, Volume 65, Issue 5, pp 839–850 | Cite as

Evaluation of the effects of nicorandil and its molecular precursor (without radical NO) on proliferation and apoptosis of 786-cell

  • Natália Aparecida de Paula
  • Andressa Megumi Niwa
  • Diogo Campos Vesenick
  • Carolina Panis
  • Rubens Cecchini
  • Ângelo de Fátima
  • Lúcia Regina Ribeiro
  • Mário Sérgio Mantovani
Original Research


Nicorandil is a nitric oxide (NO) donor used in the treatment of angina symptoms. It has also been reported to protect cells and affect the proliferation and death of cells in some tissues. The molecules that interfere with these processes can cause dysfunction in healthy tissues but can also assist in the therapy of some disorders. In this study we examined the effect of nicorandil and of the molecular precursor that does not have the NO radical (N-(beta-hydroxyethyl) nicotinamide) on the cell proliferation and death of human renal carcinoma cells (786-O) under normal oxygenation conditions. The molecular precursor was used in order to analyze the effects independents of NO. In the cytotoxicity test, nicorandil was shown to be cytotoxic at very high concentrations and it was more cytotoxic than its precursor (cytotoxic at concentrations of 2,000 and 3,000 μg/mL, respectively). We propose that the lower cytotoxicity of the precursor is due to the absence of the NO radical. In this study, the cells exposed to nicorandil showed neither statistically significant changes in cell proliferation nor increases in apoptosis or genotoxicity. The precursor generated similar results to those of nicorandil. We conclude that nicorandil causes no changes in the proliferation or apoptosis of the cell 786-O in normal oxygenation conditions. Moreover, the lack of NO radical in the precursor molecule did not show a different result, except in the cell cytotoxicity.


Nicorandil N-(beta-hydroxyethyl) nicotinamide 786-O cells Cell proliferation Apoptosis Cytotoxicity 



We would like to thank the Coordination of Improvement of Higher Education Personnel (CAPES), National Council for Scientific and Technological Development (CNPq) and Araucaria Foundation for their financial support.


  1. Ahmed LA, Salem HA, Attia AS, Agha AM (2011) Pharmacological preconditioning with nicorandil and pioglitazone attenuates myocardial ischemia/reperfusion injury in rats. Eur J Pharmacol 663:51–58CrossRefGoogle Scholar
  2. Akao M, Teshima Y, Marbán E (2002) Antiapoptotic effect of nicorandil mediated by mitochondrial ATP-sensitive potassium channels in cultured cardiac myocytes. J Am Coll Cardiol 40:803–810CrossRefGoogle Scholar
  3. Barreto RL, Correia CRD (2005) Óxido nítrico: propriedades e potenciais usos terapêuticos. Quim Nova 28:1046–1054CrossRefGoogle Scholar
  4. Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, Mueller R, Nolan T, Pfaffl MW, Shipley GL, Vandesompele J, Wittwer CT (2009) The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments. Clin Chem 55:611–622CrossRefGoogle Scholar
  5. Carreira RS, Monteiro P, Kowaltowski AJ, Gonçalves LM, Providência LA (2008) Nicorandil protects cardiac mitochondria against permeability transition induced by ischemia-reperfusion. J Bioeng Biomembr 40:95–102CrossRefGoogle Scholar
  6. Castaneda F, Rosin-Steiner S (2006) Low concentration of ethanol induce apoptosis in HepG2 cells: role of various signal transduction pathways. Int J Med Sci 3:160–167CrossRefGoogle Scholar
  7. Chen XY, Liu J, Xu KS (2009) Apoptosis of human hepatocellular carcinoma cell (HepG2) induced by cardiotoxin III through S-phase arrest. Exp Toxicol Pathol 61:307–315CrossRefGoogle Scholar
  8. Chong S, Fung HL (1991) Biochemical and pharmacological interactions between nitroglycerin and thiols. Effects of thiol structure on nitric oxide generation and tolerance reversal. Biochem Pharmacol 42:1433–1439CrossRefGoogle Scholar
  9. Dimmeler S, Zeiher AM (1997) Nitric oxide and apoptosis: another paradigm for the double-edged role of nitric oxide. Nitric Oxide 1:5–281CrossRefGoogle Scholar
  10. Eeckhout E (2003) Nicorandil: a drug for many purposes: too good to be true? Eur Heart J 24:1282–1284CrossRefGoogle Scholar
  11. Frydman A (1992) Pharmacokinetic profile of nicorandil in humans: an overview. J Cardiovasc Pharmacol 20:34–44CrossRefGoogle Scholar
  12. Griess JP (1879) Bemerkungen zu der abhandlung der H.H. Weselsky und Benedikt “ueber einige azoverbindugen”. Chem Ber 12:426–428CrossRefGoogle Scholar
  13. Hiremath JG, Valluru R, Jaiprakash N, Katta SA, Matad PP (2010) Pharmaceutical aspects of Nicorandil. Int J Pharm Pharm Sci 2:24–29Google Scholar
  14. Huang Y-H, Shang B-Y, Zhen Y-S (2005) Antitumor efficacy of lidamycin on hepatoma and active moiety of its molecule. World J Gastroenterol 11:3980–3984Google Scholar
  15. Hwang S-Y, Yoo B-C, Jung J-W, Oh E-S, Hwang J-S, Shin J-A, Kim S-Y, Cha S-H, Han I-O (2009) Induction of glioma apoptosis by microglia-secreted molecules: the role of nitric oxide and cathepsin B. Biochim Biophys Acta 1793:1656–1668CrossRefGoogle Scholar
  16. Ishii H, Toriyama T, Aoyama T, Takahashi H, Yamada S, Kasuga H, Ichimiya S, Kanashiro M, Mitsuhashi H, Maruyama S, Matsuo S, Naruse K, Matsubara T, Murohara T (2007) Efficacy of oral nicorandil in patients with end-stage renal disease: a retrospective chart review after coronary angioplasty in Japanese patients receiving hemodialysis. Clin Ther 29–1:110–122CrossRefGoogle Scholar
  17. Jefferson JA, Shankland SJ, Pichler RH (2008) Proteinuria in diabetic kidney disease: a mechanistic viewpoint. Kidney Int 74–1:22–36CrossRefGoogle Scholar
  18. Kastrati I, Edirisinghe PD, Wijewickrama GT, Thatcher GR (2010) Estrogen-induced apoptosis of breast epithelial cells is blocked by NO/cGMP and mediated by extranuclear estrogen receptors. Endocrinology 151:5206–5216CrossRefGoogle Scholar
  19. Kobayashi H (1995) A comparison between manual microscopic analysis and computerized image analysis in the single cell gel electrophoresis assay. MMS Commun 3:103–115Google Scholar
  20. Krumenacker JS, Murad F (2006) NO-cGMP signaling in development and stem cells. Mol Genet Metab 87:311–314CrossRefGoogle Scholar
  21. Liou JY, Hong HJ, Sung LC, Chao HH, Chen PY, Cheng TH, Chan P, Liu JC (2011) Nicorandil inhibits angiotensin-II-induced proliferation of cultured rat cardiac fibroblasts. Pharmacology 87:144–151CrossRefGoogle Scholar
  22. Lu C (2006) Nicorandil improves post-ischemic myocardial dysfunction in association with opening the mitochondrial KATP channels and decreasing hydroxyl radicals in isolated rat hearts. Circ J 70:1650–1654CrossRefGoogle Scholar
  23. Masri F (2010) Role of nitric oxide and its metabolites as potential markers in lung cancer. Ann Thorac Med 5:123–127CrossRefGoogle Scholar
  24. Moncada S, Palmer RMJ, Higgs EA (1991) Nitric oxide: physiology, pathophysiology, and pharmacology. Pharmacol Rev 43:109–142Google Scholar
  25. Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and citotoxicity assays. J Immunol Methods 65:55–63CrossRefGoogle Scholar
  26. Mujoo K, Sharin VG, Martin E, Choi BK, Sloan C, Nikonoff LE, Kots AY, Murad F (2010) Role of soluble guanylyl cyclase–cyclic GMP signaling in tumor cell proliferation. Nitric Oxide 22:43–50CrossRefGoogle Scholar
  27. Nagata K, Obata K, Odashima M, Yamada A, Somura F, Nishizawa T, Ichihara S, Izawa H, Iwase M, Hayakawa A, Murohara T, Yokota M (2003) Nicorandil inhibits oxidative stress-induced apoptosis in cardiac myocytes through activation of mitochondrial ATP-sensitive potassium channels and a nitrate-like effect. J Mol Cell Cardiol 35:1505–1512CrossRefGoogle Scholar
  28. Nguyen T, Brunson D, Crespi CL, Penman BW, Wishnok JS, Tannenbaum SR (1992) DNA damage and mutation in human cells exposed to nitric oxide in vitro. Proc Natl Acad Sci USA 89:3030–3034CrossRefGoogle Scholar
  29. Nishikawa S, Tatsumi T, Shiraishi J, Matsunaga S, Takeda M, Mano A, Kobara M, Keira N, Okigaki M, Takahashi T, Matsubara H (2006) Nicorandil regulates Bcl-2 family proteins and protects cardiac myocytes against hypoxia-induced apoptosis. J Mol Cell Cardiol 40:510–519CrossRefGoogle Scholar
  30. Panis C, Mazzuco TL, Costa CZF, Victorino VJ, Tatakihara VLH, Yamauchi LM, Yamada-Ogatta SF, Cecchini R, Rizzo LV, Pinge-Filho P (2010) Trypanosoma cruzi: effect of the absence of 5-lipoxygenase (5-LO)-derived leukotrienes on levels of cytokines, nitric oxide and iNOS expression in cardiac tissue in the acute phase of infection in mice. Exp Parasitol 127:58–65CrossRefGoogle Scholar
  31. Peters H, Daig U, Martini S, Rückert M, Schäper F, Liefeldt L, Krämer S, Neumayer HH (2003) NO mediates antifibrotic actions of l-arginine supplementation following induction of anti-thy1 glomerulonephritis. Kidney Int 64:509–518CrossRefGoogle Scholar
  32. Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29:2002–2007CrossRefGoogle Scholar
  33. Pfaffl MW, Horgan GW, Dempfle L (2002) Relative expression software tool (REST©) for group-wise comparison and statistical analysis of relative expression results in real-time PCR. Nucleic Acids Res 30:1–10CrossRefGoogle Scholar
  34. Rovozzo GC, Burke CN (1973) A manual of basic virological techniques. Prentice-Hall, Englewood CliffsGoogle Scholar
  35. Sato T, Sasaki N, O’Rourke B, Marbán E (2000) Nicorandil, a potent cardioprotective agent, acts by opening mitochondrial ATP—dependent potassium. Channels J Am Coll Cardiol 35:514–518CrossRefGoogle Scholar
  36. Segawa K, Minami K, Shiga Y, Shiraishi M, Sata T, Nakashima Y, Shigematsu A (2001) Inhibitory effects of nicorandil on rat mesangial cell proliferation via the protein kinase G pathway. Nephron 87:263–268CrossRefGoogle Scholar
  37. Serizawa K, Yogo K, Aizawa K, Tashiro Y, Ishizuka N (2011) Nicorandil prevents endothelial dysfunction due to antioxidative effects via normalisation of NADPH oxidase and nitric oxide synthase in streptozotocin diabetic rats. Cardiovasc Diabetol 10:105CrossRefGoogle Scholar
  38. Seth P, Fung HL (1993) Biochemical characterization of a membrane-bound enzyme responsible for generating nitric oxide from nitroglycerin in vascular smooth muscle cells. Biochem Pharmacol 46:1481–1486CrossRefGoogle Scholar
  39. Simpson D, Wellington K (2004) Nicorandil: a review of its use in the management of stable angina pectoris, including high-risk patients. Drugs 64:1941–1955CrossRefGoogle Scholar
  40. Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175:184–191CrossRefGoogle Scholar
  41. Sudo H, Hirata M, Kanada H, Yorozu K, Tashiro Y, Serizawa K-I, Yogo K, Kataoka M, Moriguchi Y, Ishizuka N (2009) Nicorandil improves glomerular injury in rats with mesangioproliferative glomerulonephritis via inhibition of proproliferative and profibrotic growth factors. J Pharmacol Sci 111:53–59CrossRefGoogle Scholar
  42. Sugaya S, Nakanishi H, Tanzawa H (2005) Down-regulation of SMT3A gene expression in association with DNA synthesis induction after X-ray irradiation in nevoid basal cell carcinoma syndrome (NBCCS) cells. Mutat Res 578:327–332CrossRefGoogle Scholar
  43. Taimor G, Hofstaetter B, Piper HM (2000) Apoptosis induction by nitric oxide in adult cardiomyocytes via cGMP-signaling and its impairment after simulated ischemia. Cardiovasc Res 45:588–594CrossRefGoogle Scholar
  44. Tice RR, Agurell E, Anderson D, Burlinson B, Hartmann A, Kobayashi H, Miyamae Y, Rojas E, Ryu J-C, Sasaki YF (2000) Single cell gel/comet assay: guideline for in vitro and in vivo genetic toxicology testing. Environ Mol Mutagen 35:206–221CrossRefGoogle Scholar
  45. Tsuboy MS, Marcarini JC, Luiz RC, Barros IB, Ferreira DT, Ribeiro LR, Mantovani MS (2010) In vitro evaluation of the genotoxic activity and apoptosis induction of the extracts of roots and leaves from the medicinal plant Coccoloba mollis (Polygonaceae). J Med Food 13:503–508CrossRefGoogle Scholar
  46. Wink DA, Vodovotz Y, Laval J, Laval F, Dewhirst MW, Mitchell JB (1998) The multifaceted roles of nitric oxide in cancer. Carcinogenesis 19:711–721CrossRefGoogle Scholar
  47. Yim CY et al (1993) Macrophage nitric oxide synthesis delays progression of ultraviolet light induced murine Skin Cancers. Cancer Res 53:5507–5511Google Scholar
  48. Zhang T, Otevrel T, Gao Z, Gao Z, Ehrlich SM, Fields JZ, Boman BM (2001) Evidence that APC regulates survivin expression: a possible mechanism contributing to the stem cell origin of colon cancer. Cancer Res 61:8664–8667Google Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Natália Aparecida de Paula
    • 1
  • Andressa Megumi Niwa
    • 1
  • Diogo Campos Vesenick
    • 1
  • Carolina Panis
    • 2
  • Rubens Cecchini
    • 2
  • Ângelo de Fátima
    • 3
  • Lúcia Regina Ribeiro
    • 4
  • Mário Sérgio Mantovani
    • 1
  1. 1.Laboratório de Genética Toxicológica, Departamento de Biologia Geral, Centro de Ciências BiológicasUniversidade Estadual de LondrinaLondrinaBrazil
  2. 2.Laboratório de Imunopatologia Experimental, Departamento de Ciências Patológicas, Centro de Ciências BiológicasUniversidade Estadual de LondrinaLondrinaBrazil
  3. 3.Departamento de QuímicaUniversidade Federal de Minas GeraisPampulha, Belo HorizonteBrazil
  4. 4.Instituto de BiociênciasUniversidade Estadual PaulistaRio ClaroBrazil

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