The Mechanism Of Hederagenin In RANKL-induced Osteoclast Formation And Bone Loss Inducd By OVX Mouse Model

AIMS: Postmenopausal osteoporosis and other osteolytic bone diseases are often caused by the elevation in osteoclastogenesis and/or increased osteoclastic bone resorption,leading to excessive bone loss.Osteoclasts are multinucleated cells derived from hematopoietic progenitor cells in the bone marrow,which are formed by fusing precursor cells and play a role in bone resorption.The maintenance of bones in adults mainly depends on the process of bone remodeling.Bone remodeling is mainly the reabsorption of bone tissue by osteoclasts,and then new bone tissue is formed in the cavity by osteoblasts to maintain the dynamic balance of bone mass.Therefore,osteoclasts are essential for normal bone development(growth and modeling),maintaining their integrity,and calcium metabolism(remodeling).Hederagenin(Hed)is a pentacyclic triterpenoid saponin extracted from various natural medicinal plants and exhibits numerous biological activities such as anti-tumor and anti-inflammatory,and may offer benefits against bone-related conditions.we evaluated the effects of Hed on osteoclast formation and bone resorption in vitro and the in vivo therapeutic benefits in the mouse model of ovariectomy(OVX)-induced bone loss.MAIN METHODS: C57 BL / 6 mouse bone marrow macrophages(BMMs)were extracted and cultured in vitro.Hed was administered at different concentrations under RANKL induction,and TRAP staining was performed to determine the effect of Hed on osteoclast formation.Fluorescent probe DCFH-DA was used to detect intracellular ROS to determine the effect of Hed on ROS content in osteoclasts.bone resorption were examined using Hydroxyapatite resorption assay and Podosomal actin belt formation assay;RT-PCR was used to quantitatively analyze the expression of specific genes in related osteoclasts.Related molecular mechanisms were determined by western blot assay.Ovx mice were constructed by bilateral ovariectomy to simulate bone loss in vivo.C57 BL / 6 mice were randomly divided into 4 groups in vivo: blank control group,surgery group(OVX group),low-dose group(OVX + low-dose Hed),high-dose group(OVX + high-dose Hed).After 6 weeks,the mouse tibia was extracted as an experimental sample.Analysis of tibia bone loss by Micro CT scan and statistical analysis.Histological staining was used to observe the metabolic toxicity of Hed on the number of osteoclasts in the tibia and major metabolic organs(heart,liver,spleen,lung and kidney).KEY FINDINGS: In vitro cellular assays showed that Hed inhibited RANKL-induced osteoclast formation and osteoclast bone(hydroxyapatite)resorption as well as marker gene expression from BMM culture.Mechanistically,Hed attenuated RANKL-induced intracellular reactive oxygen species(ROS)production,and MAPK signaling pathway(ERK and p38)activation which curbed the downstream induction of c-Fos and NFATc1.Consistent with the in vitro findings,Hed administration effectively protected OVX mice from bone loss by reducing osteoclast number and activity on bone surface.SIGNIFICANCE: our data of this experiment provide strong evidence that Hed can inhibit the formation of osteoclasts and bone resorption by inhibiting RANKL-mediated activation of the MAPK signaling pathway,for which the potential use of Hederagenin in the treatment of osteoclast-mediateed osteolytic bone diseases such as postmenopausal osteoporosis.