高糖环境诱导的过度自噬抑制鸡胚颅面部神经嵴产生

发布时间：2018-02-11 01:12

本文关键词： 颅面部神经嵴高糖环境自噬细胞凋亡鸡胚　出处：《暨南大学》2015年博士论文　论文类型：学位论文

【摘要】：目的高糖环境会导致胚胎中颅面部神经嵴参与形成的组织和器官的畸形,然而其机制仍未得以阐明。本课题利用早期鸡胚为模型,探索高糖影响颅面部神经嵴发育的细胞及其分子机制。方法鸡胚经高糖处理后,首先通过阿利新蓝/茜素红染色观察由颅面部神经嵴细胞衍生而来的颅面部骨骼。其后,利用免疫荧光,原位杂交,q RT-PCR,Western blot、透射电镜等方法对高糖环境下颅面部神经嵴细胞中关键基因表达,增殖,凋亡和自噬进行观察和检测,探索自噬对颅面部神经嵴产生的影响机制。并结合HEK293T细胞探索高糖诱导的自噬与凋亡之间的关系。结果在此研究中,我们首先观察到了高糖环境导致的由颅面部神经嵴衍生而来的鸡胚颅顶骨骼的发育缺陷,尤其是顶骨的发育缺陷。在早期鸡胚中,我们发现高糖环境抑制颅面部神经嵴的产生,而神经嵴细胞的增殖并未受到显著影响。然而高糖环境促进了颅面部神经嵴细胞的死亡。并且高糖环境能够促进HEK293T细胞中凋亡和自噬相关基因的表达,包括P53,cleaved-Caspase3、cleaved-Caspase7,LC3B,truncated-Atg5以及Beclin-1。通过微珠实验结合自噬抑制剂(氯喹)和激动剂(衣霉素)处理鸡胚,从功能上证实了自噬涉及高糖环境对神经嵴细胞产生的抑制作用。而高糖环境激活自噬主要是通过ERK信号通路而非m TOR通路实现的。结论我们的结果表明胚胎暴露于高糖环境中会抑制颅面部神经嵴的产生,这种抑制作用可能是由于高糖引起的过度自噬导致了神经嵴细胞的死亡增加。
[Abstract]:Objective High glucose environment may lead to the deformity of tissues and organs involved in the formation of craniofacial nerve crest in embryos, but its mechanism has not been elucidated. In this study, early chicken embryos were used as model. Methods after high glucose treatment, the craniofacial bone derived from cranio-facial nerve crest cells was observed by Alisin blue / alizarin red staining. The expression, proliferation, apoptosis and autophagy of key genes in cranio-facial nerve crest cells were observed and detected by immunofluorescence, in situ hybridization and Western blot, transmission electron microscopy (TEM). To explore the mechanism of autophagy on craniofacial neural crest and to explore the relationship between high glucose induced autophagy and apoptosis in combination with HEK293T cells. We first observed the developmental defects of the craniocephalic bone, especially the parietal bone, induced by high glucose environment. In the early chicken embryos, we found that high glucose environment inhibited the formation of cranio-facial neural crest. But the proliferation of neural crest cells was not significantly affected. However, high glucose environment promoted the death of craniofacial neural crest cells. Furthermore, high glucose environment promoted apoptosis and autophagy related gene expression in HEK293T cells. Including P53 cleaved-Caspase3, cleaved-cleaved-caspase7, LC3Bnctruncated-Atg5 and Beclin-1. Chicken embryos were treated with microbead test combined with autophagy inhibitor (chloroquine) and agonist (chlortetracycline). It is proved that autophagy is related to the inhibitory effect of high glucose environment on neural crest cells, while high glucose environment activates autophagy mainly through ERK signaling pathway rather than m TOR pathway. Conclusion our results indicate that embryonic burst. Exposure to high glucose conditions inhibits the production of craniofacial nerve crest. This inhibition may be due to hyperglycemic autophagy resulting in increased death of neural crest cells.
【学位授予单位】：暨南大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R321.5

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