Insights from dietary supplementation with zinc and strontium on the skeleton of zebrafish, Danio rerio (Hamilton, 1822) larvae: From morphological analysis to osteogenic markers

In order to obtain additional insights into the zinc (Zn) and strontium (Sr) effects on bone formation, the zebrafish (Danio rerio) was used as a recognized model for vertebrate skeletogenesis. The treatments began 5 days post-fertilization, using triplicates for each test condition, with 100 larvaeper replicate for the strontium trials and 80 larvae per replicate for the zinc trials. Tanks were maintained at 28 degrees C with a temperature and photoperiod control (recirculation system ZebTECH, Tecniplast). Diets for the zebrafish larvae undergoing vertebrae formation and mineralization (from 5 to 30 dpf) were supplemented with increasing concentrations of either zinc sulfate or strontium citrate, as sources of Zn or Sr, respectively, and their effects on the fish skeleton were evaluated at both histomorphological and molecular levels. Whereas high amounts of Zn (240 and 480 mg/kg) increased larvae mortality, intermediate amounts (60 to 120 mg/kg) gave similar results as the control diet regarding deformities, suggesting that these amounts are not detrimental to development of the larvae skeleton. In addition, osteogenic markers runx2, sp7 and oc were increased by Zn supplementation, indicating that appropriate amounts of Zn seem to support bone formation in zebrafish larvae. In Sr experiments, two Sr concentrations and two bio-encapsulation materials were evaluated: chitosan and ?-carrageenan. Interestingly, Sr bio-encapsulation with chitosan decreased the number of deformed larvae and the number of deformities per larvae at 2500 mg/kg, while kappa-carrageenan encapsulation failed to produce any positive effects. Furthermore, the chitosan experimental groups presented an increased expression of the osteoblast differentiation markers, sp7 and oc, suggesting that Sr promotes osteoblast differentiation and bone formation in zebrafish larvae.