Fig. 2: Alv. and Int. MKs engulf particles.

a Schematic showing the interactions between fluorescently labeled Cy5-PSs (red) and Alv. or Int. MKs (green). b Imaging using two-photon intravital microscopy (2PIVM) shows lung MKs (green) migrating to engulf Cy5-PSs (pink) in PF4-mTmG mice treated with 2 mg/kg body weight Cy5-PSs through o.p.a. for 2 h. c Fluorescent images of Alv. MKs (green) crawling towards and engulfing Cy5-PSs (yellow arrows) deposited in the alveoli. Nuclei are stained with DAPI (blue). d Intravital two-photon imaging of lung MK (green) squeezing into and out of 2 bordering alveoli possibly through the pore of Kohn (white arrow). e Flow cytometry images show that CD41⁺ MKs (yellow) from the lungs of mice treated with Cy5-PSs for 12 h contain Cy5-PSs (red). Nuclei are stained with Hoechst (blue). f Fluorescent images showing engulfed Cy5-PSs (orange) inside CD41⁺ MKs sorted from mice and Dami cells treated with 200 ng/mL Cy5-PSs for 12 h. White dotted line outlines a cell. g TEM image of lung MKs treated with Cy5-PSs for 12 h. Red arrow in the insert image indicates engulfed particles. h Flow cytometer analysis of engulfed particles in lung MKs of mice treated with Cy5-PSs after 6 and 12 h (n = 10 mice per group). N.D. not detected. i Quantification of internalized Cy5-PSs in Alv. MKs, Int. MKs and AMs from mice in h after 6 h (n = 10 mice per group). Data are presented as the mean values of 3 independent experiments ± SEM (h, i), one-way analysis of variance (ANOVA). j, k RNA-seq analyses on Int. MKs (j) and Alv. MKs (k) show significant mRNA enrichment in thrombopoiesis and immune regulatory functions, respectively. The statistical analyses were performed using Metascape, one-side Fisher’s exact test (j, k). Figure 2a, created with BioRender.com, released under a Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International license.