11,12-Epoxyeicosatrienoic acid-induced inhibition of FOXO factors promotes endothelial proliferation by down-regulating p27Kip1.
Cytochrome P450-derived epoxyeicosatrienoic acids (EETs) stimulate endothelial cell proliferation and angiogenesis. In this study, we investigated the involvement of the forkhead box, class O (FOXO) family of transcription factors and their downstream target p27Kip1 in EET-induced endothelial cell proliferation. Incubation of human umbilical vein endothelial cells with 11,12-EET induced a time- and dose-dependent decrease in p27Kip1 protein expression, whereas p21Cip1 was not significantly affected. This effect on p27Kip1 protein was associated with decreased mRNA levels as well as p27Kip1 promoter activity. 11,12-EET also stimulated the time-dependent phosphorylation of Akt and of the forkhead factors FOXO1 and FOXO3a, effects prevented by the phosphatidylinositol 3-kinase inhibitor LY 294002. Transfection of endothelial cells with either a dominant-negative or an "Akt-resistant"/constitutively active FOXO3a mutant reversed the 11,12-EET-induced down-regulation of p27Kip1, whereas transfection of a constitutive active Akt decreased p27Kip1 expression independently of the presence or absence of 11,12-EET. To determine whether these effects are involved in EET-induced proliferation, endothelial cells were transfected with the 11,12-EET-generating epoxygenase CYP2C9. Transfection of CYP2C9 elicited endothelial cell proliferation and this effect was inhibited in cells co-transfected with CYP2C9 and either a dominant-negative Akt or constitutively active FOXO3a. Reducing FOXO expression using RNA interference, on the other hand, attenuated p27Kip1 expression and stimulated endothelial cell proliferation. These results indicate that EET-induced endothelial cell proliferation is associated with the phosphatidylinositol 3-kinase/Akt-dependent phosphorylation and inactivation of FOXO factors and the subsequent decrease in expression of the cyclin-dependent kinase inhibitor p27Kip1.
Potente
Michael
M
Institut für Kardiovaskuläre Physiologie, Klinikum der J. W. G.-Universität, Theodor-Stern-Kai 7, D-60590 Frankfurt am Main, Germany.
Fisslthaler
Beate
B
Busse
Rudi
R
Fleming
Ingrid
I
D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology*; D001189:Aryl Hydrocarbon Hydroxylases / Q000378:metabolism / Q000502:physiology*; D015152:Blotting, Northern; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division; D002478:Cells, Cultured; D050760:Cyclin-Dependent Kinase Inhibitor p27; D065729:Cytochrome P-450 CYP2C9; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors*; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004730:Endothelium, Vascular / Q000166:cytology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; D005799:Genes, Dominant; D006801:Humans; D015151:Immunoblotting; D008156:Luciferases / Q000378:metabolism; D018919:Neovascularization, Physiologic; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D013997:Time Factors; D014157:Transcription Factors / Q000037:antagonists & inhibitors*; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology; D014665:Vasodilator Agents / Q000494:pharmacology
D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D004791:Enzyme Inhibitors; C085126:FOXO1 protein, human; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; D000081082:Phosphoinositide-3 Kinase Inhibitors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D014665:Vasodilator Agents; D050760:Cyclin-Dependent Kinase Inhibitor p27; C046783:11,12-epoxy-5,8,14-eicosatrienoic acid; D008156:Luciferases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; D015126:8,11,14-Eicosatrienoic Acid