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Koji Sugioka, Aya Kodama, Koji Yoshida, Kiyotaka Okada, Shunji Kusaka, Chota Matsumoto, Yoshikazu Shimomura; TGF-beta2 promotes RPE cell invasion into collagen gel by mediating urokinase-type plasminogen activator (uPA) expression. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2012.
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Transforming growth factor-beta (TGF-beta) is one of the main epithelial-mesenchymal transition (EMT)-inducing factors and TGF-beta-induced EMT promotes cell migration and invasion. In this study, we investigated the role of urokinase-type plasminogen activator (uPA) during TGF-beta-induced RPE cell invasiveness.
ARPE-19 cells were cultured and stimulated with TGF-beta2. uPA and uPA receptor (uPAR) expressions during TGF-beta-induced EMT process in ARPE-19 cells were evaluated by real-time RT-PCR and fibrin zymography. Effects of TGF-beta inhibitor (SB431542) on the expression of uPA and uPAR by TGF-beta2 stimulation were also evaluated. IAsys-binding assay was performed to investigate the binding ability of uPA to the surface of ARPE-19 cells. For invasion assay, ARPE-19 cells were seeded onto the Transwell chambers and allowed to invade into collagen matrix in the presence of TGF-beta2 alone or TGF-beta2+u-PA inhibitor.
TGF-beta2 enhances expression of uPA and uPAR in ARPE-19 cells and secretion of uPA. SB431542 significantly suppressed TGF-beta2-stimulated uPA expression and secretion but not uPAR expression in ARPE-19 cells. ARPE-19 cells pretreated and non-pretreated with TGF-beta2 bound to uPA in a time-dependent or cell-number-dependent fashion. However, the binding ability of uPA to ARPE-19 cells was enhanced more greatly with ARPE-19 cells pretreated with TGF-beta2. TGF-beta2 treatment induced the invasiveness of ARPE-19 cells into collagen gel. TGF-beta2+u-PA inhibitor treatment strongly inhibited the invasiveness of ARPE-19 cells.
TGF-beta-induced RPE cell invasiveness occurs via its regulation of uPA. These results suggest that uPA mediates TGF-beta-induced RPE cell invasion in collagen gel matrix.
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