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Morphological abnormalities of pancreas and intestine in wrn-/- larvae (A–K) Morphology of pancreatic exocrine cell in Tg(ptf1a:EGFP); casper larvae. (A) The wrn+/+ (n = 24/24) and (B) wrn-/- (n = 6/6) larvae at 5 dpf. (C) The wrn+/+ (n = 27/27), (D) wrn-/-_mild (n = 8/13), and (E) wrn-/-_severe (n = 5/13) larvae at 7 dpf. (F) The wrn+/+ (n = 47/47), (G) wrn-/-_mild (n = 8/19), and (H) wrn-/-_severe (n = 11/19) larvae at 9 dpf. (I) The wrn+/+ (n = 40/43), (J) wrn-/-_mild (n = 8/15), and (K) wrn-/-_severe (n = 7/15) larvae at 12 dpf. Scale bars, 0.5 mm. (L–N) HE staining of a paraffin section. (L) The wrn+/+ (n = 6), (M) wrn-/-_mild (n = 6), and (N) wrn-/-_severe (n = 6) larvae at 9 dpf. Dashed green boxes indicate the pancreatic exocrine region. Black and white arrowheads indicate the liver and intestine, respectively. Scale bars, 50 μm. (O) Area of GFP-positive pancreatic exocrine tissue at 5, 7, 9, and 12 dpf (corresponding to Figures 3A–3K). The number of samples. wrn+/+ (n = 24): wrn-/- (n = 6) at 5 dpf; wrn+/+ (n = 27): wrn-/-_mild (n = 8): wrn-/-_severe (n = 5) at 7 dpf; wrn+/+ (n = 47): wrn-/-_mild (n = 8): wrn-/-_severe (n = 11) at 9 dpf; wrn+/+ (n = 43): wrn-/-_mild (n = 8): wrn-/-_severe (n = 7) at 12 dpf. Statistical significance at 5 dpf was assessed using an unpaired two-tailed Student’s t test, and that at 7, 9, and 12 dpf was assessed using one-way ANOVA followed by Tukey’s honest significant difference (HSD) test. Data are presented as mean ± SD. The detailed mean ± SD are shown in Table S6. (P and Q) Quantification of average intestinal villi length (P) and intestine area (Q) of wrn+/+ (n = 6), wrn-/-_mild (n = 6), and wrn-/-_severe (n = 6) at 9 dpf (corresponding to Figures 3L–3N). Statistical significance was assessed using one-way ANOVA followed by Tukey’s HSD test. Data are presented as mean ± SD. The detailed mean ± SD are shown in Table S7. (R and S) In situ hybridization of wrn expression in wild-type sibling (WT sib) zebrafish at 5 dpf (R) and 7 dpf (S). Red, white, and green arrowheads indicate the pancreas, intestine, and liver, respectively. Scale bars, 100 μm.
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