Damage induces cell proliferation of OSN and of neutrophil precursors in the adult olfactory organ. (A–C) BrdU labeled cells (white), neutrophils (green) in whole mount OO of adult fish. Scale bars A-C = 50 μm (A). Prior to copper exposure BrdU labeling and scattered neutrophils were observed in the medial raphe (mr), sensory (ss), and non-sensory (ns) epithelia. n= 3 OE. (B) After four hours of exposure to copper, intense BrdU labeling was observed in the mr. n= 3 OE. (C) One day post recovery, neutrophils lined the lamellae and intense BrdU labeling was observed in ss and LOE-EN. n= 3 OE. (D) Section of Tg(fli1a:EGFP) adult OE showing extensions of blood vasculature (green) within the OE. Scale bar = 100 μm. n= 3 sectioned heads. (E) Significant increases in neutrophil number were observed after 4 hour copper exposure (red) in both the ns and ss epithelia when compared to control (grey). At one dpt (green) only the number of labeled ss cells remained significantly greater than controls. n= 3 OE. (F) Damage induced changes in the number of BrdU-positive cells were significant in the ss epithelia but not the ns at 4 hour copper exposure (red) and one dpt (green). n= 3 OE. (G) A small but significant increase in mpx:GFP+ cells double labeled for BrdU scored in the OE. (E-G, n=3 adult OE from different fish; Two-way ANOVA, Tukey multiple comparison test, p < 0.05). (*P, 0.05, **P, 0.01, ***P, 0.001). n= 3 OE. (H) Four hours of copper exposure (orange) and one day post-treatment (green), an increase in rounded (nt1; circularity index 0.7 or greater see Figure 5) and amoeboid (nt2; circularity index 0.4-0.6) neutrophils were observed. Two-way ANOVA, Tukey’s multiple comparison test, p < 0.05). (*P, 0.05, **P, 0.01, ***P, 0.001, ****P, 0.0001, ns: non significant).
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