Absence of feedback regulation in the synthesis of COL1A1

Aim: Recent studies have emphasized the importance of the extracellular microenvironment in modulating cell growth, motility, and signalling. In this study we have evaluated the ability of a fibroblast derived-extracellular matrix (fd-ECM) to regulate type I collagen synthesis and degradation in fibroblasts. Main methods: Fibroblasts were plated on plastic (control) or on fd-ECM and type I collagen synthesis and degradation was evaluated. MTT, western blotting, real time PCR, zymographic analysis and inhibitor assays were utilised to investigate the molecular mechanism of type I collagen regulation by the fd-ECM. Key findings: Fibroblasts plated on fd-ECM showed significant downregulation in the production of type I collagen and COL1A2 messenger ribonucleic acid (mRNA) whilst COL1A1 mRNA remained unchanged. Cells grown on fd-ECM exhibited increased matrix metalloproteases (MMPs) and their corresponding mRNAs. The use of transforming growth factor beta (TGF-beta) and MMP inhibitors showed that the excess COL1A1 polypeptide chains were degraded by the combined action of MMP-1, MMP-2, MMP-9 and cathepsins. Significance: These results show the crucial role played by proteases in regulating extracellular matrix protein levels in the feedback regulation of connective tissue gene expression. (C) 2014 Elsevier Inc. All rights reserved.