Different Capacity of Monocyte Subsets to Phagocytose Iron-Oxide Nanoparticles
Figure 1
Monocyte subsets display different capacity of phagocytosis, which results in divergent uptake of iron-oxide nanoparticles.
A) Flow cytometry dot plots of freshly isolated leucocytes from healthy donors. Monocytes are identified upon their FSC/SSC profile. Monocytes subsets are identified by their CD14/CD16 expression profile. H&E stains show monocyte morphology. B) Phagocytosis assay: flow cytometry histograms of both monocyte subsets after incubation with FITC-labeled latex beads for 2 h. Bar graph compares mean fluorescence intensity of internalized FITC-beads between both subsets, p-value shows significant difference (student's t-test). C) Bar graph depicts percentage of positively labeled alive monocyte subsets after 2h incubation with increasing concentrations of fluorescently labeled iron-oxide nanoparticles (CLIO-680), assessed by multi-color flow cytometry. D) Representative histogram shows fluorescence intensity of both monocyte subsets after labeling for 2 h with CLIO-680 at 100 µg Fe/ml. E) Quantitative immunofluorescence microscopy of FACS sorted monocytes after labeling with fluorescent iron-oxide nanoparticles (CLIO-680). Green: Clathrin, Red: CLIO-680, Blue: DAPI/nuclear staining. For co-localization analysis (upper right panel) images were taken under reproducible conditions and no additional enhancement was performed for the red and green channels. Fiducials indicate the red-green signal intensities used for determination of co-localization. White-circled areas indicate co-localization of Clathrin and CLIO-680 signal. Bar graph compares mean grey values of intracellular CLIO-680 signal between both subsets (from n = 3 different experiments with >10 of each subset analyzed per experiment).