Effect Of Basangmu Suyou Wan On Proliferation,Osteogenic Differentiation And Secretion Of Hematopoietic Regulatory Factors In Mouse MC3T3-E1 Cells

Objective:In order to provide traditional nourishing theory and clinical formula using of Tibetan medicine with biological basis,this experiment investigated the effect and mechanism of Basangmu Suyou Wan to mice osteoblast cell line MC3T3-E1 cells' proliferation,osteogenesis and hematopoietic function,and exploring the targets of “strong bones and muscles” function and promoting the recovery of hematopoietic function of “deficient” body of Basangmu Suyou Wan.Method: Mice MC3T3-E1 cells was cultured in medium containing serum from mice feed with Basangmu Suyou Wan(Ba serum),cells proliferation,ALP activity and calcification conditon were mesured by CCK-8 assay,ALP dyeing and alizarin red dyeing method.Main factors' mRNA expression of related signaling pathway such as ?-catenin,Runx2 and Osx were detected by real-time PCR.The expression level of SDF-1,GM-CSF and LIF were detected by western blot and ELISA.Treated group was compared with no mice serums' Mock group or NS serum group in the experiment.Result:1.Compared with Mock group,the OD value of cells cultured with 5% and 10% Ba serum after 48 h increased significantly(P<0.01).OD value in1% and 5% NS serum cultured group and each Ba serum cultured group were all significantly increased at 48 h and 72 h.OD value in 10% NS serum cultured group was also increased(P<0.05).Compared with the same concentration NS serum group,all OD values were increased significantly in cells cultured with 5%,10%,and 20% Ba serum at 24 h and cells cultured with 10%,20% Ba serum at 48 h,72h(P<0.01).In the Ba serum group,10% concentration and 20% concentration are compared with each other,OD value was significantly higher in cells cultured with 10% Ba serum group than 20% Ba serum group at 72h(P<0.01).2.ALP activity and calcified nodules of cells cultured in 10% Ba serum group were higher than 10% NS serum group(P<0.01).3.Runx2 mRNA expression level in cells cultured in 10% Ba serum group for 48 h was lower than 10% NS serum group,?-catenin and Osx mRNA expression level were higher than 10% NS serum group,there was no statistical significance.4.LIF and SDF-1 expression level was higher in cells cultured in 10% Ba serum group for 48 h than 10% NS serum group(P<0.01),though expression level of GM-CSF was higher in the former group,there was no statistical significance.Conclusion:1.Osteoblasts' proliferation,differentiation and calcification are increased by serum from mice feed with Basangmu Suyou Wan,which support the hypothesis that Basangmu Suyou Wan may promote the recovery of injured hematopoietic cell osteoblastic niche and the recovery of hematopoietic function.2.Serum from mice feed with Basangmu Suyou Wan increased the adhesion of hematopoietic stem cells to osteogenesis and maintain the characteristic of asymmetrical splitting,self-renewal,and simultaneous differentiation of stem cells by increasing osteoblasts secreting adhesion molecule SDF-1.3.Serum from mice feed with Basangmu Suyou Wan adjust and maintain the normal hematopoietic function by increasing osteoblasts secreting hematopoietic regulator LIF.4.osteoblasts is the link site for the efficacy of “strong bones and muscles”and nourishing “deficient” body of Basangmu Suyou Wan.