Curcumin regulates odontogenic differentiation of human dental pulp stem cells through Wnt signaling pathway

doi:10.3969/j.issn.2095-4344.1784

Abstract

Abstract:

BACKGROUND: Studies have shown that curcumin not only promotes osteogenic differentiation of adult stem cells, but also promotes proliferation and differentiation of neural stem cells by activating the Wnt signaling pathway. However, there is no report on whether curcumin can promote odontoblast the differentiation of human dental pulp stem cells by activating the Wnt signaling pathway.
OBJECTIVE: To investigate the effect of curcumin on the proliferation and odontogenic differentiation of human dental pulp stem cells and its possible mechanism.
METHODS: Human dental pulp stem cells (purchased from Shanghai Yansheng Industrial Co., Ltd., China) were divided into six groups according to different intervention methods. The normal control group was not intervened. Low-, medium- and high-concentration curcumin groups were treated with 50, 250, and 500 nmol/L curcumin, respectively. IWR-1 group was intervened with 1 μmol/L Wnt signaling pathway inhibitor IWR-1. IWR-1+curcumin group were treated with 1 μmol/L IWR-1 and 500 nmol/L curcumin. After 7 and 14 days of mineralization, the mRNA levels of Wnt5a, alkaline phosphatase and dentin sialophosphoprotein were detected by real-time quantitative PCR. The protein levels of Wnt5a, β-catenin, bone sialic acid protein, dentin sialophosphoprotein, osteocalcin and dentin matrix protein 1 were detected by western blot. The activity of alkaline phosphatase was detected by colorimetry, and the proliferation activity of the cells was detected by cell counting kit-8 method.
RESULTS AND CONCLUSION: (1) Curcumin had a time-dependent and dose-dependent effect on the proliferation of dental pulp stem cells. (2) Curcumin had a time- and dose-dependent effect on the mRNA expressions of dentin sialophosphoprotein and alkaline phosphatase, as well as the protein expressions of dentin sialophosphoprotein, dentin matrix protein 1, bone sialoprotein and osteocalcin in dental pulp stem cells. (3) Curcumin up-regulated the expressions of Wnt5a and β-catenin proteins in dental pulp stem cells in a time- and dose-dependent manner. (4) Compared with the control and IWR-1 groups, the combination of curcumin and IWR-1 significantly increased the expressions of Wnt5a mRNA and protein, dentin sialophosphoprotein, dentin matrix protein 1, and bone sialoprotein and osteocalcin proteins in human dental pulp stem cells, and significantly promoted the proliferative ability of the cells (P < 0.05). These results suggest that curcumin promotes the proliferation and odontogenic differentiation of human dental pulp stem cells by activating the Wnt signaling pathway.

Key words: dental pulp stem cells, curcumin, Wnt signaling pathway, odontogenic differentiation, osteogenic differentiation, dental regeneration, cell proliferation

CLC Number:

Chen Dongmei, Xu Lili, Zhou Jianwei.

Curcumin regulates odontogenic differentiation of human dental pulp stem cells through Wnt signaling pathway

[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(25): 4018-4024.

Figures/Tables 4

2.3 姜黄素对人牙髓干细胞成牙本质分化能力的影响 RT-qPCR检测结果显示，诱导分化7 d后，高浓度姜黄素组细胞的牙本质涎磷蛋白、碱性磷酸酶mRNA表达高于其他3组，诱导分化14 d后，中、高浓度姜黄素组细胞的牙本质涎磷蛋白、碱性磷酸酶mRNA表达高于其他2组，差异有显著性意义(P < 0.05)；与诱导分化7 d比较，中、高浓度姜黄素组诱导分化14 d后牙本质涎磷蛋白、碱性磷酸酶mRNA表达升高，差异有显著性意义(P < 0.05)。 Western blot 检测结果也显示，诱导分化7 d后，高浓度姜黄素组细胞的牙本质涎磷蛋白、牙本质基质蛋白1、骨唾液酸蛋白和骨钙蛋白表达高于其他3组，诱导分化14 d后，中、高浓度姜黄素组细胞的牙本质涎磷蛋白、牙本质基质蛋白1、骨唾液酸蛋白和骨钙蛋白表达高于其他2组，差异有显著性意义(P < 0.05)；与诱导分化7 d比较，中、高浓度姜黄素组细胞诱导分化14 d后牙本质涎磷蛋白、牙本质基质蛋白1、骨唾液酸蛋白和骨钙蛋白表达升高，差异有显著性意义(P < 0.05)，见图1。以上结果提示姜黄素对人牙髓干细胞的成牙本质分化发挥促进作用。 2.4 姜黄素对人牙髓干细胞Wnt信号通路相关蛋白表达的影响 Western blot 检测结果显示，诱导分化7，14 d后，高浓度姜黄素组细胞中Wnt5a和β-catenin蛋白的表达高于其他3组，差异有显著性意义(P < 0.05)；与诱导分化7 d比较，高浓度姜黄素组诱导分化14 d后细胞中Wnt5a和β-catenin蛋白的表达升高，差异有显著性意义(P < 0.05)，见图2。以上结果提示姜黄素可促进人牙髓干细胞Wnt信号通路相关蛋白的表达。 2.5 姜黄素对抑制Wnt信号通路的人牙髓干细胞增殖和成牙本质分化的影响为了进一步明确Wnt信号通路在姜黄素促进人牙髓干细胞增殖和成牙本质分化中的作用，应用Wnt信号通路抑制剂处理人牙髓干细胞，RT-qPCR和Western blot检测结果显示，与正常对照组比较，IWR-1组人牙髓干细胞中Wnt5a mRNA和Wnt5a、β-catenin、牙本质涎磷蛋白、牙本质基质蛋白1、骨唾液酸蛋白及骨钙蛋白的表达降低，差异有显著性意义(P < 0.05)；IWR-1+姜黄素组人牙髓干细胞中Wnt5a mRNA和Wnt5a、牙本质涎磷蛋白、牙本质基质蛋白1、骨唾液酸蛋白及骨钙蛋白的表达高于IWR-1组和正常对照组，差异有显著性意义(P < 0.05)。CCK-8检测结果显示，与正常对照组比较，IWR-1组人牙髓干细胞增殖活性降低，差异有显著性意义(P < 0.05)；与IWR-1组比较，IWR-1+姜黄素组人牙髓干细胞增殖活性增高，差异有显著性意义(P < 0.05)，见图3。以上结果提示姜黄素通过激活Wnt信号通路促进人牙髓干细胞的增殖与成牙本质分化。"

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