Fig. 3

Macrophages produce ETs during injury response in the retina. a Transmission electron microscopy of ET formation showing nuclear envelope alterations in phagocytic cells on day 1. Close-up of separation of the inner nuclear membrane (INM) from the outer nuclear membrane (ONM) and small DNA strands within the lumen between the INM and ONM. b Images show a phagocytic cell with DNA containing vesicles in the cytoplasm on day 4. Enlarged view of vesicle containing DNA strands. c Transmission electron microscopy of phagocytic cells releasing ETs on day 7. Close-up of dense DNA filaments released into the extracellular space. d Heatmaps of differentially expressed NETs-related genes in Csfr1^GFP cells. Genes were selected from KEGG pathways (mmu04613). Data are expressed as fold-changes between different time points (days 1, 3, and 7) compared to negative controls (Csfr1^EGFP cells from uninjured retinas). e Representative in vivo images of macrophages of LysM^GFP mice (left) or Cx3cr1⁺ cells (green, right) releasing extracellular DNA (red) on day 4. Extracellular DNA was labeled with intravenous injection of SYTOX Red. Arrows indicate extracellular DNA fibers, and stars are placed on the cell bodies. f Percentage of Cx3cr1⁺/SYTOX⁺ and LysM⁺/SYTOX⁺ per lesion 4 days after injury. Significant differences (****p < 0.0001) between baseline and the different time points were determined by using a two-tailed Mann–Whitney test analysis (n = 8). Field of view is approximately ≈ 575 µm