MicroRNA‑300 inhibits the growth of hepatocellular carcinoma cells by downregulating CREPT/Wnt/β‑catenin signaling

Bai,Jinping; Gao,Yingchun; Du,Yanhui; Yang,Xue; Zhang,Xinye

doi:10.3892/ol.2019.10712

MicroRNA‑300 inhibits the growth of hepatocellular carcinoma cells by downregulating CREPT/Wnt/β‑catenin signaling

Authors:
- Jinping Bai
- Yingchun Gao
- Yanhui Du
- Xue Yang
- Xinye Zhang
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Affiliations: School of Clinical Medicine, Changchun University of Chinese Medicine, Changchun, Jilin 130117, P.R. China, Quality Control Office, The Affiliated Hospital of Changchun University of Chinese Medicine, Changchun, Jilin 130000, P.R. China, Department of Geriatrics, The Affiliated Hospital of Changchun University of Chinese Medicine, Changchun, Jilin 130000, P.R. China, Department of Thyroid Head and Neck Surgery, Jilin Cancer Hospital, Changchun, Jilin 130033, P.R. China, Nursing College, Changchun University of Chinese Medicine, Changchun, Jilin 130117, P.R. China
Published online on: August 5, 2019 https://doi.org/10.3892/ol.2019.10712
Pages: 3743-3753
Copyright: © Bai et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

A number of studies have demonstrated that altered expression levels of microRNA‑300 (miR‑300) are associated with tumor progression; however, little is understood regarding the role of miR‑300 in hepatocellular carcinoma (HCC). The present study aimed to investigate the expression, biological function and potential regulatory mechanism of miR‑300 in HCC. A miR‑300 mimic and miR‑300 inhibitor were transfected into liver cancer cells using RNAiMAX reagent. The expression levels of miR and mRNA were detected by reverse transcription‑quantitative polymerase chain reaction. Protein expression levels were detected by western blot analysis. Cell growth was determined using Cell Counting Kit‑8, a colony formation assay and cell cycle assay. miRNA targeting sites were analyzed using bioinformatics analysis and dual‑luciferase reporter assay. The results revealed that miR‑300 expression was significantly decreased in HCC tissues and cell lines. In vitro experiments demonstrated that overexpression of miR‑300 could inhibit cell proliferation, colony formation and cell cycle progression of liver cancer cells. By contrast, inhibition of miR‑300 was associated with increased rates of cell proliferation, colony formation and cell cycle progression. Notably, regulation of nuclear pre‑mRNA domain‑containing protein 1B (CREPT) was identified as a putative target gene of miR‑300 by bioinformatics analysis. A luciferase reporter assay revealed that miR‑300 directly targets the 3'‑untranslated region of CREPT. Further data demonstrated that miR‑300 can regulate CREPT expression levels in liver cancer cells. Notably, miR‑300 was identified to regulate the Wnt/β‑catenin signaling pathway in liver cancer cells. The restoration of CREPT expression partially reversed the antitumor effect of miR‑300. In conclusion, the current results revealed a tumor suppressive role of miR‑300 in HCC and indicated that the underlying mechanism was associated with a regulation of CREPT. The present study suggests that miR‑300 and CREPT may serve as potential therapeutic targets for liver cancer.

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1	Forner A, Llovet JM and Bruix J: Hepatocellular carcinoma. Lancet. 379:1245–1255. 2012. View Article : Google Scholar : PubMed/NCBI
2	Yu MC and Yuan JM: Environmental factors and risk for hepatocellular carcinoma. Gastroenterology 127 (5 Suppl 1). S72–S78. 2004.
3	El-Serag HB and Rudolph KL: Hepatocellular carcinoma: Epidemiology and molecular carcinogenesis. Gastroenterology. 132:2557–2576. 2007. View Article : Google Scholar : PubMed/NCBI
4	Hanahan D and Weinberg RA: Hallmarks of cancer: The next generation. Cell. 144:646–674. 2011. View Article : Google Scholar : PubMed/NCBI
5	Maluccio M and Covey A: Recent progress in understanding, diagnosing, and treating hepatocellular carcinoma. CA Cancer J Clin. 62:394–399. 2012. View Article : Google Scholar : PubMed/NCBI
6	Hu MD, Jia LH, Liu HB, Zhang KH and Guo GH: Sorafenib in combination with transarterial chemoembolization for hepatocellular carcinoma: A meta-analysis. Eur Rev Med Pharmacol Sci. 20:64–74. 2016.PubMed/NCBI
7	Bartel DP: MicroRNAs: Genomics, biogenesis, mechanism, and function. Cell. 116:281–297. 2004. View Article : Google Scholar : PubMed/NCBI
8	Bartel DP: MicroRNAs: Target recognition and regulatory functions. Cell. 136:215–233. 2009. View Article : Google Scholar : PubMed/NCBI
9	Kloosterman WP and Plasterk RH: The diverse functions of microRNAs in animal development and disease. Dev Cell. 11:441–450. 2006. View Article : Google Scholar : PubMed/NCBI
10	Calin GA and Croce CM: MicroRNA signatures in human cancers. Nat Rev Cancer. 6:857–866. 2006. View Article : Google Scholar : PubMed/NCBI
11	Volinia S, Calin GA, Liu CG, Ambs S, Cimmino A, Petrocca F, Visone R, Iorio M, Roldo C, Ferracin M, et al: A microRNA expression signature of human solid tumors defines cancer gene targets. Proc Natl Acad Sci USA. 103:2257–2261. 2006. View Article : Google Scholar : PubMed/NCBI
12	Wei R, Huang GL, Zhang MY, Li BK, Zhang HZ, Shi M, Chen XQ, Huang L, Zhou QM, Jia WH, et al: Clinical significance and prognostic value of microRNA expression signatures in hepatocellular carcinoma. Clin Cancer Res. 19:4780–4791. 2013. View Article : Google Scholar : PubMed/NCBI
13	Callegari E, Gramantieri L, Domenicali M, D'Abundo L, Sabbioni S and Negrini M: MicroRNAs in liver cancer: A model for investigating pathogenesis and novel therapeutic approaches. Cell Death Differ. 22:46–57. 2015. View Article : Google Scholar : PubMed/NCBI
14	Gao Y, Zhang SG, Wang ZH and Liao JC: Down-regulation of miR-342-3p in hepatocellular carcinoma tissues and its prognostic significance. Eur Rev Med Pharmacol Sci. 21:2098–2102. 2017.PubMed/NCBI
15	Zhao XQ, Liang B, Jiang K and Zhang HY: Down-regulation of miR-655-3p predicts worse clinical outcome in patients suffering from hepatocellular carcinoma. Eur Rev Med Pharmacol Sci. 21:748–752. 2017.PubMed/NCBI
16	Lu D, Wu Y, Wang Y, Ren F, Wang D, Su F, Zhang Y, Yang X, Jin G, Hao X, et al: CREPT accelerates tumorigenesis by regulating the transcription of cell-cycle-related genes. Cancer Cell. 21:92–104. 2012. View Article : Google Scholar : PubMed/NCBI
17	Carvalho B, Postma C, Mongera S, Hopmans E, Diskin S, van de Wiel MA, van Criekinge W, Thas O, Matthäi A, Cuesta MA, et al: Multiple putative oncogenes at the chromosome 20q amplicon contribute to colorectal adenoma to carcinoma progression. Gut. 58:79–89. 2009. View Article : Google Scholar : PubMed/NCBI
18	Deng G, Yu M, Chen LC, Moore D, Kurisu W, Kallioniemi A, Waldman FM, Collins C and Smith HS: Amplifications of oncogene erbB-2 and chromosome 20q in breast cancer determined by differentially competitive polymerase chain reaction. Breast Cancer Res Treat. 40:271–281. 1996. View Article : Google Scholar : PubMed/NCBI
19	Zhang Y, Wang S, Kang W, Liu C, Dong Y, Ren F, Wang Y, Zhang J, Wang G, To KF, et al: CREPT facilitates colorectal cancer growth through inducing Wnt/β-catenin pathway by enhancing p300-mediated β-catenin acetylation. Oncogene. 37:3485–3500. 2018. View Article : Google Scholar : PubMed/NCBI
20	Zheng G, Li W, Zuo B, Guo Z, Xi W, Wei M, Chen P, Wen W and Yang AG: High expression of CREPT promotes tumor growth and is correlated with poor prognosis in colorectal cancer. Biochem Biophys Res Commun. 480:436–442. 2016. View Article : Google Scholar : PubMed/NCBI
21	Li W, Zheng G, Xia J, Yang G, Sun J, Wang X, Wen M, Sun Y, Zhang Z and Jin F: Cell cycle-related and expression-elevated protein in tumor overexpression is associated with proliferation behaviors and poor prognosis in non-small-cell lung cancer. Cancer Sci. 109:1012–1023. 2018. View Article : Google Scholar : PubMed/NCBI
22	Ma J, Ren Y, Zhang L, Kong X, Wang T, Shi Y and Bu R: Knocking-down of CREPT prohibits the progression of oral squamous cell carcinoma and suppresses cyclin D1 and c-Myc expression. PLoS One. 12:e01743092017. View Article : Google Scholar : PubMed/NCBI
23	Wang Y, Qiu H, Hu W, Li S and Yu J: RPRD1B promotes tumor growth by accelerating the cell cycle in endometrial cancer. Oncol Rep. 31:1389–1395. 2014. View Article : Google Scholar : PubMed/NCBI
24	Chen Z, Zhang W, Jiang K, Chen B, Wang K, Lao L, Hou C, Wang F, Zhang C and Shen H: MicroRNA-300 regulates the ubiquitination of PTEN through the CRL4B^DCAF13 E3 ligase in osteosarcoma cells. Mol Ther Nucleic Acids. 10:254–268. 2018. View Article : Google Scholar : PubMed/NCBI
25	He J, Feng X, Hua J, Wei L, Lu Z, Wei W, Cai H, Wang B, Shi W, Ding N, et al: miR-300 regulates cellular radiosensitivity through targeting p53 and apaf1 in human lung cancer cells. Cell Cycle. 16:1943–1953. 2017. View Article : Google Scholar : PubMed/NCBI
26	Yu J, Xie F, Bao X, Chen W and Xu Q: miR-300 inhibits epithelial to mesenchymal transition and metastasis by targeting Twist in human epithelial cancer. Mol Cancer. 13:1212014. View Article : Google Scholar : PubMed/NCBI
27	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
28	Jin K, Chen H, Zuo Q, Huang C, Zhao R, Yu X, Wang Y, Zhang Y, Chang Z and Li B: CREPT and p15RS regulate cell proliferation and cycling in chicken DF-1 cells through the Wnt/β-catenin pathway. J Cell Biochem. 119:1083–1092. 2018. View Article : Google Scholar : PubMed/NCBI
29	Zhang Y, Liu C, Duan X, Ren F, Li S, Jin Z, Wang Y, Feng Y, Liu Z and Chang Z: CREPT/RPRD1B, a recently identified novel protein highly expressed in tumors, enhances the β-catenin. TCF4 transcriptional activity in response to Wnt signaling. J Biol Chem. 289:22589–22599. 2014. View Article : Google Scholar : PubMed/NCBI
30	Pan XP, Wang HX, Tong DM, Li Y, Huang LH and Wang C: miRNA-370 acts as a tumor suppressor via the downregulation of PIM1 in hepatocellular carcinoma. Eur Rev Med Pharmacol Sci. 21:1254–1263. 2017.PubMed/NCBI
31	Wang R, Yu Z, Chen F, Xu H, Shen S, Chen W, Chen L, Su Q, Zhang L, Bi J, et al: miR-300 regulates the epithelial-mesenchymal transition and invasion of hepatocellular carcinoma by targeting the FAK/PI3K/AKT signaling pathway. Biomed Pharmacother. 103:1632–1642. 2018. View Article : Google Scholar : PubMed/NCBI
32	Zhou F, Li Y, Hao Z, Liu X, Chen L, Cao Y, Liang Z, Yuan F, Liu J, Wang J, et al: MicroRNA-300 inhibited glioblastoma progression through ROCK1. Oncotarget. 7:36529–36538. 2016.PubMed/NCBI
33	He FY, Liu HJ, Guo Q and Sheng JL: Reduced miR-300 expression predicts poor prognosis in patients with laryngeal squamous cell carcinoma. Eur Rev Med Pharmacol Sci. 21:760–764. 2017.PubMed/NCBI
34	Ge W, Han C, Wang J and Zhang Y: MiR-300 suppresses laryngeal squamous cell carcinoma proliferation and metastasis by targeting ROS1. Am J Transl Res. 8:3903–3911. 2016.PubMed/NCBI
35	Ma F, Wang SH, Cai Q, Jin LY, Zhou D, Ding J and Quan ZW: Long non-coding RNA TUG1 promotes cell proliferation and metastasis by negatively regulating miR-300 in gallbladder carcinoma. Biomed Pharmacother. 88:863–869. 2017. View Article : Google Scholar : PubMed/NCBI
36	Zhang JQ, Chen S, Gu JN, Zhu Y, Zhan Q, Cheng DF, Chen H, Deng XX, Shen BY and Peng CH: MicroRNA-300 promotes apoptosis and inhibits proliferation, migration, invasion and epithelial-mesenchymal transition via the Wnt/β-catenin signaling pathway by targeting CUL4B in pancreatic cancer cells. J Cell Biochem. 119:1027–1040. 2018. View Article : Google Scholar : PubMed/NCBI
37	Wang L and Yu P: miR-300 promotes proliferation and EMT-mediated colorectal cancer migration and invasion by targeting p53. Oncol Rep. 36:3225–3232. 2016. View Article : Google Scholar : PubMed/NCBI
38	Xue Z, Zhao J, Niu L, An G, Guo Y and Ni L: Up-regulation of miR-300 promotes proliferation and invasion of osteosarcoma by targeting BRD7. PLoS One. 10:e01276822015. View Article : Google Scholar : PubMed/NCBI
39	Zhang J, Luo H, Du J and Liu Y: MicroRNA-300 plays as oncogene by promoting proliferation and reducing apoptosis of liver cancer cells by targeting MDC1. Int J Clin Exp Pathol. 9:1231–1239. 2016.
40	Olatunji I: Potential application of tumor suppressor microRNAs for targeted therapy in head and neck cancer: A mini-review. Oral Oncol. 87:165–169. 2018. View Article : Google Scholar : PubMed/NCBI
41	Lo Russo G, Tessari A, Capece M, Galli G, de Braud F, Garassino MC and Palmieri D: MicroRNAs for the diagnosis and management of malignant pleural mesothelioma: A literature review. Front Oncol. 8:6502018. View Article : Google Scholar : PubMed/NCBI
42	Kuang YS, Wang Y, Ding LD, Yang L, Wang Y, Liu SH, Zhu BT, Wang XN, Liu HY, Li J, et al: Overexpression of CREPT confers colorectal cancer sensitivity to fluorouracil. World J Gastroenterol. 24:475–483. 2018. View Article : Google Scholar : PubMed/NCBI
43	Liu T, Li WM, Wang WP, Sun Y, Ni YF, Xing H, Xia JH, Wang XJ, Zhang ZP and Li XF: Inhibiting CREPT reduces the proliferation and migration of non-small cell lung cancer cells by down-regulating cell cycle related protein. Am J Transl Res. 8:2097–2113. 2016.PubMed/NCBI
44	Sun M, Si G, Sun HS and Si FC: Inhibition of CREPT restrains gastric cancer growth by regulation of cycle arrest, migration and apoptosis via ROS-regulated p53 pathway. Biochem Biophys Res Commun. 496:1183–1190. 2018. View Article : Google Scholar : PubMed/NCBI
45	Liang Z, Feng Q, Xu L, Li S and Zhou L: CREPT regulated by miR-138 promotes breast cancer progression. Biochem Biophys Res Commun. 493:263–269. 2017. View Article : Google Scholar : PubMed/NCBI