microRNA-338-3p 通过靶向 smoothened 抑制结直肠癌细胞的侵袭和迁移。

MicroRNA-338-3p inhibits colorectal carcinoma cell invasion and migration by targeting smoothened.

机构信息

*Department of General Surgery, Nanfang Hospital of Southern Medical University, Guangzhou 510515, Guangdong Province, China.

出版信息

Jpn J Clin Oncol. 2014 Jan;44(1):13-21. doi: 10.1093/jjco/hyt181. Epub 2013 Nov 25.

DOI:10.1093/jjco/hyt181

PMID:24277750

Abstract

OBJECTIVE

To investigate the regulative effect of microRNA-338-3p on colorectal carcinoma cell invasion and migration.

METHODS

The microRNA-338-3p expression pattern of colorectal carcinoma tissues and cell lines was detected by real-time reverse transcriptase polymerase chain reaction. The protein level of smoothened was detected by western blot analysis. Furthermore, colorectal carcinoma cells were pretreated with or without anti-smoothened-small interfering ribonucleic acid prior to the addition of pre-microRNA-338-3p or anti-microRNA-338-3p. The status of colorectal carcinoma cell invasion and that of migration were detected by transwell assay and wound healing assay, respectively.

RESULTS

The expression of microRNA-338-3p was significantly down-regulated in colorectal carcinoma tissues in comparison with those in the adjacent non-tumorous tissues, and the value was negatively related to advanced tumor, node, metastasis stage and local invasion. The expression of microRNA-338-3p in colorectal carcinoma cells transfected with pre-microRNA-338-3p p was significantly increased. Furthermore, over-expression of microRNA-338-3p inhibited the expression of smoothened protein in colorectal carcinoma cells, which showed obviously suppressed invasion and migration ability. The expression of microRNA-338-3p in colorectal carcinoma cells transfected with anti-microRNA-338-3p was significantly decreased. Moreover, the down-regulated expression of microRNA-338-3p caused the up-regulated expression of smoothened protein in colorectal carcinoma cells, which showed significantly enhanced invasion and migration ability. However, anti-smoothened-small interfering ribonucleic acid largely, but not completely, reversed the effects induced by blockage of microRNA-338-3p, suggesting that the regulative effect of microRNA-338-3p on colorectal carcinoma cell invasion and migration was indeed mediated by smoothened. Additionally, smoothened was identified as a direct target of microRNA-338-3p by luciferase assay.

CONCLUSIONS

MicroRNA-338-3p could inhibit colorectal carcinoma cell invasion and migration by inhibiting smoothened expression.

摘要

目的

研究 microRNA-338-3p 对结直肠癌细胞侵袭和迁移的调节作用。

方法

采用实时逆转录聚合酶链反应检测结直肠癌细胞和组织中 microRNA-338-3p 的表达模式。通过 Western blot 分析检测 smoothened 蛋白的水平。此外，在加入预 microRNA-338-3p 或抗 microRNA-338-3p 之前，用抗 smoothened-小干扰 RNA 预处理结直肠癌细胞。通过 Transwell 测定和划痕愈合试验分别检测结直肠癌细胞的侵袭和迁移状态。

结果

与邻近非肿瘤组织相比，microRNA-338-3p 在结直肠癌细胞中的表达明显下调，且与肿瘤进展、淋巴结转移、远处转移分期和局部浸润程度呈负相关。转染前 microRNA-338-3p 的结直肠癌细胞中的 microRNA-338-3p 表达明显增加。此外，过表达 microRNA-338-3p 抑制了 smoothened 蛋白在结直肠癌细胞中的表达，从而明显抑制了结直肠癌细胞的侵袭和迁移能力。转染抗 microRNA-338-3p 的结直肠癌细胞中的 microRNA-338-3p 表达明显降低。此外，下调 microRNA-338-3p 的表达导致 smoothened 蛋白在结直肠癌细胞中的表达上调，从而显著增强了结直肠癌细胞的侵袭和迁移能力。然而，抗 smoothened-小干扰 RNA 虽然在很大程度上但并非完全逆转了 microRNA-338-3p 阻断引起的作用，这表明 microRNA-338-3p 对结直肠癌细胞侵袭和迁移的调节作用确实是通过 smoothened 介导的。此外，通过荧光素酶测定鉴定 smoothened 是 microRNA-338-3p 的直接靶标。