title: Losartan metabolite EXP3179 activates Akt and endothelial nitric oxide synthase via vascular endothelial growth factor receptor-2 in endothelial cells: angiotensin II type 1 receptor-independent effects of EXP3179.
abstract: BACKGROUND
  Recent studies suggest that angiotensin type 1 receptor (AT1R) blockers have vascular protective effects beyond blood pressure lowering. Because of the importance of endothelial nitric oxide synthase (eNOS) in vascular and platelet function, we hypothesized that losartan and its metabolites would stimulate eNOS and its upstream activators Akt and phosphatidylinositol 3-kinase (PI3K).
  
  
  RESULTS
  Losartan is metabolized into EXP3174 (AT1R-blocking metabolite) and EXP3179 (no AT1R-blocking properties). Treatment of endothelial cells (ECs) with losartan and both metabolites stimulated phosphorylation of Akt and eNOS in the absence of angiotensin II. However, the magnitude for EXP3179 was much greater than EXP3174, and the EC50 was significantly lower (-logEC50, 8.2+/-0.1 versus 5.4+/-0.2 mol/L), suggesting an AT1R-independent effect. Inhibiting PI3K or vascular endothelial growth factor receptor 2 (VEGFR2) tyrosine phosphorylation abrogated EXP3179-induced eNOS phosphorylation. In endothelium of intact rat aorta, EXP3179 also stimulated Akt and eNOS phosphorylation. VEGFR2 activation was shown to be calcium and Src family kinase dependent by use of specific drug inhibitors and dominant negative kinase transfection. EXP3179 significantly inhibited tumor necrosis factor alpha-induced apoptosis by approximately 60% (from 30.1+/-5.8% to 12.2+/-2.0% TUNEL-positive cells), which was abolished by pretreatment with the PI3K inhibitor LY294002. Cleaved caspase-3 was suppressed by 48% with EXP3179.
  
  
  CONCLUSIONS
  The losartan metabolite EXP3179 stimulates eNOS phosphorylation and suppresses tumor necrosis factor alpha-induced EC apoptosis by activating the VEGFR2/PI3K/Akt pathway.
authors.affiliation: Cardiovascular Research Institute, Department of Medicine, University of Rochester, Rochester, NY 14642, USA.
authors.forename: Tetsu
authors.initials: T
authors.lastname: Watanabe
authors.forename: Jun
authors.initials: J
authors.lastname: Suzuki
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Yamawaki
authors.forename: Virendra K
authors.initials: VK
authors.lastname: Sharma
authors.forename: Shey-Shing
authors.initials: SS
authors.lastname: Sheu
authors.forename: Bradford C
authors.initials: BC
authors.lastname: Berk
mesh_terms: D047228:Angiotensin II Type 1 Receptor Blockers / Q000494:pharmacology*; D000818:Animals; D001011:Aorta; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology*; D019808:Losartan / Q000493:pharmacokinetics* / Q000031:analogs & derivatives; D009025:Morpholines / Q000494:pharmacology; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D051381:Rats; D044140:Receptor, Angiotensin, Type 1 / Q000502:physiology*; D013777:Tetrazoles / Q000494:pharmacology*; D014162:Transfection; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000502:physiology*
chemical_list: D047228:Angiotensin II Type 1 Receptor Blockers; D002867:Chromones; D004791:Enzyme Inhibitors; D007093:Imidazoles; D009025:Morpholines; D044140:Receptor, Angiotensin, Type 1; D013777:Tetrazoles; C073600:2-butyl-4-chloro-1-((2'-(1H-tetrazol-5-yl)(1,1'-biphenyl)-4-yl)methyl)-1H-imidazole-5-carboxaldehyde; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D052250:Nitric Oxide Synthase Type III; D040301:Vascular Endothelial Growth Factor Receptor-2; D051057:Proto-Oncogene Proteins c-akt; D019808:Losartan; D002118:Calcium