Kinetics of inflammatory macrophage migration to LNs. (A) FACS plots depicting the appearance of inflammatory macrophages (F4/80⁺CD36⁺ or F4/80⁺ MHC-II⁺) in the mediastinal LN over the course of 8 days after i.p. administration of thioglycollate. (B) Total LN cells (Left) or total cells with F4/80⁺ CD36⁺ cells gated out (Right) based on cell surface expression of CD11c and MHC-II. Bottom plot overlays F4/80⁺ CD36⁺ gated cells on total LN cells, plotted to show CD11c vs MHC-II. (C) Labeling of peritoneal inflammatory macrophages 24 hours after i.p. injection of the phagocytic tracer dye PKH26 in mice inflamed 2 days earlier by thioglycollate injection. (D) PKH26⁺ cells were identified in the mediastinal LN 3 days after injection of PKH26 in the peritoneal cavity (5 days after inflammation was induced by thioglycollate) and analyzed for F4/80 and CD36 cell surface expression. Far-right plot shows PKH26 levels after gating on all F4/80⁺ CD36⁺ LN cells. (E) Inflammatory macrophages were retrieved from CD45.1 mice that had been injected 1 day earlier with thioglycollate. These macrophages were adoptively transferred into CD45.2 mice at the same stage of inflammation. CD45.1⁺ inflammatory macrophages were gated in the mediastinal LN cell suspension (left FACS plot) 4 days later and analyzed for CD36 expression. (F-H) Inflammatory macrophages in the peritoneum, draining LN and omentum at different times after induction of peritonitis by thioglycollate (n = 4-6 mice per group per time point). Data are representative of at least 2 independent experiments.