Transforming Growth Factor-β1 Induces Tissue Inhibitor of Metalloproteinase-1 Expression via Activation of Extracellular Signal-Regulated Kinase and Sp1 in Human Fibrosarcoma Cells

Abstract

The net balance of matrix metalloproteinases (MMP) and tissue inhibitor of metalloproteinases (TIMP) system has been known to be a key factor in tumor cell invasion. In the present study, we investigated the molecular mechanisms of anti-invasive and antimigrative activity of transforming growth factor (TGF)-B1 on HT1080 human fibrosarcoma cells. In in vitro Matrigel invasion and Transwell migration assays, TGF-B1 dose-dependently inhibited the invasion and migration of HT1080 cells, respectively. Gelatin zymography, Western blot, and real-time PCR analysis showed that TGF-B1 enhanced the expression and secretion of MMP-2, TIMP-1, and, to a lesser degree, MMP-9 but not membrane type 1-MMP and TIMP-2. The addition of recombinant TIMP-1 protein reduced the Matrigel invasion and Transwell migration of HT1080 cells, similar to TGF-B1. Because augmentation of TIMP-1 might be the major factor for the anti-invasive and antimigrative activity of TGF-B1, we investigated possible molecular mechanisms responsible for the expression of TIMP-1 induced by TGF-B1. Treatment of HT1080 cells with TGF-B1 rapidly phosphorylated three mitogen-activated protein kinases [MAPK; extracellular signal-regulated kinase 1/2 (ERK1/2), p38, and c-Jun NH 2 -terminal kinase] and Akt. Among these kinases, the inhibition of only ERK1/2 pathway by PD98059, a specific inhibitor of MAPK/ERK kinase(MEK)-1, and transfection of dominant-negative MEK 1 effectively blocked the TIMP-1 induction by TGF-B1. Mithramycin, a specific inhibitor of Sp1 transcription factor, but not curcumin, an inhibitor of activator protein-1, and transfection of Sp1 small interfering RNA significantly inhibited the TGF-B1-induced expression of TIMP-1. In addition, electrophoretic mobility shift assay showed that TGF-B1 up-regulated Sp1 DNA-binding activity, and PD98059 and mithramycin effectively inhibited these events. Finally, pretreatment of HT1080 cells with PD98059 and mithramycin, but not curcumin, restored the invasive activity of these cells. Taken together, these data suggest that TGF-B1 modulates the net balance of the MMPs/TIMPs the systems in HT1080 cells for antiinvasion and antimigration by augmenting TIMP-1 through ERK1/2 pathway and Sp1 transcription factor.

HDAC-mediated control of ERK- and PI3K-dependent TGF-β-induced extracellular matrix-regulating genes

“…Indeed, we showed that in addition to Smad phosphorylation, TGF-β-activated MAPK and PI3K signalling with a range of kinetics; the transient activation being consistent with previous observations in C3H10T1/2 cells (Lien et al, 2006). Although we have conclusively shown that the TGF-β induction of Adam12 and Timp-1 expression is Smad-dependent, Adam12 induction has also been shown to require ERK and Akt signalling (Le Pabic et al, 2003Pabic et al, , 2005, while Timp-1 induction is reportedly dependent on ERK activation (Kwak et al, 2006). We used pharmacological inhibitors to confirm that these signalling pathways are required, and also provide evidence that p38 and JNK signalling may also be involved, although concerns about the lack of specificity of the p38 and JNK inhibitors must be considered (Bain et al, 2007).…”

Glioblastoma-secreted factors induce IGFBP7 and angiogenesis by modulating Smad-2-dependent TGF-β signaling

“…As the same pathway was responsible for Tumor cells induce IGFBP7 in endothelium A Pen et al IGFBP7 induction in HBEC, it is tempting to speculate that autocrine actions of IGFBP7 could participate in U87MG-CM-induced angiogenic responses in HBEC. TGF-b mediates the transcription of several genes related to the ECM remodeling, including SPARC and collagen I (Reed et al, 1994), MMP-2, TIMP-1 (Kwak et al, 2006), plasminogen activator inhibitor-1 (Keeton et al, 1991) and collagen IV (Poncelet and Schnaper, 1998). Secreted IGFBP7 is known to interact with ECM components (Akaogi et al, 1996;St Croix et al, 2000), and could thus participate in the TGF-b1-induced ECM turnover and angiogenesis.…”

Fibroblast Growth Factor-1 Suppresses TGF-β-Mediated Myofibroblastic Differentiation of Rat Hepatic Stellate Cells

“…MMP-9 was found in scaring areas along with the activated HSC (11). TGF-β1 stimulated expression of MMP-2 and TIMP-1 in vitro in dose dependent manner (19,20). In our hands, stimulation of the cells with TGF-β1 promoted MMP-2, MMP-9 and TIMP-1 expression and reduced the expression of TIMP-2, but no changes at the protein level were detected.…”