Objective: We previously reported that lysophosphatidic acid (LPA) stimulates cellular invasion of ovarian cancer (OVCA) cells by enhancing membrane-type-1-matrix metalloproteinase (MT1-MMP)-mediated activation of MMP2. Here, we investigate a second mechanism in which LPA enhances cellular invasion through the increased expression of IL-8, independent of the expression or activation of MMP2.
Methods: Epithelial ovarian carcinoma cells (DOV 13) were exposed to LPA (80 microM) and IL-8 (100 ng/ml) for 24 h. IL-8 expression was quantified by enzyme-linked immunosorbent assay (ELISA). Cellular invasion (Matrigel invasion), migration (colloidal gold), and urinary-type plasminogen activator (uPA) activity (colorimetric assay) were quantified. Conditioned medium was also assayed for MMP activation and expression by SDS-PAGE gelatin zymography, ELISA, and Western blotting. In addition, IL-8 neutralizing antibody and MMP inhibitors were employed.
Results: Our results found LPA to increase IL-8 expression threefold. IL-8 did not affect cellular migration, MMP2 activation, or uPA expression. However, exposure to various concentrations of IL-8 increased cellular invasiveness. Using an IL-8 blocking antibody and various MMP inhibitors, we determined that the increase in invasion was IL-8-dependent, while independent of the activation of MMP2 or MMP9. We further determined IL-8 exposure increased the expression of matrilysin (MMP7). Cells exposed to LPA and IL-8 resulted in a synergistic effect on cellular invasion. Adding the IL-8 blocking antibody, slightly decreased cellular invasion, indicating LPA in part, increases cellular invasion through the increased expression of IL-8.
Conclusion: We have identified a separate mechanism of enhanced cellular invasion, which is independent of MMP2 activation and involves the increased expression of IL-8 and subsequent increased expression of MMP7.