Yan Jin, Zhang Yifeng, She Qiang, Li Xuan, Peng Lei, Wang Xiaoyong, Liu Shiyu, Shen Xiaoran, Zhang Weifeng, Dong Yu, Lu Jieyu, Zhang Guoxin
Department of Gastroenterology, The First Affiliated Hospital with Nanjing Medical University, Nanjing, China.
The First Clinical Medical College, Nanjing Medical University, Nanjing, China.
Cell Physiol Biochem. 2017;42(6):2364-2376. doi: 10.1159/000480028. Epub 2017 Aug 18.
Long noncoding RNA (lncRNA) H19 is emerging as a vital regulatory molecule in the progression of different types of cancer and miR-675 is reported to be embedded in H19's first exon. However, their function and specific mechanisms of action have not been fully elucidated. The aim of this study was to identify a novel lncRNA-microRNA-mRNA functional network in gastric cancer.
Quantitative real-time polymerase chain reaction (qRT-PCR) was used to assess the relative expression of H19 and miR-675 in normal (GES-1) and gastric cancer cell lines (SGC-7901, SGC-7901/DDP) as well as in tumor tissues. Gain and loss of function approaches were carried out to investigate the potential roles of H19/miR-675 in cell proliferation and apoptosis. Moreover, Fas associated via death domain (FADD) was validated to be the target of miR-675 via luciferase reporter assay. Western blotting was used to evaluate the protein expression of related signaling pathway.
In our study H19 and miR-675 were increased in gastric cancer cell lines and tissues. Overexpression of H19 and miR-675 promoted cell proliferation and inhibited cell apoptosis, whereas knockdown of H19 and miR-675 inhibited these effects. By further examining the underlying mechanism, we showed that H19/miR-675 axis inhibited expression of FADD. FADD downregulation subsequently inhibited the caspase cleavage cascades including caspase 8 and caspase 3.
Taken together, our results point to a novel regulatory pathway H19/miR-675/ FADD/caspase 8/caspase 3 in gastric cancer which may be potential target for cancer therapy.
长链非编码RNA（lncRNA）H19正逐渐成为不同类型癌症进展中的重要调节分子，且据报道miR-675嵌入在H19的第一个外显子中。然而，它们的功能和具体作用机制尚未完全阐明。本研究的目的是在胃癌中鉴定一种新的lncRNA-微小RNA-信使RNA功能网络。
采用定量实时聚合酶链反应（qRT-PCR）评估H19和miR-675在正常（GES-1）和胃癌细胞系（SGC-7901、SGC-7901/DDP）以及肿瘤组织中的相对表达。采用功能获得和缺失方法研究H19/miR-675在细胞增殖和凋亡中的潜在作用。此外，通过荧光素酶报告基因检测验证死亡结构域相关蛋白（FADD）是miR-675的靶标。采用蛋白质印迹法评估相关信号通路的蛋白表达。
在我们的研究中，H19和miR-675在胃癌细胞系和组织中表达增加。H19和miR-675的过表达促进细胞增殖并抑制细胞凋亡，而敲低H19和miR-675则抑制这些作用。通过进一步研究潜在机制，我们发现H19/miR-675轴抑制FADD的表达。FADD下调随后抑制了包括半胱天冬酶8和半胱天冬酶3在内的半胱天冬酶裂解级联反应。
综上所述，我们的结果表明胃癌中存在一条新的调节途径H19/miR-675/FADD/半胱天冬酶8/半胱天冬酶3，这可能是癌症治疗的潜在靶点。
