Effects of canonical Wnt/β-catenin over expression on proliferation and osteogenic differentiation of human periodontal ligament stem cells

LI Ying; LIU Na; ZHANG Wei; GU Bin

doi:10.3969/j.issn.1672-2973.2017.04.002

Abstract：Objective:To construct Wnt/β-catenin gene overexpressed vector and to investigate the effects of its over expression on proliferation and osteogenic ability of human periodontal ligament stem cells.Methods:8 healthy premolars and third molars extracted for treatment were isolated and cultured for human healthy individuals (PDLSCs).Amplify the Wnt/β-catenin gene from PDLSCs,and cloned it into pLV-EGFP-Nvector,to construct Wnt/β-catenin overexpressed vector,then to transfect it into PDLSCs by lipofection method.Cell morphology was observed under fluorescent microscope after transfection of PDLSCs in conventional and osteogenic culture condition on the seventh day.We tested MTT results of PDLSCs.lymphoid enhancer factor-1 (LEF1),T cell factor-4 (LEF4),Runt-related transfer factor-2 (Runx2),alkaline phosphatase (ALP),human type I collagen (COL-1) and P38 mitogen activated protein kinase (P38 MAPK) were detected by real-time quantitative PCR.Results:GFP was expressed efficiently in PDLSCs after transfection.MTT results showed proliferation of PDLSCs after transfection was boosted specifically and efficiently than before.Real-time PCR results showed the expression of LEF1 mRNA was significantly increased than before,while there is no difference in TCF4.Simultaneously,Runx2,ALP,COL-1 and p38 were significantly decreased of PDLSCs transfected with Wnt/β-catenin overexpressed vector cultured in osteogenic differentiation condition.Conclusion:Classic Wnt/β-catenin pathway plays an important role in PDLSCs proliferation and osteogenic differentiation.Overexpression of the core protein Wnt/β-catenin can upregulate the expression of LEF1,promote the proliferation of PDLSCs,and inhibit the osteogenic ability of PDLSCs by down regulating the expression of RUNX2,ALP,COL-1 and P38.