Park Young Ran, Lee Soo Teik, Kim Se Lim, Zhu Shi Mao, Lee Min Ro, Kim Seong Hun, Kim In Hee, Lee Seung Ok, Seo Seung Young, Kim Sang Wook
Department of Internal Medicine, Chonbuk National University Hospital, Chonbuk National University Medical School, 20 Geonji-ro, Deokjin-gu, Jeonju, Jeonbuk 54907, Republic of Korea; Research Institute of Clinical Medicine of Chonbuk National University-Biomedical Research Institute of Chonbuk National University Hospital, Jeonju, Republic of Korea.
Department of Surgery, Chonbuk National University Hospital, Chonbuk National University Medical School, Jeonju, Republic of Korea.
Cancer Genet. 2019 Feb;231-232:22-31. doi: 10.1016/j.cancergen.2018.12.004. Epub 2018 Dec 27.
MicroRNA-9 (miR-9) has been reported to play a suppressive or promoting role according to cancer type. In this study, we investigated the effects of anoctamin-1 (ANO1) and miR-9 on colorectal cancer (CRC) cell proliferation, migration, and invasion and determined the underlying molecular mechanisms. Thirty-two paired CRC tissues and adjacent normal tissues were analyzed for ANO1 expression using quantitative real-time PCR (qRT-PCR). HCT116 cells were transiently transfected with miR-9 mimic, miR-9 inhibitor, or si-ANO1. Cell proliferation was determined by MTT, and flow cytometric analysis, while cell migration and invasion were assayed by trans-well migration and invasion assay in HCT116 cells. ANO1 was validated as a target of miR-9 using luciferase reporter assay and bioinformatics algorithms. We found that ANO1 expression was up-regulated in CRC tissues compared with adjacent normal tissues. ANO1 expression was associated with advanced tumor stage and lymph node metastasis, and there was an inverse relationship between miR-9 and ANO1 mRNA expression in CRC specimens, but no significant difference was found between miR-9 and ANO1 expression. ANO1 is a direct target of miR-9, and overexpression of miR-9 suppressed both mRNA and protein expression of ANO1 and inhibited cell proliferation, migration, and invasion of HCT116 cells. We also showed that overexpression of miR-9 suppressed expression of p-AKT, cyclin D1, and p-ERK in HCT116 cells. We conclude that miR-9 inhibits CRC cell proliferation, migration, and invasion by directly targeting ANO1, and miR-9/ANO1 could be a potential therapeutic target for CRC.
据报道，微小RNA-9（miR-9）根据癌症类型发挥抑制或促进作用。在本研究中，我们调查了anoctamin-1（ANO1）和miR-9对结直肠癌（CRC）细胞增殖、迁移和侵袭的影响，并确定了潜在的分子机制。使用定量实时PCR（qRT-PCR）分析了32对CRC组织和相邻正常组织中的ANO1表达。用miR-9模拟物、miR-9抑制剂或si-ANO1瞬时转染HCT116细胞。通过MTT和流式细胞术分析测定细胞增殖，而通过HCT116细胞的Transwell迁移和侵袭试验测定细胞迁移和侵袭。使用荧光素酶报告基因试验和生物信息学算法验证ANO1是miR-9的靶标。我们发现，与相邻正常组织相比，CRC组织中ANO1表达上调。ANO1表达与肿瘤晚期和淋巴结转移相关，并且在CRC标本中miR-9与ANO1 mRNA表达呈负相关，但miR-9与ANO1表达之间未发现显著差异。ANO1是miR-9的直接靶标，miR-9的过表达抑制了ANO1的mRNA和蛋白表达，并抑制了HCT116细胞的增殖、迁移和侵袭。我们还表明，miR-9的过表达抑制了HCT116细胞中p-AKT、细胞周期蛋白D1和p-ERK的表达。我们得出结论，miR-9通过直接靶向ANO1抑制CRC细胞的增殖、迁移和侵袭，并且miR-9/ANO1可能是CRC的潜在治疗靶点。
