Klf10沉默抑制机械力诱导的人牙周膜细胞成骨向分化

doi:10.19439/j.sjos.2017.06.006

上海口腔医学 ›› 2017, Vol. 26 ›› Issue (6): 599-604.doi: 10.19439/j.sjos.2017.06.006

Klf10沉默抑制机械力诱导的人牙周膜细胞成骨向分化

余小琴¹, 李云龙¹, 高锦瑜², 刘华¹

1.安康市人民医院口腔科,陕西安康 725000;
2.延安大学附属医院口腔医院正畸科,陕西延安 716000

收稿日期:2017-06-16 修回日期:2017-08-16 出版日期:2017-12-20 发布日期:2018-01-09
通讯作者: 高锦瑜,E-mail: jinyujyya@163.com
作者简介:余小琴（1980-）,女,学士,主治医师,E-mail: xiaoqinakkq@qq.com

Klf10 silencing inhibited mechanical force induced osteogenic differentiation of human periodontal ligament cells

YU Xiao-qin¹, LI Yun-long¹, Gao Jin-yu², LIU Hua¹

1.Department of Stomatology, Ankang People's Hospital. Ankang 725000;
2.Department of Orthodontics, Stomatological Hospital of Yanan University Affiliated Hospital. Yanan 716000, Shaanxi Province, China

Received:2017-06-16 Revised:2017-08-16 Online:2017-12-20 Published:2018-01-09

摘要/Abstract

摘要： 目的: 探讨Klf10沉默对人牙周膜细胞（human periodontal ligament cells,HPDLCs）受力后骨向分化的影响。方法: 分离培养HPDLCs,RNA干扰沉默Klf10的表达,用离心加力法对细胞加载机械力,并予hedgehog信号通路特异性激动剂purmorphamine进行干预。采用ELISA法检测碱性磷酸酶（alkaline phosphatase,ALP）活性,采用RT-PCR和Western印迹法分别检测各组细胞Klf10 mRNA和蛋白表达水平,并检测成骨标志基因Runt相关转录因子2（runt-related transcription factor 2,Runx2）、骨桥蛋白（osteopontin,OPN）和骨钙素（osteocalcin,OCN）mRNA和蛋白的表达情况;利用Western 免疫印迹法检测hedgehog信号转导通路成员胶质瘤相关癌基因同源物1（glioma-associated oncogene homolog 1,GLI1）和Ptch1（patched-1）的蛋白表达。采用SPSS 20.0软件包对数据进行统计学分析。结果: 机械力诱导6 h后,Klf10、Runx2、OPN和OCN mRNA和蛋白的表达显著升高,ALP活性升高,GLI1和Ptch1蛋白表达升高（P<0.05）。与空白组相比,转染Klf10 siRNA组Klf10 mRNA和蛋白水平显著降低（P<0.05）。同时,Klf10 siRNA显著抑制ALP活性,下调Runx2、OPN和OCN mRNA和蛋白的表达,抑制GLI1和Ptch1蛋白表达（P<0.05）。Purmorphamine显著抑制Klf10 siRNA沉默介导的成骨分化效应（P<0.05）。结论: Klf10沉默可以抑制机械力作用下HPDLCs的骨向分化,可能与调节hedgehog信号转导通路有关。

关键词: 口腔正畸, Klf10, 人牙周膜细胞, 机械力, 成骨分化

Abstract: PURPOSE: To investigate the effect of Klf10 silence on human periodontal ligament cells (HPDLCs) under mechanical force. METHODS: HPDLCs were isolated and transfected with Klf10 siRNA, and then exposed to centrifugal force for 6 h at 631 r/min. Purmorphamine, an hedgehog signaling pathway agonist, was used for intervention. The activity of alkaline phosphatase (ALP) was detected by ELISA. RT-PCR and Western blot were performed to detect the mRNA and protein expression of Klf10, Runt related transcription factor 2 (Runx2), osteopontin (OPN) and osteocalcin (OCN). The protein expression of glioma associated oncogene homolog 1 (Gli1) and patched-1 (PTCH1) was detected by Western blot. SPSS 20.0 software package was used to analyze the data. RESULTS: Mechanical force increased mRNA and protein level of Klf10, Runx2, OPN and OCN, and elevated ALP activity significantly (P<0.05). Mechanical force also upregulated the protein expression of GLI1 and Ptch1 significantly (P<0.05). Compared with the control group, Klf10 siRNA transfection significantly decreased mRNA and protein level of Klf10 (P<0.05). Klf10 siRNA significantly inhibited the activity of ALP, and downregulated mRNA and protein expression of Runx2, OPN and OCN (P<0.05). Moreover, Klf10 siRNA significantly inhibited protein expression of GLI1 and Ptch1 (P<0.05), and purmorphamine obviously inhibited the effect of Klf10 siRNA (P<0.05). CONCLUSIONS: Klf10 silencing could inhibit bone differentiation of human periodontal ligament cells under mechanical force, which may be through regulation of hedgehog signaling pathway.

Key words: Orthodontics, Klf10, Human periodontal ligament cells, Mechanical force, Osteogenic differentiation

中图分类号:

R783.5

余小琴, 李云龙, 高锦瑜, 刘华. Klf10沉默抑制机械力诱导的人牙周膜细胞成骨向分化[J]. 上海口腔医学, 2017, 26(6): 599-604.

YU Xiao-qin, LI Yun-long, Gao Jin-yu, LIU Hua. Klf10 silencing inhibited mechanical force induced osteogenic differentiation of human periodontal ligament cells[J]. Shanghai Journal of Stomatology, 2017, 26(6): 599-604.

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Klf10沉默抑制机械力诱导的人牙周膜细胞成骨向分化

Klf10 silencing inhibited mechanical force induced osteogenic differentiation of human periodontal ligament cells

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