In the current context of longer life expectancy, the prevalence of osteoporosis is increasingly important. This is why development of new strategies of prevention is highly suitable. Pomegranate seed oil (PSO) and its major component, punicic acid (a conjugated linolenic acid), have potent anti-inflammatory and anti-oxidative properties both in vitro and in vivo, two processes strongly involved in osteoporosis establishment. In this study, we demonstrated that PSO consumption (5% of the diet) improved significantly bone mineral density (240.24 +/- 11.85 vs. 203.04 +/- 34.19 mg/cm(3)) and prevented trabecular microarchitecture impairment in ovariectomized (OVX) mice C57BL/6J, compared to OVX control animals. Those findings are associated with transcriptional changes in bone tissue, suggesting involvement of both osteoclastogenesis inhibition and osteoblastogenesis improvement. In addition, thanks to an ex vivo experiment, we provided evidence that serum from mice fed PSO (5% by gavage) had the ability to significantly down-regulate the expression of specific osteoclast differentiation markers and RANK-RANKL downstream signaling targets in osteoclast-like cells (RAW264.7) (RANK: negative 0.49-fold vs. control conditions). Moreover, in osteoblast-like cells (MC3T3-E1), it elicited significant increase in alkaline phosphatase activity (+159% at day 7), matrix mineralization (+271% on day 21) and transcriptional levels of major osteoblast lineage markers involving the Wnt/beta-catenin signaling pathways. Our data also reveal that PSO inhibited pro-inflammatory factors expression while stimulating anti-inflammatory ones. These results demonstrate that PSO is highly relevant regarding osteoporosis. Indeed, it offers promising alternatives in the design of new strategies in nutritional management of age-related bone complications. (C) 2013 Elsevier Inc. All rights reserved.