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BackgroundUrokinase plasminogen activator (uPA) receptor (uPAR) is up-regulated at the invasive tumour front of human oral squamous cell carcinoma (OSCC), indicating a role for uPAR in tumour progression. We previously observed elevated expression of uPAR at the tumour-stroma interface in a mouse model for OSCC, which was associated with increased proteolytic activity. The tumour microenvironment regulated uPAR expression, as well as its glycosylation and cleavage. Both full-length- and cleaved uPAR (uPAR (II-III)) are involved in highly regulated processes such as cell signalling, proliferation, migration, stem cell mobilization and invasion. The aim of the current study was to analyse tumour associated factors and their effect on uPAR cleavage, and the potential implications for cell proliferation, migration and invasion.MethodsMouse uPAR was stably overexpressed in the mouse OSCC cell line AT84. The ratio of full-length versus cleaved uPAR as analysed by Western blotting and its regulation was assessed by addition of different protease inhibitors and transforming growth factor - β1 (TGF-β1). The role of uPAR cleavage in cell proliferation and migration was analysed using real-time cell analysis and invasion was assessed using the myoma invasion model.ResultsWe found that when uPAR was overexpressed a proportion of the receptor was cleaved, thus the cells presented both full-length uPAR and uPAR (II-III). Cleavage was mainly performed by serine proteases and urokinase plasminogen activator (uPA) in particular. When the OSCC cells were stimulated with TGF-β1, the production of the uPA inhibitor PAI-1 was increased, resulting in a reduction of uPAR cleavage. By inhibiting cleavage of uPAR, cell migration was reduced, and by inhibiting uPA activity, invasion was reduced. We could also show that medium containing soluble uPAR (suPAR), and cleaved soluble uPAR (suPAR (II-III)), induced migration in OSCC cells with low endogenous levels of uPAR.ConclusionsThese results show that soluble factors in the tumour microenvironment, such as TGF-β1, PAI-1 and uPA, can influence the ratio of full length and uPAR (II-III) and thereby potentially effect cell migration and invasion. Resolving how uPAR cleavage is controlled is therefore vital for understanding how OSCC progresses and potentially provides new targets for therapy.

Glutathione plays an important role in drug resistance of tumor cells and in their ability to resist oxidative stress. Improved salvage of glutathione can be obtained through increased activity of γ‐glutamyltransferase (GGT), which is of importance in the maintenance of cellular glutathione homeostasis. We investigated the regulation of GGT in 2 cisplatin‐resistant and 1 cisplatin‐sensitive colon carcinoma cell lines. Enzyme activity was induced in all 3 cell lines after acute exposure to cisplatin. The elevation was significantly higher in sensitive cells (3.3‐fold) than in resistant (1.6‐ to 1.7‐fold) cells. Exposure of cells to oxidative stress generated by menadione also resulted in enzyme induction but only in cisplatin‐sensitive cells. Addition of anti‐oxidants had different effects on the 2 inductions: N‐acetylcysteine blocked the induction of both cisplatin and menadione, whereas catalase and glutathione‐ester blocked only the menadione induction. Glutathione depletion alone was not sufficient to induce GGT in these cells. The data show that GGT is regulated by multiple mechanisms during anti‐tumor drug treatment and oxidative stress and that reactive oxygen species were involved in the menadione, but not cisplatin, induction of the enzyme. Int. J. Cancer 88:464–468, 2000. © 2000 Wiley‐Liss, Inc.

Injection of the rat colon carcinoma cell line CC531 into spleen of syngeneic rats results in considerable amounts of liver metastases within 14 days. We investigated whether preincubation of the cells with butyrate reduced their metastatic ability in vivo and whether this was accompanied by reduction in related properties such as secretion of metalloproteinases and their ability to withstand oxidative stress. Butyrate incubation reduced cell growth rate and initiated apoptosis in a dose- and time-related manner, but proliferation was retrieved when cultivation was continued in medium without butyrate. Splenic injection of butyrate treated, proliferating cells resulted in significantly reduced amounts of tumor mass compared to untreated cells. The butyrate treated cells were more susceptible to oxidative stress than control cells, as demonstrated by increased number of apoptotic cells and reduced cell growth after exposure to menadione. A reduction in cellular glutathione was found after prolonged incubation with butyrate. Butyrate appeared not to alter the secretion of active metalloproteinases from the cells although an apparent increase in proforms was demonstrated. Neither did butyrate alter the synthesis of metalloproteinase inhibitors. Lastly, a reduced adhesion of the tumor cells to collagen coated matrix was found after butyrate treatment. Thus, the inhibitory effects of butyrate on tumor malignancy are caused by a diversity of mechanisms.