Roles of ovalbumin on formation of central nerve system in chick embryo

卵清蛋白被归类为丝氨酸蛋白酶抑制剂（丝氨酸蛋白酶抑制剂）的同源物，但除了作为营养来源的可能贡献外，对其功能知之甚少。我们已经证明，在胚胎发育过程中，卵白蛋白的天然分子（n -卵白蛋白）被结合到组织中而不降解，同时经历构型变化，形成一种名为“hs -卵白蛋白”的热稳定形式。为了研究hs -卵清蛋白的必要性，我们建立了体外培养条件，在体外培养条件下，在7期（神经管关闭开始）从鸡蛋中分离出的小鸡胚胎可以正常发育长达24小时。当内源性残余卵清蛋白被免疫阻断时，离体胚胎变得不规则，特别是在脑组织中，很难形成正常胚胎在第9期出现的神经管闭合。神经管闭合失败称为神经管缺损（neural tube defects， NTD）。卵清蛋白抗体处理的胚胎神经管中极凋亡细胞和增殖细胞较少。抗卵清蛋白IgG处理后，在培养基中重新加入足量hs -卵清蛋白，挽救异常胚胎。hs -卵白蛋白不仅比N-卵白蛋白有更高的拯救率，而且比另一种已知的通过体外加热N-卵白蛋白人工产生的热稳定的对应物（s -卵白蛋白）也有更高的拯救率。在NTD胚中，FGF8和Slug的表达极低，而pax3的表达上调5倍。许多形态发生因子和细胞周期因子不受卵清蛋白缺失的影响。这些发现可能是第一个表明卵清蛋白是正常器官发生所必需的蛋白质。

Ovalbumin has been classified as a homolog of serine protease inhibitor (serpin), but less is known for its function except for a possible contribution as a nutrient source. We have shown that, during embryonic development, native molecules of ovalbumin (N-ovalbumin) are incorporated into tissues without degradation, while undergoing configurational changes into a heat-stable form named 'HS-cvalbumin. ' Here, to investigate the indispensability of HS-ovalbumin, we established in vitro culture conditions, under which chick embryos, isolated from eggs at stage 7, when closure of neural tube begins, could develop normally for up to 24 hours. When endogenous, remnant ovalbumin was blocked immunologically, the isolated embryos became irregular particularly in the brain tissue, hardly forming the neural tube closure which would occur in normal embryos at stage 9. The failure of closure of neural tube is so-called neural tube defects (NTD). The neural tube of the embryo treated with ovalbumin-antibody had less extremely apoptotic cells and proliferating cells. The abnormal embryos were rescued by re-adding a sufficient amount of HS-ovalbumin to the medium after treating anti-ovalbumin IgG. HS-ovalbumin showed a higher rescue rate compared not only to N- ovalbumin but also to another heat stable counterpart (S-ovalbumin) known to occur artificially by heating N-ovalbumin in vitro. In NTD embryo, expression of FGF8 and Slug has decreased extremely, while Pax 3 up-regulated 5 folds. Many morphogenic and cell cyclic factors were not affected by depletion of ovalbumin. These findings may be the first indication that ovalbumin is an essential protein involved in the normal organogenesis.