miR-149-5p通过靶向AEBP1调控胃癌细胞的迁移侵袭

miR-149-5p targets AEBP1 to inhibit migration and invasion of gastric cancer cells

摘要:

目的 探讨miR-149-5p对胃癌细胞迁移侵袭能力的影响及分子机制。方法 qRT-PCR检测胃癌细胞株和组织标本中miR-149-5p的表达，分析其表达水平与胃癌患者临床病理特征参数及预后的相关性，建立过表达和干扰miR-149-5p的胃癌细胞株，Transwell实验检测miR-149-5p表达水平对胃癌癌细胞迁移侵袭能力的影响；生物信息学网站预测miR-149-5p的靶基因，采用荧光素酶报告基因实验加以验证。结果 miR-149-5p在胃癌组织和细胞株中均低表达（均P<0.05）。miR-149-5p的表达水平与胃癌患者的浸润深度（P=0.016）、淋巴结转移（P=0.001） 和TNM分期（P=0.023） 相关。miR-149-5p低表达是胃癌患者总生存期的独立危险因素。与对照组相比，miR-149-5p mimics明显抑制胃癌细胞的迁移侵袭能力（均P<0.01），而转染miR-149-5p inhibitors后得到了相反的结果（均P<0.05）。miR-149-5p靶向调控脂肪细胞增强结合蛋白1（Adipocyte enhancer binding protein 1，AEBP1）蛋白的表达。荧光素酶报告基因实验表明，miR-149-5p明显抑制了野生型AEBP1载体的荧光素酶活性（P<0.01），而突变型的荧光素酶活性不受影响。敲低AEBP1可部分逆转下调miR-149-5p对胃癌细胞迁移侵袭能力的影响。结论 miR-149-5p在胃癌组织中低表达，通过靶向AEBP1负性调控胃癌癌细胞的迁移侵袭。

Abstract:

Objective To investigate the effects of miR-149-5p in regulating the migration and invasion ability of gastric cancer（GC） cells and the molecular mechanism.Methods The expression of miR-149-5p in GC cell lines and tissue specimens was measured by qRT-PCR to analyze the relationship between miR-149-5p expression and the clinicopathological parameters and prognosis of patients with GC. We separately overexpressed and knocked down miR-149-5p in GC cells to investigate the expression level of miR-149-5p on the migration and invasion ability of GC cells using Transwell assay. The target genes of miR-149-5p were predicted using bioinformatic tools，which were verified using luciferase reporter gene assay.Results The expression of miR-149-5p was significantly down-regulated in both GC tissues and cell lines（P<0.05）. The expression level of miR-149-5p was significantly correlated with the depth of invasion（P=0.016），lymph node metastasis（P=0.001），and TNM stage（P=0.023） of patients with GC. Low expression of miR-149-5p was an independent risk factor for the overall survival of patients with GC. Compared with the control group，miR-149-5p mimics significantly inhibited the migration and invasion ability of gastric cancer cells（P<0.01），while transfection with miR-149-5p inhibitors produced the opposite effects（P<0.05）. miR-149-5p targeted the expression of adipocyte enhancer-binding protein 1（AEBP1）. The luciferase reporter gene assay showed that miR-149-5p significantly inhibited the luciferase activity of the wild-type AEBP1 vector（P<0.01），with no effects on the luciferase activity of the mutant type. Knockdown of AEBP1 could partly reverse the effects of downregulating miR-149-5p on the migration and invasion ability of GC cells.Conclusion miR-149-5p is lowly expressed in GC tissues，which can negatively regulate the migration and invasion of GC cells by targeting AEBP1 protein expression.