IFN signaling and neutrophil degranulation transcriptional signatures are induced during SARS-CoV-2 infection

  1. Bruce A. Rosa
  2. Mushtaq Ahmed
  3. Dhiraj K. Singh
  4. José Alberto Choreño-Parra
  5. Journey Cole
  6. Luis Armando Jiménez-Álvarez
  7. Tatiana Sofía Rodríguez-Reyna
  8. Bindu Singh
  9. Olga Gonzalez
  10. Ricardo Carrion
  11. Larry S. Schlesinger
  12. John Martin
  13. Joaquín Zúñiga
  14. Makedonka Mitreva
  15. Deepak Kaushal
  16. Shabaana A. Khader

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Abstract

SARS-CoV-2 virus has infected more than 92 million people worldwide resulting in the Coronavirus disease 2019 (COVID-19). Using a rhesus macaque model of SARS-CoV-2 infection, we have characterized the transcriptional signatures induced in the lungs of juvenile and old macaques following infection. Genes associated with Interferon (IFN) signaling, neutrophil degranulation and innate immune pathways are significantly induced in macaque infected lungs, while pathways associated with collagen formation are downregulated, as also seen in lungs of macaques with tuberculosis. In COVID-19, increasing age is a significant risk factor for poor prognosis and increased mortality. Type I IFN and Notch signaling pathways are significantly upregulated in lungs of juvenile infected macaques when compared with old infected macaques. These results are corroborated with increased peripheral neutrophil counts and neutrophil lymphocyte ratio in older individuals with COVID-19 disease. Together, our transcriptomic studies have delineated disease pathways that improve our understanding of the immunopathogenesis of COVID-19.

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  1. Version published to 10.1038/s42003-021-01829-4
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    SciScore for 10.1101/2020.08.06.239798: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIACUC: The animal studies in each of the species were approved by the Animal Care and Use Committee of the Texas Biomedical Research Institute and as an omnibus Biosafety Committee protocol.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Software and Algorithms
    SentencesResources
    After adapter trimming using Trimmomatic v0.39(14), sequenced RNA-seq reads were aligned to the Macaca mulatta genome (version 10, Ensembl release 100(15)) using the STAR aligner v2.7.3a(16)
    Trimmomatic
    suggested: (Trimmomatic, RRID:SCR_011848)
    STAR
    suggested: (STAR, RRID:SCR_015899)
    All raw RNA-Seq fastq files were uploaded to the NCBI Sequence Read Archive (SRA(17)), and complete sample metadata and accession information are provided in Table S1.
    NCBI Sequence Read Archive
    suggested: (NCBI Sequence Read Archive (SRA, RRID:SCR_004891)
    Read fragments (read pairs or single reads) were quantified per gene per sample using featureCounts v1.5.1(18).
    featureCounts
    suggested: (featureCounts, RRID:SCR_012919)
    Principal components analysis also was calculated using DESeq2 output (default settings, using the top 500 most variable genes).
    DESeq2
    suggested: (DESeq, RRID:SCR_000154)
    Pathway enrichment analysis among differentially expressed gene sets of interest was performed for (a) Reactome(21) pathways, using the human orthologs as input into the WebGestalt(22) web server (p ≤ 0.05 after FDR correction, minimum 3 genes per term) and (b) KEGG(23) pathways and Gene Ontology(24) terms, using the g:profiler web server(25) which has a database of these annotations matched to macaque ENSEMBL gene IDs (p ≤ 0.05 after FDR correction, minimum 3 genes per term).
    WebGestalt(22
    suggested: None
    g:profiler
    suggested: (G:Profiler, RRID:SCR_006809)
    Mapped fragment counts, relative gene expression levels, gene annotations, and differential expression data for every macaque gene are available in Table S2, along with orthology matches to human genes retrieved from ENSEMBL(15) and identifications of differentially expressed (DE) genes belonging to enriched pathways of interest, for genes of interest in Table S3, and significant functional enrichment for Reactome, KEGG and Gene Ontology pathways, among differentially gene sets of interest in Table S4.
    KEGG
    suggested: (KEGG, RRID:SCR_012773)
    Plasma levels of interferon-gamma (IFN-γ) and vascular endothelial growth factor (VEGF), were determined by Luminex assays using the Luminex platform Bio-Plex Multiplex 200 (Bio-Rad Laboratories, Inc., Hercules, CA, USA)
    Bio-Rad Laboratories
    suggested: (Bio-Rad Laboratories, RRID:SCR_008426)

    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:
    To overcome these limitations, we have generated a nonhuman primate model (rhesus macaques) of SARS-CoV-2 infection that reflects several features of the immunopathogenesis of human COVID-19, and provides a platform to interrogate the immune pathways that mediate disease versus protection, especially in the context of young versus older hosts. In this study, we show that upregulation of pathways characteristic of neutrophil degranulation and IFN signaling are characteristic of COVID-19 disease in infected hosts. Importantly, the significantly higher induction of genes associated with Type I IFN signaling pathway and Notch signaling in young macaques infected with SARS-CoV-2 is a key determinant that distinguishes them from infected old macaques. Lungs of old macaques infected with COVID-19 however, uniquely feature downregulation of VEGF signaling pathways. Importantly, in PBMCs of humans infected with SARS-CoV-2 we found increased levels of VEGF and peripheral neutrophil counts in individuals >60 years when compared to younger individuals. These results together provide novel insights into the immunopathogenesis of COVID-19 disease, especially from the unique perceptive of age as a contributing factor. As we learn more about the pathophysiology of COVID-19, it is becoming clear that disease severity is associated with hyperinflammation which in turn induces lung and multiorgan injury and mortality via a cytokine storm (1, 2, 56). While therapeutic options that focus on immun...

    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 33. 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

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  3. Version published to 10.1101/2020.08.06.239798v1 on bioRxiv