SARS-CoV-2 infects lung epithelial cells and induces senescence and an inflammatory response in patients with severe COVID-19

  1. Konstantinos Evangelou
  2. Dimitris Veroutis
  3. Periklis G. Foukas
  4. Koralia Paschalaki
  5. Nefeli Lagopati
  6. Marios Dimitriou
  7. Angelos Papaspyropoulos
  8. Orsalia Hazapis
  9. Aikaterini Polyzou
  10. Sophia Havaki
  11. Athanassios Kotsinas
  12. Christos Kittas
  13. Athanasios G. Tzioufas
  14. Laurence de Leval
  15. Demetris Vassilakos
  16. Sotirios Tsiodras
  17. Ioannis Karakasiliotis
  18. Peter J Barnes
  19. Vassilis G. Gorgoulis

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Abstract

Rationale

SARS-CoV-2 infection of the respiratory system can progress to a life threatening multi-systemic disease, mediated via an excess of cytokines (“cytokine storm”), but the molecular mechanisms are poorly understood.

Objectives

To investigate whether SARS-CoV-2 may induce cellular senescence in lung epithelial cells, leading to secretion of inflammatory cytokines, known as the senescence-associated secretory phenotype (SASP).

Methods

Autopsy lung tissue samples from eleven COVID-19 patients and sixty age-matched non-infected controls were analysed by immunohistochemistry for SARS-CoV-2 and markers of cellular senescence (SenTraGor, p16 INK4A ) and key SASP cytokines (interleukin-1β, interleukin-6). We also investigated whether SARS-CoV-2 infection of an epithelial cell line induces senescence and cytokine secretion.

Measurements and Main Results

SARS-CoV-2 was detected by immunocytochemistry and electron microscopy predominantly in alveolar type-2 (AT2) cells, which also expressed the angiotensin-converting-enzyme 2 (ACE2), a critical entry receptor for this virus. In COVID-19 samples, AT2 cells displayed increased markers of senescence [p16 INK4A , SenTraGor staining positivity in 12±1.2% of cells compared to 1.7±0.13% in non-infected controls (p<0.001)], with markedly increased expression of interleukin-1β and interleukin-6 (p<0.001). Infection of epithelial cells (Vero E6) with SARS-CoV-2 in-vitro induced senescence and DNA damage (increased SenTraGor and γ-H2AX), and reduced proliferation (Ki67) compared to uninfected control cells (p<0.01).

Conclusions

We demonstrate that in severe COVID-19 patients, AT2 cells are infected with SARS-CoV-2 and show senescence and expression of proinflammatory cytokines. We also show that SARS-CoV-2 infection of epithelial cells may induce senescence and inflammation, indicating that cellular senescence may be an important molecular mechanism of severe COVID-19.

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  1. Version published to 10.1101/2021.01.02.424917v2 on bioRxiv
  2. ScreenIT

    SciScore for 10.1101/2021.01.02.424917: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: All protocols were approved by the Commission cantonale d’éthique de la recherche, Lausanne (CERVD) Ref 2020-01257 and the Bio-Ethics Committee of Athens Medical School.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablePatient samples: Formalin-Fixed-Paraffin-Embedded lung tissues from ten COVID-19 patients (mean±SD: 72.4±16yr; 7 males) confirmed by molecular testing, and ten age-matched non-COVID-19 lung tissues samples from previously published cohort (71.8 ± 15 yr, 5 males) were analyzed (7).

    Table 2: Resources

    No key resources detected.

    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: 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: 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.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    About SciScore

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  3. Version published to 10.1101/2021.01.02.424917v1 on bioRxiv