Preliminary analysis of scRNA sequencing of children’s lung tissues excludes the expression of SARS-CoV-2 entry related genes as the key reason for the milder syndromes of COVID-19 in children

  1. Yue Tao
  2. Ruwen Yang
  3. Chen Wen
  4. Jue Fan
  5. Jing Ma
  6. Qiao He
  7. Zhiguang Zhao
  8. Xinyu Song
  9. Hao Chen
  10. Guocheng Shi
  11. Minzhi Yin
  12. Nan Fang
  13. Hao Zhang
  14. Huiwen Chen
  15. Xi Mo

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Abstract

To explore whether the expression levels of viral-entry associated genes might contribute to the milder symptoms in children, we analyzed the expression of these genes in both children and adults’ lung tissues by single cell RNA sequencing (scRNA-seq) and immunohistochemistry (IHC). Both scRNA-seq and IHC analyses showed comparable levels of the key genes for SARS-CoV-2 entry in children and adults, including ACE2, TMPRSS2 and FURIN , suggesting that instead of lower virus intrusion rate, other factors are more likely to be the key reasons for the milder symptoms of SARS-CoV-2 infected children.

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  1. Version published to 10.1101/2020.05.25.20110890v2 on medRxiv
  2. ScreenIT

    SciScore for 10.1101/2020.05.25.20110890: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: The study was approved by the Institutional Review Board and the Ethics Committee of Shanghai Children’s Medical Center (SCMCIRB-K2020034), and written informed consent was obtained from (the parents of) each patient.
    Consent: The study was approved by the Institutional Review Board and the Ethics Committee of Shanghai Children’s Medical Center (SCMCIRB-K2020034), and written informed consent was obtained from (the parents of) each patient.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    After deparaffinization, antigen retrieval and blocking, the slides were stained using the Leica Bond (Leica Biosystems) automated staining system with the following primary antibodies for 60 min at room temperature: anti-ACE2 antibody (rabbit, ab108252 from Abcam, 1:100) and anti-TMPRSS2 antibody (rabbit, ab92323 from Abcam, 1:2000) for cohort 1; anti-ACE2 antibody (mouse, 66699-1-Ig, Proteintech, 1:1200) and anti-FURIN antibody (rabbit, 18413-1-AP, Proteintech, 1:1200) for cohort 2.
    anti-ACE2
    suggested: (Enzo Life Sciences Cat# ALX-804-722-C100, RRID:AB_11180102)
    anti-TMPRSS2
    suggested: (Abcam Cat# ab92323, RRID:AB_10585592)
    anti-FURIN
    suggested: (Proteintech Cat# 18413-1-AP, RRID:AB_2878542)
    After three washes, the slides were then incubated with HRP-conjugated goat anti-rabbit antibody (cohort 1) or biotin-conjugated anti-mouse/rabbit secondary antibody (cohort 2) for 15-30 min at room temperature.
    anti-rabbit
    suggested: None
    anti-mouse/rabbit
    suggested: (Biorbyt Cat# orb27530, RRID:AB_10954533)
    Software and Algorithms
    SentencesResources
    Briefly, FastQC (V0.11.7), fastp (1)
    FastQC
    suggested: (FastQC, RRID:SCR_014583)
    STAR aligner (2.5.3a) and featureCounts (1.6.2) were used for quality evaluation, trimming, alignment and transcript counting, respectively
    STAR
    suggested: (STAR, RRID:SCR_015899)
    featureCounts
    suggested: (featureCounts, RRID:SCR_012919)
    Adapters and poly(A) tails were trimmed before aligning read 2 to GRCh38 with Ensembl 92 annotation.
    Ensembl
    suggested: (Ensembl, RRID:SCR_002344)
    Integrated analysis of children lung data with public adult lung data: The public data of 8 healthy adults were downloaded from Gene Expression Omnibus (GSE122960)
    Gene Expression Omnibus
    suggested: (Gene Expression Omnibus (GEO, RRID:SCR_005012)
    Statistical analyses of IHC scores were performed in GraphPad Prism 8 using Mann-Whitney test with an exact p value computed.
    GraphPad Prism
    suggested: (GraphPad Prism, RRID:SCR_002798)

    Results from OddPub: Thank you for sharing your data.

    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 13. 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

    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.

  3. Version published to 10.1101/2020.05.25.20110890v1 on medRxiv