Modulation of Human Renal Cell Carcinoma 786-0 MMP-2 and MMP-9 Activity by Inhibitors and Inducers in Vitro

M.W. Roomi, N. Roomi, V. Ivanov,T. Kalinovsky, A. Niedzwiecki, M. Rath
Medical Oncology 2006, 23(2): 245-250

A hallmark of renal cell carcinoma RCC) invasion is its ability to degrade ECM by local production of gelatinase enzymes. Although many studies on RCC have demonstrated the importance of MMPs, very little information is currently known regarding the effect of inducers and inhibitors. We therefore investigated the effect of inducers and inhibitors on RCC 786-0 in vitro.

Human RCC 786-0 ATCC) was grown in RPMI medium supplemented with 10% FBS, penicillin, and streptomycin in 24-well tissue plates. At near confluence, the cells were washed with PBS; the serum free medium was incubated with various inducers: phorbol ester (PMA; tumor necrosis factor alpha (TNFα); interleukin 1-beta ( IL-1ß); and lipopolysaccharides (LPS). Cells were also incubated with inhibitors: EGCG; doxycycline; a nutrient mixture with and without PMA; retinoic acid; dexamethasone; H-7; actinomcin D; and cyclohexamide. After 24 hours, the medium was removed and analyzed for MMP-2 and MMP-9 by gelatinase zymography.

RCC 786-0 secreted two bands, a major band corresponding to MMP-2 and a faint band corresponding to MMP-9. PMA and TNF-α, with increased concentration, increased MMP-9 secretion, while IL-1ß and LPS did not significantly modify MMP-9 activity. MMP-2 secretion was not affected by any of the inducers. All the inhibitors tested without and with PMA showed a dose-dependent decrease in both MMP-2 and -9 expression.

In conclusion, this study has shown that exposure of the highly metastatic renal cell carcinoma cell line 786-0 to different growth factors increases secretion of MMP-9 but not MMP-2; MMP-2 and-9 activity was decreased by exposure to inhibitors. Further studies are in progress to confirm the role of MMP-9 on Matrigel invasion using PMA, cytokines and LPS.