Catalase expression in beta-cells leads to a reduction in beta-cell numbers in larvae. (A) Whole mount in situ hybridization (WISH) for catalase in WT and Tg(ins:catalase;ins:H2BmCherry) larvae at 5 dpf. Arrows point to the principal islet. Catalase expression was not detected in the beta-cells of WT larvae whereas Tg(ins:catalase;ins:H2BmCherry) larvae exhibit catalase transcripts. (B) Double WISH for insulin (brown) and catalase (purple) showing overlap between the insulin and catalase transcripts in Tg(ins:catalase;ins:H2BmCherry) siblings. (C) Confocal projections of the principal islets of 4 dpf WT and Tg(ins:catalase;ins:H2BmCherry) larvae. (D) Quantification of the average number of beta-cell in WT (n = 15 larvae) and Tg(ins:catalase;ins:H2BmCherry) larvae (n = 12 larvae). Tg(ins:catalase;ins:H2BmCherry) larvae exhibit a reduced beta-cell number compared to WT (p = 0,008, error bars = SEM).

Different levels of H₂O₂ induce beta-cell proliferation and quiescence. (A) Quantification of the number of Tg(ins:FUCCI-S/G2/M)⁺ beta-cells after continuous treatment with increasing concentrations of H₂O₂ from 3 to 4 dpf. The treatment with 50 μM H₂O₂ (n = 89 larvae) increased the number of Tg(ins:FUCCI-S/G2/M)⁺ beta-cells compared to controls (n = 104 larvae) (p = 0.025) whereas the treatments with 500 and 1000 μM H₂O₂ decreased the number of Tg(ins:FUCCI-S/G2/M)⁺ beta-cells (n = 38 larvae for 500 H₂O₂ μM; n = 63 larvae for controls; p = 0.03) (n = 46 larvae for 1000 μM H₂O₂; n = 78 larvae for controls; p = 0.0004). (B) Quantification of the number of Tg(ins:FUCCI-S/G2/M)⁺ beta-cells in 3 dpf larvae treated with increasing concentrations of H₂O₂ for 1.5 hours followed by a wash-out. The number of Tg(ins:FUCCI-S/G2/M)⁺ beta-cells was examined 24 hours after the initiation of the treatment. The short treatment with 200 and 500 μM H₂O₂ increased the number of Tg(ins:FUCCI-S/G2/M)⁺ beta-cells compared to controls (n = 51 larvae for 200 μM H₂O₂; n = 86 larvae for controls; p = 0.02) (n = 68 larvae for 500 μM H₂O₂; n = 96 for larvae for controls; p = 0.02). (C) Confocal projections of the principal islets of 4 dpf Tg(ins:H2B-GFP; ins:dsRED) larvae treated from 3 to 4 dpf with 50 μM H₂O₂. (D) Quantification of the average number of beta-cells marked with H2B-GFP in controls and H₂O₂-treated larvae. H₂O₂-treatement increased beta-cell number (n = 30 larvae) compared to controls (n = 26 larvae) (p = 0,046). Error bars = SEM. (E) Bar chart showing the increase in the number of INS-1 cells after H₂O₂ or NAC-treatments. Data from four independent experiments and > 3.5 × 10⁵ cells per condition were normalized to the mean of control treated cells ( = 100%) and analyzed for significance using a one-way ANOVA with Dunnett’s multiple comparison test. Error bars indicate S.E.M. Cartoons with permission by Luis Delgadillo.

High levels of H₂O₂ promote new beta-cell neogenesis and regeneration. (A) Quantification of the average number of beta-cells marked with H2B-GFP in controls (n = 22) and larvae treated with 1 mM H₂O₂ from 3 to 4 dpf (n = 23). The larvae treated with 1 mM H₂O₂ exhibit an increase in the number of beta-cells compared to controls (p = 0,04). (B) Confocal projections of the principal islet of 4 dpf WT and Tg(ins:H2B-GFP; ins:dsRED) larvae treated with vehicle or 1 mM H₂O₂ from 3 to 4 dpf. Arrowheads point to H2B-GFP⁺ dsRED⁻ cells. (C) Quantification of the number of H2B-GFP⁺ and dsRED⁻ cells in controls and H₂O₂-treated larvae. Beta-cells that are H2B-GFP⁺ but dsRED⁻ represent recently-formed cells due to the slower maturation of dsRED compared to GFP. H₂O₂-treatment increased the number H2B-GFP⁺ and dsRED⁻ beta-cells compared to controls (p = 0,004), indicating an increase in new beta-cell formation. Error bars = SEM. (D) Confocal sections of primary islets from Tg(Tp1:H2BmCherry) larvae stained for insulin (blue). Tg(Tp1:H2BmCherry) drives expression of a fluorescent protein with long half-life (H2BmCherry) in the Notch responsive cells (NRCs) in the pancreas. Beta-cells that differentiate from NRCs can retain H2BmCherry-flurescence due to perdurance. The larvae were treated with vehicle or 1 mM H₂O₂ from 3 to 4 dpf. The arrows indicate Tp1:H2BmCherry⁺ and insulin⁺ cells in the periphery of the primary islets in controls and H₂O₂-treated larvae. See Figure S6 for higher resolution images. (E) Quantification of the average number of Tp1:H2BmCherry⁺ and insulin⁺ cells in the principal islets of controls (n = 21) and H₂O₂-treated larvae (n = 19), showing an increase following the H₂O₂-treatement (p = 0,049). (F) Confocal sections of primary islets from Tg(Tp1:H2BmCherry); Tg(neurod1-GFP) larvae. The arrows indicate Tp1:H2BmCherry⁺ and GFP⁺ cells in the periphery of the primary islets in controls and H₂O₂-treated larvae. See Figure S7 for higher resolution images. (G) Quantification of the average number of Tp1:H2BmCherry⁺ and GFP⁺ cells in the principal islets of controls (n = 21) and H₂O₂-treated larvae (n = 19), showing an increase following the H₂O₂-treatement (p = 0,046). (H) Confocal projections of Tg(ins:FLAG-NTR); Tg(ins:H2B-GFP;ins:dsRED) larvae. Beta-cells were ablated by incubating larvae in MTZ at 3 dpf. Subsequently, the larvae were treated with vehicle or 1 mM H₂O₂. (I) Quantification of the average number of beta-cells in controls (n = 24) and H₂O₂-treated larvae (n = 22). H₂O₂-treatment significantly increased the number of regenerating beta-cells (p = 0,03). Note that a majority of the regenerated beta-cells are H2B-GFP⁺ and dsRED⁻. Error bars = SEM.

ZFIN is incorporating published figure images and captions as part of an ongoing project. Figures from some publications have not yet been curated, or are not available for display because of copyright restrictions.

ZFIN is incorporating published figure images and captions as part of an ongoing project. Figures from some publications have not yet been curated, or are not available for display because of copyright restrictions.