Tenascin-C (TN-C) and matrix metalloproteinases (MMPs) are highly expressed in cancer tissues and probably promote cell migration during cancer progression. TN-C and MMPs are often co-localized in areas of active tissue remodeling in pathologic conditions, suggesting reciprocal regulation. To investigate whether TN-C regulates MMPs expression in cancer cells, we first exposed mammary cancer cells derived from TN-C-deficient mice to TN-C and examined MMPs expression. TN-C was then compared with fibronectin (FN), laminin (LN), basic fibroblast growth factor (b-FGF) and transforming growth factor-beta1 (TGF-β1). Results of endpoint RT-PCR, quantitative real-time RT-PCR and gelatin zymography demonstrated that TN-C, strongly and dose dependently, upregulates MMP-9 expression in murine mammary cancer cells. TN-C weakly induced MMP-2, MMP-3 and MMP-13. FN and LN induced MMP-9 to lesser extents compared with TN-C. b-FGF had no effect on MMP-9 expression. TGF-β1 induced MMP-9 expression in a dose-dependent manner, and this induction was significantly enhanced by addition of TN-C. TN-C and TGF-β1 also upregulated MMP-9 expression in the human breast cancer cell line MDA-MB-231. Neutralization with specific anti-TGF-β1 antibody showed decreased expression of MMP-9, indicating that TGF-β controls the baseline MMP-9 expression by a direct autocrine mechanism. Under neutralization of TGF-β, addition of TN-C still upregulated MMP-9. Conversely, neutralization of endogenous TN-C (in a TN-C-positive mammary cancer cell line) downregulated TGF-β-induced MMP-9 expression. Thus, TN-C induces MMP-9 expression directly and by collaboration with TGF-β. These findings reveal a novel role of TN-C in breast cancer progression. © 2003 Wiley-Liss, Inc.