Neutrophil and macrophage populations change in the kidney field after AKI. (A-D) Tg(cdh17:mCherry); Tg(lyz:EGFP) transgenic zebrafish were used for neutrophil analyses. (E-H) Tg(cdh17:mCherry); Tg(mpeg1;dendra2) transgenic zebrafish were used for macrophage analyses. (A,E) Transgenic lines were injected with gentamicin at 3 dpf and imaged at 2 dpi. (B,F) Snapshots of live imaging of lyz+ neutrophils imaged at 1 dpi for 13.5 h (B) and mpeg1+ macrophages imaged at 2 dpi for 19.5 h (F) in a no-injury and gent-AKI setting. Arrows indicate neutrophils or macrophages adjacent to PTs. (C,G) Immunofluorescence co-stain of PTs (red or green) and neutrophils (green) (C) or macrophages (red) (G) in no-injury and gent-AKI at 3 dpi. PTs are outlined in white and adjacent leukocytes are marked with arrows. (D,H) Quantification of neutrophil (D) and macrophage (H) numbers adjacent to the PT before and after injury. MeanNoInjury5dpf=4.24 (N=34) vs Mean2dpi=7.76 (N=34) vs MeanNoInjury6dpf=5.15 (N=34) vs Mean3dpi=11.60 (N=48). Adjacent leukocytes were counted for both no injury and gent-AKI at 1, 2 and 3 dpi (D), and 2 and 3 dpi (H). Data pooled from three biological replicates are shown expressed as mean±s.e.m. One-way ANOVA: *P<0.05, **P<0.01, ***P<0.005. Scale bars: 20 μm.