induce host immunoparalysis for the benefit of COVID-19 progression.

SARS-CoV-2 infection of macrophages induces a specific M2 transcriptional program. In Covid-19 patients, monocyte subsets were decreased associated with up-expression of the immunoregulatory molecule CD163 suggesting that SARS-CoV-2 drives immune system for the benefit of Covid-19 disease progression.

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  1. Version published to 10.1093/infdis/jiab044
  2. ScreenIT

    SciScore for 10.1101/2020.09.17.300996: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Immunofluorescence: After a 24 or 48-hour infection, cells were incubated in blocking buffer (Phosphate buffer saline (PBS) supplemented with 5% FBS and 0.5% Triton X-100) for 30 minutes and washed before incubation with an anti-SARS-CoV-2 spike protein antibody (Life Technologies).
    anti-SARS-CoV-2 spike protein
    suggested: None
    Flow cytometry: PBMCs from healthy donors or Covid-19 patients were resuspended in PBS (Life Technologies) containing 5% FBS and 2mM EDTA (Sigma-Aldrich) for 20 minutes before staining using the following fluorochrome-conjugated antibodies (mouse IgG1): CD3 (UCHT1), CD20 (B9E9), CD14 (RMO52), CD16 (3G8) purchased from Beckman Coulter, Paris, France; HLA-DR (G46-6) and CD163 (GHI/61) from BD Biosciences, Le Pont de Claix, France, and appropriate isotype controls.
    mouse IgG1)
    suggested: None
    CD20
    suggested: (BioLegend Cat# 348805, RRID:AB_2889063)
    CD14
    suggested: (BioLegend Cat# 348805, RRID:AB_2889063)
    RMO52
    suggested: None
    CD16
    suggested: None
    3G8
    suggested: None
    G46-6
    suggested: None
    CD163
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Virus production and cell infection: SARS-CoV-2 strain IHU-MI3 was obtained after Vero E6 cells (American type culture collection ATCC® CRL-1586(tm)) infection in Minimum Essential Media (MEM) (Life Technologies) supplemented with 4% FBS as previously described (20).
    Vero E6
    suggested: None
    Experimental Models: Organisms/Strains
    SentencesResources
    Virus production and cell infection: SARS-CoV-2 strain IHU-MI3 was obtained after Vero E6 cells (American type culture collection ATCC® CRL-1586(tm)) infection in Minimum Essential Media (MEM) (Life Technologies) supplemented with 4% FBS as previously described (20).
    SARS-CoV-2
    suggested: None
    Software and Algorithms
    SentencesResources
    A minimum of 50,000 events were acquired for each sample using a BD Canto II instrument (BD Biosciences) and data were analyzed with FlowJo software (Tree Star, Ashland, OR).
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Statistical analysis: Statistical analysis was performed with GraphPad Prism (7.0, La Jolla, CA), using the two-way ANOVA test for viral quantification (Ct values) and transcriptional analysis, nonparametric Kruskall-Wallis test for group comparison, nonparametric Mann-Whitney U test for cytokine levels, and nonparametric t-test for flow cytometry results with monocyte populations and surface marker expression.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)
    qRT-PCR data for monocytes and macrophages, including principal component analysis (PCA) and hierarchical clustering of gene expression, were analyzed using the ClustVis webtool (23).
    ClustVis
    suggested: (ClustVis, RRID:SCR_017133)

    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: Please consider improving the rainbow (“jet”) colormap(s) used on page 24. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

    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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  3. Version published to 10.1101/2020.09.17.300996 on bioRxiv