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Prematurely dying wrn-/- zebrafish become malnourished (A and B) (A) Glucose and (B) glycogen levels in whole larvae at 9 dpf. The number of samples. wrn+/+ (n = 12); wrn-/-_mild (n = 10); wrn-/-_severe (n = 13). Statistical significance was assessed using Welch’s ANOVA followed by Games-Howell post-hoc test. Data are presented as mean ± SD. The detailed mean ± SD are shown in Table S10. (C–E) Oil Red O staining of whole-mount larvae. (C) The wrn+/+ (n = 21), (D) wrn-/-_mild (n = 20), and (E) wrn-/-_severe (n = 11) larvae at 9 dpf. Scale bars, 0.5 mm. (F) Quantification of Oil Red O signal intensity (corresponding to Figures 5C–5E). The number of samples. wrn+/+ (n = 21): wrn-/-_mild (n = 20): wrn-/-_severe (n = 11). Statistical significance was assessed using Welch’s ANOVA followed by Games-Howell post-hoc test. Data are presented as mean ± SD. The detailed mean ± SD are shown in Table S11. (G) Survival assay under normal, starved, and overfed conditions. The wrn+/+_starved (purple, n = 56) and wrn-/-_starved (orange, n = 54); the wrn+/+_overfed (blue, n = 78) and wrn-/-_overfed (green, n = 68). The wrn+/+ (black, n = 214) and wrn-/- (red, n = 133) under normal conditions (data reused from Figure 1B). Statistical significance was assessed using the log-rank test. (H–M) Morphology of pancreatic exocrine cell in Tg(ptf1a:EGFP); casper larvae at 9 dpf. (H) The wrn+/+ (n = 47/47), (I) wrn-/-_mild (n = 8/19), and (J) wrn-/-_severe (n = 11/19) larvae under normal feed conditions (data reused from Figures 3F–3H). (K) The wrn+/+ (n = 15/15), (L) wrn-/-_mild (n = 16/33), and (M) wrn-/-_severe (n = 17/33) larvae under overfeed condition. Scale bars, 0.5 mm. (N) Area of GFP-positive pancreatic exocrine tissue at 9 dpf (corresponding to Figures 5H–5M). The number of samples. wrn+/+ (n = 47): wrn-/-_mild (n = 8): wrn-/-_severe (n = 11) under normal feed condition; wrn+/+ (n = 15): wrn-/-_mild (n = 16): wrn-/-_severe (n = 17) under overfeed condition. Statistical significance was assessed using an unpaired two-tailed Student’s t test for comparisons between conditions (e.g., wrn+/+ under normal conditions vs. wrn+/+ under overfeed condition), and one-way ANOVA followed by Tukey’s HSD test for comparisons under overfeed condition (wrn+/+, wrn-/-_mild, and wrn-/-_severe). Data are presented as mean ± SD. The detailed mean ± SD are shown in Table S6.
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