eGFP expression patterns in Tg(ola.sp7:nlsGFP) zebrafish. (A), At 1 d postfertilization (dpf), GFP is weakly expressed in the posterior trunk and tail regions. (B), Until 8 dpf, GFP‐labeled osteoblasts are detectable in ceratobranchial, opercle, branchiostegal rays, and mandible of the head region. (C), The rostral skeleton, axial skeleton, fin, and tail vertebral columns are clearly labeled by eGFP in 30 dpf juvenile fish. (D), Fluorescent fish scales are visible in 42 dpf transgenic fish. CT, cleithrum; BR, branchiostegal rays; MD, mandible

Comparison of GFP‐labeled cephalic bones in Tg(ola.sp7:nlsGFP) zebrafish to alizarin red‐stained cephalic bones at 8 dpf. (A), Alizarin red stains mineralized bones in zebrafish head region, including cleithrum, opercle, branchiostegal rays, mandible, and notochord. (B), 3D‐reconstructed confocal image of Tg(ola.sp7:nlsGFP) zebrafish larvae. In comparison with alizarin red‐stained fish, eGFP is not expressed in the notochord of transgenic fish. CT, cleithrum; BR, branchiostegal rays; MD, mandible; NC, notochord

Morphologies of zebrafish larvae after Dex induction or FE treatment. In blank and DMSO‐treated control groups, zebrafish display a straight fish body (A, B). In Dex‐induced OP models, the larvae show a bent body and a curve at the link point of the head and trunk region as depicted by the red dash lines. This impairment is exacerbated by the increase in the Dex concentration from 5 μM to 20 μM (C‐E). (F), 1 μg/mL FE treatment rescues the impaired morphology observed in OP‐like fish

Dex induces OP‐like symptom in both wild‐type and Tg(ola.sp7:nlsGFP) zebrafish. (A), Dex‐treated larvae were stained by alizarin red. The stained area show the level of bone mineralization which is decreased in a dose‐dependent manner of Dex concentration from 0 to 20 μM (D). (B), 3D‐reconstructed confocal images of Dex‐treated Tg(ola.sp7:nlsGFP) zebrafish. Bone mass and density are reflected by GFP area and integrated optical density, which also presented a dose‐dependent manner of Dex concentration from 0 to 20 μM (E, F). Images in white frames are 5× magnifications relative to the red insets, where osteoblasts are visible (white arrow). Data are presented as means ± SD; *P < .05; **P < .01; ***P < .001; n = 18 fish in each group; scale bar, 100 μm

FE rescues OP‐like symptom induced by Dex. (A), Alizarin red‐stained zebrafish head regions of FE‐treated larvae. 1 μg/mL FE‐added zebrafish totally recovers the stained signal strength in comparison with GIOP model (**P < .01) (D). (B), 3D‐reconstructed confocal images of FE‐treated Tg(ola.sp7:nlsGFP) zebrafish head regions. Bone mass and bone mineral density (BMD) evaluated by fluorescent area and IOD reflect a rescued effect on 1 μg/mL FE‐treated OP‐like zebrafish (**P < .01) (E, F). Data are presented as means ± SD; **P < .01; ***P < .001; n = 18 fish in each group; scale bar, 100 μm