Effects of methopterin on osteoclasts and study of its mechanism of action

This study is to observe the effects of methopterin on the activation and bone resorption function of murine osteoclasts, which were obtained by induction from bone marrow cell and purified to the purity of 70%-80%. The mechanism underlying the inhibitory effects of methopterin on inflammatory bone destruction was explored. MTT method was used to determine the effect of methopterin on the proliferation of osteoclasts. Flow cytometric analysis was used to determine the effect of methopterin on the apoptosis of osteocalsts. TRAP stain, bone resorption lacuna stain and measurement of lacuna area were executed to determine the effects of methopterin on the activation and function of osteoclasts. ELISA method was used to determine the effect of methopterin on the MMP-9 secretion from osteoclasts. RT-PCR method was used to determine the effect of methopterin on the mRNA expression of RANK and MMP-9 in osteoclasts. The results showed that methopterin (0.1-10 μmol·L^-1) inhibited the proliferation of osteoclasts, methopterin (0.1-10 μmol·L^-1) could inhibit the activation and bone resorption function of osteoclasts and induced the apoptosis of osteoclasts. Methopterin (0.01-10 μmol·L^-1) also decreased the mRNA expression of RANK, but only at 1-10 μmol·L^-1 decreased the mRNA expression of MMP-9. These results indicated that there were intense relation between the inhibitory effects on the activation and function of osteoclasts and the inhibition of inflammatory bone destruction by methopterin.