Pf4+-macrophages are the major resource to generate second wave of pro-inflammatory factors at late stage of IAV-driven pneumonia. a Heat-maps showing the normalized expression (Z-score) of the pro-inflammatory genes in the various cells of clusters with high ratios at day 7 p.i.. b The normalized expression (UMI counts) of some significant genes with high expression in different clusters at day 7 p.i.. c Pf4+ cells in the lung after IAV infection. The tdtomato-Pf4 mice were uninfected or infected with 0.5 LD50 of influenza A/PR/8/34 (H1N1) viruses. At day 7 p.i, the tdtomato-Pf4+ cells in the lung were scanned. The nucleus was stained with DAPI (blue) and tomato-Pf4 (red) was shown in red. Scale bars, 500 μm. d At day 0 and day 7 p.i., the tdtomato-Pf4+ cells in the lung were calculated. At least 300 cells in each group from three independent assays were scored. Data are shown as the mean ± SD. **, P < 0.01 (Student t test, n = 3). e The expression of Ccl2 in Pf4+ cells. The tdTomato-Pf4 mice were infected with 0.5 LD50 of influenza A/PR/8/34 (H1N1) viruses. At day 7 p.i, the tdTomato-Pf4+ cells in the lung were stained with anti-Ccl2 antibodies (green). The nucleus was stained with DAPI (blue) and Pf4-tdTomato (red) was shown in red. Scale bars, left, 200 μm; right, 50 μm. f Heatmap showing the scaled distances calculated based on Pearson correlations for relationships between the z-score normalized mean expression profiles in all the indicated cells. g PCA plot showing the relationships among the indicated cell clusters and the megakarycytes from lung and bone mellow described by others (SRP097794, NCBI)