A human disease model of SARS-CoV-2-induced lung injury and immune responses with a microengineered organ chip

  1. Min Zhang
  2. Peng Wang
  3. Ronghua Luo
  4. Yaqing Wang
  5. Zhongyu Li
  6. Yaqiong Guo
  7. Yulin Yao
  8. Minghua Li
  9. Tingting Tao
  10. Wenwen Chen
  11. Jianbao Han
  12. Haitao Liu
  13. Kangli Cui
  14. Xu zhang
  15. Yongtang Zheng
  16. Jianhua Qin

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Abstract

Coronavirus disease 2019 (COVID-19) is a global pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that seriously endangers human health. There is an urgent need to build physiological relevant human models for deep understanding the complex organ-level disease processes and facilitating effective therapeutics for COVID-19. Here, we first report the use of microengineered alveolus chip to create a human disease model of lung injury and immune responses induced by native SARS-CoV-2 at organ-level. This biomimetic system is able to reconstitute the key features of human alveolar-capillary barrier by co-culture of alveolar epithelial and microvascular endothelial cells under microfluidic flow. The epithelial cells on chip showed higher susceptibility to SARS-CoV-2 infection than endothelial cells identified by viral spike protein expression. Transcriptional analysis showed distinct responses of two cell types to SARS-CoV-2 infection, including activated type I interferon (IFN-I) signaling pathway in epithelium and activated JAK-STAT signaling pathway in endothelium. Notably, in the presence of circulating immune cells, a series of alveolar pathological changes were observed, including the detachment of endothelial cells, recruitment of immune cells, and increased production of inflammatory cytokines (IL-6, IL-8, IL-1β and TNF-α). These new findings revealed a crucial role of immune cells in mediating lung injury and exacerbated inflammation. Treatment with antiviral compound remdesivir could suppress viral copy and alleviate the disruption of alveolar barrier integrity induced by viral infection. This bioengineered human organ chip system can closely mirror human-relevant lung pathogenesis and immune responses to SARS-CoV-2 infection, not possible by other in vitro models, which provides a promising and alternative platform for COVID-19 research and preclinical trials.

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  1. ScreenIT

    SciScore for 10.1101/2020.07.20.211789: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board Statementnot detected.
    RandomizationThe kit eliminates duplication bias in PCR and sequencing steps, using unique molecular identifier (UMI) of 8 random bases to label the pre-amplified cDNA molecules.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    After being blocked with 5% BSA in TBST buffer containing 0.05% Tween-20, the membranes were probed with the anti-ACE2 antibody (1:1000 dilution), anti-TMPRSS2 antibody (1:1000 dilution) or anti-GAPDH antibody (1:2000 dilution) at 4ū overnight.
    anti-ACE2
    suggested: None
    anti-TMPRSS2
    suggested: None
    anti-GAPDH
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Human lung microvasculature cell line HULEC-5a was purchased from Procell Corporation and were maintained in HULEC-5a growth medium (Procell, CM-0565).
    HULEC-5a
    suggested: ATCC Cat# CRL-3244, RRID:CVCL_0A11)
    Virus: A clinical isolate SARS-COV-2 strain 107 was obtained from Guangdong Provincial Center for Disease Control and Prevention, China, and propagated in Vero E6 cells.
    Vero E6
    suggested: RRID:CVCL_XD71)
    The virus titers were (infectious titers of virus) were determined by a TCID50 assay on Vero cells.
    Vero
    suggested: CLS Cat# 605372/p622_VERO, RRID:CVCL_0059)
    Software and Algorithms
    SentencesResources
    Image processing was done using ImageJ (NIH).
    ImageJ
    suggested: (ImageJ, RRID:SCR_003070)
    RNA-seq data analysis: Raw sequencing data was first filtered by Trimmomatic (version 0.36), low-quality reads were discarded and the reads contaminated with adaptor sequences were trimmed.
    Trimmomatic
    suggested: (Trimmomatic, RRID:SCR_011848)
    They were mapped to the reference genome of Homo sapiens from Ensembl database (ftp://ftp.ensembl.org/pub/release-87/fasta/homo_sapiens/dna/) using STAR software (version 2.5.3a) with default parameters.
    Ensembl
    suggested: (Ensembl, RRID:SCR_002344)
    STAR
    suggested: (STAR, RRID:SCR_015899)
    Reads mapped to the exon regions of each gene were counted by featureCounts (Subread-1.5.1; Bioconductor) and then RPKMs were calculated.
    featureCounts
    suggested: (featureCounts, RRID:SCR_012919)
    Bioconductor
    suggested: (Bioconductor, RRID:SCR_006442)
    Genes differentially expressed between groups were identified using the edgeR package (version 3.12.1).
    edgeR
    suggested: (edgeR, RRID:SCR_012802)
    Gene ontology (GO) analysis and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis for differentially expressed genes were both implemented by KOBAS software (version: 2.1.1) with a corrected P-value cutoff of 0.05 to judge statistically significant enrichment.
    KEGG
    suggested: (KEGG, RRID:SCR_012773)
    KOBAS
    suggested: (KOBAS, RRID:SCR_006350)
    Statistical analyses: Data were collected in Excel (Microsoft).
    Excel
    suggested: None

    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:
    One potential limitation of this work is the lack of human primary alveolar tissues that contains multicellular types of pneumocytes, such as alveolar epithelial type I and type II cells as existing in vivo. In addition, this model has yet to be tried for assessing more other drug candidates for COVID-19 therapeutics. Despite some limitations, the great value of the human alveolus chip is that it is capable to model human lung pathophysiology and study host-immune responses to respiratory viral infection at organ level. In combination with the existing cell-based models, this bioengineered lung infection model on chip may provide a complement to animal models for evaluating candidate drugs and repurposing approved drugs to face the crisis of SARS-CoV-2 epidemic. Collectively, this work provides a proof-of-concept to establish a microengineered human disease model in vitro that enables to closely recapitulate lung pathophysiology and immune responses to native SARS-CoV-2 infection for the first time. Comprehensive analysis of this infection model revealed new insights into the pathogenesis associated with COVID-19, in which virus-induced inflammation is a major contributor lung injury with the involvement of circulating immune cells. This human organ chip system provided a synthetic strategy to rebuild human organs and analyze pathological responses at organ level by flexibly varying system parameters, which is opening up new avenues for COVID-19 research and drug development.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

    Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on pages 19, 31 and 23. 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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  2. Version published to 10.1101/2020.07.20.211789 on bioRxiv