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• W1580212652 abstract "To determine in rabbit corneal epithelial cells in culture whether epidermal growth factor (EGF)-induced increases in prostaglandin (PG) E2 production inhibit both the extracellular signal-regulated kinase 2 (Erk-2), a mitogen-activated protein kinase (MAPK), cascade activation, and the mitogenic response to this growth factor.Serum starvation for 24 to 36 hours was used to synchronize cultures of SV40-transformed rabbit corneal epithelial (RCE) cells. The effects of exogenous PGE2, inhibition of PGE2 synthesis, and modulation of protein kinase A (PKA) activity on EGF-induced Erk-2 activation were assessed by immunoprecipitation, kinase assays, and Western blot analysis. PGE2 synthesis was measured by using enzyme-linked immunosorbent assay. [3H]-Thymidine incorporation was used to measure RCE cell proliferation rates.EGF (5 ng/ml) significantly increased PGE2 production in a time-dependent manner up to 94%+/-8% after 3 hours. EGF-induced PGE2 production was suppressed by AACOCF3, a phospholipase A2 (cPLA2) inhibitor. EGF-induced Erk-2 activation reached a maximal level at 15 minutes, followed by a decline toward the control level after 3 hours. In the presence of either PGE2 (50 microg/ml) or 8-CPT-cAMP (100 microM), the EGF-induced Erk-2 activation was lessened. PKA was activated by applications of EGF or PGE2 and suppressed by AACOCF3. On the other hand, either inhibition of PGE2 production with AACOCF3 or H-89, a PKA inhibitor, enhanced EGF-induced Erk-2 activity. Raf-1 activity was stimulated by EGF to maximal activity at 5 minutes and returned toward its control level after 60 minutes. As with the dependence of Erk-2 activity on PKA activity, in the presence of H-89, the EGF-induced Raf-1 activation was significantly enhanced. DNA synthesis was increased 59%+/-5% (n = 4) after EGF stimulation, indicating a mitogenic effect of EGF in RCE cells. Inhibition of cPLA2 activity with AACOCF3 increased DNA synthesis in RCE cells by another 64% relative to the effect of EGF alone. In contrast, with either PGE2 or 8-CPT-cAMP present the mitogenic response to EGF was totally suppressed.EGF-induced increases in PGE2 production dampened the mitogenic response to this growth factor. This suppression appears to be a consequence of PGE2-elicited increases in PKA activity, which leads to inhibition of EGF-induced activation of MAPK cascades at the level of Raf-1 and further affects downstream events including Erk-2. These results indicate that the mitogenic response to EGF in vivo in the proliferating basal cell layer may be dependent on the level of its PKA activity." @default.
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• W1580212652 date "2000-07-01" @default.
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• W1580212652 title "Inhibitory effect of PGE2 on EGF-induced MAP kinase activity and rabbit corneal epithelial proliferation." @default.
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