Interaction of ERK1/2 and Smad2/3 signaling pathways in TGF-β1-induced TIMP-3 expression in rat chondrocytes

Tissue inhibitor of metalloproteinase-3 (TIMP-3) is an important natural inhibitor of matrix metalloproteinases (MMPs) and of a disintegrin and metalloproteinase with thrombospondin motif (ADAMTs), which can cleave cartilage extracellular matrix components to cause cartilage degradation. In this study, our data suggest TGF-beta 1 induces TIMP-3 expression through activations of both the ERKI/2 and Smad2/3 signaling pathways. TGF-beta 1-stimulated TIMP-3 expression was significantly inhibited by SB525334 (TGF11 beta receptor I kinase inhibitor), accompanied by a reduction in ERKI/2 and Smad3 phosphorylation. We used P098059 (MEK inhibitor) and SIS3 (inhibitor of Smad3 phosphorylation) to investigate the respective roles of ERKI/2 and Smad2/3 signaling pathways in TGF-beta 1-induced TIMP-3 expression. The results show PD98059 treatment significantly suppressed TGF-beta 1-induced ERKI/2 phosphorylation and TIMP-3 expression. Under these conditions, the degree of Smad3 phosphorylation correlated with ERK1/2 activation, which suggests that ERK1/2 may activate Smad3 phosphorylation. SIS3 significantly inhibited TGF-beta 1-induced Smad3 phosphorylation and TIMP-3 expression. ERKI/2 phosphorylation alone had no effect on TGF-beta 1-induced TIMP-3 expression, which suggests ERKI/2 via Smad3 phosphorylation regulates TGF-beta 1 -induced TIMP-3 expression. Here, we demonstrate that ERKI/2 may be capable of activating the Smad2/3 signaling pathway to result in TGF-beta 1-induced TIMP-3 up-regulation. (C) 2014 Elsevier Inc. All rights reserved.