Bidirectional ephrinB2-EphB4 signaling regulates the osteogenic differentiation of canine periodontal ligament stem cells
Bidirectional ephrinB2-EphB4 signaling regulates the osteogenic differentiation of canine periodontal ligament stem cells
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双向ephrinB2-EphB4信号调节犬牙周膜干细胞的成骨分化
DOI：
10.3892/ijmm.2020.4473
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发表时间：
2020-01
影响因子：
5.4
通讯作者：
Gu Min
中科院分区：
文献类型：
--
作者：
Zhu Shaoyue;Liu Zongxiang;Yuan Changyong;Lin Yifan;Yang Yanqi;Wang Haiming;Zhang Chengfei;Wang Penglai;Gu Min
本研究旨在探讨ephrinB 2基因转染的犬牙周膜干细胞（cPDLSCs）对成骨分化的调控作用。用转基因空白对照绿色荧光蛋白（GFP）载体（称为载体-cPDLSC）或用NFNB 2 GFP-杀稻瘟素（称为EfnB 2-cPDLSC）转染cPDLSC。随后，通过逆转录-定量聚合酶链反应（RT-qPCR）、碱性磷酸酶（ALP）测定和茜素红S染色来评估Vector-cPDLSCs和EfnB 2-cPDLSCs的成骨分化。还评估了cPDLSC、Vector-cPDLSC和EfnB 2-cPDLSC的迁移能力。在Vector-cPDLSC和EfnB 2-cPDLSC的成骨诱导后，通过蛋白质印迹测定分析胶原蛋白I、Runt相关转录因子2、骨钙素、肝配蛋白B型受体4（EphB 4）、磷酸-EphB 4、肝配蛋白B2和磷酸肝配蛋白B2的蛋白质表达水平。RT-qPCR和Western blotting结果显示，EfnB 2-cPDLSCs中ephrinB 2的mRNA和蛋白表达水平均显著高于Vector-cPDLSCs（P0.05）。ALP和茜素红S染色测定分别显示EfnB 2-cPDLSC中ALP活性和矿化结节增加。细胞增殖和迁移试验显示，与Vector-cPDLSC相比，EfnB 2-cPDLSC表现出增强的增殖和迁移（P 0.05）。总之，本研究的结果表明，ephrinB 2基因修饰的cPDLSCs表现出增强的成骨分化，ephrinB 2反向信号传导和EphB 4正向信号传导途径在此过程中起关键作用。此外，ephrinB 2基因修饰可促进cPDLSCs的迁移和增殖。
The aim of the present study was to evaluate the effect of ephrinB2 gene-transfected canine periodontal ligament stem cells (cPDLSCs) on the regulation of osteogenic differentiation. cPDLSCs were transfected with a transgenic null-control green fluorescent protein (GFP) vector (termed Vector-cPDLSCs) or with NFNB2 GFP-Blasticidin (termed EfnB2-cPDLSCs). Subsequently, the osteogenic differentiation of Vector-cPDLSCs and EfnB2-cPDLSCs was assessed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), alkaline phosphatase (ALP) assay and Alizarin Red S staining. The migratory abilities of cPDLSCs, Vector-cPDLSCs and EfnB2-cPDLSCs were also assessed. Following osteogenic induction of Vector-cPDLSCs and EfnB2-cPDLSCs, the protein expression levels of collagen I, Runt-related transcription factor 2, osteocalcin, ephrin type-B receptor 4 (EphB4), phospho-EphB4, ephrinB2 and phosphoephrinB2 were analyzed by western blot assays. Following gene transfection, the RT-qPCR and western blotting results revealed that the mRNA and protein expression levels of ephrinB2, respectively, were significantly increased in EfnB2-cPDLSCs compared with that in Vector-cPDLSCs (P<0.05). ALP and Alizarin Red S staining assays revealed increased ALP activity and mineralization nodules, respectively, in EfnB2-cPDLSCs. Cell proliferation and migration assays revealed that EfnB2-cPDLSCs exhibited enhanced proliferation and migration compared with Vector-cPDLSCs (P<0.05). In conclusion, the findings of the current study indicated that ephrinB2 gene-modified cPDLSCs exhibited enhanced osteogenic differentiation, with the ephrinB2 reverse signaling and EphB4 forward signaling pathways serving a key role in this process. Furthermore, ephrinB2 gene modification was observed to promote the migration and proliferation of cPDLSCs.
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通讯作者：
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