Effect of the astragaloside Ⅳ on proliferation and osteogenic differentiation of human periodontal stem cells

Objective: To investigate the effects of astragaloside Ⅳ(AS-Ⅳ) on proliferation and osteogenic differentiation of human periodontal stem cells(hPDLSCs) and its mechanism. Methods: After removing one-third of the human periodontal ligament tissue,the hPDLSCs were cultured using the tissue block method.The expression of CD105,CD90,CD44,CD45,and CD34 on the surface of hPDLSCs were identifiied using flow cytometry.Alkaline phosphatase(ALP),alizarin red(ARS) and oil red O staining were used to identify the multidirection differentiation(osteogenic and lipogenic differentiation) of cells.The hPDLSCs were treated with different concentrations of AS-Ⅳ(0,20,50,100,150 μmol/L),and the proliferative activity of hPDLSCs was detected by CCK-8 method.The hPDLSCs were co-cultured with 0,20,50,100,150 μmol/L AS-Ⅳ for 7 and 14 days,and the effects of different AS-Ⅳ concentrations on osteogenic differentiation of hPDLSCs were detected by ALP staining,ALP activity detection and ARS staining.The experiment was divided into the control group and experimental group(150 μmol/L AS-Ⅳ),two groups were cultured with osteogenic induction solution for 14 days.The mRNA expression levels of ALP,Runt-related transcription factor 2(RUNX2) and osteocalcin(OCN) were detected by qRT-PCR.The expression levels of ALP,RUNX2,OCN and β-catenin proteins were detected by Western blotting. Results: The hPDLSCs possessed mesenchymal stem cell properties,the surface antigens CD105,CD44,and CD90 were positive,but the CD34 and CD45 were negative.These cells had the capacity to differentiate in multiple directions,and can differentiate into osteogenesis and adipogenesis.AS-Ⅳ could promote the osteogenic differentiation of hPDLSCs.The results of qRT-PCR and Western blotting showed that compared with the control group,the relative expression levels of ALP,RUNX2 and OCN mRNA and protein in AS-Ⅳ(150 μmol/L) group increased(P<0.05 to P<0.01),and the expression level of β-catenin protein decreased(P<0.05). Conclusions: AS-Ⅳ can promote the osteogenic differentiation of hPDLSCs,which may be achieved by inhibiting Wnt/β-catenin signaling pathway.