BACKGROUND:Overexpression of the nuclear factor I-C gene in vitro promotes the differentiation of human stem cells from apical papilla,as does the activation of the Wnt/β-catenin signaling pathway.Moreover,nuclear factor I-C regulates the Wnt/β-catenin pathway in mesenchymal stem cells.However,whether nuclear factor I-C can affect cell differentiation by activating the Wnt/β-catenin pathway inhuman stem cells from apical papilla has not been reported.OBJECTIVE:To investigate the role of nuclear factor I-C in the Wnt/β-catenin signaling pathway in regulating the differentiation of human stem cells from apical papilla.METHODS:H uman stem cells from apical papilla were cultured by the slide-covered tissue block method and lentiviral transfection overexpressing the nuclear factor I-C gene.(1)A control group,an empty viral vector group,and an overexpressed nuclear factor I-C gene group were set up.The expression ofβ-Catenin,LRP5,and TCF7L2 was detected by Western blotting.(2)The control group,empty viral vector group,overexpressed nuclear factor I-C gene group,and overexpressed nuclear factor I-C gene+DKK-1(Wnt pathway inhibitor)group were set up.Alkaline phosphatase staining and activity quantification were performed after 7 days of osteogenic induction.qPCR and Western blotting were performed to detect the expression of Runt-related transcription factor 2,dentin salivary phosphoprotein,osteocalcin mRNA,and protein after 14 days of osteogenic induction.Alizarin Red staining was used to observe the formation of mineralized nodules.RESULTS AND CONCLUSION:(1)Compared with the control and empty viral vector groups,the expression of Wnt/β-Catenin pathway-related proteins β-Catenin,LRP5,and TCF7L2 inhuman apical dentin papilla stem cells was significantly increased in the overexpressed nuclear factor I-C gene group(P＜0.01).(2)Compared with the control and empty viral vector groups,the expression of alkaline phosphatase and osteocalcin in human apical dentin papilla stem cells was significantly increased(P＜0.01);the expression levels of Runt-related transcription factor 2,dentin salivary phosphoprotein,osteocalcin mRNA and protein were significantly higher(P＜0.01),and the number of mineralized nodules was significantly increased(P＜0.01)in the overexpressed nuclear factor I-C gene group.(3)Compared with the overexpressed nuclear factor I-C gene group,the alkaline phosphatase activity and the expression of Runt-related transcription factor 2,dentin salivary phosphoprotein,osteocalcin mRNA and protein expression levels were significantly down-regulated(P＜0.05),and the number of mineralized nodules was significantly reduced(P＜0.05)in human stem cells from apical papilla of the overexpressed nuclear factor I-C gene+DKK-1 group.The results show that nuclear factor I-C can activate the Wnt/β-catenin signaling pathway in human stem cells from apical papilla and mediate the osteogenic/odontogenic differentiation of human stem cells from apical papilla.

BACKGROUND:Overexpression of the nuclear factor I-C gene in vitro promotes the differentiation of human stem cells from apical papilla,as does the activation of the Wnt/β-catenin signaling pathway.Moreover,nuclear factor I-C regulates the Wnt/β-catenin pathway in mesenchymal stem cells.However,whether nuclear factor I-C can affect cell differentiation by activating the Wnt/β-catenin pathway inhuman stem cells from apical papilla has not been reported.OBJECTIVE:To investigate the role of nuclear factor I-C in the Wnt/β-catenin signaling pathway in regulating the differentiation of human stem cells from apical papilla.METHODS:H uman stem cells from apical papilla were cultured by the slide-covered tissue block method and lentiviral transfection overexpressing the nuclear factor I-C gene.(1)A control group,an empty viral vector group,and an overexpressed nuclear factor I-C gene group were set up.The expression ofβ-Catenin,LRP5,and TCF7L2 was detected by Western blotting.(2)The control group,empty viral vector group,overexpressed nuclear factor I-C gene group,and overexpressed nuclear factor I-C gene+DKK-1(Wnt pathway inhibitor)group were set up.Alkaline phosphatase staining and activity quantification were performed after 7 days of osteogenic induction.qPCR and Western blotting were performed to detect the expression of Runt-related transcription factor 2,dentin salivary phosphoprotein,osteocalcin mRNA,and protein after 14 days of osteogenic induction.Alizarin Red staining was used to observe the formation of mineralized nodules.RESULTS AND CONCLUSION:(1)Compared with the control and empty viral vector groups,the expression of Wnt/β-Catenin pathway-related proteins β-Catenin,LRP5,and TCF7L2 inhuman apical dentin papilla stem cells was significantly increased in the overexpressed nuclear factor I-C gene group(P＜0.01).(2)Compared with the control and empty viral vector groups,the expression of alkaline phosphatase and osteocalcin in human apical dentin papilla stem cells was significantly increased(P＜0.01);the expression levels of Runt-related transcription factor 2,dentin salivary phosphoprotein,osteocalcin mRNA and protein were significantly higher(P＜0.01),and the number of mineralized nodules was significantly increased(P＜0.01)in the overexpressed nuclear factor I-C gene group.(3)Compared with the overexpressed nuclear factor I-C gene group,the alkaline phosphatase activity and the expression of Runt-related transcription factor 2,dentin salivary phosphoprotein,osteocalcin mRNA and protein expression levels were significantly down-regulated(P＜0.05),and the number of mineralized nodules was significantly reduced(P＜0.05)in human stem cells from apical papilla of the overexpressed nuclear factor I-C gene+DKK-1 group.The results show that nuclear factor I-C can activate the Wnt/β-catenin signaling pathway in human stem cells from apical papilla and mediate the osteogenic/odontogenic differentiation of human stem cells from apical papilla.

Objective:The aim of this study was to investigate the role of cyclic adenosine monophosphate (cAMP) signaling on the differentiation of stem cells from the apical papilla (SCAPs).Methods:SCAPs were cultured by enzyme digestion method.After adding activator (Forskolin) and inhibitor (H-89) into the culture of SCAPs in mineralization-inducing medium,calcium deposition was measured by alizarin red staining.QPCR was performed to measure the mRNA expression of mineralization-related genes.Results:When the cAMP signaling was activated,calcium deposition and the mRNA expression of ALP,OCN,OSX and RUNX2 increased sharply.After inhibiting the signaling,calcium deposition and the mRNA expression of ALP,OCN,OSX and RUNX2 decreased.Conclusion:The cAMP signaling may promote the odontogenic and osteogenic differentiation of SCAPs.