Li Huijuan, Jiang Li, Yu Zhangbin, Han Shuping, Liu Xuehua, Li Mengmeng, Zhu Chun, Qiao Lixing, Huang Li
Department of Pediatrics, Zhongda Hospital Affiliated to Southeast University, Nanjing, Jiangsu, China.
Department of Pediatrics, Zhongda Hospital Affiliated to Southeast University, Nanjing, Jiangsu, China.
Am J Med Sci. 2017 Dec;354(6):608-616. doi: 10.1016/j.amjms.2017.08.019. Epub 2017 Sep 1.
In previous studies, TUC40-, a new long noncoding RNA, was found to be overexpressed in human ventricular septal defect (VSD) embryonic heart samples. In this article, we carried out experiments on the P19 cell line to elucidate the effects of TUC40- overexpression on cardiomyocyte development relevant to VSD pathogenesis.
We established the overexpression cell model by plasmid transfection, and explored the expression profile of Pbx1, the sense gene of TUC40-, and the marker genes of cardiomyocyte linage commitment (Nkx2.5 and GATA4) and maturation (cardiac troponin T). In addition, we combined cell cycle and Cell Counting Kit-8 analysis to detect cell proliferation and used flow cytometry and caspase-3 assays to test apoptosis. At last, bioinformatics analysis was performed to show the possible role of TUC40-.
In the control group, Pbx1 elevated steadily during cardiomyocyte induction; whereas in the overexpression group, it showed significantly lower expression at day 6, 8 and 10 of induction. Cells in the overexpression group failed to induce cardiomyocytes indicated by GATA4 and cardiac troponin T. Proliferation was inhibited possibly owing to G2/M cell cycle arrest and the induced apoptosis rate was higher in the overexpression group.
TUC40- overexpression reduced Pbx1 expression, cardiomyocyte induction and differentiation, inhibited proliferation and promoted apoptosis. Combining the results and previous studies, we propose TUC40- as a potential pathologic factor for VSD.
在先前的研究中，发现一种新的长链非编码RNA——TUC40 -，在人类室间隔缺损（VSD）胚胎心脏样本中过表达。在本文中，我们对P19细胞系进行实验，以阐明TUC40 -过表达对与VSD发病机制相关的心肌细胞发育的影响。
我们通过质粒转染建立过表达细胞模型，并探究TUC40 -的正义基因Pbx1以及心肌细胞谱系定向（Nkx2.5和GATA4）和成熟（心肌肌钙蛋白T）的标记基因的表达谱。此外，我们结合细胞周期和细胞计数试剂盒 - 8分析来检测细胞增殖，并使用流式细胞术和caspase - 3检测来测试细胞凋亡。最后，进行生物信息学分析以显示TUC40 -的可能作用。
在对照组中，Pbx1在心肌细胞诱导过程中稳步升高；而在过表达组中，在诱导的第6、8和10天其表达显著降低。过表达组中的细胞未能诱导出由GATA4和心肌肌钙蛋白T所指示的心肌细胞。增殖可能由于G2/M期细胞周期阻滞而受到抑制，并且过表达组中诱导的凋亡率更高。
TUC40 -过表达降低了Pbx1表达、心肌细胞诱导和分化，抑制了增殖并促进了凋亡。结合这些结果和先前的研究，我们提出TUC40 -作为VSD的潜在病理因素。
