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Dan-Ning Hu, Steven A McCormick, Tommaso Vagaggini, Richard B Rosen; Effects of various MAPK and PI3K/Akt inhibitors on the constitutive secretion of VEGF by uveal melanoma cells. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5056.
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VEGF plays a critical role in angiogenesis and in the growth and metastasis of uveal melanoma (UMa). Studying the regulation of secretion of VEGF by UMa cells with MAPK and PI3K/Akt signal pathways is important for understanding the pathogenesis of and target therapy for UMa.
The constitutive levels of VEGF secretion were tested in six human immortal UMa cell lines (MP17, M21, M23, SP6.5, SP8.0 and TP3.1). Cells were seeded into 24-well plates and cultured with serum-free culture medium. After 24 hr, the conditioned culture medium was collected, and the amount of VEGF was measured using a sandwich enzyme-linked immunosorbent assay kit (R & D Systems). In a separate signal pathway study, cells were cultured siminarly with or without various signal pathway inhibitors, including LY294002 (PI3K/Akt inhibitor), UO1026 (ERK inhibitor), SP600125 (JNK inhibitor) and SB203580 (p38 MAPK inhibitor). The conditioned media were collected 24 hr later, and the amount of VEGF was measured as described above. All tests were performed in triplicate.
VEGF could be detected in the conditioned medium at a relatively high concentration from all cultured UMa cells (1989 ± 71 pg/106 cells/24 hr). Treatment of cells with PI3K/Akt inhibitor and ERK inhibitor significantly decreased VEGF levels in the conditioned media to 41% and 46% of that in controls (P < 0.05). JNK1/2 inhibitor and p38 MAPK inhibitor did not cause a significant change in the VEGF production by UMa cells (P > 0.05).
Uveal melanoma cells have a significantly constitutive secretion of VEGF. Inhibition of PI3K/Akt and ERK pathways, but not JNK1/2 and p38 MAPK pathways, decreases the constitutive secretion of VEGF by UMa. This indicates that PI3K/Akt and ERK pathways play an important role in the constitutive secretion of VEGF by UMa cells and might be considered as a target in the prevention and treatment of UMa.
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