title: Transactivation of fetal liver kinase-1/kinase-insert domain-containing receptor by lysophosphatidylcholine induces vascular endothelial cell proliferation.
abstract: Lysophosphatidylcholine (LPC), a major lipid component of oxidized low-density lipoprotein, is a bioactive lipid molecule involved in numerous biological processes including the progression of atherosclerosis. Recently orphan G protein-coupled receptors were identified as high-affinity receptors for LPC. Although several G protein-coupled receptor ligands transactivate receptor tyrosine kinases, LPC-stimulated transactivation of receptor tyrosine kinase has not yet been reported. Here we observed for the first time that LPC treatment of human umbilical vein endothelial cells (HUVECs) induces tyrosyl phosphorylation of vascular endothelial growth factor receptor 2 [fetal liver kinase-1/kinase-insert domain-containing receptor, Flk-1/KDR)]. Flk-1/KDR transactivation by LPC was inhibited by vascular endothelial growth factor receptor tyrosine kinase inhibitors, SU1498 and 4-[(4'-chloro-2'-fluoro) phenylamino]6,7-dimethoxyquinazoline (VTKi) in immunoprecipitation. Furthermore, we examined the effects of the Src family kinases inhibitors, herbimycin A and 4-amino-5-(4-chlorophenyl)-7-(t-butyl) pyrazolo[3,4-d] pyrimidine (PP2), on LPC-induced Flk-1/KDR transactivation. Results from Western blots, c-Src is involved in LPC-induced Flk-1/KDR transactivation because herbimycin A and PP2 inhibited this transactivation. Kinase-inactive (KI) Src transfection also inhibited LPC-induced Flk-1/KDR transactivation. In addition, results from Western blots, ERK1/2 and Akt, which are downstream effectors of Flk-1/KDR, were also activated by LPC, and this was inhibited by SU1498, VTKi, herbimycin A, PP2, and KI Src transfection in HUVECs. LPC-induced stimulation of HUVEC proliferation was shown to be secondary to transactivation because it was suppressed by SU1498, VTKi, herbimycin A, PP2, and KI Src transfection in dimethylthiazoldiphenyltetra-zoliumbromide assay. These findings suggest that LPC-induced Flk-1/KDR transactivation via c-Src may have important implications for the progression of atherosclerosis.
authors.affiliation: Department of Pharmacology, The University of Tokushima Graduate School of Medical Sciences, 3-18-15 Kuramoto, Tokushima 770-8503, Japan. fujita56@ri.ncvc.go.jp
authors.forename: Yoshiko
authors.initials: Y
authors.lastname: Fujita
authors.forename: Masanori
authors.initials: M
authors.lastname: Yoshizumi
authors.forename: Yuki
authors.initials: Y
authors.lastname: Izawa
authors.forename: Nermin
authors.initials: N
authors.lastname: Ali
authors.forename: Hideki
authors.initials: H
authors.lastname: Ohnishi
authors.forename: Yasuhisa
authors.initials: Y
authors.lastname: Kanematsu
authors.forename: Keisuke
authors.initials: K
authors.lastname: Ishizawa
authors.forename: Koichiro
authors.initials: K
authors.lastname: Tsuchiya
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Tamaki
mesh_terms: D050197:Atherosclerosis / Q000473:pathology; D016227:Benzoquinones; D015153:Blotting, Western; D000081247:CSK Tyrosine-Protein Kinase; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D002934:Cinnamates / Q000494:pharmacology; D018450:Disease Progression; D042783:Endothelial Cells / Q000166:cytology*; D004730:Endothelium, Vascular / Q000166:cytology*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D047029:Lactams, Macrocyclic; D008024:Ligands; D008244:Lysophosphatidylcholines / Q000378:metabolism*; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D011799:Quinazolines / Q000494:pharmacology; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D015533:Transcriptional Activation; D014162:Transfection; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics* / Q000378:metabolism*; D019061:src-Family Kinases
chemical_list: C412373:AG 1879; D016227:Benzoquinones; C508740:CB676475; D002934:Cinnamates; D047029:Lactams, Macrocyclic; D008024:Ligands; D008244:Lysophosphatidylcholines; D011743:Pyrimidines; D011799:Quinazolines; D011809:Quinones; C483044:SU 1498; D017828:Rifabutin; C020754:herbimycin; D011505:Protein-Tyrosine Kinases; D040301:Vascular Endothelial Growth Factor Receptor-2; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human