Gene expression profiling of IL-1β-stimulated synovial cells from TMJ using Gene Chip

使用基因芯片对 IL-1β 刺激的 TMJ 滑膜细胞进行基因表达谱分析

基本信息

批准号：
14571915
负责人：
OGURA Naomi
金额：
$ 2.24万
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：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

Interleukin-1β(IL-1β) has important roles in the inflammation and connective tissue destruction observed in joint diseases such as rheumatoid arthritis. IL-1β is also a key mediator of intracapsular pathologic conditions of the temporomandibular joint(TMJ), including disk displacement/internal derangement and osteoarthritis. To identify putative IL-1β-responsive genes from arthritic disease tissues, we investigated the effects of IL-1β on gene expression of 8,793 genes in synovial fibroblasts from five TMJ patients. <Methods>Synovial fibroblasts were obtained using an outgrowth method with synovial tissue from a patient who underwent arthrotomy of the TMJ. Total RNA of synovial fibroblasts, incubated with or without i□unit/ml of IL-1β for 4 hours, was extracted using the RNeasy kit. Gene expression pro filing was used GeneChip (HG Focus Array,8,763 genes).Hybridization data were analyzed using Gene Springs^<TM> software. Gene expression levels were also confirmed by real time-PCR. <results>Significant changes between control and IL-1β-treated cells (p<0.05) were detected in 170 genes:139 up-regulated genes and 31 down-regulated genes. The most enhanced gene by IL-1β was CCL20, which is one of chemokines. The 170 IL-1β-responsive genes included 9 chemokines. Real-time PCR confirms that mRNA levels of these chemokines increase after IL-1β treatment. In addition, the effects of IL-1β were investigated using Ingenuity Pathway Analysis. We found that IL-1β affects the expression of several genes in the NFκB signaling pathway and were able to confirm that mRNA levels of NFκB1 and IκBα increase after IL-1β treatment by endpoint-PCR and real time-PCR analysis. The results suggest that IL-1β-responsive genes play important roles in progression of inflammation and destruction of joint components.

白介素-1β（IL-1β）在炎症和连接的组织破坏中具有重要作用，在类风湿关节炎等关节疾病中观察到。 IL-1β也是颞下颌关节（TMJ）的囊内病理状况的关键介体，包括椎间盘位移/内部进化和骨关节炎。为了从关节炎疾病时机中鉴定出假定的IL-1β反应性基因，我们研究了IL-1β对来自五名TMJ患者的滑膜成纤维细胞中8,793个基因基因表达的影响。 <methods>滑膜成纤维细胞是使用带有滑膜组织进行TMJ手术的滑膜组织的出生方法获得的。使用RNeasy试剂盒提取了带或不带I□单位/mL IL-1β的单位组合成纤维细胞的总RNA，持续4小时。使用Gene表达Pro归档（HG焦点阵列，8,763个基因）。使用基因弹簧^<tm>软件分析了杂交数据。基因表达水平也通过实时PCR证实。在170个基因中检测到对照和IL-1β处理的细胞之间的显着变化（P <0.05）：139个上调的基因和31个下调的基因。 IL-1β最大增强的基因是CCL20，它是趋化因子之一。 170 IL-1β反应基因9趋化因子。实时PCR证实IL-1β处理后这些趋化因子的mRNA水平升高。另外，使用Ingenuity途径分析研究了IL-1β的效果。我们发现IL-1β会影响NFκB信号通路中几个基因的表达，并能够确认通过端点PCR和实时PCR分析，IL-1β处理后NFκB1和IκBα的mRNA水平升高。结果表明，IL-1β响应基因在注射和破坏关节成分方面起着重要作用。