Activation of synovial fibroblasts by laminin-1 and transforming growth factor beta induces expression of stromelysins independently of TNFα, IL-1β or NF-κB

Recently it was shown that attachment of synovial fibroblasts (SF) from rheumatoid arthritis patients to laminin-111 (LM-111) induced an elevated expression of stromelysin-1 (MMP-3). We therefore investigated the regulation of additional matrix metalloproteinases (MMPs) and their specific tissue inhibitors of matrix metalloproteinases (TIMPs) by attachment to LM-111 in the presence of transforming growth factor beta (TGFβ). Changes in steady-state mRNA levels encoding TIMPs and MMPs were investigated by quantitative RT-PCR. Production of MMPs and cytokines was monitored by a multiplexed immunoarray or by ELISA. Signal transduction pathways were studied by immunoblotting. Attachment of SF to LM-111 in the presence of TGFβ induced significant increases in stromelysin-1 mRNA (12.35-fold, P < 0.001) and protein (mean 62 ng/ml, sixfold, P < 0.008). Expression of stromelysin-2 (MMP-10) mRNA (11.68-fold, P < 0.05) and protein (54 ng/ml, 20-fold, P ≥ 0.02) was significantly activated as well. All other TIMPs and MMPs investigated failed to show this LM-111-facilitated TGFβ response. Induction of stromelysin-1 and stromelysin-2 was associated with the activation of transcription factors c-fos and Egr-1, but phosphorylation of NF-κB was not observed. Further, LM-111-activated and TGFβ-activated SF failed to produce remarkable amounts of IL-1β or TNFα. We conclude that costimulation of synovial fibroblasts by LM-111 together with TGFβ suffices to induce significant expression of MMP-3 and MMP-10 by SF and that this induction is independent of phosphorylation of NF-κB.