Figure 6

Treatment with rHSP47 ameliorates zebrafish p3h1^–/– larvae bone mineralization.

(A) Upon mechanical chorion removal, embryos from prolyl 3-hydroxylase 1–knockout (p3h1^–/–) zebrafish were treated every other day with a 4-hour pulse of 0.5 μM rHSP47 or with placebo. At 11 dpf, the skeletons of larvae were stained with alizarin red in order to assess the rHSP47 effect on mineralization. (B) Analysis of the fish morphology was carried out measuring the snout-operculum length, the height at eye, and the snout-operculum length/height-at-eye ratio (n ≥ 40). (C) Lateral and ventral images of p3h^–/– larvae, treated with rHSP47 or placebo, upon mineral staining by alizarin red. (D–F) Bone mineralization level analyzed in rHSP47-treated and untreated fish. Three classes of mineralization were defined, namely, no mineralization, incomplete, and complete, based on the level of alizarin red staining. Biological replicates (n ≥ 41) were performed. (G) Total larvae mineral content was extracted and quantified by spectrophotometric analyses. Biological replicates (n = 32) were performed. Error bars indicate SD. Mann-Whitney Wilcoxon test was applied. Statistical analyses details are reported in Supplemental Table 2. *P ≤ 0.05.