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Transforming growth factor beta (TGF$\beta$) plays a central role in morphogenesis, growth, and cell differentiation. This cytokine is particularly important in cartilage where it regulates cell proliferation and extracellular matrix synthesis. While the action of TGF$\beta$ on chondrocyte metabolism has been extensively catalogued, the modulation of specific genes that function as mediators of TGF$\beta$ signalling is poorly defined. In the current study, elements of the Smad component of the TGF$\beta$ intracellular signalling system and TGF$\beta$ receptors were characterised in human chondrocytes upon TGF$\beta$1 treatment. Human articular chondrocytes were incubated with TGF$\beta$1. Then, mRNA and protein levels of TGF$\beta$ receptors and Smads were analysed by RT-PCR and western blot analysis. The role of specific protein 1 (Sp1) was investigated by gain and loss of function (inhibitor, siRNA, expression vector). We showed that TGF$\beta$1 regulates mRNA levels of its own receptors, and of Smad3 and Smad7. It modulates TGF$\beta$ receptors post-transcriptionally by affecting their mRNA stability, but does not change the Smad-3 and Smad-7 mRNA half-life span, suggesting a potential transcriptional effect on these genes. Moreover, the transcriptional factor Sp1, which is downregulated by TGF$\beta$1, is involved in the repression of both TGF$\beta$ receptors but not in the modulation of Smad3 and Smad7. Interestingly, Sp1 ectopic expression permitted also to maintain a similar expression pattern to early response to TGF$\beta$ at 24 hours of treatment. It restored the induction of Sox9 and COL2A1 and blocked the late response (repression of aggrecan, induction of COL1A1 and COL10A1). These data help to better understand the negative feedback loop in the TGF$\beta$ signalling system, and enlighten an interesting role of Sp1 to regulate TGF$\beta$ response.

A significant association between a polymorphism in the D repeat of the gene encoding asporin and osteoarthritis, the most frequent of articular diseases, has been recently reported. The goal of the present study was to investigate the expression of this new class I small leucine-rich proteoglycan (SLRP) in human articular chondrocytes. First, we studied the modulation of asporin (ASPN) expression by cytokines by Western blot and reverse transcription-polymerase chain reaction. Interleukin-1$\beta$ and tumor necrosis factor-$\alpha$ downregulated ASPN, whereas transforming growth factor-$\beta$1 (when incubated in a serum-free medium) upregulated it. Similarly to proinflammatory cytokines, chondrocyte dedifferentiation induced by a successive passages of cells was accompanied by a decreased asporin expression, whereas their redifferentiation by three-dimensional culture restored its expression. Finally, we found an important role of the transcription factor Sp1 in the regulation of ASPN expression. Sp1 ectopic expression increased ASPN mRNA level and promoter activity. In addition, using gene reporter assay and electrophoretic mobility shift assay, we showed that Sp1 mediated its effect through a region located between -473 and -140 bp upstream of the transcription start site in ASPN gene. In conclusion, this report is the first study on the regulation of asporin expression by different cytokines in human articular chondrocytes. Our data indicate that the expression of this gene is finely regulated in cartilage and suggest a major role of Sp1.