The Effects of mTOR-Akt Interactions on Anti-apoptotic Signaling in Vascular Endothelial Cells

Recent studies have determined that mTOR mediates the activation of the protein kinase Akt in several cell types, but little is known about the association between mTOR and Akt in vascular endothelial cells. Furthermore, the functional significance of mTOR/Akt signaling has not been characterized in the endothelium. In these studies we treated endothelial cells with the mTOR inhibitor rapamycin, and we found that it decreases Akt phosphorylation and activity, as determined by phosphorylation of its substrate glycogen synthase kinase-3. This effect of rapamycin on Akt phosphorylation could not be demonstrated in endothelial cells transfected with a rapamycin-resistant mTOR construct. Also, in the presence of rapamycin, vascular endothelial growth factor, tumor necrosis factor, and insulin failed to phosphorylate Akt, further indicating that mTOR regulates Akt activation in endothelial cells. The activation of Akt is well established to mediate pro-survival signals. In part this is mediated via the phosphorylation and inactivation of the pro-apoptotic Akt substrates Foxo1 and Foxo3a. We find that rapamycin totally blocks vascular endothelial growth factor and Akt-inducible phosophorylation of these transcription factors in endothelial cells. Furthermore, inhibition of Akt activity by rapamycin increased the number of endothelial cells undergoing apoptosis after serum withdrawal as well as after stimulation by vascular endothelial growth factor or tumor necrosis factor. Taken together these observations demonstrate first, that mTOR regulates the phosphorylation and activation of Akt in endothelial cells and, second, that a major effect of mTOR inhibition in endothelial cells is to suppress Akt-inducible pro-survival signals. Recent studies have determined that mTOR mediates the activation of the protein kinase Akt in several cell types, but little is known about the association between mTOR and Akt in vascular endothelial cells. Furthermore, the functional significance of mTOR/Akt signaling has not been characterized in the endothelium. In these studies we treated endothelial cells with the mTOR inhibitor rapamycin, and we found that it decreases Akt phosphorylation and activity, as determined by phosphorylation of its substrate glycogen synthase kinase-3. This effect of rapamycin on Akt phosphorylation could not be demonstrated in endothelial cells transfected with a rapamycin-resistant mTOR construct. Also, in the presence of rapamycin, vascular endothelial growth factor, tumor necrosis factor, and insulin failed to phosphorylate Akt, further indicating that mTOR regulates Akt activation in endothelial cells. The activation of Akt is well established to mediate pro-survival signals. In part this is mediated via the phosphorylation and inactivation of the pro-apoptotic Akt substrates Foxo1 and Foxo3a. We find that rapamycin totally blocks vascular endothelial growth factor and Akt-inducible phosophorylation of these transcription factors in endothelial cells. Furthermore, inhibition of Akt activity by rapamycin increased the number of endothelial cells undergoing apoptosis after serum withdrawal as well as after stimulation by vascular endothelial growth factor or tumor necrosis factor. Taken together these observations demonstrate first, that mTOR regulates the phosphorylation and activation of Akt in endothelial cells and, second, that a major effect of mTOR inhibition in endothelial cells is to suppress Akt-inducible pro-survival signals. Growth factors that are essential for angiogenesis, defined as the formation of new blood vessels from preexisting ones, induce protective genes in endothelial cells (EC) 3The abbreviations used are: EC, endothelial cell(s); VEGF, vascular endothelial growth factor; TNF, tumor necrosis factor; RRmTOR, rapamycin-resistant mTOR; WT, wild type; CHAPS, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid; p, phosphorylated. 3The abbreviations used are: EC, endothelial cell(s); VEGF, vascular endothelial growth factor; TNF, tumor necrosis factor; RRmTOR, rapamycin-resistant mTOR; WT, wild type; CHAPS, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid; p, phosphorylated. (1Dimmeler S. Zeiher A.M. Circ. Res. 2000; 87: 434-439Crossref PubMed Scopus (359) Google Scholar). Analysis of the signaling pathways that facilitate growth factor-mediated EC survival have demonstrated a critical function for the serine/threonine kinase Akt in this response (2Shiojima I. Walsh K. Circ. Res. 2002; 90: 1243-1250Crossref PubMed Scopus (846) Google Scholar). For example, vascular endothelial growth factor (VEGF), insulin, and ligation of the Tie2 receptor by angiopoietin-1, which all promote angiogenesis, induce Akt-dependent signals (3Gerber H.P. McMurtrey A. Kowalski J. Yan M. Keyt B.A. Dixit V. Ferrara N. J. Biol. Chem. 1998; 273: 30336-30343Abstract Full Text Full Text PDF PubMed Scopus (1741) Google Scholar, 4Hermann C. Assmus B. Urbich C. Zeiher A.M. Dimmeler S. Arterioscler. Thromb. Vasc. Biol. 2000; 20: 402-409Crossref PubMed Scopus (196) Google Scholar, 5Papapetropoulos A. Fulton D. Mahboubi K. Kalb R.G. O'Connor D.S. Li F. Altieri D.C. Sessa W.C. J. Biol. Chem. 2000; 275: 9102-9105Abstract Full Text Full Text PDF PubMed Scopus (548) Google Scholar). Akt is evolutionarily conserved and in mammalian cells consists of three highly homologous isoforms that share more than 80% of their amino acid sequence. Moreover, intracellular signals leading to Akt activation are conserved across species (6Testa J.R. Tsichlis P.N. Oncogene. 2005; 24: 7391-7393Crossref PubMed Scopus (231) Google Scholar, 7Woodgett J.R. Curr. Opin. Cell Biol. 2005; 17: 150-157Crossref PubMed Scopus (309) Google Scholar). The binding of cytokines and growth factors to vascular EC increases phosphatidylinositol 3-kinase activity, resulting in the production of phosphatidylinositol 3,4,5-triphosphates and the recruitment and activation of phosphoinositide-dependent kinase 1 within the cell membrane. Akt is phosphorylated by phosphoinositide-dependent kinase 1 (PDK1) at a threonine residue (Thr-308) in the activation loop and by another putative PDK2 kinase at a serine residue (Ser-473) in the carboxyl-terminal domain. Although several candidates have been proposed to function as the putative phosphoinositide-dependent kinase 2 (PDK2) kinase, including PDK1, integrin-linked kinase 1, ataxia telangiectasia mutant, and DNA-dependent protein kinase, recent studies have demonstrated that the rapamycin-insensitive Sin-1·Rictor·mTOR complex functions as PDK2 (8Sarbassov D.D. Guertin D.A. Ali S.M. Sabatini D.M. Science. 2005; 307: 1098-1101Crossref PubMed Scopus (5223) Google Scholar, 9Hresko R.C. Mueckler M. J. Biol. Chem. 2005; 280: 40406-40416Abstract Full Text Full Text PDF PubMed Scopus (509) Google Scholar, 10Jacinto E. Facchinetti V. Liu D. Soto N. Wei S. Jung S.Y. Huang Q. Qin J. Su B. Cell. 2006; 127: 125-137Abstract Full Text Full Text PDF PubMed Scopus (1139) Google Scholar). Upon activation Akt mediates pro-survival and anti-apoptosis in part via the phosphorylation and inhibition of the Bcl-2 homolog BAD, the phosphorylation and inactivation of the FoxO subfamily of forkhead transcription factors, and the transcriptional co-activator Yes-associated protein (11Datta S.R. Dudek H. Tao X. Masters S. Fu H. Gotoh Y. Greenberg M.E. Cell. 1997; 91: 231-241Abstract Full Text Full Text PDF PubMed Scopus (4936) Google Scholar, 12Brunet A. Bonni A. Zigmond M.J. Lin M.Z. Juo P. Hu L.S. Anderson M.J. Arden K.C. Blenis J. Greenberg M.E. Cell. 1999; 96: 857-868Abstract Full Text Full Text PDF PubMed Scopus (5412) Google Scholar, 13Basu S. Totty N.F. Irwin M.S. Sudol M. Downward J. Mol. Cell. 2003; 11: 11-23Abstract Full Text Full Text PDF PubMed Scopus (643) Google Scholar). In addition, Akt mediates cell proliferation and migration in part via the phosphorylation and activation of the endothelial nitricoxide synthase and glycogen synthase kinase-3β known to be functional in cell cycle progression as well as via the inactivation of p21 and p27 (14Morales-Ruiz M. Fulton D. Sowa G. Languino L.R. Fujio Y. Walsh K. Sessa W.C. Circ. Res. 2000; 86: 892-896Crossref PubMed Scopus (349) Google Scholar, 15Fosbrink M. Niculescu F. Rus V. Shin M.L. Rus H. J. Biol. Chem. 2006; 281: 19009-19018Abstract Full Text Full Text PDF PubMed Scopus (71) Google Scholar, 16Shin I. Yakes F.M. Rojo F. Shin N.Y. Bakin A.V. Baselga J. Arteaga C.L. Nat. Med. 2002; 8: 1145-1152Crossref PubMed Scopus (683) Google Scholar, 17Zhou B.P. Liao Y. Xia W. Spohn B. Lee M.H. Hung M.C. Nat. Cell Biol. 2001; 3: 245-252Crossref PubMed Scopus (914) Google Scholar, 18Diehl J.A. Cheng M. Roussel M.F. Sherr C.J. Genes Dev. 1998; 12: 3499-3511Crossref PubMed Scopus (1858) Google Scholar). Rapamycin is a well established immunosuppressive agent that has recently been found to possess anti-angiogenic activities (19Vignot S. Faivre S. Aguirre D. Raymond E. Ann. Oncol. 2005; 16: 525-537Abstract Full Text Full Text PDF PubMed Scopus (597) Google Scholar, 20Sawyers C.L. Cancer Cell. 2003; 4: 343-348Abstract Full Text Full Text PDF PubMed Scopus (178) Google Scholar). Studies of its mechanism of action have shown that when bound to its intracellular receptor, the immunophilin FK506-binding protein-12, rapamycin interacts and inhibits the formation of a complex composed of mTOR, raptor, and mLST8 (called mTORC1) (21Chiu M.I. Katz H. Berlin V. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 12574-12578Crossref PubMed Scopus (411) Google Scholar). Inhibition of mTORC1 results in the hypophosphorylation of p70 S6 kinase and 4E-BP1, which are involved in the control of the translation initiation, ribosome biogenesis, and other growth and proliferation events (22Burnett P.E. Barrow R.K. Cohen N.A. Snyder S.H. Sabatini D.M. Proc. Natl. Acad. Sci. U. S. A. 1998; 95: 1432-1437Crossref PubMed Scopus (932) Google Scholar, 23Sarbassov dos D. Ali S.M. Sabatini D.M. Curr. Opin. Cell Biol. 2005; 17: 596-603Crossref PubMed Scopus (1314) Google Scholar). Another mTOR complex consisting of mTOR, rictor, Sin1, and mLST8 (called mTORC2) has been found to regulate Akt activity and actin polymerization (8Sarbassov D.D. Guertin D.A. Ali S.M. Sabatini D.M. Science. 2005; 307: 1098-1101Crossref PubMed Scopus (5223) Google Scholar, 10Jacinto E. Facchinetti V. Liu D. Soto N. Wei S. Jung S.Y. Huang Q. Qin J. Su B. Cell. 2006; 127: 125-137Abstract Full Text Full Text PDF PubMed Scopus (1139) Google Scholar, 24Jacinto E. Loewith R. Schmidt A. Lin S. Ruegg M.A. Hall A. Hall M.N. Nat. Cell Biol. 2004; 6: 1122-1128Crossref PubMed Scopus (1681) Google Scholar, 25Sarbassov D.D. Ali S.M. Kim D.H. Guertin D.A. Latek R.R. Erdjument-Bromage H. Tempst P. Sabatini D.M. Curr. Biol. 2004; 14: 1296-1302Abstract Full Text Full Text PDF PubMed Scopus (2148) Google Scholar). Although mTORC2 is rapamycin-insensitive, in several studies it has been found that prolonged exposure to rapamycin may inhibit mTORC2 function by blocking the assembly of the newly synthesized complex within the cell (26Sarbassov dos D. Ali S.M. Sengupta S. Sheen J.H. Hsu P.P. Bagley A.F. Markhard A.L. Sabatini D.M. Mol. Cell. 2006; 22: 159-168Abstract Full Text Full Text PDF PubMed Scopus Google Scholar). of intracellular mTOR by rapamycin may in inhibition of mTORC2 and, inhibition of phosphorylation and activation of In this we have and signaling pathways in EC rapamycin to regulate mTOR We find that a major effect of rapamycin is to inhibit Akt phosphorylation and Moreover, we find that the of rapamycin to inhibit the migration and proliferation of EC is of its on mTORC2 and to the by used for Akt, Akt, and all from Cell from and from The rapamycin-resistant mTOR to the by A. H. Science. 1997; PubMed Scopus Google and the wild from D. The of Akt by the with Akt as a R. J. F. A. S. Res. 2004; PubMed Scopus (196) Google Scholar). insulin from and a from Cell endothelial cells from as M.A. Thromb. 3: Google and in with serum EC growth factor, 2 EC used for the between 2 and transfected with the Akt, a of the kinase, and the to the and as J.A. M. D.M. S. J. 2004; PubMed Scopus Google cells for of the In all we used as a control for the Akt or mTOR with and with 1 1 and at for and of on with and a for 1 with in for 1 and with the and with a and the by in cell treated with rapamycin or for in and in CHAPS, 1 1 at for and with of protein with of for at with protein and in in protein and for on a and by by the of serum to the cells treated with rapamycin, or as cells and EC and in The cells in in and by cell In EC transfected with Akt, or with as Akt kinase activity in Akt kinase EC treated with rapamycin for the and cells in to the of protein with the in the Akt with 1 of glycogen synthase kinase-3β in the and on a and phosphorylation of glycogen synthase by as A. C. Ruegg C. Nat. Med. 2001; PubMed Scopus Google Scholar). the of a with and with serum EC used or transfected with or the and in cell for The cells or treated with rapamycin for the migration EC to the of the and after the in and in cells on the of the by in three transfected with or as cells in cell in serum for in the or presence of of EC by in the of cell a cell by of Rapamycin Akt and in that rapamycin Akt activity by Akt phosphorylation of the R.C. Mueckler M. J. Biol. Chem. 2005; 280: 40406-40416Abstract Full Text Full Text PDF PubMed Scopus (509) Google Scholar, dos D. Ali S.M. Sengupta S. Sheen J.H. Hsu P.P. Bagley A.F. Markhard A.L. Sabatini D.M. Mol. Cell. 2006; 22: 159-168Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, K. D. G. M. C. J. I. J.A. Lin M.I. Walsh K. A.M. D.M. M. Sessa W.C. 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Cell. 2006; 22: 159-168Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, F. A. J. Biol. Chem. 2001; Full Text Full Text PDF PubMed Google Scholar, L.S. Sci. 2005; Full Text Full Text PDF PubMed Scopus Google Scholar). as this exposure to rapamycin mTORC1 and in hypophosphorylation of in EC In these studies rapamycin phosphorylation which we to that phosphorylation is for phosphorylation and, for Akt kinase activity by J.R. Mol. Cell. Biol. 2002; 22: PubMed Scopus Google Scholar, J. P. V. D. B.A. D. Mol. Cell. 2002; Full Text Full Text PDF PubMed Scopus Google the phosphorylation of and is for the activation of signaling M. B. P. N. Cohen P. B.A. J. PubMed Scopus Google we to prolonged exposure of EC to rapamycin Akt kinase EC treated with rapamycin for and Akt activity a kinase activity in we found that exposure to rapamycin increased Akt activity, but for than Akt activity are with observations by in 1, and B. further that mTOR mediates Akt phosphorylation in EC, we transfected EC with a a rapamycin-resistant of mTOR A. H. Science. 1997; PubMed Scopus Google and we treated these transfected cells with rapamycin in we found that of in EC the effect of rapamycin on the phosphorylation of to rapamycin mTORC2 in EC, by we the and association between mTOR and in or EC we found that mTOR a complex with in EC and that rapamycin of Taken together these that prolonged exposure of EC to rapamycin Akt phosphorylation and its kinase activity and that this effect is mediated the inhibition of Rapamycin and Akt cytokines as and growth factors as and insulin are well established to induce Akt activity and its signaling (3Gerber H.P. McMurtrey A. Kowalski J. Yan M. Keyt B.A. Dixit V. Ferrara N. J. Biol. Chem. 1998; 273: 30336-30343Abstract Full Text Full Text PDF PubMed Scopus (1741) Google Scholar, 4Hermann C. Assmus B. Urbich C. Zeiher A.M. Dimmeler S. Arterioscler. Thromb. Vasc. Biol. 2000; 20: 402-409Crossref PubMed Scopus (196) Google Scholar, J. Biol. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar). We to rapamycin could inhibit and growth factor-mediated phosphorylation of EC treated for with rapamycin exposure to VEGF, TNF, or insulin, and Akt phosphorylation by the phosphorylation of the and when EC to all three we found that this effect totally by of the cells with are that rapamycin has to inhibit and growth factor-mediated in Rapamycin in we activation of EC results in on known Akt substrates as its pro-survival is well established to EC from undergoing and this function is to be mediated by Akt (3Gerber H.P. McMurtrey A. Kowalski J. Yan M. Keyt B.A. Dixit V. Ferrara N. J. Biol. 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Cell. 1999; 96: 857-868Abstract Full Text Full Text PDF PubMed Scopus (5412) Google Scholar). we found that the phosphorylation of Foxo1 and in of cells with rapamycin as well as Foxo1 and phosphorylation and We these to that the of rapamycin to inhibit Akt phosphorylation in EC has to inhibit the pro-survival function of growth factors as and inhibition of the FoxO pro-apoptotic is of we treated EC with rapamycin and apoptosis in the or presence of serum in in EC, serum withdrawal in of cells undergoing apoptosis after of cells undergoing apoptosis after and after In in the presence of rapamycin, apoptosis of EC after and in with this we by that the inhibition of Akt, and phosphorylation for and after prolonged with rapamycin further effect of Akt on cell we transfected EC with in which the and of Akt are to a of the kinase (called M. B. P. N. Cohen P. B.A. J. PubMed Scopus Google Scholar). we transfected EC with a wild Akt or determined by EC and treated with rapamycin, and after cell determined by with its known pro-survival we found that the of EC with wild Akt apoptosis after serum but that rapamycin apoptosis in these cells of the not apoptosis by serum but the pro-apoptotic function of of EC with the rapamycin-resistant mTOR cells from the pro-apoptotic of rapamycin We these observations to that the of rapamycin to inhibit the assembly of the mTORC2 complex and the inhibition of Akt phosphorylation and activity results in apoptosis of or survival of EC of EC with or in the or presence of of EC determined by and cell in we that and EC from undergoing apoptosis and that this effect totally by we transfected EC with the of Akt in we that this rapamycin from apoptosis in the presence of these results demonstrate that signals are critical for the survival of EC and for from Inhibition of EC and by that mTOR mediates Akt-dependent activation in signals resulting from have been to be functional in EC proliferation and migration (14Morales-Ruiz M. Fulton D. Sowa G. Languino L.R. Fujio Y. Walsh K. Sessa W.C. Circ. Res. 2000; 86: 892-896Crossref PubMed Scopus (349) Google Scholar, 15Fosbrink M. Niculescu F. Rus V. Shin M.L. Rus H. J. Biol. Chem. 2006; 281: 19009-19018Abstract Full Text Full Text PDF PubMed Scopus (71) Google Scholar). the effect of mTORC2 on this response is not We proliferation and migration EC transfected with the of Akt We found that of in in EC proliferation and that rapamycin this response the of on EC migration by rapamycin Taken these that and in EC are by mTORC1 of In this we that mTOR regulates Akt activity in We found that rapamycin serum apoptosis and the protective of and growth in mediated via inhibition of Akt by inhibition of mTORC2 complex Rapamycin the proliferation and migration of EC, which a of inhibition of The between Akt and mTOR and N. Cancer Cell. 2005; 8: Full Text Full Text PDF PubMed Scopus Google Scholar). mTORC2 is composed of a protein complex and mTOR that regulates the phosphorylation of Akt via the residue in the domain. Although mTORC2 is to rapamycin, is in with (26Sarbassov dos D. Ali S.M. Sengupta S. Sheen J.H. Hsu P.P. Bagley A.F. Markhard A.L. Sabatini D.M. Mol. Cell. 2006; 22: 159-168Abstract Full Text Full Text PDF PubMed Scopus Google Scholar, K. D. G. M. C. J. I. J.A. Lin M.I. Walsh K. A.M. D.M. M. Sessa W.C. Cancer Cell. 2006; Full Text Full Text PDF PubMed Scopus Google that of mTOR within a cell by rapamycin inhibit mTORC2 complex In mTORC1 mediates of Akt but a loop to inhibit phosphatidylinositol Akt that this loop in EC in as as we found that exposure of EC to rapamycin inhibits signals Akt In exposure of EC to rapamycin inhibits the association between mTOR and and inhibits Akt phosphorylation (8Sarbassov D.D. Guertin D.A. Ali S.M. Sabatini D.M. 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We find that rapamycin inhibits signaling in EC, that its effect as anti-angiogenic agent may to its to inhibit and Furthermore, we find that rapamycin inhibits the functions of Akt and EC to serum the of rapamycin on EC are to its to inhibit several Akt-inducible Akt is as a major pro-survival and its to cells from undergoing apoptosis is well established (3Gerber H.P. McMurtrey A. Kowalski J. Yan M. Keyt B.A. Dixit V. Ferrara N. J. Biol. Chem. 1998; 273: 30336-30343Abstract Full Text Full Text PDF PubMed Scopus (1741) Google Scholar, S.R. Dudek H. Tao X. Masters S. Fu H. Gotoh Y. Greenberg M.E. Cell. 1997; 91: 231-241Abstract Full Text Full Text PDF PubMed Scopus (4936) Google Scholar, 12Brunet A. Bonni A. Zigmond M.J. Lin M.Z. Juo P. Hu L.S. Anderson M.J. Arden K.C. Blenis J. Greenberg M.E. Cell. 1999; 96: 857-868Abstract Full Text Full Text PDF PubMed Scopus (5412) Google Scholar, J. Cell Dev. Biol. 2004; PubMed Scopus Google Scholar). The FoxO subfamily of forkhead transcription factors as well as are known substrates of Akt that in part facilitate its Foxo1 and are known to be by EC M. Urbich C. K. C. A. R. Zeiher A.M. Dimmeler S. J. 2005; PubMed Scopus Google Scholar). The stimulation of EC with results in the phosphorylation and inactivation of factors in EC, indicating a major for these genes in EC this the of of has been found to inhibit EC survival S. K.C. K. C. Walsh K. W.C. Arterioscler. Thromb. Vasc. Biol. 2004; 24: PubMed Scopus (196) Google Scholar). We that of EC with rapamycin hypophosphorylation of as well as phosphorylation of Foxo1 and Although we could not in EC by not we found that rapamycin the phosphorylation and inactivation of another the protein not these are that mTOR may regulate EC survival via Akt-inducible inactivation of the FoxO transcription with this are and vascular M. Urbich C. K. C. A. R. Zeiher A.M. Dimmeler S. J. 2005; PubMed Scopus Google Scholar, K. Y. M. K. K. H. S. K. K. N. N. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). Akt in EC migration (14Morales-Ruiz M. Fulton D. Sowa G. Languino L.R. Fujio Y. Walsh K. Sessa W.C. Circ. Res. 2000; 86: 892-896Crossref PubMed Scopus (349) Google we that the inhibition of EC migration by rapamycin may be of its effect on We found that of a of Akt in EC proliferation and which by the effect of rapamycin on cell inhibition of p70 S6 kinase and Li F. J.A. Blenis J. Huang S. Oncogene. 2006; PubMed Scopus Google Scholar). the inhibition of EC proliferation by rapamycin of its to inhibit the phosphorylation of Akt and is with the of mTOR to control cell cycle progression p70 S6 kinase and D.C. C.J. C. Blenis J. Mol. Cell. Biol. 2004; 24: PubMed Scopus Google Scholar). rapamycin inhibits activation of Akt, we the that it inhibits migration and proliferation via inhibition of mTORC1 and mTORC2 in this it is that it recently found that of rapamycin in inhibit the phosphorylation of K. D. G. M. C. J. I. J.A. Lin M.I. Walsh K. A.M. D.M. M. Sessa W.C. Cancer Cell. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). In these studies mTOR as a kinase in EC that is of critical in EC and cell We that rapamycin inhibits the phosphorylation and activation of Akt in EC as well as Akt-inducible that mTOR is a kinase involved in Akt-inducible EC activation and survival results are the to demonstrate that the inhibition of Akt-inducible signals in EC by rapamycin results in The of rapamycin to inhibit these signals this agent as a to EC in We for critical of the and we of for We the at for in for the of endothelial and we and for for these we and for