title: Both Kdr and Flt1 play a vital role in hypoxia-induced Src-PLD1-PKCγ-cPLA(2) activation and retinal neovascularization.
abstract: To understand the mechanisms of Src-PLD1-PKCγ-cPLA2 activation by vascular endothelial growth factor A (VEGFA), we studied the role of Kdr and Flt1. VEGFA, while having no effect on Flt1 phosphorylation, induced Kdr phosphorylation in human retinal microvascular endothelial cells (HRMVECs). Depletion of Kdr attenuated VEGFA-induced Src-PLD1-PKCγ-cPLA2 activation. Regardless of its phosphorylation state, downregulation of Flt1 also inhibited VEGFA-induced Src-PLD1-PKCγ-cPLA2 activation, but only modestly. In line with these findings, depletion of either Kdr or Flt1 suppressed VEGFA-induced DNA synthesis, migration, and tube formation, albeit more robustly with Kdr downregulation. Hypoxia induced tyrosine phosphorylation of Kdr and Flt1 in mouse retina, and depletion of Kdr or Flt1 blocked hypoxia-induced Src-PLD1-PKCγ-cPLA2 activation and retinal neovascularization. VEGFB induced Flt1 tyrosine phosphorylation and Src-PLD1-PKCγ-cPLA2 activation in HRMVECs. Hypoxia induced VEGFA and VEGFB expression in retina, and inhibition of their expression blocked hypoxia-induced Kdr and Flt1 activation, respectively. Furthermore, depletion of VEGFA or VEGFB attenuated hypoxia-induced Src-PLD1-PKCγ-cPLA2 activation and retinal neovascularization. These findings suggest that although VEGFA, through Kdr and Flt1, appears to be the major modulator of Src-PLD1-PKCγ-cPLA2 signaling in HRMVECs, facilitating their angiogenic events in vitro, both VEGFA and VEGFB mediate hypoxia-induced Src-PLD1-PKCγ-cPLA2 activation and retinal neovascularization via activation of Kdr and Flt1, respectively.
authors: Department of Physiology, University of Tennessee Health Science Center, Memphis, TN 38163, USA.
authors: Nikhlesh K
authors: NK
authors: Singh
authors: Dale E
authors: DE
authors: Hansen
authors: Venkatesh
authors: V
authors: Kundumani-Sridharan
authors: Gadiparthi N
authors: GN
authors: Rao
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002465:Cell Movement; D002478:Cells, Cultured; D004261:DNA Replication; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D005260:Female; D005455:Fluorescent Antibody Technique; D054513:Group IV Phospholipases A2 / Q000378:metabolism*; D006801:Humans; D000860:Hypoxia; D051379:Mice; D008810:Mice, Inbred C57BL; D010739:Phospholipase D / Q000378:metabolism*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D015861:Retinal Neovascularization; D014443:Tyrosine / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D042561:Vascular Endothelial Growth Factor B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D040281:Vascular Endothelial Growth Factor Receptor-1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D034741:RNA, Small Interfering; D042461:Vascular Endothelial Growth Factor A; D042561:Vascular Endothelial Growth Factor B; C467480:vascular endothelial growth factor A, mouse; C467482:vascular endothelial growth factor B, mouse; D014443:Tyrosine; C087857:protein kinase C gamma; C501163:Flt1 protein, mouse; D040281:Vascular Endothelial Growth Factor Receptor-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D016392:Proto-Oncogene Proteins pp60(c-src); D011493:Protein Kinase C; D054513:Group IV Phospholipases A2; D010739:Phospholipase D; C106801:phospholipase D1