Abstract：
OBJECTIVE: To assess the effect of Tideglusib and CHIR99021 small molecules on the odontogenic differentiation potential of human dental pulp stem cells (hDPSCs) via Wnt/β-catenin pathway activation.
METHODS: hDPSCs were isolated from impacted third molars indicated for extraction and were characterized by flow cytometry. hDPSCs were then induced to differentiate into odontogenic lineage in the presence of Tideglusib and CHIR99021. Odontogenic differentiation was evaluated using Alizarin Red stain and RT-PCR for expression of odontogenic specific differentiation markers: DSPP, DMP1, ALP, OPN, and RUNX2 in relation to undifferentiated cells. RT-PCR was also conducted to assess the expression of Wnt/β-catenin pathway activation marker (AXIN2). One-way ANOVA Kruskal-Wallis test was used for statistical analysis.
RESULTS: Wnt/β-catenin pathway was successfully activated by Tideglusib and CHIR99021 in hDPSCs where AXIN2 was significantly upregulated. Successful odontogenic differentiation was confirmed by Alizarin Red staining of calcified nodules. RT-PCR for odontogenic differentiation markers DSPP, DMP1, and RUNX expression by hDPSCs induced by CHIR99021 was higher than that expressed by hDPSCs induced by Tideglusib, whereas expression of OPN and ALP was higher in Tideglusib-induced cells than in CHIR99021-induced cells.
CONCLUSIONS: Both small molecules successfully induced odontogenic differentiation of hDPSCs through Wnt/β-catenin pathway activation.
CONCLUSIONS: These findings suggest that Tideglusib and CHIR99021 can be applied clinically in pulp regeneration to improve strategies for vital pulp regeneration and to promote dentine repair.
METHODS: hDPSCs were isolated from impacted third molars indicated for extraction and were characterized by flow cytometry. hDPSCs were then induced to differentiate into odontogenic lineage in the presence of Tideglusib and CHIR99021. Odontogenic differentiation was evaluated using Alizarin Red stain and RT-PCR for expression of odontogenic specific differentiation markers: DSPP, DMP1, ALP, OPN, and RUNX2 in relation to undifferentiated cells. RT-PCR was also conducted to assess the expression of Wnt/β-catenin pathway activation marker (AXIN2). One-way ANOVA Kruskal-Wallis test was used for statistical analysis.
RESULTS: Wnt/β-catenin pathway was successfully activated by Tideglusib and CHIR99021 in hDPSCs where AXIN2 was significantly upregulated. Successful odontogenic differentiation was confirmed by Alizarin Red staining of calcified nodules. RT-PCR for odontogenic differentiation markers DSPP, DMP1, and RUNX expression by hDPSCs induced by CHIR99021 was higher than that expressed by hDPSCs induced by Tideglusib, whereas expression of OPN and ALP was higher in Tideglusib-induced cells than in CHIR99021-induced cells.
CONCLUSIONS: Both small molecules successfully induced odontogenic differentiation of hDPSCs through Wnt/β-catenin pathway activation.
CONCLUSIONS: These findings suggest that Tideglusib and CHIR99021 can be applied clinically in pulp regeneration to improve strategies for vital pulp regeneration and to promote dentine repair.
摘要：
目的：探讨Tideglusib和CHIR99021小分子通过Wnt/β-catenin通路激活对人牙髓干细胞（hDPSCs）牙源性分化潜能的影响。
方法：hDPSC从指示提取的撞击第三磨牙中分离，并通过流式细胞术进行表征。然后在Tideglusib和CHIR99021的存在下诱导hDPSC分化成牙源性谱系。使用茜素红染色和RT-PCR评估牙源性分化，以表达牙源性特异性分化标志物：DSPP，DMP1,ALP,OPN,和RUNX2与未分化细胞有关。还进行了RT-PCR以评估Wnt/β-连环蛋白途径激活标志物(AXIN2)的表达。单因素方差分析Kruskal-Wallis检验用于统计分析。
结果：在AXIN2显著上调的hDPSC中，Tideglusib和CHIR99021成功激活了Wnt/β-catenin通路。通过钙化结节的茜素红染色证实了成功的牙源性分化。牙源性分化标志物DSPP的RT-PCR,CHIR99021诱导的hDPSCs的DMP1和RUNX表达高于Tideglusib诱导的hDPSCs表达，而Tideglusib诱导的细胞中OPN和ALP的表达高于CHIR99021诱导的细胞。
结论：两种小分子均通过激活Wnt/β-catenin通路成功诱导hDPSCs牙源性分化。
结论：这些发现表明，Tideglusib和CHIR99021可以在临床上应用于牙髓再生，以改善重要牙髓再生策略并促进牙本质修复。
方法：hDPSC从指示提取的撞击第三磨牙中分离，并通过流式细胞术进行表征。然后在Tideglusib和CHIR99021的存在下诱导hDPSC分化成牙源性谱系。使用茜素红染色和RT-PCR评估牙源性分化，以表达牙源性特异性分化标志物：DSPP，DMP1,ALP,OPN,和RUNX2与未分化细胞有关。还进行了RT-PCR以评估Wnt/β-连环蛋白途径激活标志物(AXIN2)的表达。单因素方差分析Kruskal-Wallis检验用于统计分析。
结果：在AXIN2显著上调的hDPSC中，Tideglusib和CHIR99021成功激活了Wnt/β-catenin通路。通过钙化结节的茜素红染色证实了成功的牙源性分化。牙源性分化标志物DSPP的RT-PCR,CHIR99021诱导的hDPSCs的DMP1和RUNX表达高于Tideglusib诱导的hDPSCs表达，而Tideglusib诱导的细胞中OPN和ALP的表达高于CHIR99021诱导的细胞。
结论：两种小分子均通过激活Wnt/β-catenin通路成功诱导hDPSCs牙源性分化。
结论：这些发现表明，Tideglusib和CHIR99021可以在临床上应用于牙髓再生，以改善重要牙髓再生策略并促进牙本质修复。
