Tumor Biology

, Volume 35, Issue 7, pp 7063–7072 | Cite as

Nicotine upregulates microRNA-21 and promotes TGF-β-dependent epithelial-mesenchymal transition of esophageal cancer cells

  • Yi Zhang
  • Tiecheng Pan
  • Xiaoxuan Zhong
  • Cai Cheng
Research Article


A consistent positive association between cigarette smoking and the human esophageal cancer has been confirmed all over the world. However, details in the association need to be more focused on and be identified. Recently, aberrantly expressed microRNAs (miRNAs) have been shown to be promising biomarkers for understanding the tumorigenesis of a wide array of human cancers, including the esophageal cancer, and the deregulation on the epithelial to mesenchymal transition (EMT) by miRNAs is involved in the tumorigenesis. In present study, we were going to identify the role of nicotine-induced miR-21 in the EMT of esophageal cells. We found that there was an overexpression of miR-21 in esophageal specimens, having an association with cigarette smoking, and the upregulation of miR-21 was also induced by nicotine in esophageal carcinoma cell line, EC9706. Moreover, the upregulated miR-21 by nicotine promoted EMT transforming growth factor beta (TGF-β) dependently. Thus, the present study reveals a novel oncogenic role of nicotine in human esophageal cancer.


Nicotine Epithelial-mesenchymal transition microRNA-21 TGF-β 



This study was supported by the grant of outstanding Henan province science and technology innovation talent project (114200510007).


  1. 1.
    Ferlay J, Shin HR, Bray F, Forman D, Mathers C, Parkin DM. Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. Int J Cancer J Int Cancer. 2010;127:2893–917.CrossRefGoogle Scholar
  2. 2.
    Botterweck AA, Schouten LJ, Volovics A, Dorant E, van Den Brandt PA. Trends in incidence of adenocarcinoma of the oesophagus and gastric cardia in ten European countries. Int J Epidemiol. 2000;29:645–54.CrossRefPubMedGoogle Scholar
  3. 3.
    Fernandes ML, Seow A, Chan YH, Ho KY. Opposing trends in incidence of esophageal squamous cell carcinoma and adenocarcinoma in a multi-ethnic Asian country. Am J Gastroenterol. 2006;101:1430–6.CrossRefPubMedGoogle Scholar
  4. 4.
    Wei WQ, Yang J, Zhang SW, Chen WQ, Qiao YL. Analysis of the esophageal cancer mortality in 2004–2005 and its trends during last 30 years in China. Zhonghua Yu Fang Yi Xue Za Zhi Chin J Prev Med. 2010;44:398–402.Google Scholar
  5. 5.
    Lee YC, Cohet C, Yang YC, Stayner L, Hashibe M, Straif K. Meta-analysis of epidemiologic studies on cigarette smoking and liver cancer. Int J Epidemiol. 2009;38:1497–511.CrossRefPubMedGoogle Scholar
  6. 6.
    Terry PD, Rohan TE. Cigarette smoking and the risk of breast cancer in women: a review of the literature. Cancer Epidemiol Biomark Prev Publ Am Assoc Cancer Res Am Soc Prev Oncol. 2002;11:953–71.Google Scholar
  7. 7.
    Dische S, Saunders MI, Lee M, Bennett MH. Cigarette smoking and cancer of bladder and lung. Br Med J. 1976;2:1174–5.PubMedCentralCrossRefPubMedGoogle Scholar
  8. 8.
    Tai SY, Wu IC, Wu DC, Su HJ, Huang JL, Tsai HJ, et al. Cigarette smoking and alcohol drinking and esophageal cancer risk in Taiwanese women. World J Gastroenterol WJG. 2010;16:1518–21.CrossRefPubMedGoogle Scholar
  9. 9.
    Oze I, Matsuo K, Ito H, Wakai K, Nagata C, Mizoue T, et al. Research group for the D. evaluation of cancer prevention strategies in J. cigarette smoking and esophageal cancer risk: an evaluation based on a systematic review of epidemiologic evidence among the Japanese population. Jpn J Clin Oncol. 2012;42:63–73.CrossRefPubMedGoogle Scholar
  10. 10.
    Kimm H, Kim S, Jee SH. The independent effects of cigarette smoking, alcohol consumption, and serum aspartate aminotransferase on the alanine aminotransferase ratio in Korean men for the risk for esophageal cancer. Yonsei Med J. 2010;51:310–7.PubMedCentralCrossRefPubMedGoogle Scholar
  11. 11.
    Ishiguro S, Sasazuki S, Inoue M, Kurahashi N, Iwasaki M, Tsugane S, et al. Effect of alcohol consumption, cigarette smoking and flushing response on esophageal cancer risk: a population-based cohort study (JPHC study). Cancer Lett. 2009;275:240–6.CrossRefPubMedGoogle Scholar
  12. 12.
    Gao YT, McLaughlin JK, Blot WJ, Ji BT, Benichou J, Dai Q, et al. Risk factors for esophageal cancer in Shanghai, China. I. Role of cigarette smoking and alcohol drinking. Int J Cancer J Int Cancer. 1994;58:192–6.CrossRefGoogle Scholar
  13. 13.
    Abrams JA, Lee PC, Port JL, Altorki NK, Neugut AI. Cigarette smoking and risk of lung metastasis from esophageal cancer. Cancer Epidemiol Biomark Prev Publ Am Assoc Cancer Res Am Soc Prev Oncol. 2008;17:2707–13.CrossRefGoogle Scholar
  14. 14.
    Au WW, Su D, Yuan J. Cigarette smoking in China: public health, science, and policy. Rev Environ Health. 2012;27:43–9.CrossRefPubMedGoogle Scholar
  15. 15.
    Schaal C, Chellappan SP. Nicotine-mediated cell proliferation and tumor progression in smoking-related cancers. Mol Cancer Res MCR. 2014;12:14–23.CrossRefPubMedGoogle Scholar
  16. 16.
    Brown KC, Perry HE, Lau JK, Jones DV, Pulliam JF, Thornhill BA, et al. Nicotine induces the up-regulation of the α7-nicotinic receptor (α7-nAChR) in human squamous cell lung cancer cells via the Sp1/GATA protein pathway. J Biol Chem. 2013;288:33049–59.PubMedCentralCrossRefPubMedGoogle Scholar
  17. 17.
    Cucina A, Dinicola S, Coluccia P, Proietti S, D'Anselmi F, Pasqualato A, et al. Nicotine stimulates proliferation and inhibits apoptosis in colon cancer cell lines through activation of survival pathways. J Surg Res. 2012;178:233–41.CrossRefPubMedGoogle Scholar
  18. 18.
    Al-Wadei MH, Al-Wadei HA, Schuller HM. Effects of chronic nicotine on the autocrine regulation of pancreatic cancer cells and pancreatic duct epithelial cells by stimulatory and inhibitory neurotransmitters. Carcinogenesis. 2012;33:1745–53.PubMedCentralCrossRefPubMedGoogle Scholar
  19. 19.
    Jensen K, Afroze S, Munshi MK, Guerrier M, Glaser SS. Mechanisms for nicotine in the development and progression of gastrointestinal cancers. Transl Gastrointest Cancer. 2012;1:81–7.PubMedCentralPubMedGoogle Scholar
  20. 20.
    Zou W, Zou Y, Zhao Z, Li B, Ran P. Nicotine-induced epithelial-mesenchymal transition via Wnt/β-catenin signaling in human airway epithelial cells. Am J Physiol Lung Cell Mol Physiol. 2013;304:L199–209.CrossRefPubMedGoogle Scholar
  21. 21.
    Dasgupta P, Rizwani W, Pillai S, Kinkade R, Kovacs M, Rastogi S, et al. Nicotine induces cell proliferation, invasion and epithelial-mesenchymal transition in a variety of human cancer cell lines. Int J Cancer J Int Cancer. 2009;124:36–45.CrossRefGoogle Scholar
  22. 22.
    Liu Y, Liu BA. Enhanced proliferation, invasion, and epithelial-mesenchymal transition of nicotine-promoted gastric cancer by periostin. World J Gastroenterol WJG. 2011;17:2674–80.CrossRefPubMedGoogle Scholar
  23. 23.
    Shin VY, Jin HC, Ng EK, Sung JJ, Chu KM, Cho CH. Activation of 5-lipoxygenase is required for nicotine mediated epithelial-mesenchymal transition and tumor cell growth. Cancer Lett. 2010;292:237–45.CrossRefPubMedGoogle Scholar
  24. 24.
    Ambros V. MicroRNA pathways in flies and worms: growth, death, fat, stress, and timing. Cell. 2003;113:673–6.CrossRefPubMedGoogle Scholar
  25. 25.
    Bartel DP. MicroRNAs: target recognition and regulatory functions. Cell. 2009;136:215–33.PubMedCentralCrossRefPubMedGoogle Scholar
  26. 26.
    Brennecke J, Hipfner DR, Stark A, Russell RB, Cohen SM. Bantam encodes a developmentally regulated microRNA that controls cell proliferation and regulates the proapoptotic gene hid in Drosophila. Cell. 2003;113:25–36.CrossRefPubMedGoogle Scholar
  27. 27.
    Reinhart BJ, Slack FJ, Basson M, Pasquinelli AE, Bettinger JC, Rougvie AE, et al. The 21-nucleotide let-7: RNA regulates developmental timing in Caenorhabditis elegans. Nature. 2000;403:901–6.CrossRefPubMedGoogle Scholar
  28. 28.
    Jay C, Nemunaitis J, Chen P, Fulgham P, Tong AW. miRNA profiling for diagnosis and prognosis of human cancer. DNA Cell Biol. 2007;26:293–300.CrossRefPubMedGoogle Scholar
  29. 29.
    Yu SL, Chen HY, Yang PC, Chen JJ. Unique microRNA signature and clinical outcome of cancers. DNA Cell Biol. 2007;26:283–92.CrossRefPubMedGoogle Scholar
  30. 30.
    Mani SA, Guo W, Liao MJ, Eaton EN, Ayyanan A, Zhou AY, et al. The epithelial-mesenchymal transition generates cells with properties of stem cells. Cell. 2008;133:704–15.PubMedCentralCrossRefPubMedGoogle Scholar
  31. 31.
    Gupta PB, Chaffer CL, Weinberg RA. Cancer stem cells: mirage or reality? Nat Med. 2009;15:1010–2.CrossRefPubMedGoogle Scholar
  32. 32.
    Zhang B, Zhang Z, Xia S, Xing C, Ci X, Li X, et al. KLF5 activates microRNA 200 transcription to maintain epithelial characteristics and prevent induced epithelial-mesenchymal transition in epithelial cells. Mol Cell Biol. 2013;33:4919–35.PubMedCentralCrossRefPubMedGoogle Scholar
  33. 33.
    Ding X, Park SI, McCauley LK, Wang CY. Signaling between transforming growth factor β (TGF-β) and transcription factor SNAI2 represses expression of microRNA miR-203 to promote epithelial-mesenchymal transition and tumor metastasis. J Biol Chem. 2013;288:10241–53.PubMedCentralCrossRefPubMedGoogle Scholar
  34. 34.
    Yu F, Jiao Y, Zhu Y, Wang Y, Zhu J, Cui X, et al. MicroRNA 34c gene down-regulation via DNA methylation promotes self-renewal and epithelial-mesenchymal transition in breast tumor-initiating cells. J Biol Chem. 2012;287:465–73.PubMedCentralCrossRefPubMedGoogle Scholar
  35. 35.
    Maegdefessel L, Azuma J, Toh R, Deng A, Merk DR, Raiesdana A, et al. MicroRNA-21 blocks abdominal aortic aneurysm development and nicotine-augmented expansion. Sci Transl Med. 2012;4:122ra122.CrossRefGoogle Scholar
  36. 36.
    Katsuno Y, Lamouille S, Derynck R. TGF-β signaling and epithelial-mesenchymal transition in cancer progression. Curr Opin Oncol. 2013;25:76–84.CrossRefPubMedGoogle Scholar
  37. 37.
    Fuxe J, Karlsson MC. TGF-β-induced epithelial-mesenchymal transition: a link between cancer and inflammation. Semin Cancer Biol. 2012;22:455–61.CrossRefPubMedGoogle Scholar
  38. 38.
    Bhagat TD, Zhou L, Sokol L, Kessel R, Caceres G, Gundabolu K, et al. miR-21 mediates hematopoietic suppression in MDS by activating TGF-β signaling. Blood. 2013;121:2875–81.PubMedCentralCrossRefPubMedGoogle Scholar
  39. 39.
    Di Bernardini E, Campagnolo P, Margariti A, Zampetaki A, Karamariti E, Hu Y, et al. Endothelial lineage differentiation from induced pluripotent stem cells is regulated by microRNA-21 and transforming growth factor β2 (TGF-β2) pathways. J Biol Chem. 2014;289:3383–93.PubMedCentralCrossRefPubMedGoogle Scholar
  40. 40.
    Foroni C, Broggini M, Generali D, Damia G. Epithelial-mesenchymal transition and breast cancer: role, molecular mechanisms and clinical impact. Cancer Treat Rev. 2012;38:689–97.CrossRefPubMedGoogle Scholar
  41. 41.
    Wang T, Xuan X, Pian L, Gao P, Xu H, Zheng Y, et al. Notch-1-mediated esophageal carcinoma EC-9706 cell invasion and metastasis by inducing epithelial-mesenchymal transition through Snail. Tumour Biol J Int Soc Oncodevelopmental Biol Med. 2013;35(2):1193–201.CrossRefGoogle Scholar
  42. 42.
    Li Y, Fu L, Li JB, Qin Y, Zeng TT, Zhou J, Zeng ZL, Chen J, Cao TT, Ban X, Qian C, Cai Z, Xie D, Huang P, Guan XY. Increased expression of EIF5A2, via hypoxia or gene amplification, contributes to metastasis and angiogenesis of esophageal squamous cell carcinoma. Gastroenterology 2014.Google Scholar
  43. 43.
    Min S, Xiaoyan X, Fanghui P, Yamei W, Xiaoli Y, Feng W. The glioma-associated oncogene homolog 1 promotes epithelial–mesenchymal transition in human esophageal squamous cell cancer by inhibiting E-cadherin via Snail. Cancer Gene Ther. 2013;20:379–85.CrossRefPubMedGoogle Scholar
  44. 44.
    Natsuizaka M, Kinugasa H, Kagawa S, Whelan KA, Naganuma S, Subramanian H, et al. IGFBP3 promotes esophageal cancer growth by suppressing oxidative stress in hypoxic tumor microenvironment. Am J Cancer Res. 2014;4:29–41.PubMedCentralPubMedGoogle Scholar
  45. 45.
    Han M, Wang Y, Liu M, Bi X, Bao J, Zeng N, et al. MiR-21 regulates epithelial-mesenchymal transition phenotype and hypoxia-inducible factor-1α expression in third-sphere forming breast cancer stem cell-like cells. Cancer Sci. 2012;103:1058–64.CrossRefPubMedGoogle Scholar
  46. 46.
    Han M, Liu M, Wang Y, Mo Z, Bi X, Liu Z, et al. Re-expression of miR-21 contributes to migration and invasion by inducing epithelial-mesenchymal transition consistent with cancer stem cell characteristics in MCF-7 cells. Mol Cell Biochem. 2012;363:427–36.CrossRefPubMedGoogle Scholar
  47. 47.
    Han M, Liu M, Wang Y, Chen X, Xu J, Sun Y, et al. Antagonism of miR-21 reverses epithelial-mesenchymal transition and cancer stem cell phenotype through AKT/ERK1/2 inactivation by targeting PTEN. PLoS One. 2012;7:e39520.PubMedCentralCrossRefPubMedGoogle Scholar
  48. 48.
    Pan X, Wang ZX, Wang R. MicroRNA-21: a novel therapeutic target in human cancer. Cancer Biol Ther. 2010;10:1224–32.CrossRefPubMedGoogle Scholar
  49. 49.
    Asangani IA, Rasheed SA, Nikolova DA, Leupold JH, Colburn NH, Post S, et al. MicroRNA-21 (miR-21) post-transcriptionally downregulates tumor suppressor Pdcd4 and stimulates invasion, intravasation and metastasis in colorectal cancer. Oncogene. 2008;27:2128–36.CrossRefPubMedGoogle Scholar
  50. 50.
    Lou Y, Yang X, Wang F, Cui Z, Huang Y. MicroRNA-21 promotes the cell proliferation, invasion and migration abilities in ovarian epithelial carcinomas through inhibiting the expression of PTEN protein. Int J Mol Med. 2010;26:819–27.CrossRefPubMedGoogle Scholar
  51. 51.
    Connolly EC, Van Doorslaer K, Rogler LE, Rogler CE. Overexpression of miR-21 promotes an in vitro metastatic phenotype by targeting the tumor suppressor RHOB. Mol Cancer Res MCR. 2010;8:691–700.CrossRefPubMedGoogle Scholar
  52. 52.
    Zhu S, Wu H, Wu F, Nie D, Sheng S, Mo YY. MicroRNA-21 targets tumor suppressor genes in invasion and metastasis. Cell Res. 2008;18:350–9.CrossRefPubMedGoogle Scholar
  53. 53.
    Livak KJ, Schmittgen TD. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 2001;25:402–8.CrossRefPubMedGoogle Scholar
  54. 54.
    Kim SW, Li Z, Moore PS, Monaghan AP, Chang Y, Nichols M, et al. A sensitive non-radioactive northern blot method to detect small RNAs. Nucleic Acids Res. 2010;38:e98.PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2014

Authors and Affiliations

  • Yi Zhang
    • 1
  • Tiecheng Pan
    • 2
  • Xiaoxuan Zhong
    • 1
  • Cai Cheng
    • 2
  1. 1.Department of SurgeryThe Third Affiliated Hospital of Jianghan UniversityWuhanChina
  2. 2.Department of Cardiothoracic Surgery, Tongji Hospital of Tongji Medical CollegeHuazhong University of Science and TechnologyWuhanChina

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