The Roles Of Curculigoside On The Differentiation Of Osteoblasts And The Treatment Of Osteoporosis

Osteoporosis is a systematic disease,which is characterized by the loss of bone and the destruction of bone tissues.As the aging of the society,more people are suffering from osteoporosis.The cause of the osteoporosis is the imbalance between osteoclastic bone resorption and osteoblastic bone formation,with the bone resorption exceeding bone formation,leading to bone loss.As aging proceeds,reactive oxygen species produced by the metabolism of mitochondria accumulate,while the function of anti-oxidation is deceased,leading to excessive oxidative stress response within the cells.This has been identified as one of the mechanisms of the osteoporosis.Nowadays,the drugs used for treating osteoporosis are designed focusing on promoting osteoblastic bone formation or inhibiting osteoclastic bone resorption.Among these drugs,parathyroid hormone（PTH）is the only one that has been approved by U.S.Food and Drug Administration（FDA）.PTH can promote bone formation,however,it can also cause osteosarcoma,limiting its application clinically.Therefore,people then focused on natural compounds.It is reported that a plethora natural compounds can regulate the differentiation of osteoblasts.The combination of epimedium and curculigo can significantly increased the bone mineral density in ovariectomized rats,exhibiting an obvious anti-osteoporosis pharmacological effects.Curculigoside is the extract from curculigo,with biological functions.The aim of this study is to explore the roles of curculigoside on the differentiation of osteoblastic cell line MG-63 and the treatment of osteoporosis in the mouse model.In Chapter 2,MTT was used to detect the cell viability of MG-63after treatment with dexamethasone or H₂O₂.There were 6 groups:control group,curculigoside only group（5.0μmol/L curculigoside）,dexamethasone or H₂O₂ treated group,low dose group（dexamethasone or H₂O₂ treated+1.0μmol/L curculigoside）,medium dose（dexamethasone or H₂O₂ treated+2.5μmol/L curculigoside）,and high dose（dexamethasone or H₂O₂ treated+5μmol/L curculigoside）.The results showed that the cell viability in the dexamethasone or H₂O₂ treated group was decreased,compared with the control group.While after treatment with different doses of curculigoside,the cell viability was recovered.Then total protein was extracted after treatment with curculigoside for 1 d,7 d,and 14 d,respectively.Western blot was used to detect the protein levels representing the differentiation of osteoblasts.The results showed that osteoblastic markers（i.e.collagen type I,Osterix,integrinβ1 and osteocalcin）were elevated at different time points.These result suggested that curculigoside could promote the proliferation and differentiation of MG-63 in vitro.In Chapter 3,we established an osteoporosis mouse model by intraperitoneal injection of dexamethasone.There were 4 groups:control group,osteoporosis group（treated by intraperitoneal injection of dexamethasone）,low dose of curculigoside group（treated by dexamethasone and 5 mg/kg curculigoside）and high dose of curculigoside group（treated by dexamethasone and 45 mg/kg curculigoside）.Eight weeks later,the mice were sacrificed.The distal part of the femurs and the proximal part of the tibia were scanned by micro-CT to detect the bone mass and bone mineral density.Histology and HE staining was performed to observe bone mass and the number of osteoclasts.ELISA was used to detect the concentration of molecules related with oxidation in the serum.The results showed that in osteoporosis group,bone mass and bone mineral density were decreased,the number of osteoclasts were increased,the concentration of oxide molecules in the serum were elevated,whereas the concentration of anti-oxidation molecules were decreased,compared with the control group.In the curculigoside treatment groups,including low lose and high lose,the above indices were all recovered.These results suggested that curculigoside could promote osteoblastic bone formation and inhibit osteoclastic bone resorption in the dexamethasone-induced osteoporotic mouse model,and that curculigoside exhibited pharmacological effects for dexamethasone-induced osteoporosis.