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Characterization of cultured synovial cells from human TMJ

人颞下颌关节培养滑膜细胞的表征

基本信息

批准号：
11672017
负责人：
OGURA Naomi
金额：
$ 2.37万
依托单位：
Nihon University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11672017/
关键词：
Cultured synovial cells TMJ IL-1β TNF-α Plasminogen activator IL-6 IL-8 MCP-1 顎関節症ヒト顎関節滑膜細胞

项目摘要

The aims of the present study were, firstly, to isolate and to characterize culture synovial cells obtained from human TMJ specimens, and secondly, to investigate the productions of cytokines and enzymes in those cells.<Methods> Synovial cells were isolated from TMJ synovial tissue using an out growth method and then cultured in Ham F12 containing 20% or 10% fetal bovine serum. Synovial cells were examined cell-specific markers using immunocytochemistry. In the next, synovial cells were treated with IL-1β or TNF-α, which was increased in synovial fluids from patients with TMJ disorders. Activity of plasminogen activator (PA), which is associated with extracellular matrix degradation, was measured by enzyme assay. Amounts of IL-6, IL-8 and macrophage attractant protein (MCP)-1 were measured by ELISA. Gene expressions of these cytokines were also investigated using RTPCR method.<Rersults> Synovial cells were positive for fibroblast-specific marker and vimentin. In contrast, the cells were negative for macrophage marker, HLA class II and dendritic cell marker. Then, synovial cells were treated with or without IL-1β. IL-1β increased IL-6, IL-8 and MCP-1 production in synovial cells in a time- and dose-dependent manner. IL-1β stimulated gene expression of these cytokines in them. IL-1β also enhanced PA activity in synovial cells. In the next, the cells were treated with TNF-α. TNF-α enhanced productions and gene expressions of IL-8 and MCP-1.<Conclusions> We found that cultured synovial cells might provide important advantages for studies of the cellular and molecular mechanisms in TMJ. In addition, PA activity and IL-6, IL-8, and MCP-1 productions may be increased in synovial cells when IL-1β and TNF-α levels are increased in synovial fluids from patients with TMJ disorders.

本研究的目的首先是对从人颞下颌关节标本中获得的滑膜细胞进行分离和表征，其次是研究这些细胞中细胞因子和酶的产生。<方法>采用外生长法从TMJ滑膜组织中分离滑膜细胞，然后在含有20%或10%胎牛血清的Ham F12中培养。使用免疫细胞化学检查滑膜细胞的细胞特异性标记。接下来，用 IL-1β 或 TNF-α 处理滑膜细胞，TMJ 疾病患者滑液中的 IL-1β 或 TNF-α 含量有所增加。通过酶测定来测量与细胞外基质降解相关的纤溶酶原激活剂（PA）的活性。通过ELISA测量IL-6、IL-8和巨噬细胞吸引蛋白(MCP)-1的量。采用RTPCR方法检测这些细胞因子的基因表达。<结果>滑膜细胞成纤维细胞特异性标志物和波形蛋白呈阳性。相反，细胞的巨噬细胞标记、HLA II 类和树突状细胞标记呈阴性。然后，用或不用IL-1β处理滑膜细胞。 IL-1β以时间和剂量依赖性方式增加滑膜细胞中IL-6、IL-8和MCP-1的产生。 IL-1β 刺激其中这些细胞因子的基因表达。 IL-1β 还增强滑膜细胞中的 PA 活性。接下来，用 TNF-α 处理细胞。 TNF-α增强IL-8和MCP-1的产生和基因表达。<结论>我们发现培养的滑膜细胞可能为研究TMJ的细胞和分子机制提供重要的优势。此外，当 TMJ 疾病患者滑液中的 IL-1β 和 TNF-α 水平增加时，滑膜细胞中的 PA 活性以及 IL-6、IL-8 和 MCP-1 的产生可能会增加。