The nature of infected cells was characterized in a reconstructed human bronchial epithelium model with cells placed at the air/liquid interface differentiated into a pseudostratified epithelium. Immunofluorescence confocal imaging revealed orthogonal sections of ciliated cells labeled for β-tubulin IV (red), basal cells labeled for cytokeratin-5 (yellow), nuclei labeled for DNA (Hoechst, blue), and infected cells labeled for the SARS-CoV-2 spike (green). The green line corresponds to the autofluorescence of the insert membrane. Four days after infection, the spike+ cells showed weaker or absent β-tubulin IV staining, indicating a loss of motile cilia. SARS-CoV-2 particles were not released directly from cilia, but rather from de-ciliated areas close to the plasma membrane. Interestingly, some spike-negative basal cells appeared to rise through the pseudostratified epithelium in infected samples suggesting an epithelial response to virally induced damage. Taken together, the study showed that ciliated epithelial cells were the main target of SARS-CoV-2. Infection had several consequences, including a temporary decrease in epithelial barrier function with disruption of tight junctions, a loss of the ciliary layer associated with damaged ciliated cells, and a temporary increase in apoptotic cells. The transcription factor Foxj1, a master regulator of ciliogenesis, was downregulated prior to extensive cilia loss. A mucociliary clearance assay showed that the motile function of cilia was compromised. Furthermore, epithelial defense mechanisms, like interferon production, ramped up only after the initiation of cilia damage. The loss of motile cilia also took place in vivo, as shown in the Syrian hamster model of SARS-CoV-2 infection.
In conclusion, a decrease in cilia movement could slow the clearance of viral particles and facilitate their dissemination to deeper regions of the airways. This process could self-perpetuate, with cycles of localized cilia destruction facilitating SARS-CoV-2 progression towards increasingly more distal regions, until the virus reaches the alveoli and triggers pneumocyte damage. The impairment of mucociliary clearance may also facilitate the spread of other respiratory pathogens and increase the risk of secondary infections in COVID-19 patients.
Robinot, R. et al.:
SARS-CoV-2 infection induces the dedifferentiation of multiciliate cells and impairs mucociliary clearance
Nature Communications (2021) vol. 12, article 4354.
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