Department of Clinical Laboratory, The Affiliated Yantai Yuhuangding Hospital of Qingdao University, Yantai, China.
Eur Rev Med Pharmacol Sci. 2019 Jul;23(14):6131-6138. doi: 10.26355/eurrev_201907_18426.
The study aimed to explore whether microRNA-155 and FOXP3 could regulate invasive and migratory capacities of colorectal cancer (CRC) cells by mediating Zinc finger E-box binding homeobox 2 (ZEB2) expression.
Dual-luciferase reporter gene assay was performed to detect the binding condition between microRNA-155, FOXP3, and ZEB2. Protein and mRNA levels of ZEB2 in CRC cells were detected after overexpression of microRNA-155 and FOXP3 by Western blot and quantitative Real Time-Polymerase Chain Reaction (qRT-PCR), respectively. In vitro experiments were conducted using HCT116 and SW620 cell lines. We first detected expression levels of microRNA-155, FOXP3, and ZEB2 in the normal colorectal epithelial cell line (NCM460) and CRC cell lines (HCT116 and SW620) by qRT-PCR. Protein expressions of ZEB2, E-cadherin, and vimentin in WT, LV-GFP, and LV-FOXP3 groups were detected. Wound healing assay and transwell assay were conducted to determine the regulatory effects of microRNA-155 and FOXP3 on invasive and migratory capacities of CRC cells, respectively.
Dual-luciferase reporter gene assay found that FOXP3 binds to the promoter and intron regions of ZEB2, and microRNA-155 binds to the 3'UTR region of wild-type ZEB2. Overexpression of FOXP3 downregulated mRNA and protein levels of ZEB2. ZEB2 was highly expressed, whereas microRNA-155 and FOXP3 were lowly expressed in HCT116 and SW620 cells than NCM460 cells. MicroRNA-155 overexpression upregulated E-cadherin and downregulated vimentin in CRC cells. Overexpression of FOXP3 and microRNA-155 inhibited invasive and migratory capacities of CRC cells.
MicroRNA-155 and FOXP3 can jointly regulate ZEB2 expression, thereby inhibiting the migration and invasion of colorectal cancer cells.
本研究旨在探讨 microRNA-155 和 FOXP3 是否能够通过调节锌指 E 盒结合同源盒 2（ZEB2）的表达来调节结直肠癌（CRC）细胞的侵袭和迁移能力。
采用双荧光素酶报告基因检测法检测 microRNA-155、FOXP3 与 ZEB2 的结合情况。通过 Western blot 和定量实时聚合酶链反应（qRT-PCR）分别检测过表达 microRNA-155 和 FOXP3 后 CRC 细胞中 ZEB2 的蛋白和 mRNA 水平。使用 HCT116 和 SW620 细胞系进行体外实验。首先通过 qRT-PCR 检测正常结直肠上皮细胞系（NCM460）和 CRC 细胞系（HCT116 和 SW620）中 microRNA-155、FOXP3 和 ZEB2 的表达水平。检测 WT、LV-GFP 和 LV-FOXP3 组中 ZEB2、E-钙黏蛋白和波形蛋白的蛋白表达。通过划痕愈合实验和 Transwell 实验分别检测 microRNA-155 和 FOXP3 对 CRC 细胞侵袭和迁移能力的调节作用。
双荧光素酶报告基因检测发现，FOXP3 结合 ZEB2 的启动子和内含子区域，microRNA-155 结合野生型 ZEB2 的 3'UTR 区域。FOXP3 的过表达下调了 ZEB2 的 mRNA 和蛋白水平。HCT116 和 SW620 细胞中 ZEB2 表达较高，而 microRNA-155 和 FOXP3 表达较低，NCM460 细胞中 ZEB2 表达较低。microRNA-155 过表达上调 CRC 细胞中的 E-钙黏蛋白，下调波形蛋白。FOXP3 和 microRNA-155 的过表达抑制了 CRC 细胞的侵袭和迁移能力。
microRNA-155 和 FOXP3 可以共同调节 ZEB2 的表达，从而抑制结直肠癌细胞的迁移和侵袭。
