Deep immune profiling of the maternal-fetal interface with mild SARS-CoV-2 infection

  1. Suhas Sureshchandra
  2. Michael Z Zulu
  3. Brianna Doratt
  4. Allen Jankeel
  5. Delia Tifrea
  6. Robert Edwards
  7. Monica Rincon
  8. Nicole E. Marshall
  9. Ilhem Messaoudi

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Abstract

Pregnant women are an at-risk group for severe COVID-19, though the majority experience mild/asymptomatic disease. Although severe COVID-19 has been shown to be associated with immune activation at the maternal-fetal interface even in the absence of active viral replication, the immune response to asymptomatic/mild COVID-19 remains unknown. Here, we assessed immunological adaptations in both blood and term decidua from 9 SARS-exposed pregnant women with asymptomatic/mild disease and 15 pregnant SARS-naive women. In addition to selective loss of tissue-resident decidual macrophages, we report attenuation of antigen presentation and type I IFN signaling but upregulation of inflammatory cytokines and chemokines in blood monocyte derived decidual macrophages. On the other hand, infection was associated with remodeling of the T cell compartment with increased frequencies of activated CD69+ tissue-resident T cells and decreased abundance of Tregs. Interestingly, frequencies of cytotoxic CD4 and CD8 T cells increased only in the blood, while CD8 effector memory T cells were expanded in the decidua. In contrast to decidual macrophages, signatures of type I IFN signaling were increased in decidual T cells. Finally, T cell receptor diversity was significantly reduced with infection in both compartments, albeit to a much greater extent in the blood. The resulting aberrant immune activation in the placenta, even with asymptomatic disease may alter the exquisitely sensitive developing fetal immune system, leading to long-term adverse outcomes for offspring.

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  1. ScreenIT

    SciScore for 10.1101/2021.08.23.457408: (What is this?)

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

    Table 1: Rigor

    EthicsIRB: Study Participants and Experimental Design: This study was approved by University of California Irvine Institutional Review Boards (Protocol ID HS# 2012-8716) and Oregon and Health Sciences University (eIRB# 22713 and 16328).
    Consent: Informed consent was obtained from all enrolled subjects.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    For the innate panel, the following antibodies were used: CD3 (SP34, BD Pharmingen) and CD20 (2H7, BioLegend) for the exclusion of T & B lymphocytes, respectively.
    CD3
    suggested: (BD Biosciences Cat# 556610, RRID:AB_396483)
    CD20
    suggested: None
    For the adaptive panel, the following antibodies were used: CD4 (OKT4, BioLegend), CD8b (2ST8.5H7, Beckman Coulter), CD45RA (HI100, TONBO Biosciences), CCR7 (G043H7, BD Biosciences), CD19 (HIB19, BioLegend), IgD (IA6-2, BioLegend), CD27 (M-T271, BioLegend), KLRG1 (SA231A2, BioLegend) and PD-1 (Eh12.2h7, BioLegend).
    CD4
    suggested: (BioLegend Cat# 391503, RRID:AB_2721611)
    CD8b
    suggested: (Abcam Cat# ab34397, RRID:AB_2291359)
    CD45RA
    suggested: None
    CCR7
    suggested: None
    CD19
    suggested: None
    HIB19
    suggested: None
    CD27
    suggested: None
    KLRG1
    suggested: None
    SA231A2
    suggested: None
    Decidua Immunophenotyping: 1-2 × 106 fresh decidual leukocytes were washed with PBS and stained using the following cocktail of antibodies: CD45 (HI30, Biolegend), CD66b (G10F5, BioLegend),
    CD45
    suggested: None
    HI30
    suggested: None
    CD66b
    suggested: None
    Samples were washed thoroughly with a cell staining buffer (1X PBS with 0.5% BSA), and Fc blocked for 10 minutes (Human TruStain FcX, Biolegend), and incubated with a cocktail containing CD3-PE (SP34, BD Pharmingen), 0.5 ug each of oligo tagged CD4 (TotalSeq™-C0072, Biolegend), CD8 (TotalSeq™-C0046, Biolegend), CCR7 (TotalSeq™-C0148, Biolegend), CD45RA (TotalSeq™-C0063, Biolegend), CD69 (TotalSeq™-C0146, Biolegend), CD103 (TotalSeq™-C0145, Biolegend), PD-1(TotalSeq™-C0088, Biolegend), CD25 (TotalSeq™-C0085, Biolegend), and unique 5’ hashing antibody (TotalSeq™-C0251, C0254, C0256, and C0260, Biolegend) for an additional 30 minutes at 4C.
    TotalSeq™-C0072
    suggested: None
    CD8
    suggested: (BD Biosciences Cat# 346048, RRID:AB_400455)
    TotalSeq™-C0046
    suggested: None
    TotalSeq™-C0063
    suggested: None
    CD69
    suggested: (BD Biosciences Cat# 558063, RRID:AB_2275535)
    TotalSeq™-C0146
    suggested: None
    CD103
    suggested: None
    TotalSeq™-C0145
    suggested: None
    PD-1
    suggested: None
    TotalSeq™-C0088 , Biolegend) , CD25
    suggested: None
    TotalSeq™-C0085
    suggested: None
    TotalSeq™-C0251
    suggested: None
    C0254
    suggested: (Assay Biotech Cat# C0254, RRID:AB_10684215)
    C0256
    suggested: (Assay Biotech Cat# C0256, RRID:AB_10684216)
    C0260
    suggested: (Assay Biotech Cat# C0260, RRID:AB_10684622)
    Software and Algorithms
    SentencesResources
    For the adaptive panel, the following antibodies were used: CD4 (OKT4, BioLegend), CD8b (2ST8.5H7, Beckman Coulter), CD45RA (HI100, TONBO Biosciences), CCR7 (G043H7, BD Biosciences), CD19 (HIB19, BioLegend), IgD (IA6-2, BioLegend), CD27 (M-T271, BioLegend), KLRG1 (SA231A2, BioLegend) and PD-1 (Eh12.2h7, BioLegend).
    BD Biosciences
    suggested: (BD Biosciences, RRID:SCR_013311)
    Data were analyzed using FlowJO (Ashland OR).
    FlowJO
    suggested: (FlowJo, RRID:SCR_008520)
    Data normalization and variance stabilization were performed on the integrated object using the NormalizeData and ScaleData functions in Seurat where a regularized negative binomial regression corrected for differential effects of mitochondrial and ribosomal gene expression levels.
    Seurat
    suggested: (SEURAT, RRID:SCR_007322)

    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:
    We acknowledge the limitations of our study that include a small sample size, which combined with the wide window of infectivity during pregnancy, precluded the assessment of gestational age on infection outcomes. Given the limited amount of blood obtained, we were unable to conduct a comprehensive measurement of antigen specific T and B cell responses. Finally, it remains unclear if the placenta harbors virus-specific T cells that migrate from blood. Importantly, while there is no evidence of vertical transmission (24), enrichment of activated T cells and loss of regulatory tissue resident macrophages (dMac1) and Tregs with infection skews the balance of decidual immune cells towards a proinflammatory state. The resulting aberrant immune activation in the placenta and misfiring of local cytokine networks can potentially contribute to pregnancy complications. Additionally, the developing fetal immune system is acutely sensitive to inflammation and stress, and it remains to be seen if such exposures, even with mild maternal infection can have long-term consequences on immunity in the offspring.

    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.

  2. Version published to 10.1101/2021.08.23.457408v1 on bioRxiv