Department of Obstetrics, Affiliated Hospital of Weifang Medical University, Weifang, China.
Eur Rev Med Pharmacol Sci. 2020 Feb;24(4):1682-1687. doi: 10.26355/eurrev_202002_20342.
To investigate the biological effect of long non-coding ribonucleic acid (lncRNA) lung cancer-associated transcript 1 (LUCAT1) in the development of ovarian cancer.
Real-time quantitative polymerase chain reaction (RT-qPCR) was utilized to detect the expression levels of lncRNA LUCAT1 in three human ovarian cancer cell lines (CaoV-3, SK-OV-3 and HO-8910) and the normal human ovarian surface epithelial cell line (IOSE80). Small interfering RNAs against lncRNA LUCAT1 (si-LUCAT1) were transfected into SK-OV-3 cells. Transfection efficiency of si-LUCAT1 was verified via RT-qPCR. Cell Counting Kit-8 (CCK-8) and colony formation assays were performed to test the effect of silencing lncRNA LUCAT1 on SK-OV-3 cell proliferation. The apoptosis was measured by flow cytometry. The miRcode database was searched to predict potential microRNAs (miRNAs) binding lncRNA LUCAT1. It was found that lncRNA LUCAT1 contained a highly conserved binding site of miR-199a-5p in the 3'-untranslated region (3'-UTR). Subsequently, the targeting relationship between them was determined through Dual-Luciferase reporter gene assay and RT-qPCR analysis.
LncRNA LUCAT1 was highly expressed in three human ovarian cancer cell lines compared to that in normal ovarian surface epithelial cell line (p<0.05). The cell proliferation rate in SK-OV-3 cells with lncRNA LUCAT1 knockdown was remarkably lower in comparison to that in control group. Moreover, colony formation assay also revealed that the number of cell clones decreased significantly after knockdown of lncRNA LUCAT1 compared to that in control group (p<0.05). In addition, the apoptosis rate was distinctly elevated in the lncRNA LUCAT1 silencing group (p<0.05). Furthermore, a highly conserved binding site of miR-199a-5p was found in the 3'-UTR of lncRNA LUCAT1. Dual-Luciferase reporter gene assay exhibited that the Luciferase activity of LUCAT1-wt was significantly reduced after overexpression of miR-199a-5p (p<0.05), while that of LUCAT1-mut was unchangeable. Further analysis via RT-qPCR suggested that miR-199a-5p overexpression significantly decreased the expression level of lncRNA LUCAT1 (p<0.05).
LncRNA LUCAT1 is overexpressed in ovarian cancer cells, which may target miR-199a-5p to exert its effects on driving the malignant development of ovarian cancer.
研究长链非编码核糖核酸（lncRNA）肺癌相关转录本 1（LUCAT1）在卵巢癌发展中的生物学效应。
利用实时定量聚合酶链反应（RT-qPCR）检测三种人卵巢癌细胞系（CaoV-3、SK-OV-3 和 HO-8910）和正常人类卵巢表面上皮细胞系（IOSE80）中 lncRNA LUCAT1 的表达水平。转染针对 lncRNA LUCAT1 的小干扰 RNA（si-LUCAT1）至 SK-OV-3 细胞。通过 RT-qPCR 验证 si-LUCAT1 的转染效率。细胞计数试剂盒-8（CCK-8）和集落形成实验用于检测沉默 lncRNA LUCAT1 对 SK-OV-3 细胞增殖的影响。通过流式细胞术测量细胞凋亡。搜索 miRcode 数据库以预测潜在的 microRNA（miRNA）与 lncRNA LUCAT1 的结合。发现 lncRNA LUCAT1 在 3'-非翻译区（3'-UTR）中含有 miR-199a-5p 的高度保守结合位点。随后，通过双荧光素酶报告基因检测和 RT-qPCR 分析确定它们之间的靶向关系。
与正常卵巢表面上皮细胞系相比，三种人卵巢癌细胞系中 lncRNA LUCAT1 的表达水平显著升高（p<0.05）。与对照组相比，lncRNA LUCAT1 敲低的 SK-OV-3 细胞的增殖率明显降低。此外，与对照组相比，lncRNA LUCAT1 敲低后细胞克隆数显著减少（p<0.05）。此外，lncRNA LUCAT1 沉默组的细胞凋亡率明显升高（p<0.05）。此外，在 lncRNA LUCAT1 的 3'-UTR 中发现了 miR-199a-5p 的高度保守结合位点。双荧光素酶报告基因检测显示，过表达 miR-199a-5p 后 LUCAT1-wt 的荧光素酶活性显著降低（p<0.05），而 LUCAT1-mut 则不变。进一步通过 RT-qPCR 分析表明，miR-199a-5p 过表达显著降低了 lncRNA LUCAT1 的表达水平（p<0.05）。
lncRNA LUCAT1 在卵巢癌细胞中高表达，可能通过靶向 miR-199a-5p 发挥作用，促进卵巢癌的恶性发展。
