Inhibition of osteoclast bone resorption activity through osteoprotegerin-induced damage of the sealing zone.
Inhibition of osteoclast bone resorption activity through osteoprotegerin-induced damage of the sealing zone.
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DOI：
10.3892/ijmm.2014.1846
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发表时间：
2014-09
影响因子：
5.4
通讯作者：
R. Song;Jianhong Gu;Xuezhong Liu;Jiaqiao Zhu;Qi-chang Wang;Q. Gao;Jiaming Zhang;L. Cheng;Xishuai Tong;Xinyi Qi;Yan Yuan;Zongping Liu
中科院分区：
文献类型：
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作者：
R. Song;Jianhong Gu;Xuezhong Liu;Jiaqiao Zhu;Qi-chang Wang;Q. Gao;Jiaming Zhang;L. Cheng;Xishuai Tong;Xinyi Qi;Yan Yuan;Zongping Liu
骨重建取决于破骨细胞介导的骨吸收和成骨细胞介导的骨生成之间的动态平衡。封闭区是破骨细胞特异性的细胞骨架结构，其完整性对于破骨细胞介导的骨吸收至关重要。迄今为止，研究主要集中在骨保护素（OPG）通过OPG/核因子κ B配体（RANKL）/RANK系统的受体激活剂诱导的破骨细胞分化抑制，该系统影响破骨细胞的骨吸收。然而，OPG对密封区的影响迄今尚未报道。在这项研究中，通过霍夫曼调制对比（HMC）显微镜和共聚焦激光扫描显微镜观察密封区的形成。通过免疫荧光染色、HMC显微镜、定量逆转录聚合酶链反应（RT-qPCR）、Western印迹分析和扫描电镜检查OPG对现有封闭带和破骨细胞介导的骨吸收活性的影响，以及参与封闭带形成的基因的调节作用。第5天形成封闭带，细胞边缘有带状突起，细胞核散在分布，无丝状伪足。未处理对照组的凝闭区完整。然而，在OPG处理组（20 ng/ml）中观察到密封区缺陷，在40和80 ng/ml OPG处理组中不存在该结构。在破骨细胞的基底面和孔面之间呈散在或成簇分布。此外，在20 ng/ml OPG处理组中未检测到吸收陷窝，表明破骨细胞介导的骨吸收活性丧失。OPG处理导致Arhgef 8/Net 1和DOCK 5 Rho鸟嘌呤核苷酸交换因子（RhoGEFs）、18种RhoGTP酶中的10种（RhoA、RhoB、cdc 42 v1、cdc 42 v2、RhoU/RhoCh 1、RhoF/Rif、Rac 2、RhoG、Rnd 1和RhoBTB 1）、ROCK 1和ROCK 2的表达显著降低。总之，足体分布受到OPG诱导的RhoGT信号通路基因表达抑制的影响。这导致密封区的损伤或破坏，从而抑制破骨细胞介导的骨吸收活性。
Bone remodeling is dependent on the dynamic equilibrium between osteoclast-mediated bone resorption and osteoblast-mediated osteogenesis. The sealing zone is an osteoclast-specific cytoskeletal structure, the integrity of which is critical for osteoclast-mediated bone resorption. To date, studies have focused mainly on the osteoprotegerin (OPG)‑induced inhibition of osteoclast differentiation through the OPG/receptor activator of the nuclear factor kappa-B ligand (RANKL)/RANK system, which affects the bone resorption of osteoclasts. However, the effects of OPG on the sealing zone have not been reported to date. In this study, the formation of the sealing zone was observed by Hoffman modulation contrast (HMC) microscopy and confocal laser scanning microscopy. The effects of OPG on the existing sealing zone and osteoclast-mediated bone resorption activity, as well as the regulatory role of genes involved in the formation of the sealing zone were examined by immunofluorescence staining, HMC microscopy, quantitative reverse transcription polymerase chain reaction (RT-qPCR), western blot analysis and scanning electron microscopy. The sealing zone was formed on day 5, with belt-like protuberances at the cell edge and scattered distribution of cell nuclei, but no filopodia. The sealing zone was intact in the untreated control group. However, defects in the sealing zone were observed in the OPG-treated group (20 ng/ml) and the structure was absent in the groups treated with 40 and 80 ng/ml OPG. The podosomes showed a scattered or clustered distribution between the basal surface of the osteoclasts and the well surface. Furthermore, resorption lacunae were not detected in the 20 ng/ml OPG-treated group, indicating the loss of osteoclast-mediated bone resorption activity. Treatment with OPG resulted in a significant decrease in the expression of Arhgef8/Net1 and DOCK5 Rho guanine nucleotide exchange factors (RhoGEFs), 10 of 18 RhoGTPases (RhoA, RhoB, cdc42v1, cdc42v2, RhoU/Wrch1, RhoF/Rif, Rac2, RhoG, Rnd1 and RhoBTB1), ROCK1 and ROCK2. In conclusion, podosome distribution was affected by the OPG-induced inhibition of the expression of genes in the RhoGTPase signaling pathway. This resulted in damage to or destruction of the sealing zone, thus inhibiting osteoclast-mediated bone resorption activity.