Fig. 1: Lung MKs accumulate more rapidly in the lung than other immune cells upon particle exposure.

a Number of AM, DC, NE, MO, MK, Baso and Eosin in the BALF increased at 6, 12, and 24 h hpi of CB particles (n = 4 mice per group). b Ratio of the cumulative increase of different cells (1-7) in BALF of treated mice in a relative to untreated control. Horizontal dotted line shows a ratio of 1. Ratio in each group is the mean values of 3 independent experiments. c Immunofluorescent images (left) and PF4⁺CD41⁺DAPI⁺ cell counts (right) show lung sections from mice post 24 h-exposure had more PF4⁺CD41⁺DAPI⁺ MKs (white arrow) than untreated control (n = 6 mice per group). d Fluorescent images of lung sections from naïve PF4-mTmG mice showing resident and crawling MKs (green) in the Alv. and Int. region (red, inside dotted line). e 3D fluorescent imaging of lung sections from untreated PF4-mTmG mice showing MK resident in Alv. wall. f Giemsa-Wright stained image of an isolated CD41⁺ Alv. MKs with the classical hyperchromatic nucleus and granular cytoplasm. g Fluorescent images of Alv. MKs (CD41⁺CD42b⁺ Hoechst⁺) obtained from BALF through magnetic bead sorting. h Fluorescence analysis of the percentage of Alv. MKs and Int. MKs in the whole lung from untreated PF4-mTmG mice (n = 3 mice per group). i Venn diagram showing the gene signature of Alv. and Int. MKs. Numbers represent number of genes. j Heatmap of the enriched pathways for genes expressed in Alv. and Int. MKs. k 3D fluorescent images showing accumulated PF4⁺GFP⁺MKs (green) and platelets (green) in lung sections from PF4-mTmG mice treated with CB particles for 24 h. l H&E and Masson stained images of lung sections from mice exposed to CB particles for 24 h show infiltration of inflammatory cells (blue arrows) and fiber deposition in alveoli (red arrows) and capillaries (green arrowheads). Data are presented as the mean values of 3 independent experiments ± SEM (a, c and h), P-value relative to 0 hpi (a) and control (c and h) by two-side student’s t-test.