Xanthine oxidase interaction with vascular endothelial growth factor in human endothelial cell angiogenesis.
OBJECTIVE
Reduced capillary density occurs early in cardiovascular diseases. Oxidant stress is implicated in endothelial apoptosis. We investigated the effects of xanthine oxidase (XO) on endothelial survival signaling: protein kinase B/Akt, its cross-talk with p38 MAPK and apoptosis pathways, and its effect on vascular tube formation in vascular endothelial growth factor (VEGF)-simulated human umbilical vein cells.


METHODS
We studied primary cultured human endothelial cells from the umbilical cord. Reactive oxygen species (ROS) production was detected by dihydroethidium staining, cell-signaling pathways by western blots, cell survival by western blots, and nuclear chromatin and angiogenesis response by MTT proliferation assay and three-dimensional Matrigel cultures.


RESULTS
Exogenous XO increased cellular ROS production and caused superoxide-dependent inhibition of Akt phosphorylation and enhancement of p38 MAPK phosphorylation in a time-and dose-dependent manner. In contrast, application of the XO inhibitor oxypurinol or allopurinol inhibited VEGF-stimulated Akt phosphorylation, indicating that endogenous XO promotes VEGF-induced endothelial cell (EC) survival signaling. Exogenous XO induced activation of caspase-3 and reduced expression of the anti-apoptosis protein Bcl-2. Exogenous XO also reduced EC viability, proliferation, and vascular tube formation by p38 MAPK-dependent, phosphoinositide 3-kinase (PI3-K) reversible mechanisms; whereas VEGF promoted EC survival by PI3-K-dependent, p38 MAPK-independent effects.


CONCLUSIONS
Exogenous XO activity is an important contributor to endothelial mechanisms for microvascular rarefaction, by modulation of cell survival signaling pathways; however, endogenous XO is necessary for maintaining EC survival.
Kou
Baijun
B
Department of Clinical Pharmacology & Therapeutics, Clinical Science Research Institute, Warwick Medical School, University of Warwick, UK. koubaijun@yahoo.com
Ni
Jinsong
J
Vatish
Manu
M
Singer
Donald R J
DR
D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology* / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D008833:Microcirculation / Q000502:physiology; D018919:Neovascularization, Physiologic / Q000502:physiology*; D010117:Oxypurinol / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism* / Q000494:pharmacology; D019820:Xanthine / Q000378:metabolism / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D004791:Enzyme Inhibitors; D017382:Reactive Oxygen Species; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D019820:Xanthine; D014969:Xanthine Oxidase; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D048051:p38 Mitogen-Activated Protein Kinases; D010117:Oxypurinol