Protein kinase C mediates basic fibroblast growth factor protection of endothelial cells against radiation-induced apoptosis.
Basic fibroblast growth factor (bFGF) was found to protect bovine aortic endothelial cells against the lethal effects of ionizing radiation by inhibiting the programmed cell death (apoptosis) induced in these cells by radiation exposure. The involvement of the bFGF receptor tyrosine kinase in this function was demonstrated by abrogation of the radioprotective effect of bFGF by a specific inhibitor of the bFGF receptor tyrosine kinase, the tyrphostin AG213. The downstream signaling after stimulation of the bFGF receptor tyrosine kinase in bovine aortic endothelial cells involved translocation of the alpha isotype of cytoplasmic protein kinase C (PKC) into the membrane and its activation within 30 s after bFGF stimulation. The involvement of PKC in the radioprotective effect conferred by bFGF was suggested by the demonstration that nonspecific PKC activation by short-term exposure (30 min) to the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA; 30 ng/ml) mimicked the radioprotective effect of bFGF. Furthermore, treatment of the cells with the PKC inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (20 microM) abrogated the radioprotective effect of bFGF, as was observed after the depletion of cellular PKC by overnight preincubation with high-dose TPA (200 nM). Agarose gel electrophoresis of DNA extracted from irradiated bovine aortic endothelial cells showed that both TPA (30 ng/ml; 30 min) and bFGF (1 ng/ml) inhibited the apoptotic degradation of DNA induced in these cells by radiation exposure (500 cGy). Both the bFGF- and the TPA-mediated inhibition of apoptosis could be reversed by the PKC inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (20 microM). These data demonstrate the involvement of PKC in the inhibition of radiation-induced apoptosis by bFGF and the rescue of endothelial cells from this mode of radiation-induced cell death.
Haimovitz-Friedman
A
A
Department of Radiation Oncology, Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
Balaban
N
N
McLoughlin
M
M
Ehleiter
D
D
Michaeli
J
J
Vlodavsky
I
I
Fuks
Z
Z
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002417:Cattle; D002462:Cell Membrane / Q000201:enzymology; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000528:radiation effects; D004790:Enzyme Induction / Q000187:drug effects; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D007527:Isoenzymes / Q000032:analysis; D051379:Mice; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000502:physiology*; D011829:Radiation Dosage; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; D016222:Fibroblast Growth Factor 2; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate