Chen Yao, Wang Guang, Wang Xiao-yu, Ma Zheng-lai, Chen You-peng, Chuai Manli, von Websky Karoline, Hocher Berthold, Yang Xuesong
Key Laboratory for Regenerative Medicine of the Ministry of Education, Division of Histology and Embryology, Medical College, Guangzhou, China.
Cell Physiol Biochem. 2014;34(3):804-17. doi: 10.1159/000363044. Epub 2014 Aug 20.
BACKGROUND/AIMS: Excess maternal salt intake during pregnancy may alter fetal development. However, our knowledge on how an increased salt intake during pregnancy influences fetal eye development is limited. In this study, we investigated the effects of high-salt treatment on the developing eyes in chick embryos, especially focusing on the development of the retina and the lens.
5.5-day chick embryos were exposed to 280mosm/l (n=17), or 300mosm/l (n=16) NaCl. The treated embryos were then incubated for 96 hours before they were fixed with 4% paraformaldehyde for H&E staining, whole-mount embryo immunostaining and TUNEL staining. BrdU and PH3 incorporation experiments were performed on the chick embryos after high-salt treatment. RT-PCR analyses were conducted from chick retina tissues.
We demonstrated that high-salt treatment altered the size of eyes in chick embryos, induced malformation of the eyes and impaired the development of the lens and the retina. We found an impaired expression of Paired box 6 (PAX6) and neuronal cells in the developing retina as revealed by neurofilament immunofluorescent staining. There was a reduction in the number of BrdU-positive cells and PH3-positive cells in the retina, indicating an impaired cell proliferation with high-salt treatment. High-salt treatment also resulted in an increased number of TUNEL-positive cells in the retina, indicating a higher amount of cell death. RT-PCR data displayed that the expression of the pro-apoptotic molecule nerve growth factor (NGF) in chick retina was increased and CyclinD1 was reduced with high-salt treatment. The size of the lens was reduced and Pax6 expression in the lens was significantly inhibited. High salt-treatment was detrimental to the migration of neural crest cells.
Taken together, our study demonstrated that high-salt exposure of 5.5-day chick embryos led to an impairment of retina and lens development, possibly through interfering with Pax6 expression.
背景/目的：孕期母亲盐摄入量过多可能会改变胎儿发育。然而，我们对于孕期盐摄入量增加如何影响胎儿眼睛发育的了解有限。在本研究中，我们调查了高盐处理对鸡胚发育中眼睛的影响，尤其关注视网膜和晶状体的发育。
将5.5日龄的鸡胚暴露于280mosm/l（n = 17）或300mosm/l（n = 16）的氯化钠中。然后将处理后的胚胎孵育96小时，之后用4%多聚甲醛固定，用于苏木精-伊红染色、全胚胎免疫染色和TUNEL染色。在高盐处理后的鸡胚上进行BrdU和PH3掺入实验。从鸡视网膜组织进行逆转录-聚合酶链反应分析。
我们证明高盐处理改变了鸡胚眼睛的大小，诱导眼睛畸形，并损害了晶状体和视网膜的发育。通过神经丝免疫荧光染色发现，发育中的视网膜中配对盒6（PAX6）和神经元细胞的表达受损。视网膜中BrdU阳性细胞和PH3阳性细胞数量减少，表明高盐处理会损害细胞增殖。高盐处理还导致视网膜中TUNEL阳性细胞数量增加，表明细胞死亡数量增加。逆转录-聚合酶链反应数据显示，高盐处理后鸡视网膜中促凋亡分子神经生长因子（NGF）的表达增加，细胞周期蛋白D1减少。晶状体大小减小，晶状体中Pax6表达受到显著抑制。高盐处理对神经嵴细胞的迁移有害。
综上所述，我们的研究表明，5.5日龄鸡胚暴露于高盐环境会导致视网膜和晶状体发育受损，可能是通过干扰Pax6表达实现的。
