Abstract:

The major and active constituents of the genus Epimedium are flavonoids, which can prevent bon e loss in ovariectonized rats. Icarrin is a flavonoid isolated from Herba Epimed ii. By enzymatic and acidic hydrolysis of icarrin, two metabolites (icariside Ⅱ and icaritin) can be obtained. Here we report the effects of icarrin and its tw o metabolites on the bone formation and bone resorption in vitro.Three cell models were used. Primary osteoblastic cells were isolated from neona tal Wistar rat calvarias. Icarrin and its two metabolites increased the viabilit y of osteoblasts significantly. Besides, alkaline phosphatase (ALP) activity and osteocalcin (OC) synthesis of osteoblasts also increased, as well as the ALP, O C and collagen mRNA expression. Since ALP and OC are phenotypic markers for earl y-stage differentiated osteoblasts and terminally differentiated osteoblasts re spectively, our findings indicate that Icarrin and its metabolites stimulate ost eoblasts differentiation at various stages (from osteoprogenitors to terminally differentiated osteoblasts). The effects of Icarrin and its metabolites on osteoclast-like cells were obtain ed by coculturing bone marrow with osteoblasts for 8 days in α-MEM containing 10^-8mol/L 1.25(OH)₂D₃. The osteoclast-like cell formation was estimat ed by staining for tartrate-resistant acid phosphatase (TRACP), a marker enzyme of osteoclast. Icarrin and its metabolites significantly inhibited osteoclast-like cell formation, and elevated the mRNA levels of OPG and RANKL dose-depend e ntly, resulting in enlarged ratio of the two as revealed by RT-PCR. Hence it is likely that icarrin and its metabolites regulate bone resorption, at least in part, via its effects on OPG and RANKL expression.Mature osteoclasts were isolated from the long bones of neonatal Japanese white rabbits (60-90g). The surface area of bone resorption lacunae was measured by p hotomicrography and image analysis. Actin microfilaments were detected by bindin g of rhodamine-conjugated phalloidin (R-PHD) to F-actin. Icarrin and its meta bolites showed the potency to withdraw the actin ring, consistent with the inhib ited bone resorption of osteoclasts. In conclusion, icarrin and its metabolites can not only interfere with the diffe rentiation of osteoclast-like cells and inhibit bone resorption ability of matu re osteoclasts, but also promote bone formation.