The effects of icariine concentration on osteoclasts bone resorption induced by titanium particles in vitro.

In artificial joint replacement, osteoclast bone resorption induced by wear debris of the implant is a main reason for aseptic loosening. To extend the life of the prosthesis, detailed mechanisms of aseptic loosening and the ways to prevent it should be explored. The aim of this study was to investigate the in vitro effect of icariine on the bone resorption of osteoclasts induced by titanium particles. Macrophage colony stimulating factor (M-CSF) and receptor activator of NF-kB ligand (RANKL) were used to generate osteoclasts from RAW264.7 precursors. The proliferation of RAW264.7 precursors in the presence of different doses of icariine was evaluated by MTT assay. The cells were treated with titanium particles, titanium particles with icariine and culture medium only (control), respectively. At 48 h after treatment, the expression level of receptor activator of NF-kB (RANK) was detected by ELISA, and messenger RNA (mRNA) levels of tartrate-resistant acid phosphatase (TRAP), matrix metalloproteinase 9 (MMP-9), carbonic anhydrase II (CAII) and Cathepsin K (CtsK) were determined by real-time polymerase chain reaction. Western blot was applied to analyze the expression levels of TRAP, RANK and CtsK. In addition, bone chips were cultured in the above conditions, and Toluidine blue staining was then employed to calculate the number and area of resorption pits in the bone chips. After treatment with icariine, expression level of RANK was significantly decreased in the RAW264.7 cell that induced by titanium particle and its cultural medium, mRNA and protein levels of TRAP, CAII, MMP-9 and CtsK were reduced as well. In addition, the numbers of bone resorption pits and areas on bone slices were both reduced by icariine challenging. Icariine could inhibit bone resorption of osteoclast induced by titanium particle, and it might be used as a promising drug for treating of aseptic loosening.