Flamme I, Schulze-Osthoff K, Jacob H J
Institute of Anatomy, Ruhr-University Bochum, FRG.
Development. 1991 Mar;111(3):683-90. doi: 10.1242/dev.111.3.683.
The chorioallantoic membrane (CAM) is one of the most vascularized tissues in the chicken embryo. Capillary growth proceeds until day 10 of development and thereafter abruptly regresses. As it is generally accepted that the formation of new blood vessel is regulated by growth factors, we have investigated the presence of angiogenic and mitogenic factors in the chicken chorioallantois. In the present study, we show that chorioallantoic fluid (CAF) contains angiogenic substances that are probably synthesized in the CAM or the embryonic kidney. When applied in the chorioallantoic membrane assay, CAF from 9 day chicken embryos elicits a strong angiogenic response. This angiogenic activity of CAF is associated with pronounced mitogenic effects in vitro. Comparison of different embryonic fluids reveals that mitogenic activity is particularly evident in the CAF but not detectable in embryonic serum and amnion fluid. Expression of mitogenic activity is found to be temporally correlated with vascular growth in the CAM. High activity is detected in CAF prior to day 10 and then sharply decreases, thus preceding termination of capillary growth by one day. Heparin-sepharose affinity chromatography suggests that the biological activities of CAF probably correspond to the presence of acidic and basic fibroblast growth factor (aFGF and bFGF). In Western blot analyses of CAF, an immunoreactive bFGF-like protein of about 17 x 10(3) Mr is recognized by a monospecific anti-bFGF antiserum. This protein elutes at 2.4 M NaCl from the heparin-sepharose. The mitogenic activity of the CAF can be specifically blocked by the anti-bFGF antibody indicating bFGF to be the active mitogenic principle of the CAF. These results strongly suggest that basic and probably acidic FGF play an important role in the regulation of chorioallantoic vascular growth.
绒毛尿囊膜（CAM）是鸡胚中血管化程度最高的组织之一。毛细血管生长持续到发育第10天，此后突然消退。由于普遍认为新血管的形成受生长因子调节，我们研究了鸡绒毛尿囊中血管生成和促有丝分裂因子的存在情况。在本研究中，我们发现绒毛尿囊液（CAF）含有可能在CAM或胚胎肾中合成的血管生成物质。当应用于绒毛尿囊膜试验时，来自9日龄鸡胚的CAF可引发强烈的血管生成反应。CAF的这种血管生成活性与体外明显的促有丝分裂作用相关。对不同胚胎液的比较显示，促有丝分裂活性在CAF中尤为明显，但在胚胎血清和羊水液中未检测到。发现促有丝分裂活性的表达与CAM中的血管生长在时间上相关。在第10天之前在CAF中检测到高活性，然后急剧下降，从而比毛细血管生长终止提前一天。肝素 - 琼脂糖亲和层析表明，CAF的生物学活性可能与酸性和碱性成纤维细胞生长因子（aFGF和bFGF）的存在相对应。在CAF 的蛋白质免疫印迹分析中，一种约17×10（3）Mr的免疫反应性bFGF样蛋白被单特异性抗bFGF抗血清识别。该蛋白在2.4 M NaCl浓度下从肝素 - 琼脂糖上洗脱。CAF 的促有丝分裂活性可被抗bFGF抗体特异性阻断，表明bFGF是CAF的活性促有丝分裂成分。这些结果强烈表明碱性和成纤维细胞生长因子可能在绒毛尿囊血管生长的调节中起重要作用。
