Osteoclast-like Cells in Early Zebrafish Embryos

OBJECTIVE Genes involved in bone and tissue remodelling in the vertebrates include matrix metalloproteinase-9 (mmp-9), receptor activator of necrosis factor κ-β (rank), cathepsin-k (Ctsk) and tartrate-resistant acid phosphatase (TRAcP). We examine whether these markers are expressed in cells of zebrafish embryos of 1-5 days post fertilization. We also examine adult scales, which are known to contain mature osteoclasts, for comparison. MATERIALS AND METHODS In this experimental study, in situ hybrdisation, histochemistry and serial plastic and paraffin sectioning were used to analyse marker expression. RESULTS We found that mmp-9 mRNA, TRAcP enzyme and Ctsk YFP protein were expressed in haematopoietic tissues and in the cells scattered sparsely in the embryo. Ctsk and rank mRNA were both expressed in the branchial skeleton and in the developing pectoral fin. In these skeletal structures, histology showed that the expressing cells were located around the developing cartilage elements, in the parachondral tissue. In a transgenic zebrafish line with YFP coupled to Ctsk promoter, Ctsk expressing cells were found around pharyngeal skeletal elements. To see whether we could activate osteoclasts, we exposed prim-6 zebrafish embryos to a mixture of 1 µM dexamethasone and 1 µM vitaminutes D3. These compounds, which are known to trigger osteoclastogenensis in cell cultures, lead to an increase in intensity of Ctsk YFP expression around the skeletal elements. CONCLUSION Our findings show that cells expressing a range of osteoclast markers are present in early larvae and can be activated by the addition of osteoclastogenic compounds.