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P. Logan, J.-C. A. Marshall, B. F. Fernandes, S. Bakalian, C. Martins, M. N. Burnier, Jr.; MMP-2 and MMP-9 Secretion by Human Uveal Melanoma Cell Lines in Response to Different Growth Factors and Chemokines. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4766.
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Secretion of matrix metalloproteinases (MMPs) by tumor cells plays a critical role in tumor progression. In the primary tumor, cells must produce MMPs in order to degrade the extracellular matrix to create room for growth. Tumor cells must also produce MMPs in order to intravasate from the primary tumor into the blood stream, and to extravasate at distant, metastatic locations. The primary site for metastatic growth in uveal melanoma patients is the liver and the production of MMPs is essential in allowing disseminated tumor cells to exit the blood stream and migrate into the liver parenchyma.
A specialized MMP Gelatinase Assay was used to determine the production of MMP-2 and MMP-9. Six different factors were used to induce MMP secretion: IL-6, HGF, IL-8, Gro-Alpha, TNF-Alpha, IGF-1. Five primary uveal melanoma cell lines were incubated with three concentrations of each factor for 48 hours. Supernatants were collected and the MMP Gelatinase Assay was used to quantify the secretion of MMP-2 and MMP-9. Each cell line was then incubated in IL-8 for 48 hours and allowed to invade an artificial matrigel membrane to determine if increases in MMP-2 and MMP-9 secretion correlated with increased invasive ability of the cell lines.
Results were analyzed as percentage of negative control. The highest increase in MMP secretion was induced by IL-8 in the 92.1 cell line (387% of negative control). In addition, IL-8 induced the largest average increase across all cell lines (177% of negative control) followed closely by IL-6 (136% of negative control). Each cell line produced a statistically significant increase in MMP-2 and MMP-9 production in response to at least one concentration of each factor. The two most aggressive cell lines (92.1 and SP6.5) produced the largest average MMP secretion in response to all factors (182% and 132%, respectively). The results of the invasion assay mimicked the results of the MMP Assay: the largest increases in MMP-2 and MMP-9 secretion in response to IL-8 correlated with the greatest ability to invade. Furthermore, the addition of IL-8 increased the ability of our cell lines to invade when compared to negative controls.
This study demonstrates the ability of five uveal melanoma cell lines to produce MMP-2 and MMP-9 in response to six different growth factors and chemokines, particularly IL-8 and IL-6. Considering that MMPs play a multi-faceted role in tumor progression these factors present as potential therapeutic targets in order to prevent crucial steps in the metastatic cascade.
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