Phenotypic alterations in Caki-1 cells as a consequence of TIMP-1 overexpression

Maintenance of the extracellular matrix (ECM) is important for tissue integrity and cellular physiology. Normal ECM turnover is regulated by a balance between matrix metalloproteinases and their inhibitors, the tissue inhibitors of metalloproteinases (TIMPs). In metastasis, this balance favours increased ECM degradation. The objective of this study was to determine the effects of TIMP-1 overexpression on the metastatic process. To this end, we stably transfected a renal carcinoma cell line, Caki-1, with TIMP-1, using a pRc/CMV expression plasmid and LIPOFECTAMINE transfection reagent. The resultant clones displayed increased adhesion on the ECM substratum, including collagen type IV and laminin, and altered invasive capacity through fibronectin and Matrigel, dependent upon the level of TIMP-1 expression. These changes were not due to altered integrin expression, as assessed by flow cytometry. As well as protease inhibitory activity, TIMPs can influence cell proliferation and cell survival. The TIMP-1 clones displayed no changes in proliferation under normal growth conditions, compared with Caki-1 cells. However, under reduced serum conditions, the TIMP-1 clones had a greater percentage of cells in both S (P<0.05) and G(2)/M (P<0.005) phases and less cells in G(0)/G(1) (P<0.001) of the cell cycle than Caki-1 cells. The results confirm a dual role for TIMP-1 in invasion and metastasis, and provide further clues behind the molecular mechanisms in these processes.