Pulmonary infection by SARS-CoV-2 induces senescence accompanied by an inflammatory phenotype in severe COVID-19: possible implications for viral mutagenesis.

Konstantinos Evangelou,Orsalia Hazapis,Vassilis G Gorgoulis,Christos Kittas,Athanasios G Tzioufas,Bindu Konda,Barry R Stripp,Ioannis Karakasiliotis,Koralia Paschalaki,Marios Dimitriou,Peter J Barnes,Aikaterini Polyzou,Sotirios Tsiodras,Athanassios Kotsinas,Angelos Papaspyropoulos,Demetris Vassilakos,Sophia Havaki,Dimitris Veroutis,Giovanni Blandino,Argyris Papantonis,Laurence De Leval,Nefeli Lаgopati ,Periklis Foukas

doi:10.1183/13993003.02951-2021

Abstract

BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection of the respiratory system can progress to a multisystemic disease with aberrant inflammatory response. Cellular senescence promotes chronic inflammation, named senescence-associated secretory phenotype (SASP). We investigated whether coronavirus disease 2019 (COVID-19) is associated with cellular senescence and SASP.MethodsAutopsy lung tissue samples from 11 COVID-19 patients and 43 age-matched non-COVID-19 controls with similar comorbidities were analysed by immunohistochemistry for SARS-CoV-2, markers of senescence and key SASP cytokines. Virally induced senescence was functionally recapitulated in vitro, by infecting epithelial Vero-E6 cells and a three-dimensional alveosphere system of alveolar type 2 (AT2) cells with SARS-CoV-2 strains isolated from COVID-19 patients.ResultsSARS-CoV-2 was detected by immunocytochemistry and electron microscopy predominantly in AT2 cells. Infected AT2 cells expressed angiotensin-converting enzyme 2 and exhibited increased senescence (p16INK4A and SenTraGor positivity) and interleukin (IL)-1β and IL-6 expression. In vitro, infection of Vero-E6 cells with SARS-CoV-2 induced senescence (SenTraGor), DNA damage (γ-H2AX) and increased cytokine (IL-1β, IL-6, CXCL8) and apolipoprotein B mRNA-editing (APOBEC) enzyme expression. Next-generation sequencing analysis of progenies obtained from infected/senescent Vero-E6 cells demonstrated APOBEC-mediated SARS-CoV-2 mutations. Dissemination of the SARS-CoV-2-infection and senescence was confirmed in extrapulmonary sites (kidney and liver) of a COVID-19 patient.ConclusionsWe demonstrate that in severe COVID-19, AT2 cells infected by SARS-CoV-2 exhibit senescence and a proinflammatory phenotype. In vitro, SARS-CoV-2 infection induces senescence and inflammation. Importantly, infected senescent cells may act as a source of SARS-CoV-2 mutagenesis mediated by APOBEC enzymes. Therefore, SARS-CoV-2-induced senescence may be an important molecular mechanism of severe COVID-19, disease persistence and mutagenesis.

Highlights

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) that primarily affects the respiratory system
Electron microscopy analysis in representative COVID-19 cases confirmed the presence of virus within alveolar type 2 (AT2) cells (Figure 1C) and high magnification revealed virions in the proximity of the endoplasmic reticulum, indicating their likely assembly and budding, as well as virions residing in cytoplasmic vesicles, implying their transfer and release into the extracellular space
We have demonstrated the presence of SARS-CoV-2 in AT2 cells of patients who died from COVID-19 using our novel and a commercial anti-SARS-CoV-2 antibody against its spike protein and by electron microscopy

Summary

Introduction

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus disease 2019 (COVID-19) that primarily affects the respiratory system. Viral infection results in the activation of complex innate and adaptive immune responses that are orchestrated sequentially, involving several cell types and inflammatory mediators [5, 6]. We investigated whether COVID-19 disease is associated with cellular senescence and SASP. Virally-induced senescence was functionally recapitulated in vitro, by infecting epithelial Vero-E6 cells and a three-dimensional alveosphere system of alveolar type 2 (AT2) cells with SARSCoV-2 strains isolated from COVID-19 patients. Infection of Vero-E6 cells with SARSCoV-2 induced senescence (SenTraGorTM), DNA damage (γ-H2AX) and increased cytokine (IL-1β, IL-6, CXCL8) and Apolipoprotein B mRNA-editing (APOBEC) enzyme expression. Conclusions: We demonstrate that in severe COVID-19, AT2 cells infected by SARS-CoV-2 exhibit senescence and a proinflammatory phenotype. SARS-CoV2-induced senescence may be an important molecular mechanism of severe COVID-19, disease persistence and mutagenesis

Methods

Results

Conclusion