By means of a multidisciplinary range of scientific and technical expertise that covers fish nutrition and metabolism, aquafeed technology, in vitro cell culture, biochemical and histochemical bone characterization and molecular expression of skeletal genes, the FATTYBONE project generated new knowledge on the mechanisms underlying the nutritional regulation of bone remodelling in juvenile gilthead seabream, the major farmed species in the Mediterranean region.

SCIENTIFIC HIGHLIGHTS
Polyunsaturated fatty acids (i.e. AA, EPA and DHA) are effectors of fish bone cell lines, altering cell morphology, proliferation, mineralization and gene expression patterns when added to culture medium. PUFA effects were molecule‐specific (DHA and AA/EPA have opposite effects on osteoblast mineralization), process‐specific (additive effect on osteoblasts proliferation and compensatory effect during mineralization) and in some cases, target gene‐specific (apparent opposite regulation of TNAP, BMP2 and COX2 gene expression by AA/EPA and DHA). We also demonstrated the suitability of our in vitro cell systems to get insights into mineralization‐related effects of PUFAs in vivo and eventually to evaluate the effects in bone of the replacement of fish oils by vegetable oil sources in fish feeds. Some of the results support previous findings in established mammalian cell systems, some are contradictory; cellular response to fatty acids may depend on the maturation of the cell but also on the species at the origin of the cell lines. The differential effect of bovine and fish sera on cell growth and ECM mineralization clearly indicates the presence in the serum of proliferative and mineralogenic factors, which are logically more potent in serum used to grow cells from the same taxon (e.g. fish serum with fish cells and mammalian serum with mammalian cells). Therefore, it supports the idea that mammalian serum is not totally adequate to culture fish cells and that the successful development of fish in vitro cell systems to study vertebrate development will ideally require the availability of commercial fish serum. Future experiments should aim at identifying these serum factors and determining their mechanisms of action.

Our work confirms that dietary lipids modulate the bone metabolism in fish, and demonstrates that dietary DHA is negatively correlated with bone resorption status. Additionally, dietary vegetable oils as replacers of fish oil influence the bone metabolic status of seabream juveniles and cause a tendency towards reduced vertebrae mineralization. These results should be viewed in more global perspective of practical feed formulation, in which both fish meal and fish oil are being simultaneously replaced by vegetable‐based ingredients. Such formulation practices comprise not only changes on the dietary fatty acid supply, but also inherent changes on selected minerals, vitamins, phytosterols, phytoestrogens and cholesterol levels, all substances with a clear association to bone remodeling processes.