Circular RNA CircHIPK3 Promotes NCI-H1299 and NCI-H2170 Cell Proliferation through miR-379 and its Target IGF1
- 生命科学－已发表论文 
背景与目的已有的研究证明:环状RNA是一类在哺乳动物中普遍存在的具有稳定闭合环状结构的内源性RNA分子。环状RNA; circHIPK3(circular RNA HIPK3, circHIPK3)在肝细胞癌(hepatocellular carcinoma,; HCC)中表达水平较高,促进肝癌细胞生长。但是其在非小细胞肺癌(non-small cell lung cancer,; NSCLC)中的作用及其调控机制尚无文献报道。本研究拟探讨环状RNA; circHIPK3对NSCLC细胞系NCI-H1299和NCI-H2170细胞增殖的影响,并进一步研究其调控的分子机制。方法Real-time; PCR法检测circHIPK3在NSCLC各细胞系中的表达水平。CCK-8实验和克隆形成实验检测过量表达和干扰circHIPK3对细胞增殖的影响; 。双荧光素酶报告基因实验分别检验miR-379与circHIPK3及miR-379与IGF1 mRNA的结合情况。Western; blot和ELISA检测细胞内外的IGF1蛋白表达水平。结果环状RNA; circHIPK3在6株NSCLC细胞株中均有表达,其中H1299表达最低,H2170表达最高,通过转染过表达的circHIPK3可显著促进NC; I-H1299细胞增殖,干扰circHIPK3可显著抑制NCI-H2170细胞增殖。miR-379可与circHIPK3及IGF1; mRNA直接结合。过表达circHIPK3导致IGF1表达量上调,干扰circHIPK3能够下调IGF1表达水平,转入miR-379; mimics恢复了circHIPK3稳转细胞株IGF1表达水平的上调及细胞增殖表型。结论环状RNA; circHIPK3在NSCLC细胞系NCI-H1299及NCI-H2170中可通过miR-379调控IGF1表达促进细胞增殖,环状RNA; circHIPK3可能成为非小细胞肺癌治疗的新靶点。Background and objective It has been proven that the circular RNA,; possessing a stable covalently closed continuous loop, is a type of RNA; molecule which is expressed widespread in mammals. The circular RNA; circHIPK3 is abundantly expressed in hepatocellular carcinoma (HCC) and; promotes tumourgenesis. However, a role for circHIPK3 has not been; systematically examined in non-small cell lung cancer (NSCLC). In this; study, we investigated whether circHIPK3 has an effect on cell; proliferation in the NSCLC cell lines NCI-H1299 and NCI-H2170 and the; underlying molecular mechanisms. Methods The expression of circHIPK3 was; measured by real-time PCR in NSCLC cell lines. Nuclear mass separation; experiment was used to detect the location of circHIPK3 in cells. The; gain and loss function experiments were used to examine the; proliferation of NCI-H1299 and NCI-H2170 cells by Cell Counting Kit-8; (CCK-8) and the colony formation assays. Then, circHIPK3 was cloned into; the downstream of the luciferase reporter gene which activity was; detected to verify whether miR-379 could bind with circHIPK3 or IGF1; mRNA. The protein level of IGF1 was detected by Western blot and ELISA; in circHIPK3 overexpressed/knock-down NCI-H1299 and NCI-H2170 cells.; Results CircHIPK3 was generally expressed in six kinds of NSCLC cells; lines we detected, and the expression level was highest in H2170 and; lowest in H1299. Overexpression of circHIPK3 obviously promoted; NCI-H1299 cell proliferation and knock-down of circHIPK3 inhibited; NCI-H2170 cell proliferation. In the luciferase assay, miR-379 was; observed to sequester circHIPK3 and IGF1 mRNA with potential binding; sites. Furthermore, we found that the overexpression of circHIPK3 could; increase the expression levels of IGF1 and knockdown reduced it.; Moreover, up-regulation of miR-379 rescued the phenotype induced by; overexpression of circHIPK3. Conclusion CircHIPK3 could promote cell; proliferation by a circHIPK3/miR-379 pathway in NCI-H1299 and NCI-H2170; cells and might be a potential tumor biomarker for NSCLC.