长链非编码RNARUNX1-IT1通过靶向miR-21对结直肠癌细胞迁移和侵袭的影响

doi:10.3969/j.issn.1004-616x.2022.05.008

摘要/Abstract

摘要： 目的：探讨长链非编码 RNA(lncRNA)RUNX1-IT1在结直肠癌中的表达及其对结直肠癌细胞侵袭和迁移的影响机制。方法：收集2017年1月—2019年1月于河北北方学院附属第一医院进行根治性手术切除的结直肠癌组织标本62例及其相应的癌旁正常组织标本，采用荧光定量PCR(qPCR)法检测结直肠癌组织和其相应的癌旁组织中lncRNA RUNX1-IT1的表达。并培养人结直肠癌细胞系(SW480、SW620、HCT-116)和人正常结直肠上皮细胞系(FHC)，qPCR检测细胞系中 lncRNA RUNX1-IT1和 miR-21的表达水平，选择最适细胞系用于后续实验。通过上调或下调SW480细胞中lncRNA RUNX1-IT1和miR-21的表达后，采用qPCR检测lncRNA RUNX1-IT1和miR-21的表达水平，双荧光素酶报告基因实验验证lncRNA RUNX1-IT1和miR-21的靶向关系，Transwell实验检测SW480细胞侵袭和迁移能力。结果：qPCR检测结果显示，与癌旁正常组织比较，lncRNA RUNX1-IT1在结直肠癌组织中表达降低(P<0.05)，而miR-21在结直肠癌组织中表达升高(P<0.05)，且lncRNA RUNX1-IT1与miR-21在结直肠癌组织中的表达呈负相关(r=-0.275，P=0.031)。与正常结直肠 FHC细胞相比，lncRNA RUNX1-IT1在 3种结直肠癌细胞系中的表达水平均显著降低(均为P<0.05)，而miR-21均显著升高(均为P<0.05)，且SW480细胞最明显，故后续采用SW480细胞系进行实验。双荧光素酶报告基因实验证实lncRNA RUNX1-IT1靶向调节miR-21的表达；与对照组相比，过表达lncRNA RUNX1-IT1可抑制SW480细胞侵袭和迁移能力(P<0.05)；抑制 miR-21表达可抑制 SW480细胞侵袭和迁移能力(P<0.05)；上调 miR-21表达可逆转过表达 lncRNARUNX1-IT1对 SW480细胞侵袭和迁移的抑制作用(P<0.05)。结论：LncRNA RUNX1-IT1在结直肠癌中表达下调，lncRNARUNX1-IT1通过靶向miR-21调控SW480细胞侵袭和迁移。

关键词: 结直肠癌, 长链非编码RNA, RUNX1-IT1, 侵袭, 迁移

Abstract: OBJECTIVE: To investigate expression of long-chain noncoding RNA (lncRNA) RUNX1-IT1 in colorectal cancer,and its mechanisms on invasion and migration of colorectal cancer cells.METHODS: Sixty-two colorectal cancer tissue samples and their corresponding adjacent tissues were collected from the First Affiliated Hospital of Hebei North University from January 2017 to January 2019.Expressions of lncRNA RUNX1-IT1 in both tissues were detected using fluorescence quantitative PCR (qPCR).The human colorectal cancer cell lines (SW480,SW620,HCT-116) and the human normal colorectal epithelial cell lines (FHC) were cultured.Expression levels of lncRNA RUNX1-IT1 and miR-21 in the cell lines were detected by qPCR.The most suitable cell lines were selected for subsequent investigations.After up-regulating or down-regulating the expression of lncRNA RUNX1-IT1 and miR-21 in SW480 cells,their expression levels were detected by qPCR,the targeting relationship between lncRNA RUNX1-IT1 and miR-21 was verified by the double luciferase reporter gene assay.The invasion and migration abilities of SW480 cells were detected by Transwell assay.RESULTS: The results of qPCR showed that the expression of lncRNA RUNX1-IT1 was lower in colorectal cancer tissues than that in adjacent tissues (P<0.05),while the expression of miR-21 was higher in colorectal cancer tissues (all P<0.05).There was a negative correlation between the expression of lncRNA RUNX1-IT1 and miR-21 in colorectal cancer tissues (r=-0.275,P=0.031).Compared with normal colorectal FHC cells,the expression levels of lncRNA RUNX1-IT1 in the three colorectal cancer cell lines were decreased significantly (all P<0.05),while that for miR-21 was increased significantly (all P<0.05).These effects were most obvious in the SW480 cells.Therefore,SW480 cell lines were used for subsequent experiments.The dual luciferase reporter assay confirmed that lncRNA RUNX1-IT1 targeted to regulate the expression of miR-21.Compared with the control group,overexpression of lncRNA RUNX1-IT1 inhibited the invasion and migration of SW480 cells (P<0.05).Inhibition of miR-21 expression reduced the invasion and migration of SW480 cells (P<0.05).Up-regulation of miR-21 expression inversely reversed the inhibitory effect of lncRNA RUNX1-IT1 on the invasion and migration of SW480 cells (P<0.05).CONCLUSION: LncRNA RUNX1-IT1 expression was down-regulated in colorectal cancers and it regulated SW480 cell invasion and migration by targeting miR-21.

Key words: colorectal cancer, long chain noncoding RNA, RUNX1-IT1, attack, transfer

中图分类号:

R735.3

梁峰, 张玮, 王胜杰, 武雪亮, 岳磊, 张迎春. 长链非编码RNARUNX1-IT1通过靶向miR-21对结直肠癌细胞迁移和侵袭的影响[J]. 癌变·畸变·突变, 2022, 34(5): 378-383,387.

LIANG Feng, ZHANG Wei, WANG Shengjie, WU Xueliang, YUE Lei, ZHANG Yingchun. Effects of long-chain noncoding RNA RUNX1-IT1 on migration and invasion of colorectal cancer cells[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2022, 34(5): 378-383,387.

参考文献

[1] THANIKACHALAM K, KHAN G. Colorectal cancer and nutrition[J]. Nutrients, 2019, 11(1):164.
[2] DEKKER E, TANIS P J, VLEUGELS J L A, et al. Colorectal cancer[J]. Lancet, 2019, 394(10207):1467-1480.
[3] WANG L Y, CHO K B, LI Y, et al. Long noncoding RNA (lncRNA)-mediated competing endogenous RNA networks provide novel potential biomarkers and therapeutic targets for colorectal cancer[J]. Int J Mol Sci, 2019, 20(22):5758.
[4] BHAN A, SOLEIMANI M, MANDAL S S. Long noncoding RNA and cancer:a new paradigm[J]. Cancer Res, 2017, 77(15):3965-3981.
[5] MCCABE E M, RASMUSSEN T P. lncRNA involvement in cancer stem cell function and epithelial-mesenchymal transitions[J]. Semin Cancer Biol, 2021, 75:38-48.
[6] CHAN J J, TAY Y. Noncoding RNA:RNA regulatory networks in cancer[J]. Int J Mol Sci, 2018, 19(5):1310.
[7] SHI J X, ZHONG X, SONG Y X, et al. Long non-coding RNA RUNX1-IT1 plays a tumour-suppressive role in colorectal cancer by inhibiting cell proliferation and migration[J]. Cell Biochem Funct, 2019, 37(1):11-20.
[8] BAIDOUN F, ELSHIWY K, ELKERAIE Y, et al. Colorectal cancer epidemiology:recent trends and impact on outcomes[J]. Curr Drug Targets, 2021, 22(9):998-1009.
[9] SIEGEL R L, JAKUBOWSKI C D, FEDEWA S A, et al. Colorectal cancer in the young:epidemiology, prevention, management[J]. Am Soc Clin Oncol Educ Book, 2020, 40((4)):1-14.
[10] LA VECCHIA S, SEBASTIÁN C. Metabolic pathways regulating colorectal cancer initiation and progression[J]. Semin Cell Dev Biol, 2020, 98:63-70.
[11] SINHA R. Colorectal cancer[J]. Clin Radiol, 2021, 76(12):870.
[12] CHI, WANG, WANG, et al. Long non-coding RNA in the pathogenesis of cancers[J]. Cells, 2019, 8(9):1015.
[13] REGGIARDO R E, MAROLI S V, KIM D H. LncRNA biomarkers of inflammation and cancer[J]. Adv Exp Med Biol, 2022, 1363:121-145.
[14] HUANG F J, LIU Y L, WANG J, et al. LncRNA RUNX1-IT1 affects the differentiation of Th1 cells by regulating NrCAM transcription in Graves' disease[J]. Cell Cycle, 2022, 21(9):921-933.
[15] LIANG M L, WANG H B, LIU C, et al. LncRNA RUNX1-IT1 is downregulated in endometrial cancer and binds to miR-21 precursor to suppress its maturation[J]. Cancer Manag Res, 2020, 12:13451-13459.
[16] MIAO Y J, LI T, LIU Y F, et al. The molecular mechanism of long non-coding ribonucleic acid (lncRNA) RUNX1-IT1 promotes the proliferation and stemness of lung cancer cells[J]. Transl Cancer Res, 2021, 10(11):4884-4893.
[17] SUN L K, WANG L, CHEN T X, et al. LncRNA RUNX1-IT1 which is downregulated by hypoxia-driven histone deacetylase 3 represses proliferation and cancer stem-like properties in hepatocellular carcinoma cells[J].Cell Death Dis, 2020, 11(2):1-15.
[18] YAN P H, WANG L, CHEN H, et al. LncRNA RUNX1-IT1 inhibits proliferation and promotes apoptosis of hepatocellular carcinoma by regulating MAPK pathways[J]. Eur Rev Med Pharmacol Sci, 2019, 23(19):8287-8294.
[19] WU Z. miR-195 connects lncRNA RUNX1-IT1 and cyclin D1 to regulate the proliferation of glioblastoma cells[J]. Int J Neurosci, 2021:1-6.
[20] SINGH A, SINGH A K, GIRI R, et al. The role of microRNA-21 in the onset and progression of cancer[J]. Future Med Chem, 2021, 13(21):1885-1906.
[21] ZHANG Z, WANG J M, WANG X F, et al. microRNA-21 promotes proliferation, migration, and invasion of cervical cancer through targeting TIMP3[J]. Arch Gynecol Obstet, 2018, 297(2):433-442.
[22] REN J, KUANG T H, CHEN J, et al. The diagnostic and prognostic values of microRNA-21 in patients with gastric cancer:a meta-analysis[J]. Eur Rev Med Pharmacol Sci, 2017, 21(1):120-130.
[23] CRETOIU D. miR-21 regulates growth and EMT in lung cancer cells via PTEN Akt GSK3 beta signaling[J]. Front Biosci, 2019, 24(8):1426-1439.