T cell responses to SARS-CoV-2 in people with and without neurologic symptoms of long COVID

  1. Lavanya Visvabharathy
  2. Barbara A. Hanson
  3. Zachary S. Orban
  4. Patrick H. Lim
  5. Nicole M. Palacio
  6. Millenia Jimenez
  7. Jeffrey R. Clark
  8. Edith L. Graham
  9. Eric M. Liotta
  10. George Tachas
  11. Pablo Penaloza-MacMaster
  12. Igor J. Koralnik

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Abstract

Many people experiencing long COVID syndrome, or post-acute sequelae of SARS-CoV-2 infection (PASC), suffer from debilitating neurologic symptoms (Neuro-PASC). However, whether virus-specific adaptive immunity is affected in Neuro-PASC patients remains poorly understood. We report that Neuro-PASC patients exhibit distinct immunological signatures composed of elevated humoral and cellular responses toward SARS-CoV-2 Nucleocapsid protein at an average of 6 months post-infection compared to healthy COVID convalescents. Neuro-PASC patients also had enhanced virus-specific production of IL-6 from and diminished activation of CD8 + T cells. Furthermore, the severity of cognitive deficits or quality of life disturbances in Neuro-PASC patients were associated with a reduced diversity of effector molecule expression in T cells but elevated IFN-γ production to the C-terminal domain of Nucleocapsid protein. Proteomics analysis showed enhanced plasma immunoregulatory proteins and reduced pro-inflammatory and antiviral response proteins in Neuro-PASC patients compared with healthy COVID convalescents, which were also correlated with worse neurocognitive dysfunction. These data provide new insight into the pathogenesis of long COVID syndrome and a framework for the rational design of predictive biomarkers and therapeutic interventions.

One Sentence Summary

Adaptive immunity is altered in patients with neurologic manifestations of long COVID.

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  1. Version published to 10.1101/2021.08.08.21261763v4 on medRxiv
  2. Version published to 10.1101/2021.08.08.21261763v3 on medRxiv
  3. ScreenIT

    SciScore for 10.1101/2021.08.08.21261763: (What is this?)

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

    Table 1: Rigor

    EthicsIRB: Ethics Statement: This study was approved by the Northwestern University Institutional Review Board (Koralnik Lab, IRB STU00212583).
    Consent: Informed consent was obtained from all enrolled participants.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    ELISA: Antigen-specific total antibody titers were measured by ELISA as described previously (Dangi et al., 2020; Palacio et al., 2020).
    ELISA: Antigen-specific total
    suggested: None
    Antigen-specific
    suggested: None
    Plates were washed three times with wash buffer followed by addition of secondary antibody conjugated to horseradish peroxidase, goat anti-human IgG (H + L) (Jackson ImmunoResearch) diluted in blocking solution (1:1000) and 100 µl/well was added and incubated for 60 min at room temperature.
    anti-human IgG
    suggested: None
    Recombinant DNA
    SentencesResources
    Vector pCAGGS containing the SARS-related coronavirus 2, Wuhan-Hu-1 spike glycoprotein gene (soluble, stabilized), NR-52394 and receptor binding domain (RBD), NR-52309, nucleocapsid gene NR-53507.
    pCAGGS
    suggested: RRID:Addgene_18926)
    Software and Algorithms
    SentencesResources
    The PBMC layer was collected and washed 2x in sterile PBS before red blood cell lysis with ACK buffer (Quality Biologicals).
    Quality Biologicals
    suggested: None
    Data was acquired on a BD FACSymphony Spectral analyzer and analyzed using FlowJo v10 (BD Biosciences) and SPICE-Pestle (Roederer et al., 2011).
    FlowJo
    suggested: (FlowJo, RRID:SCR_008520)
    Quantification and Statistical Analysis: Statistical tests to determine significance are described in figure legends and conducted largely in Prism (GraphPad).
    Prism
    suggested: (PRISM, RRID:SCR_005375)
    GraphPad
    suggested: (GraphPad Prism, RRID:SCR_002798)
    For pie graphs or heatmaps generated using SPICE analysis, statistics were determined by Permutation test following unstimulated background subtraction, with additional thresholding of 0.03% to account for noise, using SPICE-Pestle.
    SPICE
    suggested: (SPICE, RRID:SCR_016603)
    Clinical data were collected and managed using REDCap electronic data capture tools hosted at Northwestern University Feinberg School of Medicine (Harris et al., 2009)
    REDCap
    suggested: (REDCap, RRID:SCR_003445)

    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:
    Limitations of study: One limitation of our study is the relatively small sample size of unvaccinated neuro-PASC patients. This was due to the wide implementation of SARS-CoV-2 vaccines in the Chicago area soon after beginning study enrollment. Another limitation was not being able to control for time of sample collection with respect to date of COVID-19 symptom onset. As it is possible that neuro-PASC could be the result of a persistent infection, further investigations would require testing of potential cryptic reservoirs, including stool samples from CN patients.

    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

    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.

  4. Version published to 10.1101/2021.08.08.21261763v1 on medRxiv
  5. Version published to 10.1101/2021.08.08.21261763v2 on medRxiv