Selectively expressing SARS-CoV-2 Spike protein S1 subunit in cardiomyocytes induces cardiac hypertrophy in mice

  1. Steven G. Negron
  2. Chase W. Kessinger
  3. Bing Xu
  4. William T. Pu
  5. Zhiqiang Lin

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Abstract

Cardiac injury is common in hospitalized COVID-19 patients and portends poorer prognosis and higher mortality. To better understand how SARS-CoV-2 (CoV-2) damages the heart, it is critical to elucidate the biology of CoV-2 encoded proteins, each of which may play multiple pathological roles. For example, CoV-2 Spike glycoprotein (CoV-2-S) not only engages ACE2 to mediate virus infection, but also directly impairs endothelial function and can trigger innate immune responses in cultured murine macrophages. Here we tested the hypothesis that CoV-2-S damages the heart by activating cardiomyocyte (CM) innate immune responses. HCoV-NL63 is another human coronavirus with a Spike protein (NL63-S) that also engages ACE2 for virus entry but is known to only cause moderate respiratory symptoms. We found that CoV-2-S and not NL63-S interacted with Toll-like receptor 4 (TLR4), a crucial pattern recognition receptor that responsible for detecting pathogen and initiating innate immune responses. Our data show that the S1 subunit of CoV-2-S (CoV-2-S1) interacts with the extracellular leucine rich repeats-containing domain of TLR4 and activates NF-kB. To investigate the possible pathological role of CoV-2-S1 in the heart, we generated a construct that expresses membrane-localized CoV-2-S1 (S1-TM). AAV9-mediated, selective expression of the S1-TM in CMs caused heart dysfunction, induced hypertrophic remodeling, and elicited cardiac inflammation. Since CoV-2-S does not interact with murine ACE2, our study presents a novel ACE2-independent pathological role of CoV-2-S, and suggests that the circulating CoV-2-S1 is a TLR4-recognizable alarmin that may harm the CMs by triggering their innate immune responses.

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    SciScore for 10.1101/2021.06.20.448993: (What is this?)

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

    Table 1: Rigor

    EthicsIACUC: Experimental animals: All animal procedures were approved by the Institute Animal Care and Use Committees of Masonic Medical Research Institute.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Experimental Models: Cell Lines
    SentencesResources
    Luciferase reporter assay: HEK 293T cells were cultured in 24 well plates for luciferase assay.
    HEK 293T
    suggested: None

    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: We detected the following sentences addressing limitations in the study:
    Limitation of the current study: In severe COVID-19 patients, cardiac injury is likely a result of acute pneumonias (Jirak et al., 2021; Metkus et al., 2021). Since our AAV only express S1-TM in the CMs, our system is not suitable for studying the pathogenesis of severe COVID-19 related cardiac injury. Rather, this AAV system can be used for investigating the pathogenesis of myocardial injury without COVID-19 acute respiratory distress, and it is also a powerful platform for dissecting the pathological roles of CoV-2 encoded genes in the heart or other organs. In addition, the current study did not test whether CoV-2-S impairs CMs electrophysiology, a crucial question to be addressed in future studies.

    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.

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

    About SciScore

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  2. Version published to 10.1101/2021.06.20.448993v1 on bioRxiv