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Figure 1
(A) Blood glucose level (mg/ml) of adult Tg(ins:NTR-P2A-mCherry) control fish (CTL, 66 ± 15 mg/dl), 3 days (510 ± 126 mg/dl) and 20 days post treatment (dpt) (117 ± 29 mg/dl) with the NFP prodrug to trigger β-cell ablation. The huge rise of glycemia at three dpt confirms the efficiency of ablation. One-way ANOVA Kruskal-Wallis test (with Dunn’s multiple comparisons); Mean ± SD; **p < 0.005, ****p < 0.0001. (See Figure 1—source data 1). (B) Immunolabeling of β- and δ-cells with anti-INS (red) and anti-SST (green), respectively, on paraffin sections through the main islet of Tg(ins:NTR-P2A-mCherry) adult fish in control condition (CTL) and at 20 dpt. In CTL islet, no appreciable overlap between the two markers can be detected while broad colabeling is observed at 20 dpt and represented by many yellow cells (arrowheads). (C) Whole mount immunodetection of β- and sst1.1+ cells in the main islet of adult Tg(sst1.1:GFP);Tg(ins:NTR-P2A-mCherry) fish by labeling with anti-GFP marking sst1.1-expressing cells and anti-mCherry for β-cells. Both cell types show no or very few overlapping in CTL fish. At 3 and 20 dpt, many double GFP+ mCherry + cells are observed (yellow cells, arrowheads). Bright mCherry+ β-cell debris are detectable at 3 dpt (white asterisk). (D–G) Quantification of the GFP+, mCherry+ (β-cells) and double GFP+ mCherry + cells detected by FACS in the main islets of Tg(sst1.1:GFP);Tg(ins:NTR-P2A-mCherry) CTL fish and following β-cell ablation (3 and 20 dpt), based on fluorescence analysis shown in Figure 1—figure supplement 2. (D) Total islet cell number in CTL, 3 dpt, and 20 dpt islets. (E) CTL islets contain 3277 ± 1220 mCherry+ (GFP-) β-cells. At 3 dpt, ablated β-cells represent 105 ± 70 cells and were even more scarce at 20 dpt (14 cells). (F) Double GFP+ mCherry + bihormonal cells represent 135 ± 45 cells in CTL islets, 1411 ± 421 cells at 3 dpt and 1409 ± 655 cells at 20 dpt. (G) GFP+ (mCherry-) cells represent 2833 ± 615 cells in CTL islets. One-way ANOVA Kruskal-Wallis test (with Dunn’s multiple comparison); ns, not significant, *p < 0.05, **p < 0.005, ***p < 0.0005, ****p < 0.0001; Mean ± SD (See Figure 1—source data 2).
Most new ins + cells after ablation and regeneration in zebrafish are Ins + Sst1.1+ bihormonal cells.