机械力作用下人牙周膜细胞Osterix mRNA和蛋白的表达

Expression of Osterix mRNA and protein levels in cultured human periodontal ligament cells after mechanical stimulation

目的研究在机械力作用下人牙周膜细胞内Osterix(Osx) mRNA和蛋白的表达变化,探讨Osx与正畸牙周组织骨改建的关系。方法组织块法培养人牙周膜细胞,采用离心加力装置对细胞分别加载1、2、4、6、8、12h的机械力。逆转录聚合酶链反应(RT-PCR)、Western blot及细胞免疫荧光化学技术分别检测不同时间点Osx mRNA和蛋白的表达变化及其细胞定位。结果在正常人牙周膜细胞中,Osx mRNA表达微弱,蛋白未见表达;在机械力加载4h后,Osx mRNA表达开始明显增强(P<0.01),蛋白呈现弱表达(P<0.05);加载8h时,OsxmRNA和蛋白表达显著增强(P<0.01);持续增加至加力12h。同时,加力4h后,少量细胞的胞质内开始呈现微弱的绿色荧光;12h后,阳性表达主要集中在胞核内。结论机械力可诱导人牙周膜细胞Osx表达增强及活化。Osx可能参与了细胞内生物力学信号的转导,从而可能在正畸牙周组织的骨改建过程中发挥着重要作用。

Objective To examine the expression of Osterix（Osx）mRNA and protein after application of mechanical force on human periodontal ligament cells（HPDLCs）, and to investigate the role of Osx in orthodontic alveolar bone remodeling. Methods HPDLCs were isolated and cultured in vitro with explant method. Approximately 2.5×105 cells were seeded onto six-well cell culture plates and then were exposed to centrifugal force for 1, 2, 4, 6, 8 or 12 h at 631 r.min-1. The expression of Osx mRNA and protein was measured by reverse transcription-polymerase chain raction （RT-PCR）and Western blot respectively. Immunofluorescence assay was used to detect the expression and subcellular localization of Osx protein by green fluorescence. Results At the initial time point, Osx mRNA had a weak expression and protein was not detected. Under the mechanical stimulation, both mRNA and protein levels of Osx were upregulated in a time-dependent manner. Furthermore, Osx protein was translocated gradually from the cytosol into the cell nuclei. Conclusion The expression and activation of Osx were enhanced by mechanical stress in HPDLCs, which indicates that Osx may play an important role in HPDLCs osteogenic differentiation and periodontal tissue remodeling induced by mechanical stress.