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Fig. 2
Macrophages are required for fin regeneration and blastemal cell proliferation in a stage-dependent manner. (a) Schedule of macrophage (Mφ) depletion using L-clodronate injections. Macrophages were ablated using early L-clodronate injections (L-clodronate 1; no macrophages at the wound) or using injection of L-clodronate at later stages (L-clodronate 2; no more recruited M2-like macrophages from 24 hpA). (b and e) Consequences of macrophage depletion on caudal fin regeneration. (b) To deplete all macrophages from the early stage of regeneration, Tg(mpeg1:mCherry-F) were injected with L-clodronate (or L-clo) or L-PBS (control) at 48 hpf and fin were transected at 72 hpf (L-clodronate 1). (e) To deplete late-recruited macrophages, Tg(mpeg1:mCherry-F) were injected with L-clodronate or L-PBS at 6 hpA (L-clodronate 2). Fin images are representative overlays of mCherry fluorescence and transmitted light acquisitions at 3 dpA. Scale bar=100 μm. Dotted lines outline the fin and dashed arrows, the position of the initial amputation. (c and f) Corresponding quantification of the regenerated fin length at 3 dpA after L-clodronate 1 treatment (c) and L-clodronate 2 treatment (f) in indicated conditions (mean±S.E.M., **P<0.01 and ***P<0.001). (d–g) Blastema cell proliferation at 24 hpA after L-clodronate 1 (d) and 2 (g) treatments in indicated conditions. Mitotic cells were detected using an anti-phosphorylated histone H3 (PH3) antibody (Nlarvae=10–15, average value of cut/uncut ratio±S.E.M, *P<0.05)