Wnt inhibition blocks new nephron formation. Gentamicin-injured adult zebrafish were treated with either DMSO or 5 µM of the Wnt inhibitors IWR1 or IWP2 in system water starting at 1 dpi. Whole-mount in situ hybridization showing the trunk kidney region at 7 dpi. (A-C) Control-injected and DMSO-treated kidneys do not express markers of new nephrons. (D-F) Injury induces cell aggregates expressing lhx1a, fzd9b and lef1. Nephron aggregate formation is blocked by Wnt inhibition using IWR1 (G-I) or IWP2 (J-L). Scale bars: 0.2 mm. (M-P) Quantification of Wnt inhibitor effects on lhx1a+ aggregates and gene expression. (M) Percentage of lhx1a+ aggregates/mm2 of kidney calculated using ImageJ. n=3-6 fish for each condition, as indicated by individual graph symbols. (N-P) qPCR quantification of lhx1a, fzd9b and lef1 mRNA in kidney tissue harvested 7 d after gentamicin injury, n=6 for each condition. *P<0.05 calculated using Student's unpaired two-tailed t-test. Data are mean±s.d.
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