Ivnitski-Steele Irena D, Walker Mary K
College of Pharmacy, University of New Mexico, Albuquerque, New Mexico 87131, USA.
Birth Defects Res A Clin Mol Teratol. 2003 Jul;67(7):496-503. doi: 10.1002/bdra.10074.
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.
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.
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.
These data suggest that TCDD inhibits early coronary vascular outgrowth via a VEGF-dependent mechanism.
我们之前证明环境污染物2,3,7,8-四氯二苯并对二恶英（TCDD）可在体内减少鸡胚冠状动脉血管的发育。在本研究中，我们评估了TCDD是否抑制冠状动脉内皮管形成和生长的早期事件，以及这种抑制是否通过血管内皮生长因子（VEGF）依赖性机制发生。
在孵化第0天，用对照物（玉米油）或TCDD（0.3 pmol TCDD/克）处理受精鸡蛋。在胚胎第6天，当冠状动脉成血管细胞存在但尚未组装成内皮管时，将心脏心室外植体培养在三维胶原凝胶上。通过免疫组织化学鉴定从外植体迁移出的内皮细胞，并对内皮管的数量和长度进行定量。此外，在孵化第6天和第8天，分别通过逆转录聚合酶链反应（RT-PCR）和酶联免疫吸附测定（ELISA）测量心脏VEGF mRNA和蛋白。
与对照组相比，TCDD外植体中的内皮管长度和数量显著减少（分别为40%±1.7%和36%±3%）。重组外源性VEGF以及用CoCl2或10% O2进行的低氧刺激显著增加了TCDD培养物中生长出的管的长度和数量，并且这种刺激被VEGF中和抗体阻断。相反，VEGF中和抗体仅在对照培养物中减少了管的长度和数量，对TCDD外植体的管生长没有抑制作用。最后，与对照组相比，TCDD处理胚胎的心脏中VEGF mRNA和蛋白均显著减少。
这些数据表明TCDD通过VEGF依赖性机制抑制早期冠状动脉血管生长。
