南方医科大学学报

南方医科大学学报 ›› 2016, Vol. 36 ›› Issue (07): 984-.

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人甲状旁腺激素（1-34）在人成骨肉瘤细胞中对基质Gla蛋白及Wnt/β-catenin信号通路的调节作用

胡亚莉，张洁，符刘晨，杨雅   

  • 出版日期:2016-07-20 发布日期:2016-07-20

Effect of parathyroid hormone (1-34) on expression of matrix Gla protein and Wnt/β catenin signaling pathways in MG63 cell lines

  • Online:2016-07-20 Published:2016-07-20

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摘要： 目的观察人甲状旁腺激素（PTH）的N端活性片段PTH（1-34）对人成骨肉瘤细胞MG63 基质Gla 蛋白（MGP）表达的影 响，以及PTH通过Wnt/β-catenin 信号通路介导MGP表达调控的作用，探讨PTH（1-34）在防治骨质疏松症作用中可能的分子 机制。方法（1）用不同浓度PTH（1-34）（10-9、10-8、10-7 mol/L），单独或联合Wnt/β-catenin信号通路抑制剂DKK-1（200 ng/mL）干预 MG63 细胞；（2）检测碱性磷酸酶（ALP）染色及活力测定用于判断细胞分化情况；（3）MGP及Wnt/β-catenin 信号通路各组分 的mRNA及蛋白的表达分别采用实时定量RT-PCR 及Western blotting 方法。结果（1）PTH（1-34）上调MGP基因的表达， MGP mRNA的表达分别是对照组的2.56 倍、4.14 倍、7.81 倍（P<0.05 或P<0.01），且呈剂量依赖性增高；（2）PTH增强MG63 细胞ALP活力，Dkk-1 抑制ALP活性，Dkk-1 与PTH联用部分阻断PTH上调ALP活力（P<0.05）；（3）PTH上调MGP及Wnt/ β-catenin 信号通路中相关因子LRP5、β-catenin、Runx2 mRNA及蛋白水平，其mRNA分别是对照组的2.65、4.01、3.48 倍（P< 0.05 或<0.01）；DKK-1 对PTH（1-34）促MGP表达没有影响，但大部分阻断了Wnt/β-catenin 信号通路中相关因子的表达。结 论PTH（1-34）能明显上调MGP及Wnt/β-catenin 信号通路中相关因子的表达，Wnt/β-catenin 信号通路和MGP在PTH调节骨 代谢中起重要作用。

Abstract: Objective To observe the effect of parathyroid hormone (PTH)(1-34) on the expression of matrix Gla protein (MGP) and Wnt/β-catenin signaling pathway and elucidate the possible molecular mechanism of PTH (1-34) in the prevention and treatment of osteoporosis. Methods MG63 cells treated with PTH (1-34) at 10-9, 10-8, and 10-7 mol/L, alone or in combination with Wnt/beta-catenin signaling pathway inhibitors DKK-1 (200 ng/ml) were examined for mRNA and protein expressions related with Wnt/β-catenin signaling with real-time PCR and Western blotting. The cell differentiation after the treatment was assessed with alkaline phosphatase (ALP) staining and cell viability assay. Results PTH (1-34) significantly increased the expression of MGP in a dose-dependent manner in MG63 cells (P<0.05 or P<0.01). PTH treatment obviously enhanced ALP activity in the cells, and this effect was suppressed by DKK-1. Combined treatment with DKK-1 partially blocked PTH-induced enhancement of ALP activity (P<0.05). PTH promoted the expression of MGP and enhanced LRP5, β-catenin, and Runx2 expressions in Wnt/β-catenin signaling pathway at both protein and mRNA levels (P<0.05 or P<0.01). DKK-1 partially blocked the effect of PTH (1-34) on Wnt/β-catenin signaling pathway (P<0.05) without affecting MGP expression. Conclusion PTH (1-34) significantly increases the expressions of MGP and proteins in the Wnt/β-catenin signaling pathway. Wnt/β-catenin signaling pathway and MGP mediate the regulation of osteogenosis by PTH.