Andriamanalijaona R, Felisaz N, Kim S-J, King-Jones K, Lehmann M, Pujol J-P, Boumediene K
Laboratory of Connective Tissue Biochemistry, Caen, France.
Arthritis Rheum. 2003 Jun;48(6):1569-81. doi: 10.1002/art.11020.
Interleukin-1 (IL-1) and transforming growth factor beta1 (TGFbeta1) play major roles in osteoarticular diseases, exerting opposite effects on both the catabolism and anabolism of cartilage matrix. Previous findings suggest that IL-1 and TGFbeta1 could function in a feedback interaction. However, the effect exerted by IL-1 on expression of TGFbeta by articular chondrocytes is, so far, poorly understood. The present study was carried out to determine the influence of IL-1beta on the expression of TGFbeta1 by bovine articular chondrocytes (BACs) in primary culture.
BAC primary cultures were treated with IL-1beta, and TGFbeta1 messenger RNA (mRNA) steady-state levels and protein expression were measured by real-time reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. Transient transfection of TGFbeta1 gene promoter constructs was performed to delineate the DNA sequences that mediate the IL-1beta effect. Electrophoretic mobility shift assays (EMSAs) and supershift analysis were used to characterize the transcription factors binding to these sequences.
Cultured BACs responded to IL-1beta exposure by exhibiting an increase of TGFbeta1 expression at both the mRNA and protein levels. The effect was found to be mediated by a major 80-bp sequence located between -732 and -652 upstream of the transcription initiation site. EMSA and supershift analysis revealed that the transcription factors activator protein 4 (AP-4) and AP-1 specifically bound to the -720/-696 part of this sequence under IL-1beta treatment. Overexpression of AP-4 in the BAC cultures resulted in stimulation of the transcriptional activity of the -732/+11 TGFbeta1 promoter construct through the same IL-1beta-responsive element.
IL-1beta induces an increase of TGFbeta1 in articular chondrocytes through activation of AP-4 and AP-1 binding to the TGFbeta1 gene promoter. These findings may help us understand the role of IL-1beta in the disease process. Notwithstanding its deleterious effect on cartilage, IL-1 could initiate the repair response displayed by injured cartilage in the early stages of osteoarthritis through its ability to enhance TGFbeta1 expression by local chondrocytes.
白细胞介素-1（IL-1）和转化生长因子β1（TGFβ1）在骨关节疾病中起主要作用，对软骨基质的分解代谢和合成代谢产生相反的影响。先前的研究结果表明，IL-1和TGFβ1可能在反馈相互作用中发挥作用。然而，迄今为止，IL-1对关节软骨细胞中TGFβ表达的影响尚不清楚。本研究旨在确定IL-1β对原代培养的牛关节软骨细胞（BACs）中TGFβ1表达的影响。
用IL-1β处理BAC原代培养物，分别通过实时逆转录-聚合酶链反应和酶联免疫吸附测定法测量TGFβ1信使核糖核酸（mRNA）的稳态水平和蛋白质表达。进行TGFβ1基因启动子构建体的瞬时转染，以确定介导IL-1β作用的DNA序列。采用电泳迁移率变动分析（EMSA）和超迁移分析来表征与这些序列结合的转录因子。
培养的BACs对IL-1β刺激的反应是在mRNA和蛋白质水平上TGFβ1表达均增加。发现该效应由位于转录起始位点上游-732至-652之间的一个主要80碱基对序列介导。EMSA和超迁移分析显示，在IL-1β处理下，转录因子激活蛋白4（AP-4）和AP-1特异性结合该序列的-720/-696部分。在BAC培养物中过表达AP-4导致通过相同的IL-1β反应元件刺激-732/+11 TGFβ1启动子构建体的转录活性。
IL-1β通过激活AP-4和AP-1与TGFβ1基因启动子的结合，诱导关节软骨细胞中TGFβ1增加。这些发现可能有助于我们理解IL-1β在疾病过程中的作用。尽管IL-1对软骨有有害作用，但它可以通过增强局部软骨细胞中TGFβ1表达的能力，在骨关节炎早期启动受损软骨的修复反应。
