Eficacy of Asiaticoside on Proliferation and Differentiation of Osteoblasts Induced by Lipopolysaccharide
Objective:To investigate the efficacy of asiaticoside(AS)on proliferation and differentiation of MC3T3-E1 osteoblasts induced by lipopolysaccharide(LPS).Methods:MC3T3-E1 osteoblasts were first treated with 2 mg/L LPS,and then LPS induced MC3T3-E1 osteoblasts were treated with 5,10,20,40,and 80 μmol/L asiaticoside.Cell activity was measured and the optimal drug concentration was selected.MC3T3-E1 osteoblasts were separated into control group,li-popolysaccharide group(LPS group),low,medium,and high concentration asiaticoside groups(AS-L,AS-M,AS-H group),and high concentration asiaticoside+CXCR4 inhibitor AMD3100 group(AS-H+AMD3100 group).Hoechst 33258 staining was applied to observe the morphology of the nucleus;alkaline phosphatase(ALP)staining and ALP activ-ity were applied to detect the osteogenic ability of cells;alizarin red staining method was applied to detect the formation of calcium nodules;immunoblotting was applied to detect the expression of differentiation related biomarkers such as collagen type Ⅰ(COL-Ⅰ),osteopontin(OPN),osteocalcin(OCN),and SDF-1 and CXCR4.Results:10,20,and 40 pmol/L AS were selected as drug concentrations for AS-L,AS-M,and AS-H groups in subsequent experiments;compared with the control group,the nucleus of MC3T3-E1 osteoblasts in the LPS group showed pyknosis and deep staining,showing a obvi-ous apoptotic morphology,the ALP activity,calcium nodule formation,the expression of COL-Ⅰ,OPN,OCN,SDF-1,CX-CR4 proteins were greatly decreased(P＜0.05).The nuclear pyknosis and deep staining of MC3T3-E1 osteoblasts in AS-L,AS-M,and AS-H groups gradually decreased compared with the LPS group,the number of apoptotic cells gradually decreased,the ALP activity,calcium nodule formation,the expression of COL-Ⅰ,OPN,OCN,SDF-1,CXCR4 proteins were obviously increased in sequence(P＜0.05).The nuclei of MC3T3-E1 osteoblasts in the AS-H+AMD3100 group showed increased pyknosis and deep staining compared with the AS-H group,the apoptotic morphological cells obviously increased,the ALP activity,calcium nodule formation,the expression of COL-Ⅰ,OPN,OCN,SDF-1,and CXCR4 proteins were greatly reduced(P＜0.05).Conclusion:Asiaticoside can promote the proliferation and differentiation of MC3T3-E1 osteoblasts by activating SDF-1/CXCR4 signaling pathway.
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