Fig. 3. SARS-CoV-2 provoked a senescence-like phenotype in hBOs and patients with post-acute COVID-19.
a, hBOs infected with SARS-CoV-2 at m.o.i. 0.1 were cultured for 6 days and then subjected to immunofluorescence staining for SARS-CoV-2 N protein (CoV2-NP (red), p16INK4a (green) and DAPI (blue)). Representative images of days 1, 3 and 6 are shown. The histograms indicate the percentages of cells expressing COV2-NP (top) or p16INK4a (bottom). b,c, hBOs infected with SARS-CoV-2 at m.o.i. 0.1 were treated with a TNF-α inhibitor (TNFi) (500 μg ml−1 etanercept) from day 3 after infection. Immunofluorescence staining for p16INK4a (green) and DAPI (blue) (b) or phospho-p38 (red) and DAPI [blue] (c) is shown. The histograms indicate the intensity of p16INK4a signals or phospho-p38 signals normalized by DAPI count. d, Single-cell RNA transcriptomic analysis of lung tissue from patients with severe COVID-19 (ref. 28). Epithelial cells population of uniform manifold approximation and projection (UMAP) plots were divided into healthy controls (25 and 52 years old) and patients with COVID-19 (28, 54 and 57 years old), and the distribution of cells expressing senescent cell marker (CDKN2A) and SASP factor genes is shown. Arrowheads indicate a basal cell population. For all graphs, error bars indicate mean ± s.d. of biological triplicate measurements. Scale bars, 50 μm (a–c). Statistical significance was determined by two-way ANOVA followed by Sidak’s multiple comparison test (a) or one-way ANOVA followed by Dunnett’s multiple comparison test (b,c). a.u., arbitrary units.