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投稿时间：2018-08-23 网络发布日期：2019-03-22
投稿时间：2018-08-23 网络发布日期：2019-03-22
中文摘要: 目的 探讨微小核糖核酸(micro RNA,miR)-452靶向PH结构域富含亮氨酸重复序列的蛋白磷酸酶1(PHLPP1)抑制结直肠癌细胞的增殖、迁移和侵袭的作用。方法 选取142例结直肠癌组织及癌旁组织。体外培养结直肠癌细胞株SW480细胞 (SW480癌细胞组),并以miR-452模拟物(miR-452模拟物组)、miR-452 抑制剂(miR-452抑制剂组)转染SW480细胞。噻唑蓝(MTT)法检测各组细胞活力、癌细胞单克隆形成数目。流式细胞术检测凋亡率及细胞周期,Transwell细胞迁移实验检测穿膜细胞数,实时荧光定量聚合酶链反应(qRT-PCR)检测miR-452-5、PHLPP1 mRNA表达水平,酶联免疫吸附法测定培养液中基质金属蛋白酶-9(MMP-9)、细胞周期蛋白(Cyclin)D1、血管内皮生长因子(VEGF)在培养液中水平。结果 结直肠癌组miR-452-5p相对表达水平(1.25±0.65)低于癌旁组(3.76±0.43,t=17.04,P<0.01)。依miR-452 模拟物组→SW480癌细胞组→miR-452 inhibitor组之序,癌细胞单克隆形成数目、穿膜数(P<0.01)升高,细胞凋亡率、G1期比例降低(P<0.01);miR-452-5p、PHLPP1 mRNA表达水平降低(P<0.01),MMP-9、Cyclin D1、VEGF蛋白表达水平升高(P<0.01)。miR-452与PHLPP1表达水平呈明显正相关(P<0.01),与MMP-9、Cyclin D1、VEGF表达水平分别呈负相关(P<0.01)。结论 miR-452通过靶向降低PHLPP1 的表达而抑制结直肠癌细胞增殖、迁移浸润,其机制与miR-452诱导结直肠癌细胞低表达MMP-9、Cyclin D1、VEGF有关。
中文关键词: 微小核糖核酸A-452 PH结构域富含亮氨酸重复序列的蛋白磷酸酶1 结直肠癌细胞 增殖 迁移 侵袭
Abstract:Objective To investigate the inhibition effect of microRNA (miR)-452 on proliferation, migration and invasion of colorectal cancer cells by regulating PH domain leucine-rich-repeats protein phosphatase 1 (PHLPP1). Methods The colorectal cancer tissues and paracancerous tissues of 142 patients with colorectal cancer were selected.The colorectal cancer cell line SW480 cells were cultured in vitro (SW480 cancer cell group), and transfecting SW480 cells with miR-452 mimics (miR-452 mimics group)and miR-452 inhibitor (miR-452 inhibitor group).Thiazolam blue (MTT) method was used to detect cell viability, number of monoclonal formation of cancer cells.Flow cytometry was used to detect apoptosis rate and cell cycle. Transwell cell migration assay was used to detect number of penetrating cells.Real-time fluorescence quantitative polymerase chain reaction PCR (qRT-PCR) was used to detect expression levels of miR-452 and PHLPP1 mRNAs.Enzyme-linked immunosorbent assay was used to detect matrix metalloproteinase-9 (MMP-9), Cyclin D1 and vascular endothelial growth factor (VEGF) levels in culture medium. Results The relative expression level of miR-452-5p in colorectal cancer group (1.25±0.65) was lower than that in adjacent cancer group (3.76±0.43, t=17.04, P<0.01).According to the sequence of miR-452 mimic group, SW480 cancer cell group and miR-452 inhibitor group, the number of monoclonal formation and the number of penetrating cells increased (P<0.01), cell apoptosis rate and G1 phase proportion decreased (P<0.01); the expression levels of miRNA-452-5p and PHLPP1 mRNA decreased (P<0.01); the expression levels of MMP-9, Cyclin D1 and VEGF increased (P<0.01).The expression of miR-452 was significantly positively correlated with PHLPP1 mRNA (P<0.01)and was significantly negatively correlated with MMP-9, Cyclin D1 and VEGF respectively (P<0.01). Conclusion miR-452 inhibits the proliferation, migration and invasion of colorectal cancer cells through targeting PHLPP1 for reducing its expression, and its mechanism may be related to the low expression of MMP-9, Cyclin D1 and VEGF in colorectal cancer cells induced by miR-452.
keywords: MicroRNA-452 PH domain leucine-rich-repeats protein phosphatase 1 Colorectal cancer cell Proliferation Migration Invasion
文章编号： 中图分类号：R 735.3+5 文献标志码：A
基金项目:广东省医学科学技术研究基金项目（A2017415）
| 作者 | 单位 |
| 黄仲根1 | 1.广州市白云区第二人民医院外科,广东 广州 510450 |
| 徐波2 | 2.广州市第一人民医院外科,广东 广州 510180 |
| 沈飞2 | 2.广州市第一人民医院外科,广东 广州 510180 |
| Author Name | Affiliation |
| HUANG Zhong-gen*, XU Bo, SHEN Fei | *Department of Surgery, Baiyun District Second People′s Hospital of Guangzhou, Guangzhou, Guangdong 510450, China |
引用文本：
黄仲根,徐波,沈飞.miR-452靶向PHLPP1抑制结直肠癌细胞的增殖、迁移和侵袭[J].中国临床研究,2019,32(3):340-345.
黄仲根,徐波,沈飞.miR-452靶向PHLPP1抑制结直肠癌细胞的增殖、迁移和侵袭[J].中国临床研究,2019,32(3):340-345.