Wu Rongqiang, Xue Peng, Wan Yu, Wang Shizhong, Gu Meng
Medical Research Center, The Affiliated Changzhou No.2 People's Hospital of Nanjing Medical University, Changzhou, 213003, China.
Department of Pediatrics, Changzhou Children's Hospital, Nantong Medical University, Nantong City, China.
In Vitro Cell Dev Biol Anim. 2018 Sep;54(8):600-609. doi: 10.1007/s11626-018-0284-0. Epub 2018 Aug 15.
Uc.40 is a long noncoding RNA that is highly conserved among different species, although its function is unknown. It is highly expressed in abnormal human embryonic heart. We previously reported that overexpression of uc.40 promoted apoptosis and inhibited proliferation of P19 cells, and downregulated PBX1, which was identified as a potential target gene of uc.40. The current study evaluated the effects of uc40-siRNA-44 (siRNA against uc.40) on the differentiation, proliferation, apoptosis, and mitochondrial function in P19 cells, and investigated the relationship between uc.40 and PBX1 in cardiomyocytes. The uc.40 silencing expression was confirmed by quantitative real-time polymerase chain reaction (RT-PCR). Observation of morphological changes in transfected P19 cells during different stages of differentiation revealed that uc40-siRNA-44 increased the number of cardiomyocyes. There was no significant difference in the morphology or time of differentiation between the uc40-siRNA-44 group and the control group. uc40-siRNA-44 significantly promoted proliferation of P19 cells and inhibited serum starvation-induced apoptosis. There was no significant difference in mitochondrial DNA copy number or cellular ATP level between the two groups, and ROS levels were significantly decreased in uc40-siRNA-44-transfected cells. The levels of PBX1 and myocardial markers of differentiation were examined in transfected P19 cells; uc40-siRNA-44 downregulated myocardial markers and upregulated PBX1 expression. These results suggest that uc.40 may play an important role during the differentiation of P19 cells by regulation of PBX1 to promote proliferation and inhibit apoptosis. These studies provide a foundation for further study of uc.40/PBX1 in cardiac development.
Uc.40是一种长链非编码RNA，在不同物种间高度保守，但其功能尚不清楚。它在异常的人类胚胎心脏中高表达。我们之前报道过，uc.40的过表达促进了P19细胞的凋亡并抑制其增殖，还下调了PBX1，而PBX1被确定为uc.40的一个潜在靶基因。当前研究评估了uc40-siRNA-44（针对uc.40的小干扰RNA）对P19细胞分化、增殖、凋亡及线粒体功能的影响，并研究了uc.40与心肌细胞中PBX1的关系。通过定量实时聚合酶链反应（RT-PCR）证实了uc.40的沉默表达。观察转染后的P19细胞在不同分化阶段的形态变化发现，uc40-siRNA-44增加了心肌细胞的数量。uc40-siRNA-44组与对照组在形态或分化时间上没有显著差异。uc40-siRNA-44显著促进了P19细胞的增殖并抑制了血清饥饿诱导的凋亡。两组之间线粒体DNA拷贝数或细胞ATP水平没有显著差异，而在转染了uc40-siRNA-44的细胞中ROS水平显著降低。检测了转染后的P19细胞中PBX1和心肌分化标志物的水平；uc40-siRNA-44下调了心肌标志物并上调了PBX1的表达。这些结果表明，uc.40可能通过调节PBX1在P19细胞分化过程中发挥重要作用，以促进增殖并抑制凋亡。这些研究为进一步研究uc.40/PBX1在心脏发育中的作用奠定了基础。
