de Araujo Rui Mauricio Santos, Oba Yasuo, Kuroda Shingo, Tanaka Eiji, Moriyama Keiji
Department of Orthodontics and Dentofacial Orthopedics, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto-cho, Tokushima 770-8504, Japan.
Department of Orthodontics and Dentofacial Orthopedics, Institute of Health Biosciences, The University of Tokushima Graduate School, 3-18-15 Kuramoto-cho, Tokushima 770-8504, Japan.
Arch Oral Biol. 2014 Feb;59(2):187-92. doi: 10.1016/j.archoralbio.2013.11.010. Epub 2013 Nov 25.
RhoE and regulator of G-proteins signalling (RGS) 2 were identified as the up-regulated genes in human periodontal ligament (PDL) cells under compression. RhoE belongs to the Rho GTPase family, and RGS2, a novel family of GTPase-activating proteins, turns off the G-protein signalling. Rho family proteins have recently been known to regulate actin cytoskeleton dynamics in various cell types. In this study, we investigated the involvement of RhoE and RGS2 in the regulation of actin filament organization in the PDL cells under mechanical stress.
Human PDL cells were cultured and subjected to a static compressive force (3.0g/cm(2)) for 48h. To observe changes in the actin cytoskeleton and the expression of RhoE and RGS2 in response to mechanical stress, immunofluorescence analysis was performed. To examine the role of RhoE and RGS2 in actin filament organization, cells were transfected with antisense S-oligonucleotides (ODNs) to RhoE and RGS2.
Compressive force caused a loss and disassembly of actin stress fibres leading to cell spreading. Immunocytochemical study revealed that RhoE and RGS2 expressions were induced by mechanical stress and localized in the perinuclear and in the cell membrane, respectively. The impaired formation of stress fibres caused by compressive forces was recovered by treatment with antisense S-ODN to RhoE to the control levels. However, addition of antisense S-ODN to RGS2 did not affect the stress fibre formation.
These results indicate that the loss and disassembly of stress fibres due to mechanical stress are mediating RhoE signalling, without the exertion of RGS2.
RhoE和G蛋白信号调节剂（RGS）2被鉴定为受压人牙周膜（PDL）细胞中上调的基因。RhoE属于Rho GTP酶家族，而RGS2是一种新型的GTP酶激活蛋白家族，可关闭G蛋白信号传导。最近已知Rho家族蛋白可调节多种细胞类型中的肌动蛋白细胞骨架动力学。在本研究中，我们调查了RhoE和RGS2在机械应力下对PDL细胞中肌动蛋白丝组织调节的参与情况。
培养人PDL细胞并对其施加3.0g/cm²的静态压缩力48小时。为了观察肌动蛋白细胞骨架的变化以及RhoE和RGS2在机械应力响应中的表达，进行了免疫荧光分析。为了研究RhoE和RGS2在肌动蛋白丝组织中的作用，用针对RhoE和RGS2的反义硫代寡核苷酸（ODN）转染细胞。
压缩力导致肌动蛋白应力纤维丧失和解聚，导致细胞铺展。免疫细胞化学研究表明，RhoE和RGS2的表达由机械应力诱导，分别定位于核周和细胞膜。用针对RhoE的反义硫代ODN处理可将压缩力引起的应力纤维形成受损恢复到对照水平。然而，添加针对RGS2的反义硫代ODN并不影响应力纤维的形成。
这些结果表明，机械应力导致的应力纤维丧失和解聚是由RhoE信号介导的，而RGS2未发挥作用。
