Vascular endothelial growth factor rescues 2,3,7,8-tetrachlorodibenzo-p-dioxin inhibition of coronary vasculogenesis.
BACKGROUND
We previously demonstrated that the environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) reduces coronary vascular development in chick embryos in vivo. In the current study, we assessed whether TCDD inhibits early events in coronary endothelial tube formation and outgrowth, and whether this inhibition occurs through a vascular endothelial growth factor (VEGF)-dependent mechanism.


METHODS
Fertile chicken eggs were treated with control (corn oil) or TCDD (0.3 pmol TCDD/g) on incubation day 0. On embryonic day 6, cardiac ventricle explants were cultured on a three-dimensional collagen gel, when coronary angioblasts are present, but prior to their assembly into endothelial tubes. Endothelial cells migrating out from explants were identified by immunohistochemistry, and endothelial tube number and length were quantitated. In addition, on incubation days 6 and 8, cardiac VEGF mRNA and protein were measured by reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively.


RESULTS
Endothelial tube length and number were significantly reduced (40% +/- 1.7% and 36% +/- 3%, respectively) in TCDD explants, compared to controls. Recombinant exogenous VEGF, as well as hypoxic stimulation with CoCl2 or 10% O2, significantly increased the length and number of outgrowing tubes in TCDD cultures, and this stimulation was prevented by a VEGF neutralizing antibody. In contrast, VEGF neutralizing antibody reduced the length and number of tubes only in control cultures, and had no inhibitory effect on tube outgrowth from TCDD explants. Finally, hearts from TCDD-treated embryos exhibited a significant reduction in both VEGF mRNA and protein, compared to controls.


CONCLUSIONS
These data suggest that TCDD inhibits early coronary vascular outgrowth via a VEGF-dependent mechanism.
Ivnitski-Steele
Irena D
ID
College of Pharmacy, University of New Mexico, Albuquerque, New Mexico 87131, USA.
Walker
Mary K
MK
D000818:Animals; D002642:Chick Embryo; D003314:Corn Oil / Q000008:administration & dosage; D003331:Coronary Vessels / Q000187:drug effects / Q000196:embryology*; D004730:Endothelium, Vascular / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006321:Heart / Q000187:drug effects / Q000196:embryology*; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D009924:Organ Culture Techniques; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity*; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013723:Teratogens / Q000633:toxicity*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000494:pharmacology*
D000072317:Polychlorinated Dibenzodioxins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D013723:Teratogens; D042461:Vascular Endothelial Growth Factor A; D003314:Corn Oil