ErbB信号通路基因GAB1、EPS15异常表达与心肌细胞凋亡及心脏发育异常的相关性研究

Excessive cardiomyocyte apoptosis is related directly to the onset of congenital heart disease. Diabetes mellitus in pregnancy is one of the maternal diseases that cause congenital birth defects in infants. Excessive cardiomyocyte apoptosis induced by hyperglycemia plays a critical role in cardiac malformations. However, the underlying molecular mechanism remains unknown. In our study, we analyzed genome-wide expression profiling in the cardiomyocytes treated with or without high-glucose. Microarray analysis revealed that a total of 364 genes changes in mRNA expression. Further bioinformatics analysis revealed that some of genes which involved in ErbB signaling pathway such as GAB1 and EPS15 were differently expressed in apoptotic cardiomyocyte. Based on our finding and related studies, we intend to perform further research on the molecular mechanism of GAB1 and EPS15 on high-glucose induced cardiomyocyte apoptosis. Furthermore, pregnant Rats with diabetic mellitus were generated to investigate the association between abnormal expression of ErbB/PI3K signaling pathway related genes and cardiac malformations. In conclusion, our study discovered a series of novel genes that related with apoptosis from genome-wide expression profiling of cardiomyocyte. We also investigated the detailed molecular mechanism in cardiomyocyte apoptosis and diabetic embryopathy. These results provided experimental evidence that hyperglycemia could induce apoptosis and affect the normal development of heart. We also provide some theoretical foundation for potential etiology of congenital heart disease genesis.

心肌细胞的异常凋亡与先天性心脏病发生密切相关，临床研究表明母亲妊娠期糖尿病是儿童先天性心脏病发生的病因之一, 主要原因可能由于高糖诱导心肌细胞过度凋亡导致胎儿心脏发育异常,但其具体致病机制仍不很清楚。我们课题组建立了高糖诱导的人心肌细胞凋亡模型，通过表达谱基因芯片技术,筛选出364个差异表达基因，生物信息学分析发现ErbB通路相关基因GAB1和EPS15在凋亡心肌细胞中表达异常。本项目拟在此基础上深入研究GAB1和EPS15及ErbB/PI3K等通路在高糖诱导心肌细胞凋亡中的作用机制，并建立高血糖妊娠大鼠模型，进一步明确GAB1和EPS15异常表达与胚胎心脏发育异常的相关性。本课题从心肌细胞的转录基因组筛选出与心肌细胞凋亡相关的GAB1和EPS15等一系列基因，并深入研究其作用机制，为高糖引起心肌细胞凋亡进而影响心脏发育提供了新的实验基础，也为先天性心脏病致病机制研究，提供新理论依据。

先天性心脏病(简称先心病)是由环境因素和遗传因素共同作用所导致的多基因复杂疾病。临床研究发现，妊娠期高血糖可导致胎儿出现各种各样的先天畸形，其中最主要的一类为心脏缺陷，但是，目前有关宫内高糖环境引起胚胎心脏发育异常作用机制的研究并不多见。近期研究发现高糖诱导心肌细胞过度凋亡很可能是导致胎儿心脏发育异常的重要原因之一,但其具体致病机制仍不很清楚。本课题通过建立细胞和动物模型, 结合表达谱芯片数据和分子细胞学相关实验技术手段, 深入探索高糖环境对心肌细胞的状态和心脏发育的影响。通过实验研究发现， GAB1、ROCK1、EPS15、 CITED2和 NOXA等基因在凋亡的心肌细胞中表达异常。具体体现为：在高糖环境下，心肌细胞中ROCK1、EPS15和NOXA表达升高，GAB1、 CITED2和XIAP表达降低。通过western blotting，qRT-PCR, TUNEL，Co-IP等细胞分子生物学相关技术进行深入研究发现, ROCK1介导的p53路径和GAB1介导的PI3K/Akt通路参与调节高糖诱导心肌细胞凋亡这一重要过程。在完成本课题的工作中还发现, 除了高浓度葡萄糖引起细胞凋亡之外，中浓度葡萄糖也对心肌细胞有重要影响,中浓度葡萄糖可以诱导心肌细胞发生肥大，我们对此深入进行机制研究，发现SOX9基因和Nodal通路参与心肌细胞肥大的发生。这部分工作的补充更为全面揭示不同浓度的葡萄糖对心肌细胞效应作用及相关机制。本项目的完成为妊娠期高血糖致胎儿心脏发育异常的病因探索提供重要的实验基础，也为先天性心脏病致病机制研究，提供新理论依据，也为相关分子标志物的研发提供前期的实验依据。