Skip to main content

Advertisement

SpringerLink
Log in

Overexpression of nicotinamide N-methyltransferase in HSC-2 OSCC cell line: effect on apoptosis and cell proliferation

  • Original Article
  • Published:
Clinical Oral Investigations Aims and scope Submit manuscript

Abstract

Objectives

Oral squamous cell carcinoma (OSCC) is the most common malignancy of oral cavity. Despite advances in therapeutic approaches, the 5-year survival rate for oral cancer has not improved in the last three decades. Therefore, new molecular targets for early diagnosis and treatment of OSCC are needed. In the present study, we focused on the enzyme nicotinamide N-methyltransferase (NNMT). We have previously shown that enzyme expression is upregulated in OSCC and NNMT knockdown in PE/CA PJ-15 cells significantly decreased cell growth in vitro and tumorigenicity in vivo.

Material and methods

To further explore the role of the enzyme in oral cancer cell metabolism, HSC-2 cells were transfected with the NNMT expression vector (pcDNA3-NNMT) and the effect of enzyme upregulation on cell proliferation was evaluated by MTT assay. Subsequently, we investigated at molecular level the role of NNMT on apoptosis and cell proliferation, by exploring the expression of β-catenin, survivin, and Ki-67 by real-time PCR. Moreover, we performed immunohistochemistry on 20 OSCC tissue samples to explore the expression level of NNMT and survivin ΔEx3 isoform.

Results

Enzyme upregulation significantly increased cell growth in vitro. Moreover, a positive correlation between NNMT and survivin ΔEx3 isoform expression levels was found both in HSC-2 cells and in OSCC tissue samples.

Conclusion

Taken together, our results indicate a possible involvement of NNMT in the proliferation and tumorigenic capacity of OSCC cells and seem to suggest that the enzyme could represent a potential target for the treatment of oral cancer.

Clinical relevance

The involvement of NNMT in cell growth and anti-apoptotic mechanisms seems to suggest that this enzyme could be a new therapeutic target to improve the survival of OSCC patients.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1: NNMT expression level measurements.
Fig. 2: Evaluation of cell proliferation.
Fig. 3: Expression of β-catenin, survivin isoforms, and Ki-67.
Fig. 4: Expression of NNMT and survivin-ΔEx3 in tissue samples.

Similar content being viewed by others

References

  1. Warnakulasuriya S (2009) Global epidemiology of oral and oropharyngeal cancer. Oral Oncol 45:309–316

    Article  Google Scholar 

  2. Petti S (2009) Lifestyle risk factors for oral cancer. Oral Oncol 45:340–350

    Article  Google Scholar 

  3. Gibson MK, Forastiere AA (2006) Reassessment of the role of induction chemotherapy for head and neck cancer. Lancet Oncol 7:565–574

    Article  Google Scholar 

  4. Aksoy S, Szumlanski CL, Weinshilboum RM (1994) Human liver nicotinamide N-methyltransferase. cDNA cloning, expression, and biochemical characterization. J Biol Chem 269:14835–14840

    PubMed  Google Scholar 

  5. Peng Y, Sartini D, Pozzi V, Wilk D, Emanuelli M, Yee VC (2011) Structural basis of substrate recognition in human nicotinamide N-methyltransferase. Biochem 50:7800–7808

    Article  Google Scholar 

  6. Pozzi V, Sartini D, Morganti S, Giuliante R, Di Ruscio G, Santarelli A, Rocchetti R, Rubini C, Tomasetti M, Giannatempo G, Orlando F, Provinciali M, Lo Muzio L, Emanuelli M (2013) RNA-mediated gene silencing of nicotinamide N-methyltransferase is associated with decreased tumorigenicity in human oral carcinoma cells. PLoS One 8:e71272

    Article  Google Scholar 

  7. Sartini D, Muzzonigro G, Milanese G, Pierella F, Rossi V, Emanuelli M (2006) Identification of nicotinamide N-methyltransferase as a novel tumor marker for renal clear cell carcinoma. J Urol 176:2248–2254

    Article  Google Scholar 

  8. Sartini D, Santarelli A, Rossi V, Goteri G, Rubini C, Ciavarella D, Lo Muzio L, Emanuelli M (2007) Nicotinamide N-methyltransferase upregulation inversely correlates with lymph node metastasis in oral squamous cell carcinoma. Mol Med 13:415–421

    Article  Google Scholar 

  9. Sartini D, Muzzonigro G, Milanese G, Pozzi V, Vici A, Morganti S, Rossi V, Mazzucchelli R, Montironi R, Emanuelli M (2013) Upregulation of tissue and urinary nicotinamide N-methyltransferase in bladder cancer: potential for the development of a urine-based diagnostic test. Cell Biochem Biophys 65:473–483

    Article  Google Scholar 

  10. Sartini D, Morganti S, Guidi E, Rubini C, Zizzi A, Giuliante R, Pozzi V, Emanuelli M (2013) Nicotinamide N-methyltransferase in non-small cell lung cancer: promising results for targeted anti-cancer therapy. Cell Biochem Biophys 67:865–873

    Article  Google Scholar 

  11. Emanuelli M, Santarelli A, Sartini D, Ciavarella D, Rossi V, Pozzi V, Rubini C, Lo Muzio L (2010) Nicotinamide N-Methyltransferase upregulation correlates with tumour differentiation in oral squamous cell carcinoma. Histol Histopathol 25:15–20

    PubMed  Google Scholar 

  12. Sartini D, Pozzi V, Renzi E, Morganti S, Rocchetti R, Rubini C, Santarelli A, Lo Muzio L, Emanuelli M (2012) Analysis of tissue and salivary nicotinamide N-methyltransferase in oral squamous cell carcinoma: basis for the development of a noninvasive diagnostic test for early-stage disease. Biol Chem 393:505–511

    Article  Google Scholar 

  13. Balducci E, Emanuelli M, Raffaelli N, Ruggieri S, Amici A, Magni G, Orsomando G, Polzonetti V, Natalini P (1995) Assay methods for nicotinamide mononucleotide adenylyltransferase of wide applicability. Anal Biochem 228:64–68

    Article  Google Scholar 

  14. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254

    Article  Google Scholar 

  15. Wu Y, Siadaty MS, Berens ME, Hampton GM, Theodorescu D (2008) Overlapping gene expression profiles of cell migration and tumor invasion in human bladder cancer identify metallothionein 1E and nicotinamide N-methyltransferase as novel regulators of cell migration. Oncogene 27:6679–6689

    Article  Google Scholar 

  16. Tang SW, Yang TC, Lin WC, Chang WH, Wang CC, Lai MK, Lin JY (2011) Nicotinamide N-methyltransferase induces cellular invasion through activating matrix metalloproteinase-2 expression in clear cell renal cell carcinoma cells. Carcinogenesis 32:138–145

    Article  Google Scholar 

  17. Yu T, Wang YT, Chen P, Li YH, Chen YX, Zeng H, Yu AM, Huang M, Bi HC (2015) Effects of nicotinamide N-methyltransferase on PANC-1 cells proliferation, metastatic potential and survival under metabolic stress. Cell Physiol Biochem 35:710–721

    Article  Google Scholar 

  18. Parsons RB, Aravindan S, Kadampeswaran A, Evans EA, Sandhu KK, Levy ER, Thomas MG, Austen BM, Ramsden DB (2011) The expression of nicotinamide N-methyltransferase increases ATP synthesis and protects SH-SY5Y neuroblastoma cells against the toxicity of complex I inhibitors. Biochem J 436:145–155

    Article  Google Scholar 

  19. Zhang J, Wang Y, Li G, Yu H, Xie X (2014) Down-regulation of nicotinamide N-methyltransferase induces apoptosis in human breast cancer cells via the mitochondria-mediated pathway. PLoS One 9:e89202

    Article  Google Scholar 

  20. Xie X, Yu H, Wang Y, Zhou Y, Li G, Ruan Z, Li F, Wanga X, Liu H, Zhang J (2014) Nicotinamide N-methyltransferase enhances the capacity of tumorigenesis associated with the promotion of cell cycle progression in human colorectal cancer cells. Arch Biochem Biophys 564:52–66

    Article  Google Scholar 

  21. Sartini D, Seta R, Pozzi V, Morganti S, Rubini C, Zizzi A, Tomasetti M, Santarelli L, Emanuelli M (2015) Role of nicotinamide N-methyltransferase in non-small cell lung cancer: in vitro effect of shRNA-mediated gene silencing on tumourigenicity. Biol Chem 396:225–234

    Article  Google Scholar 

  22. Pozzi V, Sartini D, Rocchetti R, Santarelli A, Rubini C, Morganti S, Giuliante R, Calabrese S, Di Ruscio G, Orlando F, Provinciali M, Saccucci F, Lo Muzio L, Emanuelli M (2015) Identification and characterization of cancer stem cells from head and neck squamous cell carcinoma cell lines. Cell Physiol Biochem 36:784–798

    Article  Google Scholar 

  23. De Maria S, Pannone G, Bufo P, Santoro A, Serpico R, Metafora S, Rubini C, Pasquali D, Papagerakis SM, Staibano S, De Rosa G, Farina E, Emanuelli M, Santarelli A, Mariggiò MA, Lo Russo L, Lo Muzio L (2009) Survivin gene-expression and splicing isoforms in oral squamous cell carcinoma. J Cancer Res Clin Oncol 135:107–116

    Article  Google Scholar 

  24. Sah NK, Seniya C (2015) Survivin splice variants and their diagnostic significance. Tumour Biol 36:6623–6631

    Article  Google Scholar 

  25. Lo Muzio L, Staibano S, Pannone G, Mignogna MD, Mariggiò A, Salvatore G, Chieffi P, Tramontano D, De Rosa G, Altieri DC (2001) Expression of the apoptosis inhibitor survivin in aggressive squamous cell carcinoma. Exp Mol Pathol 70:249–254

    Article  Google Scholar 

  26. Ge QX, Li YY, Nie YQ, Zuo WG, Du YL (2013) Expression of survivin and its four splice variants in colorectal cancer and its clinical significances. Med Oncol 30:535

    Article  Google Scholar 

  27. Marusawa H, Matsuzawa S, Welsh K, Zou H, Armstrong R, Tamm I, Reed JC (2003) HBXIP functions as a cofactor of survivin in apoptosis suppression. EMBO J 22:2729–2240

    Article  Google Scholar 

  28. Song Z, Yao X, Wu M (2003) Direct interaction between survivin and Smac/DIABLO is essential for the anti-apoptotic activity of surviving during taxol-induced apoptosis. J Biol Chem 278:23130–23140

    Article  Google Scholar 

  29. Asanuma K, Moriai R, Yajima T, Yagihashi A, Yamada M, Kobayashi D, Watanabe N (2000) Survivin as a radioresistance factor in pancreatic cancer. Jpn J Cancer Res 91:1204–1209

    Article  Google Scholar 

  30. Wang L, Zhang GM, Feng ZH (2003) Down-regulation of survivin expression reversed multidrug resistance in adriamycin-resistant HL-60/ADR cell line. Acta Pharmacol Sin 24:1235–1240

    PubMed  Google Scholar 

  31. Martinez A, Bellosillo B, Bosch F, Ferrer A, Marce S, Villamor N, Ott G, Montserrat E, Campo E, Colomer D (2004) Nuclear survivin expression in mantle cell lymphoma is associated with cell proliferation and survival. Am J Pathol 164:501–510

    Article  Google Scholar 

  32. Mahotka C, Krieg T, Krieg A, Wenzel M, Suschek CV, Heydthausen M, Gabbert HE, Gerharz CD (2002) Distinct in vivo expression patterns of survivin splice variants in renal cell carcinomas. Int J Cancer 100:30–36

    Article  Google Scholar 

  33. Mahotka C, Wenzel M, Springer E, Gabbert HE, Gerharz CD (1999) Survivin-deltaEx3 and survivin-2B: two novel splice variants of the apoptosis inhibitor survivin with different antiapoptotic properties. Cancer Res 59:6097–6102

    PubMed  Google Scholar 

  34. Waligórska-Stachura J, Andrusiewicz M, Sawicka-Gutaj N, Biczysko M, Jankowska A, Kubiczak M, Czarnywojtek A, Wrotkowska E, Ruchała M (2014) Survivin delta Ex3 overexpression in thyroid malignancies. PLoS One 9:e100534

    Article  Google Scholar 

Download references

Funding

This study was supported by grants from Marche Polytechnic University.

Author information

Author notes

  1. Riccardo Seta and Marco Mascitti contributed equally to this work.

Authors and Affiliations

  1. Department of Clinical Sciences, Polytechnic University of Marche, Via Ranieri 65, 60131, Ancona, Italy

    Riccardo Seta, Marco Mascitti, Roberto Campagna, Davide Sartini, Stefania Fumarola, Andrea Santarelli, Monia Cecati & Monica Emanuelli

  2. Department of Clinical and Experimental Medicine, University of Foggia, 71121 – 71122, Foggia, Italy

    Michele Giuliani & Lorenzo Lo Muzio

Authors
  1. Riccardo Seta
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Marco Mascitti
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Roberto Campagna
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Davide Sartini
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Stefania Fumarola
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Andrea Santarelli
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Michele Giuliani
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Monia Cecati
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Lorenzo Lo Muzio
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Monica Emanuelli
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Monica Emanuelli.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Informed consent

For this type of study, formal consent is not required.

Additional information

This paper is dedicated to the memory of my husband, Andrea L. Tranquilli (M. Emanuelli)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Seta, R., Mascitti, M., Campagna, R. et al. Overexpression of nicotinamide N-methyltransferase in HSC-2 OSCC cell line: effect on apoptosis and cell proliferation. Clin Oral Invest 23, 829–838 (2019). https://doi.org/10.1007/s00784-018-2497-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00784-018-2497-8

Keywords

Search

Navigation