Megakaryocytes (MKs) play key roles in regulating bone metabolism.To test the roles of MK-secreted factors, we investigated whether MK and promegakaryocyte (pro-MK) conditioned media (CM) may affect bone formation and resorption. K562 cell lines were differentiated into mature MKs. Mouse bone marrow macrophages were differentiated into mature osteoclasts, and MC3T3-E1 cells were used for osteoblastic experiments. Bone formation was determined by a calvaria bone formation assay in vivo. Micro-CT analyses were performed in the femurs of ovariectomized female C57B/L6 and Balb/c nude mice after intravenous injections of MK or pro-MK CM. MK CM significantly reduced in vitro bone resorption, largely due to suppressed osteoclastic resorption activity. Compared with pro-MK CM, MK CM suppressed osteoblastic differentiation, but stimulated its proliferation, resulting in stimulation of calvaria bone formation. In ovariectomized mice, treatment with MK CM for 4 weeks significantly increased trabecular bone mass parameters, such as bone volume fraction and trabecular thickness, in nude mice, but not in C57B/L6 mice. In conclusion, MKs may secrete anti-resorptive and anabolic factors that affect bone tissue, providing a novel insight linking MKs and bone cells in a paracrine manner. New therapeutic agents against metabolic bone diseases may be developed from MK-secreted factors.Bone metabolism is regulated mainly by the action of bone-resorbing osteoclasts and bone-forming osteoblasts. Increased bone resorption and/or decreased bone formation can lead to reduced bone mass and quality, resulting in high fracture risk. Typical conditions that induce this imbalance include estrogen deficiency and immobilization 1,2 . On the contrary, decreased bone resorption and/or increased formation with pharmacological interventions can reverse these imbalances. Synchronously inhibiting bone resorption and stimulating bone formation are regarded as an ideal therapeutic strategy against metabolic bone diseases, such as osteoporosis.Bone marrow, where most osteoclasts and osteoblasts exist, also contains many types of hematopoietic cells. The coexistence of the cells in the bone marrow allows these cells to influence one another by cell-to-cell contact or in a paracrine manner. Specifically, megakaryocytes (MKs), as one of hematopoietic cell residing in bone marrow, have been intriguing in the field of bone research. MKs, which are polyploid cells derived from MK/erythroid progenitors, generate platelets, which contribute to hemostasis and produce a number of growth factors 3-5 . Interestingly, estrogen deficiency and immobilization reduced the number of MKs 6,7 , and estrogen treatment increased MK number in postmenopausal women 8 . In addition, MK-related disorders are associated with osteosclerosis 9-11 . Thus, MKs may have a critical role in bone metabolism.Actually, it has been reported that MKs may act on both bone resorption and bone formation. A mouse model with increased MKs showed decreased osteoclast number and bone resorption 12 . ...