ABSTRACT: BACKGROUND: Metastasis is the main factor responsible for death in breast cancer patients. Matrix metalloproteinases (MMPs) and their inhibitors, known as tissue inhibitors of MMPs (TIMPs), and the membrane-associated MMP inhibitor (RECK), are essential for the metastatic process. We have previously shown a positive correlation between MMPs and their inhibitors expression during breast cancer progression; however, the molecular mechanisms underlying this coordinate regulation remain unknown. In this report, we investigated whether TGF-?1 could be a common regulator for MMPs, TIMPs and RECK in human breast cancer cell models. METHODS: The mRNA expression levels of TGF-? isoforms and their receptors were analyzed by qRT-PCR in a panel of five human breast cancer cell lines displaying different degrees of invasiveness and metastatic potential. The highly invasive MDA-MB-231 cell line was treated with different concentrations of recombinant TGF-?1 and also with pharmacological inhibitors of p38 MAPK and ERK1/2. The migratory and invasive potential of these treated cells were examined in vitro by transwell assays. RESULTS: In general, TGF-?2, T?RI and T?RII are over-expressed in more aggressive cells, except for T?RI, which was also highly expressed in ZR-75-1 cells. In addition, TGF-?1-treated MDA-MB-231 cells presented significantly increased mRNA expression of MMP-2, MMP-9, MMP-14, TIMP-2 and RECK. TGF-?1 also increased TIMP-2, MMP-2 and MMP-9 protein levels but downregulated RECK expression. Furthermore, we analyzed the involvement of p38 MAPK and ERK1/2, representing two well established Smad-independent pathways, in the proposed mechanism. Inhibition of p38MAPK blocked TGF-?1-increased mRNA expression of all MMPs and MMP inhibitors analyzed, and prevented TGF-?1 upregulation of TIMP-2 and MMP-2 proteins. Moreover, ERK1/2 inhibition increased RECK and prevented the TGF-?1 induction of pro-MMP-9 and TIMP-2 proteins. TGF-?1-enhanced migration and invasion capacities were blocked by p38MAPK, ERK1/2 and MMP inhibitors. CONCLUSION: Altogether, our results support that TGF-?1 modulates the mRNA and protein levels of MMPs (MMP-2 and MMP-9) as much as their inhibitors (TIMP-2 and RECK). Therefore, this cytokine plays a crucial role in breast cancer progression by modulating key elements of ECM homeostasis control. Thus, although the complexity of this signaling network, TGF-?1 still remains a promising target for breast cancer treatment.
