长链非编码 RNA NNT-AS1 通过下调 miR-203 促进胆管癌细胞增殖和上皮-间充质转化。
Long non-coding RNA NNT-AS1 promotes cholangiocarcinoma cells proliferation and epithelial-to-mesenchymal transition through down-regulating miR-203.
机构信息
Emergency Intensive Care Unit, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China.
Department of Cardiology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China.
出版信息
Aging (Albany NY). 2020 Feb 5;12(3):2333-2346. doi: 10.18632/aging.102747.
BACKGROUND
Cholangiocarcinoma (CCA) is a serious malignant tumor. Long non-coding RNA NNT-AS1 (NNT-AS1) takes crucial roles in several tumors. So, we planned to research the roles and underlying mechanism of NNT-AS1 in CCA.
RESULTS
NNT-AS1 overexpression was appeared in CCA tissues and cell lines. Proliferation was promoted by NNT-AS1 overexpression in CCLP1 and TFK1 cells. Besides, NNT-AS1 overexpression reduced E-cadherin level and raised levels of N-cadherin, vimentin, Snail and Slug. However, the opposite trend was occurred by NNT-AS1 knockdown. Further, NNT-AS1 overexpression promoted phosphatidylinositol 3 kinase (PI3K)/AKT and extracellular signal-regulated kinase (ERK)1/2 pathways. MiR-203 was sponged by NNT-AS1 and miR-203 mimic reversed the above promoting effects of NNT-AS1. Additionally, insulin-like growth factor type 1 receptor (IGF1R) and zinc finger E-box binding homeobox 1 (ZEB1) were two potential targets of miR-203.
CONCLUSION
NNT-AS1 promoted proliferation, EMT and PI3K/AKT and ERK1/2 pathways in CCLP1 and TFK1 cells through down-regulating miR-203.
METHODS
CCLP1 and TFK1 cells were co-transfected with pcDNA-NNT-AS1 and miR-203 mimic. Bromodeoxyuridine (BrdU), flow cytometry, quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot were employed to detect roles and mechanism of NNT-AS1. Interaction between NNT-AS1 and miR-203 or miR-203 and target genes was examined through luciferase activity experiment.
背景
胆管癌（CCA）是一种严重的恶性肿瘤。长链非编码 RNA NNT-AS1（NNT-AS1）在几种肿瘤中发挥着关键作用。因此，我们计划研究 NNT-AS1 在 CCA 中的作用和潜在机制。
结果
NNT-AS1 在 CCA 组织和细胞系中呈现高表达。在 CCLP1 和 TFK1 细胞中，NNT-AS1 的过表达促进了细胞增殖。此外，NNT-AS1 的过表达降低了 E-钙黏蛋白的水平，提高了 N-钙黏蛋白、波形蛋白、Snail 和 Slug 的水平。然而，NNT-AS1 敲低则出现相反的趋势。进一步研究发现，NNT-AS1 的过表达促进了磷脂酰肌醇 3 激酶（PI3K）/蛋白激酶 B（AKT）和细胞外信号调节激酶（ERK）1/2 通路。NNT-AS1 可以吸附 miR-203，miR-203 模拟物逆转了 NNT-AS1 的上述促进作用。此外，胰岛素样生长因子 1 受体（IGF1R）和锌指 E 盒结合同源盒 1（ZEB1）是 miR-203 的两个潜在靶点。
结论
NNT-AS1 通过下调 miR-203 促进了 CCLP1 和 TFK1 细胞的增殖、上皮间质转化（EMT）以及 PI3K/AKT 和 ERK1/2 通路。
方法
用 pcDNA-NNT-AS1 和 miR-203 模拟物共转染 CCLP1 和 TFK1 细胞。采用溴脱氧尿苷（BrdU）掺入实验、流式细胞术、实时定量聚合酶链反应（qRT-PCR）和蛋白质印迹法检测 NNT-AS1 的作用和机制。通过荧光素酶活性实验检测 NNT-AS1 与 miR-203 或 miR-203 与靶基因之间的相互作用。
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