miR-21 increases the programmed cell death 4 gene-regulated cell proliferation in head and neck squamous carcinoma cell lines

Sun,Zhifeng; Li,Suping; Kaufmann,Andreas  M.; Albers,Andreas  E.

doi:10.3892/or.2014.3456

miR-21 increases the programmed cell death 4 gene-regulated cell proliferation in head and neck squamous carcinoma cell lines

Authors:
- Zhifeng Sun
- Suping Li
- Andreas M. Kaufmann
- Andreas E. Albers
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Affiliations: Department of Otolaryngology, Head and Neck Surgery, Charité-Universitätsmedizin Berlin, Berlin, Germany, Clinic for Gynecology, The Affiliated Hospital of Hubei Nationalities University, Enshi, Hubei, P.R. China, Clinic for Gynecology, Charité-Universitätsmedizin Berlin, Berlin, Germany
Published online on: September 1, 2014 https://doi.org/10.3892/or.2014.3456
Pages: 2283-2289

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Abstract

MicroRNAs (miRs) are small non-coding RNAs that regulate the translation of many genes in normal and cancer cells where they are frequently dysregulated promoting tumor progression. Several studies have illustrated the potential of manipulating miR expression in cancer research and therapy. The aim of the present study was to investigate expression patterns of a panel of miRs in head and neck squamous cell carcinoma (HNSCC) shown to be relevant in other carcinomas and to elucidate their role if dysregulated. We performed analysis of miR‑21, -200c, -138-1, -138-2, -25 and -34 expression by qRT-PCR in 6 HNSCC cell lines and computerized search for genetic targets of dysregulated miRNA-21 (miR‑21). Lipofection of mock and anti-miR-21 and determination of expression efficiencies and final programmed cell death 4 (PDCD4) expression were carried out by luciferase assay and western blotting. MTT assay was used to measure cell proliferation and flow cytometry was performed for cell cycle analysis. Expression of miR-21 was most prominently upregulated in the HNSCC cell lines, particularly in UM-SCC11B (6.45±0.25-fold, P<0.05) and UM-SCC9 (4.35±0.22-fold, P<0.05) as compared to primary epidermal keratinocytes used as control. The expression levels of the other miRs showed no difference except for miR-34 and -138-1 each in one cell line. Subsequent transfection of precursor miR-21 stimulated proliferation while anti-miR-21 inhibited proliferation of both cell lines. PDCD4 was identified with software designed for this purpose as potential target gene of miR-21. Subsequently, its role in HNSCC lines was experimentally confirmed by regulation of PDCD4 transfecting miR-21 mimics and anti-miR-21. Finally, we showed that PDCD4 is negatively regulated by miR-21 at the post-transcriptional level via binding to the 3'-untranslated region of PDCD4 mRNA. A role of upregulated miR-21 and reduced PDCD4 stimulating the proliferation was demonstrated in HNSCC lines and, in turn, transfection of anti-miR-21 upregulating PDCD4 reduced the cellular division rate. We explored miR-21 and PDCD4 expression as markers of progression and prognosis and for a potential translational value in the development of agents slowing growth of HNSCC and other carcinomas useful in palliative therapy or as a component of multi-modality treatments.

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1	Jemal A, Bray F, Center MM, Ferlay J, Ward E and Forman D: Global cancer statistics. CA Cancer J Clin. 61:69–90. 2011. View Article : Google Scholar
2	Cheng ZX, Sun B, Wang SJ, et al: Nuclear factor-κB-dependent epithelial to mesenchymal transition induced by HIF-1α activation in pancreatic cancer cells under hypoxic conditions. PLoS One. 6:e237522011.
3	Semenza GL: HIF-1 and tumor progression: pathophysiology and therapeutics. Trends Mol Med. 8(Suppl 4): S62–S67. 2002. View Article : Google Scholar : PubMed/NCBI
4	Imai T, Horiuchi A, Wang C, et al: Hypoxia attenuates the expression of E-cadherin via up-regulation of SNAIL in ovarian carcinoma cells. Am J Pathol. 163:1437–1447. 2003. View Article : Google Scholar : PubMed/NCBI
5	Lu J, Getz G, Miska EA, et al: MicroRNA expression profiles classify human cancers. Nature. 435:834–838. 2005. View Article : Google Scholar : PubMed/NCBI
6	Volinia S, Calin GA, Liu CG, 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
7	Rosenfeld N, Aharonov R, Meiri E, et al: MicroRNAs accurately identify cancer tissue origin. Nat Biotechnol. 26:462–469. 2008. View Article : Google Scholar : PubMed/NCBI
8	Chu Y, Zhu H, Lv L, Zhou Y and Huo J: MiRNAs in oesophageal squamous cancer. Neth J Med. 71:69–75. 2013.PubMed/NCBI
9	Janiszewska J, Szaumkessel M and Szyfter K: microRNAs are important players in head and neck carcinoma: a review. Crit Rev Oncol Hematol. 88:716–728. 2013. View Article : Google Scholar : PubMed/NCBI
10	Wu Q, Wang C, Lu Z, Guo L and Ge Q: Analysis of serum genome-wide microRNAs for breast cancer detection. Clin Chim Acta. 413:1058–1065. 2012. View Article : Google Scholar : PubMed/NCBI
11	Liang H, Gong F, Zhang S, Zhang CY, Zen K and Chen X: The origin, function, and diagnostic potential of extracellular microRNAs in human body fluids. Wiley Interdiscip Rev RNA. 5:285–300. 2014. View Article : Google Scholar : PubMed/NCBI
12	Endo K, Weng H, Kito N, Fukushima Y and Iwai N: miR-216a and miR-216b as markers for acute phased pancreatic injury. Biomed Res. 34:179–188. 2013. View Article : Google Scholar : PubMed/NCBI
13	Olivieri F, Rippo MR, Procopio AD and Fazioli F: Circulating inflamma-miRs in aging and age-related diseases. Front Genet. 4:1212013.
14	de Yébenes VG, Bartolomé-Izquierdo N and Ramiro AR: Regulation of B-cell development and function by microRNAs. Immunol Rev. 253:25–39. 2013.PubMed/NCBI
15	Stefani G: Roles of microRNAs and their targets in cancer. Expert Opin Biol Ther. 7:1833–1840. 2007. View Article : Google Scholar : PubMed/NCBI
16	Yang J, Hao Y and Xi JJ: Therapeutic application of microRNAs against human cancers. J Lab Autom. 18:30–33. 2013. View Article : Google Scholar : PubMed/NCBI
17	Farazi TA, Spitzer JI, Morozov P and Tuschl T: miRNAs in human cancer. J Pathol. 223:102–115. 2011. View Article : Google Scholar
18	Lo WL, Yu CC, Chiou GY, et al: MicroRNA-200c attenuates tumour growth and metastasis of presumptive head and neck squamous cell carcinoma stem cells. J Pathol. 223:482–495. 2011. View Article : Google Scholar : PubMed/NCBI
19	Liu X, Jiang L, Wang A, Yu J, Shi F and Zhou X: MicroRNA-138 suppresses invasion and promotes apoptosis in head and neck squamous cell carcinoma cell lines. Cancer Lett. 286:217–222. 2009. View Article : Google Scholar : PubMed/NCBI
20	Chen HC, Chen GH, Chen YH, et al: MicroRNA deregulation and pathway alterations in nasopharyngeal carcinoma. Br J Cancer. 100:1002–1011. 2009. View Article : Google Scholar : PubMed/NCBI
21	Xu X, Chen Z, Zhao X, et al: MicroRNA-25 promotes cell migration and invasion in esophageal squamous cell carcinoma. Biochem Biophys Res Commun. 421:640–645. 2012. View Article : Google Scholar : PubMed/NCBI
22	Gao W, Xu J, Liu L, Shen H, Zeng H and Shu Y: A systematic-analysis of predicted miR-21 targets identifies a signature for lung cancer. Biomed Pharmacother. 66:21–28. 2012. View Article : Google Scholar : PubMed/NCBI
23	Schee K, Boye K, Abrahamsen TW, Fodstad Ø and Flatmark K: Clinical relevance of microRNA miR-21, miR-31, miR-92a, miR-101, miR-106a and miR-145 in colorectal cancer. BMC Cancer. 12:5052012. View Article : Google Scholar : PubMed/NCBI
24	Young MR, Santhanam AN, Yoshikawa N and Colburn NH: Have tumor suppressor PDCD4 and its counteragent oncogenic miR-21 gone rogue? Mol Interv. 10:76–79. 2010. View Article : Google Scholar : PubMed/NCBI
25	Allgayer H: Pdcd4, a colon cancer prognostic that is regulated by a microRNA. Crit Rev Oncol Hematol. 73:185–191. 2010. View Article : Google Scholar : PubMed/NCBI
26	Fassan M, Pizzi M, Battaglia G, et al: Programmed cell death 4 (PDCD4) expression during multistep Barrett’s carcinogenesis. J Clin Pathol. 63:692–696. 2010.PubMed/NCBI
27	Fassan M, Pizzi M, Giacomelli L, et al: PDCD4 nuclear loss inversely correlates with miR-21 levels in colon carcinogenesis. Virchows Arch. 458:413–419. 2011. View Article : Google Scholar : PubMed/NCBI
28	Chen C, Ridzon DA, Broomer AJ, et al: Real-time quantification of microRNAs by stem-loop RT-PCR. Nucleic Acids Res. 33:e1792005. View Article : Google Scholar : PubMed/NCBI
29	Wolf JS, Chen Z, Dong G, et al: IL (interleukin)-1α promotes nuclear factor-κB and AP-1-induced IL-8 expression, cell survival, and proliferation in head and neck squamous cell carcinomas. Clin Cancer Res. 7:1812–1820. 2001.
30	Itani S, Kunisada T, Morimoto Y, et al: MicroRNA-21 correlates with tumorigenesis in malignant peripheral nerve sheath tumor (MPNST) via programmed cell death protein 4 (PDCD4). J Cancer Res Clin Oncol. 138:1501–1509. 2012. View Article : Google Scholar : PubMed/NCBI
31	Horiuchi A, Iinuma H, Akahane T, Shimada R and Watanabe T: Prognostic significance of PDCD4 expression and association with microRNA-21 in each Dukes’ stage of colorectal cancer patients. Oncol Rep. 27:1384–1392. 2012.PubMed/NCBI
32	Wang J and Zhang Y: Expression of programmed cell death 4 protein is closely correlated with laryngeal squamous cell carcinomas. Lin Chung Er Bi Yan Hou Tou Jing Wai Ke Za Zhi. 25:539–541. 2011.(In Chinese).
33	Bartel DP: MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 116:281–297. 2004. View Article : Google Scholar : PubMed/NCBI
34	Iorio MV, Ferracin M, Liu CG, et al: MicroRNA gene expression deregulation in human breast cancer. Cancer Res. 65:7065–7070. 2005. View Article : Google Scholar : PubMed/NCBI
35	Chan JA, Krichevsky AM and Kosik KS: MicroRNA-21 is an antiapoptotic factor in human glioblastoma cells. Cancer Res. 65:6029–6033. 2005. View Article : Google Scholar : PubMed/NCBI
36	Shi GH, Ye DW, Yao XD, et al: Involvement of microRNA-21 in mediating chemo-resistance to docetaxel in androgen-independent prostate cancer PC3 cells. Acta Pharmacol Sin. 31:867–873. 2010. View Article : Google Scholar : PubMed/NCBI
37	Xu YZ, Xi QH, Ge WL and Zhang XQ: Identification of serum microRNA-21 as a biomarker for early detection and prognosis in human epithelial ovarian cancer. Asian Pac J Cancer Prev. 14:1057–1060. 2013. View Article : Google Scholar : PubMed/NCBI
38	Giovannetti E, Funel N, Peters GJ, et al: MicroRNA-21 in pancreatic cancer: correlation with clinical outcome and pharmacologic aspects underlying its role in the modulation of gemcitabine activity. Cancer Res. 70:4528–4538. 2010. View Article : Google Scholar : PubMed/NCBI
39	Feng YH, Wu CL, Tsao CJ, et al: Deregulated expression of sprouty2 and microRNA-21 in human colon cancer: correlation with the clinical stage of the disease. Cancer Biol Ther. 11:111–121. 2011. View Article : Google Scholar : PubMed/NCBI
40	Zhang BG, Li JF, Yu BQ, Zhu ZG, Liu BY and Yan M: microRNA-21 promotes tumor proliferation and invasion in gastric cancer by targeting PTEN. Oncol Rep. 27:1019–1026. 2012.PubMed/NCBI
41	Selaru FM, Olaru AV, Kan T, et al: MicroRNA-21 is overexpressed in human cholangiocarcinoma and regulates programmed cell death 4 and tissue inhibitor of metalloproteinase 3. Hepatology. 49:1595–1601. 2009. View Article : Google Scholar : PubMed/NCBI
42	Meng F, Henson R, Wehbe-Janek H, Ghoshal K, Jacob ST and Patel T: MicroRNA-21 regulates expression of the PTEN tumor suppressor gene in human hepatocellular cancer. Gastroenterology. 133:647–658. 2007. View Article : Google Scholar : PubMed/NCBI
43	Tran N, McLean T, Zhang X, et al: MicroRNA expression profiles in head and neck cancer cell lines. Biochem Biophys Res Commun. 358:12–17. 2007. View Article : Google Scholar
44	Huang S, Li XQ, Chen X, Che SM, Chen W and Zhang XZ: Inhibition of microRNA-21 increases radiosensitivity of esophageal cancer cells through phosphatase and tensin homolog deleted on chromosome 10 activation. Dis Esophagus. 26:823–831. 2013. View Article : Google Scholar
45	Schetter AJ, Leung SY, Sohn JJ, et al: MicroRNA expression profiles associated with prognosis and therapeutic outcome in colon adenocarcinoma. JAMA. 299:425–436. 2008. View Article : Google Scholar : PubMed/NCBI
46	Dillhoff M, Liu J, Frankel W, Croce C and Bloomston M: MicroRNA-21 is overexpressed in pancreatic cancer and a potential predictor of survival. J Gastrointest Surg. 12:2171–2176. 2008. View Article : Google Scholar : PubMed/NCBI
47	Asangani IA, Rasheed SA, Nikolova DA, et al: MicroRNA-21 (miR-21) post-transcriptionally downregulates tumor suppressor Pdcd4 and stimulates invasion, intravasation and metastasis in colorectal cancer. Oncogene. 27:2128–2136. 2008. View Article : Google Scholar : PubMed/NCBI
48	Feber A, Xi L, Luketich JD, et al: MicroRNA expression profiles of esophageal cancer. J Thorac Cardiovasc Surg. 135:255–260. 2008. View Article : Google Scholar
49	Fulci V, Chiaretti S, Goldoni M, et al: Quantitative technologies establish a novel microRNA profile of chronic lymphocytic leukemia. Blood. 109:4944–4951. 2007. View Article : Google Scholar : PubMed/NCBI
50	Iorio MV, Visone R, Di Leva G, et al: MicroRNA signatures in human ovarian cancer. Cancer Res. 67:8699–8707. 2007. View Article : Google Scholar : PubMed/NCBI
51	Markou A, Tsaroucha EG, Kaklamanis L, Fotinou M, Georgoulias V and Lianidou ES: Prognostic value of mature microRNA-21 and microRNA-205 overexpression in non-small cell lung cancer by quantitative real-time RT-PCR. Clin Chem. 54:1696–1704. 2008. View Article : Google Scholar
52	Yang HS, Jansen AP, Nair R, et al: A novel transformation suppressor, Pdcd4, inhibits AP-1 transactivation but not NF-κB or ODC transactivation. Oncogene. 20:669–676. 2001.PubMed/NCBI
53	Jansen AP, Camalier CE, Stark C and Colburn NH: Characterization of programmed cell death 4 in multiple human cancers reveals a novel enhancer of drug sensitivity. Mol Cancer Ther. 3:103–110. 2004.PubMed/NCBI
54	Jansen AP, Camalier CE and Colburn NH: Epidermal expression of the translation inhibitor programmed cell death 4 suppresses tumorigenesis. Cancer Res. 65:6034–6041. 2005. View Article : Google Scholar : PubMed/NCBI
55	Göke A, Göke R, Knolle A, et al: DUG is a novel homologue of translation initiation factor 4G that binds eIF4A. Biochem Biophys Res Commun. 297:78–82. 2002.PubMed/NCBI
56	Yang HS, Jansen AP, Komar AA, et al: The transformation suppressor Pdcd4 is a novel eukaryotic translation initiation factor 4A binding protein that inhibits translation. Mol Cell Biol. 23:26–37. 2003. View Article : Google Scholar : PubMed/NCBI
57	Göke R, Barth P, Schmidt A, Samans B and Lankat-Buttgereit B: Programmed cell death protein 4 suppresses CDK1/cdc2 via induction of p21^Waf1/Cip1. Am J Physiol Cell Physiol. 287:C1541–C1546. 2004.PubMed/NCBI
58	Frankel LB, Christoffersen NR, Jacobsen A, Lindow M, Krogh A and Lund AH: Programmed cell death 4 (PDCD4) is an important functional target of the microRNA miR-21 in breast cancer cells. J Biol Chem. 283:1026–1033. 2008. View Article : Google Scholar : PubMed/NCBI
59	Xu L, Dai WQ, Xu XF, Wang F, He L and Guo CY: Effects of multiple-target anti-microRNA antisense oligodeoxyribonucleotides on proliferation and migration of gastric cancer cells. Asian Pac J Cancer Prev. 13:3203–3207. 2012. View Article : Google Scholar : PubMed/NCBI
60	Lennox KA and Behlke MA: A direct comparison of anti-microRNA oligonucleotide potency. Pharm Res. 27:1788–1799. 2010. View Article : Google Scholar : PubMed/NCBI