Mediation of interleukin-1beta-induced transforming growth factor beta1 expression by activator protein 4 transcription factor in primary cultures of bovine articular chondrocytes: possible cooperation with activator protein 1.

ABSTRACT

OBJECTIVE: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.