| Objectives:Interlenkin-34(IL-34)is recently identified as a novel cytokine,substituting for the function of macrophage colony stimulating factor(M-CSF),a pivotal osteoclastogenic factor involved in bone-relevant diseases such as osteomyelitis of the jaws.The aim of the present study was to explore the potential mechanism of IL-34 in RANKL-induced osteoclasts formation.Methods:IL-34 concentrations in supernatants of osteoclasts induced by M-CSF+RANKL in bone marrow macrophages(BMMs)were assessed by ELISA assay.The effects of IL-34 and different concentrations of AG490 on survival BMMs were assessed by CCK-8 assay.The expressions of osteoclast-related genes Ctsk,NFATc1,TRAP were analyzed by real-time PCR.Osteoclastogenesis was evaluated by tartrate-resistant acid phosphatase(TRAP),DAPI and hematoxylin staining.Bone resorption assay and Von Kossa staining were used to observe the activity of osteoclasts.The protein expressions including STAT3 phosphorylation,STAT3 and Smad7 were analyzed by Western blot.Results:IL-34 alone not only significantly maintained the survival of bone marrow macrophages,but also enhanced the expression of osteoclast-related genes TRAP,Ctsk and NFATc1 as well as the TRAP positive multinucleated cells combined with RANKL,which could be reversed by AG490,whereas AG490 had no manifest impact on the group M-CSF plus RANKL.In addition,RANKL alone promoted TRAP positive multinucleated cells at later stage(Days 10-14).Furthermore,the protein expression of p-STAT3 in BMMs was enhanced by IL-34 combined with RANKL compared to RANKL alone;AG490 inhibited expression of p-SATA3 in protein level in IL-34 plus RANKL group,meanwhile exerting a significant increase in the expression of Smad7.Conclusions:This study demonstrated for the first that IL-34 may have a crucial role in RANKL-induced osteoclastogenesis by promoting the proliferation and differentiation of bone marrow macrophages,stimulating the expression of p-STAT3 and inhibiting the expression of Smad7 in the absence of M-CSF.Further in vivo investigations are necessary to explore this hypothesis. |
