Inhibitory effect of menaquinone-7 (vitamin K2) on osteoclast-like cell formation and osteoclastic bone resorption in rat bone tissues in vitro

The effect of menaquinone-7 (MK-7; vitamin K2) on osteoclast-like cell formation and osteoclastic bone resorption in rat femoral tissues in vitro was investigated. The bone marrow cells were cultured for 7 days in a a-minimal essential medium (alpha-MEM) containing a well-known bone resorbing agent [parathyroid hormone (1-34) (PTH) or prostaglandin E2 (PGE2)] with an effective concentration. Osteoclast-like cells were estimated by staining for tartrate-resistant Acid Phosphatase (TRACP), a marker Enzyme of osteoclasts. The presence of PTH (10(-8) M) or PGE2 (10(-6) M) induced a remarkable increase in osteoclast-like multinucleated cells. These increases were significantly inhibited by MK-7 (10(-8) - 10(-5) M). MK-7 (10(-7) and 10(-6) M) significantly inhibited phorbol 12-myristate 13-acetate-induced osteoclast-like cell formation, whereas MK-7 did not inhibit dibutyryl cyclic adenosine monophosphate (DcAMP) (10(-5) M)-induced osteoclast-like cell formation. These results suggest that the inhibitory action of MK-7 is partly involved in protein kinase C signaling. The bone cells isolated from rat femoral tissues were cultured for 48 h in an alpha-MEM containing either vehicle or MK-7 (10(-8) - 10(-5) M). The presence of MK-7 (10(-6) and 10(-5) M) caused a significant decrease in the number of mature osteoclasts. Such a decrease was also seen in the presence of Calcitonin (10(-10) - 10(-8) M), DcAMP (10(-6) and 10(-5) M), or calcium chloride (10(-4) and 10(-3) M). The effect of MK-7 (10(-6) M) in decreasing the number of osteoclasts was not further enhanced in the presence of Calcitonin (10(-8) M), DcAMP (10(-5) M), or calcium chloride (10(-3) M), and was completely abolished by the presence of dibucaine (10(-6) M) or staurosporine (10(-7) M), which are inhibitors of Ca2+-dependent protein kinases. These results suggested that MK-7 has a suppressive effect on osteoclasts. Moreover, the femoral-metaphyseal tissues obtained from rats were cultured for 48 h in Dulbecco's modified Eagle's medium containing either vehicle, PTH (10(-7) M), orPGE2 (10(-5) M) in the absence or presence of MK-7 (10(-7) - 10(-5) M). The presence of PTH or PGE2 induced a significant decrease in bone calcium content. These decreases were significantly blocked by MK-7 (10(-7) - 10(-5) M). This study demonstrates that MK-7 has an inhibitory effect on osteoclastic bone resorption in vitro.