miR-122-5p靶向PDX1抑制结直肠癌细胞上皮-间质转化的研究

张静; 赵振伟; 郭学海; 王海娟; 杨瑞英

miR-122-5p靶向PDX1抑制结直肠癌细胞上皮-间质转化的研究

miR-122-5p targeting PDX1 to inhibit epithelial-mesenchymal transition of colorectal cancer cells

摘要

摘要:
目的： 探究miR-122-5p调控胰腺-十二指肠同源盒1（PDX1）表达对结直肠癌细胞上皮-间质转化（EMT）的影响。
方法： 收集43例结直肠癌病人的结直肠癌组织和癌旁组织样本，体外培养人正常结肠上皮细胞系NCM460及结直肠癌细胞系HCT116、SW620、SW480，qRT-PCR、Western blotting法检测miR-122-5p和PDX1蛋白表达。双荧光素酶报告基因实验检测SW480细胞中miR-122-5p与PDX1的靶向关系。将SW480细胞分为Control组、模拟物（mimic）NC组、miR-122-5p mimic组、miR-122-5p mimic + pcDNA3.1组、miR-122-5p mimic + pcDNA3.1-PDX1组，qRT-PCR法检测各组SW480细胞miR-122-5p、PDX1 mRNA表达；Transwell实验、划痕实验分别检测SW480细胞侵袭、迁移能力；Western blotting检测SW480细胞PDX1、基质金属蛋白酶（MMP）2、MMP9、E-钙黏附蛋白（E-cadherin）、波形蛋白（Vimentin）蛋白表达。
结果： 结直肠癌组织和细胞中miR-122-5p呈低表达，PDX1蛋白呈高表达（P ＜ 0.01）。结直肠癌组织miR-122-5p、PDX1表达与肿瘤分化程度、TNM分期和淋巴结转移明显相关（P ＜ 0.01）。SW480细胞中miR-122-5p与PDX1可能存在靶向调控关系（P ＜ 0.01）。与Control组比，miR-122-5p mimic组SW480细胞侵袭细胞数、迁移率、PDX1 mRNA和蛋白以及MMP9、MMP2、Vimentin蛋白表达均明显降低（P ＜ 0.01），miR-122-5p、E-cadherin蛋白表达均明显增加（P ＜ 0.01）；与miR-122-5p mimic组比，miR-122-5p mimic + pcDNA3.1-PDX1组SW480细胞侵袭细胞数、迁移率、PDX1 mRNA和蛋白以及MMP9、MMP2、Vimentin蛋白表达均明显增加（P ＜ 0.01），E-cadherin蛋白表达降低（P ＜ 0.01）。
结论： miR-122-5p可通过靶向抑制PDX1表达来抑制结直肠癌细胞EMT，并降低细胞侵袭及迁移能力。

Abstract:
Objective To investigate the eeffect of miR-122-5p on the expression of pancreatic duodenal homeobox-1 (PDX1) on epithelial-mesenchymal transition (EMT) of colorectal cancer cells.
Methods Colorectal cancer tissue and adjacent tissue samples from 43 patients with colorectal cancer were collected. Human normal colon epithelial cell lines NCM460 and colorectal cancer cell lines HCT116, SW620 and SW480 were cultured in vitro. The expressions of miR-122-5p and PDX1 protein were detected by realtime-PCR and Western blotting. Dual-luciferase reporter gene assay was performed to measure the targeting relationship between miR-122-5p and PDX1 in SW480 cells. SW480 cells were separated into the ccontrol group, mimic NC group, miR-122-5p mimic group, miR-122-5p mimic + pcDNA3.1 group, and miR-122-5p mimic + pcDNA3.1-PDX1 group. Realtime-PCR method was performed to detect the expressions of miR-122-5p and PDX1 mRNA in SW480 cells in each group. Transwell assay and scratch assay were performed to measure the invasion and migration abilities of SW480 cells, respectively. Western blotting was performed to measure the expressionsof PDX1, matrix metalloproteinase (MMP)2, MMP9, E-cadherin and Vimentin proteins in SW480 cells.
Results The expression of miR-122-5p was lower in colorectal cancer tissues and cells, and the expression of PDX1 protein was higher (P ＜ 0.01). The expressions of miR-122-5p and PDX1 in colorectal cancer were significantly correlated with the degree of tumor differentiation, TNM stage and lymph node metastasis (P ＜ 0.01). There may be targeting regulatory relationship between miR-122-5p and PDX1 in SW480 cell (P ＜ 0.01). Compared with control group, the number of invasive cells, migration rate, the expression of PDX1 mRNA and protein,MMP9, MMP2 and Vimentin protein of SW480 cells in miR-122-5p mimic group were greatly decreased (P ＜ 0.01), while the expressions of miR-122-5p and E-cadherin protein were greatly increased (P ＜ 0.01). Compared with miR-122-5p mimic group, the number of invasive cells, migration rate, the expression of PDX1 mRNA and protein,MMP9, MMP2 and vimentin protein of SW480 cells in miR-122-5p mimic + pcDNA3.1-PDX1 group were greatly increased (P ＜ 0.01), while the expression of E-cadherin protein was greatly decreased (P ＜ 0.01).
Conclusion miR-122-5p can inhibit EMT of colorectal cancer cells by targeting and inhibiting PDX1 expression, which reduces the cell invasion and migration ability.

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