Effect Of Oleuropein On Osteoclast Differentiation And Bone Resoroption

Osteoporosis,tumor bone metastasis and aseptic loosening of artificial joint are common diseases of bone destruction.Osteoclasts are the only cells in human body to absorb bone,so targeted inhibition of osteoclast differentiation is the most important direction to treat these diseases.However,treatment of these diseases has obvious side effects.It is urgent to develop a targeted therapy for osteoclasts.Natural plant extracts have relatively effective and less side effects,which provide innovative ideas for the treatment of bone destruction related diseases.Oleuropein is one of the most important phenolic compounds in olive oil extract,with multi-purpose pharmacological effects such as atherosclerosis,anti-oxidation and anti-inflammatory effects.It has been reported that oleuropein could enhance osteoblastogenesis and inhibits adipogenesis,but the effects and molecular mechanism of oleuropein on osteoclasts are not clear.In this study,we mainly studied the effects of oleuropein on osteoclast differentiation,bone resorption and signaling pathway.Experiment methods include tartrate resistant acid phosphatase(TRAP)staining,apoptosis experiment,bone pits experiment,Western blot.Bone marrow macrophages were cultured with three variable concentrations(12.5μM,25μM,50μM)of oleuropein based on RANKL-induced osteoclast system in vitro.Multinucleated cells were detected by tartrate resistant acid phosphatase.TRAP staining and TRAP-positive cells were observed.RAW264.7cells were induced by RANKL with variable concentrations of oleuropein,the cell apoptosis was analyzed by flow cytometry.Osteoclastic resorption pits that formed on the bone slices were observed with SEM,the areas of osteoclastic bone resorption were determined using computer image process.The osteoclast-specific protein expression of CTSK,NFATC1 and phosphorylated p38 was measured by Western blotting.Results The RANKL-induced osteoclastogenesis was significantly inhibited by oleuropein in dose-dependent manner.Morover,the protein expression level of CTSK,a osteoclast-specific protein could be down-regulated by oleuropein.Oleuropein had no obvious effect on apoptosis in osteoclast cells.Oleuropein could markedly decrease the area of bone resorption pits.Oleuropein down-regulated the protein expression of NFATC1 and phosphorylated p38.Our finding suggested The RANKL-induced osteoclastogenesis and bone resorption were inhibited by oleuropein in vitro,which may be associated with the down-regulation of p38/MAPK signaling pathway.