Modulation of transforming growth factor beta signalling pathway genes by transforming growth factor beta in human osteoarthritic chondrocytes: involvement of Sp1 in both early and late response cells to transforming growth factor beta

摘要

Transforming growth factor beta (TGF$\beta$) plays a central role inmorphogenesis, growth, and cell differentiation. This cytokine is particularlyimportant in cartilage where it regulates cell proliferation and extracellularmatrix synthesis. While the action of TGF$\beta$ on chondrocyte metabolism hasbeen extensively catalogued, the modulation of specific genes that function asmediators of TGF$\beta$ signalling is poorly defined. In the current study,elements of the Smad component of the TGF$\beta$ intracellular signallingsystem and TGF$\beta$ receptors were characterised in human chondrocytes uponTGF$\beta$1 treatment. Human articular chondrocytes were incubated withTGF$\beta$1. Then, mRNA and protein levels of TGF$\beta$ receptors and Smadswere analysed by RT-PCR and western blot analysis. The role of specific protein1 (Sp1) was investigated by gain and loss of function (inhibitor, siRNA,expression vector). We showed that TGF$\beta$1 regulates mRNA levels of its ownreceptors, and of Smad3 and Smad7. It modulates TGF$\beta$ receptorspost-transcriptionally by affecting their mRNA stability, but does not changethe Smad-3 and Smad-7 mRNA half-life span, suggesting a potentialtranscriptional effect on these genes. Moreover, the transcriptional factorSp1, which is downregulated by TGF$\beta$1, is involved in the repression ofboth TGF$\beta$ receptors but not in the modulation of Smad3 and Smad7.Interestingly, Sp1 ectopic expression permitted also to maintain a similarexpression pattern to early response to TGF$\beta$ at 24 hours of treatment. Itrestored the induction of Sox9 and COL2A1 and blocked the late response(repression of aggrecan, induction of COL1A1 and COL10A1). These data help tobetter understand the negative feedback loop in the TGF$\beta$ signallingsystem, and enlighten an interesting role of Sp1 to regulate TGF$\beta$response.