Molecular biological analysis of bone metabolism by compressive mechanical stress in human synovial cells of the temporomandibular joint

人颞下颌关节滑膜细胞压缩机械应力对骨代谢的分子生物学分析

基本信息

批准号：
18592196
负责人：
TAKANO Hiroshi
金额：
$ 1.9万
依托单位：
Akita University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18592196/
关键词：
dentistry cell tissue mechanical stress

项目摘要

1. OBJECTIVES: We investigated the effects of compressive mechanical stress on osteoclastogenesis of synovial cells to clarify the mechanism of osteoclast formation by those cells in temporomandibular joint (TMJ) disorders. STUDY DESIGN: Synovial cells were isolated from rat knee joints and continuously compressed using a conventional method. The expression of receptor activator nuclear factor kappaB ligand (RANKL) mRNA and protein in synovial cells was analyzed by reverse transcriptase-polymerase chain reaction, immunoblotting, and immunofluorescence staining. Mouse bone marrow cells were cultured with synovial cells for 7 days to detect osteoclasts. RESULTS: The expressions of RANKL mRNA and protein in synovial cells were increased with compressive force. When mouse bone marrow cells were cultured with continuously compressed synovial cells, tartrate-resistant acid phosphatase-positive multinucleated cells were formed. Osteoprotegerin completely inhibited osteoclast formation induced … More by culturing with compressed synovial cells. CONCLUSION: Our results indicated that the expression of RANKL in compressed synovial cells enhanced osteoclast formation, whereas continuous compressive force may induce osteoclastic bone destruction in the TMJ.2. OBJECTIVE: Although biochemical studies have examined the synovial fluid (SF) of patients with temporomandibular joint (TMJ) disorders (TMDs), the details of the molecular mechanism of bone destruction and remodeling remain unknown. In this study, we induced and characterized osteoclast-like cells from the SF of patients with TMD and investigated the participation of these cells in the pathogenesis of TMD. METHODS: We collected SF cells from patients with TMD after a pumping procedure, cultured osteoclast-like cells, and examined their characteristics, including osteoclast markers and bone resorption activities. In addition, we obtained fibroblastic cells from the SF of TMD patients by continuous sub-culturing. Using these fibroblastic cells, we examined fibroblast markers using immunocytochemical staining and analyzed the receptor activator of nuclear-factor-kappaB ligand (RANKL) mRNA levels. Detection of soluble form of RANKL (sRANKL) in the SF was measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: Osteoclast-like cells were induced from the SF cells of patients with TMD by adding recombinant human (rh) macrophage colony stimulating factor (M-CSF) and either 1, 25-dihydroxy vitamin D3 [1, 25 (OH) 2D3] or prostaglandin E2 (PGE2). These multinucleated giant cells were positive for tartrate-resistant acid phosphatase (TRAP) and had the ability to absorb bone. The fibroblastic cells from the SF of TMD patients were positive for fibroblast markers and RANKL mRNA was up-regulated. Detection of sRANKL in SF of patient group was significantly higher than control group. CONCLUSION: The results suggest that the joint-infiltrating SF cells from TMD patients play important roles in the pathogenesis of these disorders, which is characterized by progressive bone destruction or remodeling. Less

1。目标：我们研究了压缩机械应力对滑膜细胞破骨细胞发生的影响，以阐明颞下颌关节（TMJ）疾病中这些细胞形成破骨细胞的机制。研究设计：从大鼠膝关节中分离出滑细胞，并使用常规方法连续压缩。通过逆转录酶 - 聚合酶链反应，免疫印迹和免疫荧光染色，分析了滑膜细胞中受体激活核因子Kappab配体（RANKL）mRNA和蛋白质中的表达。将小鼠骨髓细胞用滑膜细胞培养7天，以检测破骨细胞。结果：滑膜细胞中RANKL mRNA和蛋白质的表达通过压缩力增加。当小鼠骨髓细胞与连续压缩滑膜细胞培养时，形成了耐锈酸磷酸酶阳性多核细胞。骨蛋白蛋白蛋白蛋白完全抑制破骨细胞的形成……更多通过与压缩滑膜聚类聚类。结论：我们的结果表明，压缩滑膜中RANKL的表达增强了破骨细胞的形成，而连续的压缩力可能会诱导TMJ.2中的破骨细胞破坏。目的：尽管生化研究已经检查了颞下颌关节（TMJ）疾病（TMD）患者的滑液（SF），但骨破坏和重塑的分子机制的细节仍然未知。在这项研究中，我们诱导并表征了来自TMD患者SF的破骨细胞样细胞，并研究了这些细胞参与TMD的发病机理。方法：我们在抽水手术后，从TMD患者，培养的破骨细胞样细胞中收集了SF细胞，并检查了其特征，包括破骨细胞标记和骨修复活动。此外，我们通过连续亚培养从TMD患者的SF中获得了成纤维细胞。使用这些成纤维细胞细胞，我们使用免疫细胞化学染色检查了成纤维细胞标记，并分析了核因子-Kappab配体（RANKL）mRNA水平的受体激活剂。通过酶联免疫吸附测定（ELISA）测量SF中RANKL（SRANKL）的固体形式的检测。结果：通过添加重组人（RH）巨噬细胞刺激因子（M-CSF）和1、25-二羟基维生素D3 [1，25（OH）2D3]或Prostaglandin E2（PGE2），通过添加重组人（RH）巨噬细胞刺激因子（M-CSF）和1，25-二羟基羟基维生素D3 [1，25-二羟基维生素D3），从TMD患者的SF细胞中诱导破骨细胞样细胞。这些多核巨细胞对耐锈蚀酸磷酸酶（TRAP）呈阳性，并具有吸收骨骼的能力。来自TMD患者SF的成纤维细胞的成纤维细胞标记为阳性，RANKL mRNA被上调。患者组SF中SRANKL的检测显着高于对照组。结论：结果表明，来自TMD患者的联合浸入SF细胞在这些疾病的发病机理中起着重要作用，这是表征的。通过进行性骨骼破坏或重塑。较少的