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

作　　者：余小琴 李云龙 高锦瑜[2] 刘华 YU Xiao-qin;LI Yun-long;Gao Jin-yu;LIU Hua(Department of Stomatology, A nkang People ＂s Hospital. Ankang 725000;Department of Orthodontics, Stomatological Hospital of Yanan University Affiliated Hospital. Yanan 716000, Shaanxi Province, China)

机构地区：[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

出　　处：《上海口腔医学》2017年第6期599-604,共6页Shanghai Journal of Stomatology

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 m RNA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)m RNA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 m RNA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 si RNA组Klf10 m RNA和蛋白水平显著降低(P<0.05)。同时,Klf10 si RNA显著抑制ALP活性,下调Runx2、OPN和OCN m RNA和蛋白的表达,抑制GLI1和Ptch1蛋白表达(P<0.05)。Purmorphamine显著抑制Klf10 si RNA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。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.

关 键 词：口腔正畸 Klf10 人牙周膜细胞 机械力 成骨分化 

分 类 号：R783.5[医药卫生—口腔医学]