SARS-CoV-2 Spike Glycoprotein S1 Induces Neuroinflammation in BV-2 Microglia

  1. Olumayokun A. Olajide
  2. Victoria U. Iwuanyanwu
  3. Oyinkansola D. Adegbola
  4. Alaa A. Al-Hindawi

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Abstract

In addition to respiratory complications produced by SARS‐CoV‐2, accumulating evidence suggests that some neurological symptoms are associated with the disease caused by this coronavirus. In this study, we investigated the effects of the SARS‐CoV‐2 spike protein S1 stimulation on neuroinflammation in BV-2 microglia. Analyses of culture supernatants revealed an increase in the production of TNF-α, IL-6, IL-1β and iNOS/NO. S1 also increased protein levels of phospho-p65 and phospho-IκBα, as well as enhanced DNA binding and transcriptional activity of NF-κB. These effects of the protein were blocked in the presence of BAY11-7082 (1 µM). Exposure of S1 to BV-2 microglia also increased the protein levels of NLRP3 inflammasome and enhanced caspase-1 activity. Increased protein levels of p38 MAPK was observed in BV-2 microglia stimulated with the spike protein S1 (100 ng/ml), an action that was reduced in the presence of SKF 86,002 (1 µM). Results of immunofluorescence microscopy showed an increase in TLR4 protein expression in S1-stimulated BV-2 microglia. Furthermore, pharmacological inhibition with TAK 242 (1 µM) and transfection with TLR4 small interfering RNA resulted in significant reduction in TNF-α and IL-6 production in S1-stimulated BV-2 microglia. These results have provided the first evidence demonstrating S1-induced neuroinflammation in BV-2 microglia. We propose that induction of neuroinflammation by this protein in the microglia is mediated through activation of NF-κB and p38 MAPK, possibly as a result of TLR4 activation. These results contribute to our understanding of some of the mechanisms involved in CNS pathologies of SARS-CoV-2.

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  1. Version published to 10.1007/s12035-021-02593-6
  2. Version published to 10.1101/2020.12.29.424619v3 on bioRxiv
  3. Version published to 10.1101/2020.12.29.424619v2 on bioRxiv
  4. ScreenIT

    SciScore for 10.1101/2020.12.29.424619: (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.

    Table 2: Resources

    Antibodies
    SentencesResources
    The following antibodies were used: rabbit anti-iNOS (Abcam), rabbit anti-phospho-p65 (Cell Signalling Technology), rabbit anti-phospho-IκBα (Santa Cruz Biotechnology), rabbit anti-NLRP3 (Abcam), and rabbit anti-phospho-p38 (Cell Signalling Technology) antibodies.
    rabbit anti-iNOS ( Abcam)
    suggested: None
    anti-iNOS
    suggested: None
    rabbit
    suggested: None
    rabbit anti-phospho-IκBα ( Santa Cruz Biotechnology) , rabbit anti-NLRP3 ( Abcam) , and rabbit anti-phospho-p38 ( Cell Signalling Technology ) antibodies
    suggested: None
    rabbit anti-phospho-IκBα ( Santa Cruz Biotechnology)
    suggested: None
    anti-phospho-IκBα ( Santa Cruz Biotechnology)
    suggested: None
    rabbit anti-NLRP3 ( Abcam)
    suggested: None
    rabbit anti-phospho-p38 ( Cell Signalling Technology ) antibodies
    suggested: None
    anti-phospho-p38 ( Cell Signalling Technology )
    suggested: None
    Thereafter, cells were washed with PBS and incubated with anti-rabbit HRP secondary antibody for 2 h at room temperature.
    anti-rabbit HRP
    suggested: None
    Software and Algorithms
    SentencesResources
    Statistical analysis were conducted using the GraphPad Prism software (version 9).
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)

    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.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  5. Version published to 10.1101/2020.12.29.424619v1 on bioRxiv