Circulating multimeric immune complexes drive immunopathology in COVID-19

  1. Jakob Ankerhold
  2. Sebastian Giese
  3. Philipp Kolb
  4. Andrea Maul-Pavicic
  5. Reinhard E. Voll
  6. Nathalie Göppert
  7. Kevin Ciminski
  8. Clemens Kreutz
  9. Achim Lother
  10. Ulrich Salzer
  11. Wolfgang Bildl
  12. Tim Welsink
  13. Nils G. Morgenthaler
  14. Andrea Busse Grawitz
  15. Daniela Huzly
  16. Martin Schwemmle
  17. Hartmut Hengel
  18. Valeria Falcone

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Abstract

A dysregulated immune response with high levels of SARS-CoV-2 specific IgG antibodies characterizes patients with severe or critical COVID-19. Although a robust IgG response is traditionally considered to be protective, excessive triggering of activating Fc-gamma-receptors (FcγRs) could be detrimental and cause immunopathology. Here, we document that patients who develop soluble circulating IgG immune complexes (sICs) during infection are subject to enhanced immunopathology driven by FcγR activation. Utilizing cell-based reporter systems we provide evidence that sICs are predominantly formed prior to a specific humoral response against SARS-CoV-2. sIC formation, together with increased afucosylation of SARS-CoV-2 specific IgG eventually leads to an enhanced CD16 (FcγRIII) activation of immune cells reaching activation levels comparable active systemic lupus erythematosus (SLE) disease. Our data suggest a vicious cycle of escalating immunopathology driven by an early formation of sICs in predisposed patients. These findings reconcile the seemingly paradoxical findings of high antiviral IgG responses and systemic immune dysregulation in severe COVID-19.

Clinical implications

The identification of sICs as drivers of an escalating immunopathology in predisposed patients opens new avenues regarding intervention strategies to alleviate critical COVID-19 progression.

A vicious cycle of immunopathology in COVID-19 patients is driven by soluble multimeric immune complexes (sICs) . SARS-CoV-2 infection triggers sIC formation in prone individuals. Activation of FcγRIII/CD16 expressing immune cells by sICs precedes a humoral response to SARS-CoV2 infection. sICs and infection add to IgG afucosylation, further enhancing FcγRIII/CD16 activation by opsonized targets. High inflammation induces further sIC mediated immune cell activation ultimately leading to an escalating immunopathology.

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  1. Version published to 10.1101/2021.06.25.449893v4 on bioRxiv
  2. Version published to 10.1101/2021.06.25.449893v3 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2021.06.25.449893: (What is this?)

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

    Table 1: Rigor

    EthicsIACUC: Ethics: The protocol of this study conforms to the ethical guidelines of the 1975 Declaration of Helsinki and was approved by the institutional ethical committee of the University of Freiburg (EK 153/20).
    IRB: Ethics: The protocol of this study conforms to the ethical guidelines of the 1975 Declaration of Helsinki and was approved by the institutional ethical committee of the University of Freiburg (EK 153/20).
    Consent: Written informed consent was obtained from participants and the study was conducted according to federal guidelines, local ethics committee regulations (Albert-Ludwigs-Universität, Freiburg, Germany: No. F-2020-09-03-160428 and no. 322/20) All data associated with this study are present in the paper or Supplementary Materials.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line AuthenticationContamination: All cell lines were routinely tested for mycoplasma.
    Authentication: Assumptions about variance heterogeneity and normal distribution were checked by visual inspection of diagnostic plots.

    Table 2: Resources

    Antibodies
    SentencesResources
    Quantitation of antigen-specific IgG amount: In order to determine the relative S1- and N-SARS-CoV-2 specific IgG antibody concentration of the generated eluates, S1- and N-ELISA were performed by the anti-SARS-CoV-2 ELISA (IgG) Euroimmune Kit (Euroimmune, Lübeck, Germany) and anti-N SARS-CoV-2 IgG ELISA (recomWell SARS-CoV-2 IgG Kit (Mikrogen Diagnostik GmbH, Neuried, Germany) as aformentioned.
    antigen-specific IgG
    suggested: None
    N-SARS-CoV-2 specific IgG
    suggested: None
    anti-SARS-CoV-2 ELISA ( IgG
    suggested: None
    anti-N SARS-CoV-2 IgG
    suggested: None
    SARS-CoV-2 IgG
    suggested: None
    Briefly 250 µl serum was incubated overnight at 4°C with 5 µg of biotinylated anti-RBD-specific TRES-1-224.2.19 mouse monoclonal antibody or TRES-II-480 (isotype control) (kind gift of H.M. Jäck, Erlangen) before addition of streptavidin magnetic beads.
    anti-RBD-specific TRES-1-224.2.19
    suggested: None
    TRES-II-480
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Briefly, VeroE6 cells were seeded in 12-well plates at a density of 2.8×105 cells/well 24 h prior to infection.
    VeroE6
    suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
    Experimental Models: Organisms/Strains
    SentencesResources
    Peak lists were generated with ProteoWizard msConvert (http://proteowizard.sourceforge.net/; version 3.0.11098), linear shift mass recalibrated (after a preliminary database search) using in-house developed software and searched against a database containing the SARS-CoV-2 UniProtKB reference proteome (proteome ID: UP000464024), all human UniProtKB/Swiss-Prot entries, and optionally (to reduce the number of incorrectly assigned matches) selected bacterial proteins (finally the Pseudomonas fluorescens (strain SBW25) reference proteome; proteome ID: UP000002332) with Mascot 2.6.2 (
    SBW25
    suggested: None
    Recombinant DNA
    SentencesResources
    To verify Benzonase activity in the presence of human serum, 3 µg of pIRES-eGFP plasmid DNA (Addgene) were digested with 250 U of Benzonase.
    pIRES-eGFP
    suggested: None
    Software and Algorithms
    SentencesResources
    Peak lists were generated with ProteoWizard msConvert (http://proteowizard.sourceforge.net/; version 3.0.11098), linear shift mass recalibrated (after a preliminary database search) using in-house developed software and searched against a database containing the SARS-CoV-2 UniProtKB reference proteome (proteome ID: UP000464024), all human UniProtKB/Swiss-Prot entries, and optionally (to reduce the number of incorrectly assigned matches) selected bacterial proteins (finally the Pseudomonas fluorescens (strain SBW25) reference proteome; proteome ID: UP000002332) with Mascot 2.6.2 (
    ProteoWizard
    suggested: (ProteoWizard, RRID:SCR_012056)
    UniProtKB
    suggested: (UniProtKB, RRID:SCR_004426)
    Mascot
    suggested: (Mascot, RRID:SCR_014322)

    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 found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

    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.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

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  4. Version published to 10.1101/2021.06.25.449893v2 on bioRxiv
  5. Version published to 10.1101/2021.06.25.449893v1 on bioRxiv