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

卵清蛋白已被归类为丝氨酸蛋白酶抑制剂(Serpin)的同系物，但人们对其功能知之甚少，除了可能作为营养来源的贡献外。我们已经证明，在胚胎发育过程中，天然的卵清蛋白分子(N-卵清蛋白)在经历构型变化时不会降解地被结合到组织中，形成一种热稳定的形式，称为HS-Cvalb。在这里，为了探讨HS-卵清蛋白的必要性，我们建立了体外培养条件，在此条件下，从7期卵子中分离出的鸡胚胎，当神经管开始关闭时，可以正常发育长达24小时。当免疫阻断内源性残留卵白蛋白时，分离的胚胎变得不规则，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。神经管闭合的失败被称为神经管缺陷(NTD)。经卵白蛋白抗体处理的胚胎神经管内极度凋亡细胞和增殖细胞较少。在处理抗卵白蛋白Ig G后，通过在培养液中再次添加足量的HS-卵白蛋白，挽救了异常胚胎。HS-卵白蛋白不仅比N-卵白蛋白具有更高的救援率，而且与另一种已知的人工加热N-卵白蛋白而产生的热稳定性类似物(S-卵白蛋白)相比，具有更高的救援率。在NTD胚胎中，FGF8和Slug的表达显著降低，而Pax3的表达上调了5倍。许多形态发生因子和细胞周期因子不受卵清蛋白耗竭的影响。这些发现可能是第一个表明卵清蛋白是参与正常器官发生的必要蛋白质。

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.