Medical Oncology

, 32:266 | Cite as

Effects of X-radiation on lung cancer cells: the interplay between oxidative stress and P53 levels

  • Fernando Mendes
  • Tiago Sales
  • Cátia Domingues
  • Susann Schugk
  • Ana Margarida Abrantes
  • Ana Cristina Gonçalves
  • Ricardo Teixo
  • Rita Silva
  • João Casalta-Lopes
  • Clara Rocha
  • Mafalda Laranjo
  • Paulo César Simões
  • Ana Bela Sarmento Ribeiro
  • Maria Filomena Botelho
  • Manuel Santos Rosa
Original Paper
First Online:
Received:
Accepted:

Abstract

Lung cancer (LC) ranks as the most prevalent and deadliest cause of cancer death worldwide. Treatment options include surgery, chemotherapy and/or radiotherapy, depending on LC staging, without specific highlight. The aim was to evaluate the effects of X-radiation in three LC cell lines. H69, A549 and H1299 cell lines were cultured and irradiated with 0.5–60 Gy of X-radiation. Cell survival was evaluated by clonogenic assay. Cell death and the role of reactive oxygen species, mitochondrial membrane potential, BAX, BCL-2 and cell cycle were analyzed by flow cytometry. Total and phosphorylated P53 were assessed by western blotting. Ionizing radiation decreases cell proliferation and viability in a dose-, time- and cell line-dependent manner, inducing cell death preferentially by apoptosis with cell cycle arrest. These results may be related to differences in P53 expression and oxidative stress response. The results obtained indicate that sensibility and/or resistance to radiation may be dependent on molecular LC characteristics which could influence response to radiotherapy and treatment success.

Keywords

Radiotherapy Lung cancer Oxidative stress P53 
This is a preview of subscription content, log in to check access.

Notes

Acknowledgments

The authors would like to thank the Portuguese Foundation for Science and Technology for the award Portugal (Strategic Project PEst-C/SAU/UI3282/2013 and UID/NEU/04539/2013), COMPETE-FEDER.

Compliance with ethical standards

Conflict of interest

None.

References

  1. 1.
    Dela CS, Cruz LT, Tanoue RA. Matthay, Lung cancer: epidemiology, etiology, and prevention. Clin Chest Med. 2011;32:605–44. doi: 10.1016/j.ccm.2011.09.001.CrossRefGoogle Scholar
  2. 2.
    Torre LA, Bray F, Siegel RL, Ferlay J, Lortet-Tieulent J, Jemal A. Global cancer statistics, 2012. CA Cancer J Clin. 2015;65:87–108. doi: 10.3322/caac.21262.CrossRefPubMedGoogle Scholar
  3. 3.
    Mendes F, Antunes C, Abrantes AM, Gonçalves A, Nobre-Gois I, Sarmento-Ribeiro AB, et al. Lung cancer: the immune system and radiation. Br J Biomed Sci. 2015;72:78–84.PubMedGoogle Scholar
  4. 4.
    Rosell R, Karachaliou N. Lung cancer in 2014: optimizing lung cancer treatment approaches. Nat Rev Clin Oncol. 2014;12(2014):75–6. doi: 10.1038/nrclinonc.2014.225.CrossRefPubMedGoogle Scholar
  5. 5.
    Verma V. Lung cancer: implementing lung-cancer screening-oncological “grey areas”. Nat Rev Clin Oncol. 2015;12:256–7. doi: 10.1038/nrclinonc.2015.65.CrossRefPubMedGoogle Scholar
  6. 6.
    Muaddi H, Chowdhury S, Vellanki R, Zamiara P, Koritzinsky M. Contributions of AMPK and p53 dependent signaling to radiation response in the presence of metformin. Radiother Oncol. 2013;108:446–50. doi: 10.1016/j.radonc.2013.06.014.CrossRefPubMedGoogle Scholar
  7. 7.
    Bieging KT, Mello SS, Attardi LD. Unravelling mechanisms of p53-mediated tumour suppression. Nat Rev Cancer. 2014;14:359–70. doi: 10.1038/nrc3711.PubMedCentralCrossRefPubMedGoogle Scholar
  8. 8.
    Lee YS, Oh J-HH, Yoon S, Kwon M-SS, Song C-WW, Kim K-HH, et al. Differential gene expression profiles of radioresistant non-small-cell lung cancer cell lines established by fractionated irradiation: tumor protein p53-inducible protein 3 confers sensitivity to ionizing radiation. Int J Radiat Oncol Biol Phys. 2010;77:858–66. doi: 10.1016/j.ijrobp.2009.12.076.CrossRefPubMedGoogle Scholar
  9. 9.
    Stuschke M, Sak A, Wurm R, Sinn B, Wolf G, Stüben G, et al. Radiation-induced apoptosis in human non-small-cell lung cancer cell lines is secondary to cell-cycle progression beyond the G2-phase checkpoint. Int J Radiat Biol. 2002;78:807–19. doi: 10.1080/0955300021014890.CrossRefPubMedGoogle Scholar
  10. 10.
    Andreo P, Burns DT, Hohlfeld S, Huq MS, Kanai T, Laitano F, et al. Absorbed dose determination in external beam radiotherapy: an international code of practice for dosimetry based on standards of absorbed dose to water. Vienna: IAEA; 2011.Google Scholar
  11. 11.
    Gibbons JP. Monitor unit calculations for external photon and electron beams. In: AAPM annual meeting refresher course, Salt Lake City; 2001. p. 1–10.Google Scholar
  12. 12.
    Franken NAP, Rodermond HM, Stap J, Haveman J, van Bree C. Clonogenic assay of cells in vitro. Nat Protoc. 2006;1:2315–9. doi: 10.1038/nprot.2006.339.CrossRefPubMedGoogle Scholar
  13. 13.
    Abrantes AM, Serra MES, Gonçalves AC, Rio J, Oliveiros B, Laranjo M, et al. Hypoxia-induced redox alterations and their correlation with 99mTc-MIBI and 99mTc-HL-91 uptake in colon cancer cells. Nucl Med Biol. 2010;37:125–32. doi: 10.1016/j.nucmedbio.2009.11.001.CrossRefPubMedGoogle Scholar
  14. 14.
    Mamede AC, Pires AS, Abrantes AM, Tavares SD, Gonçalves AC, Casalta-Lopes JE, et al. Cytotoxicity of ascorbic acid in a human colorectal adenocarcinoma cell line (WiDr): in vitro and in vivo studies. Nutr Cancer. 2012;64:1049–57. doi: 10.1080/01635581.2012.713539.CrossRefPubMedGoogle Scholar
  15. 15.
    Serra AC, RochaGonsalves AMDA, Laranjo M, Abrantes AM, Gonçalves AC, Sarmento-Ribeiro AB, et al. Synthesis of new 2-galactosylthiazolidine-4-carboxylic acid amides. Antitumor evaluation against melanoma and breast cancer cells. Eur J Med Chem. 2012;53:398–402. doi: 10.1016/j.ejmech.2012.04.003.CrossRefPubMedGoogle Scholar
  16. 16.
    Sales TLF. Efeitos da radiação ionizante em linhas celulares de cancro do pulmão de não pequenas células: estudos in vitro. Coimbra: University of Coimbra; 2014.Google Scholar
  17. 17.
    Gonçalves AC, Alves V, Silva T, Carvalho C, De Oliveira CR, Sarmento-Ribeiro AB. Oxidative stress mediates apoptotic effects of ascorbate and dehydroascorbate in human Myelodysplasia cells in vitro. Toxicol Vitr. 2013;27:1542–9. doi: 10.1016/j.tiv.2013.03.009.CrossRefGoogle Scholar
  18. 18.
    Almeida S, Sarmento-ribeiro AB, Januário C, Rego AC, Oliveira CR. Evidence of apoptosis and mitochondrial abnormalities in peripheral blood cells of Huntington’ s disease patients. Biochem Biophys Res Commun. 2008;374:599–603. doi: 10.1016/j.bbrc.2008.07.009.CrossRefPubMedGoogle Scholar
  19. 19.
    Santos K, Laranjo M, Abrantes AM, Brito AF, Gonçalves C, Sarmento Ribeiro AB, et al. Targeting triple-negative breast cancer cells with 6,7-bis(hydroxymethyl)-1H,3H-pyrrolo[1,2-c]thiazoles. Eur J Med Chem. 2014;79:273–81. doi: 10.1016/j.ejmech.2014.04.008.CrossRefPubMedGoogle Scholar
  20. 20.
    Han JY, Chung YJ, Park SW, Kim JS, Rhyu MG, Kim HK, et al. The relationship between cisplatin-induced apoptosis and p53, bcl-2 and bax expression in human lung cancer cells. Korean J Intern Med. 1999;14:42–52.PubMedCentralCrossRefPubMedGoogle Scholar
  21. 21.
    Lien J-C, Huang C-C, Lu T-J, Tseng C-H, Sung P-J, Lee H-Z, et al. Naphthoquinone derivative PPE8 induces endoplasmic reticulum stress in p53 null H1299 cells. Oxid Med Cell Longev. 2015;2015:453679. doi: 10.1155/2015/453679.PubMedCentralPubMedGoogle Scholar
  22. 22.
    Würtzen PA, Pedersen LO, Poulsen HS, Claesson MH. Specific killing of P53 mutated tumor cell lines by a cross-reactive human HLA-A2-restricted P53-specific CTL line. Int J Cancer. 2001;93:855–61.CrossRefPubMedGoogle Scholar
  23. 23.
    Balça-Silva J, Neves SS, Gonçalves AC, Abrantes AM, Casalta-Lopes J, Botelho MF, et al. Effect of miR-34b overexpression on the radiosensitivity of non-small cell lung cancer cell lines. Anticancer Res. 2012;32:1603–10.PubMedGoogle Scholar
  24. 24.
    Chen X, Liao C, Chu Q, Zhou G, Lin X, Li X, et al. Dissecting the molecular mechanism of ionizing radiation-induced tissue damage in the feather follicle. PLoS ONE. 2014;9:e89234. doi: 10.1371/journal.pone.0089234.PubMedCentralCrossRefPubMedGoogle Scholar
  25. 25.
    Valko M, Rhodes CJ, Moncol J, Izakovic M, Mazur M. Free radicals, metals and antioxidants in oxidative stress-induced cancer. Chem Biol Interact. 2006;160:1–40. doi: 10.1016/j.cbi.2005.12.009.CrossRefPubMedGoogle Scholar
  26. 26.
    Thannickal VJ, Fanburg BL. Reactive oxygen species in cell signaling. Am J Physiol Lung Cell Mol Physiol. 2000;279:L1005–28. doi: 10.1164/rccm.2206007.PubMedGoogle Scholar
  27. 27.
    Yamamori T, Yasui H, Yamazumi M, Wada Y, Nakamura Y, Nakamura H, et al. Ionizing radiation induces mitochondrial reactive oxygen species production accompanied by upregulation of mitochondrial electron transport chain function and mitochondrial content under control of the cell cycle checkpoint. Free Radic Biol Med. 2012;53:260–70. doi: 10.1016/j.freeradbiomed.2012.04.033.CrossRefPubMedGoogle Scholar
  28. 28.
    Montero J, Dutta C, van Bodegom D, Weinstock D, Letai A. p53 regulates a non-apoptotic death induced by ROS. Cell Death Differ. 2013;20:1465–74. doi: 10.1038/cdd.2013.52.PubMedCentralCrossRefPubMedGoogle Scholar
  29. 29.
    Lao-Sirieix P, Brais R, Lovat L, Coleman N, Fitzgerald RC. Cell cycle phase abnormalities do not account for disordered proliferation in Barrett’s carcinogenesis. Neoplasia. 2004;6:751–60. doi: 10.1593/neo.04280.PubMedCentralCrossRefPubMedGoogle Scholar
  30. 30.
    Gadhikar MA, Sciuto MR, Alves MVO, Pickering CR, Osman AA, Neskey DM, et al. Chk1/2 inhibition overcomes the cisplatin resistance of head and neck cancer cells secondary to the loss of functional p53. Mol Cancer Ther. 2013;12:1860–73. doi: 10.1158/1535-7163.MCT-13-0157.PubMedCentralCrossRefPubMedGoogle Scholar
  31. 31.
    Yang L, Zhou Y, Li Y, Zhou J, Wu Y, Cui Y, et al. Mutations of p53 and KRAS activate NF-κB to promote chemoresistance and tumorigenesis via dysregulation of cell cycle and suppression of apoptosis in lung cancer cells. Cancer Lett. 2014;. doi: 10.1016/j.canlet.2014.12.003.Google Scholar
  32. 32.
    Holbrook LAC, Butler RA, Cashon RE, Van Beneden RJ. Soft-shell clam (Mya arenaria) p53: a structural and functional comparison to human p53. Gene. 2009;433:81–7. doi: 10.1016/j.gene.2008.11.029.PubMedCentralCrossRefPubMedGoogle Scholar
  33. 33.
    She QB, Bode AM, Ma WY, Chen NY, Dong Z. Resveratrol-induced activation of p53 and apoptosis is mediated by extracellular-signal-regulated protein kinases and p38 kinase. Cancer Res. 2001;61:1604–10.PubMedGoogle Scholar
  34. 34.
    Wu GS. The functional interactions between the p53 and MAPK signaling pathways. Cancer Biol Ther. 2004;3:156–61.CrossRefPubMedGoogle Scholar
  35. 35.
    Henness S, Davey MW, Harvie RM, Davey RA. Fractionated irradiation of H69 small-cell lung cancer cells causes stable radiation and drug resistance with increased MRP1, MRP2, and topoisomerase IIalpha expression. Int J Radiat Oncol Biol Phys. 2002;54:895–902. doi: 10.1016/S0360-3016(02)03037-7.CrossRefPubMedGoogle Scholar
  36. 36.
    Frutuoso C, Silva M, Amaral N. Valor prognóstico das proteínas p53, C-erB-2 E Ki67 no carcinoma do ovário. Acta Med Port. 2001;14:277–83.PubMedGoogle Scholar
  37. 37.
    Sousa H, Santos AM, Pinto D, Medeiros R. Is there a biological plausability for p53 codon 72 polymorphism influence on cervical cancer development? Acta Med Port. 2011;24:127–34.PubMedGoogle Scholar
  38. 38.
    Fridman JS, Lowe SW. Control of apoptosis by p53. Oncogene. 2003;22:9030–40. doi: 10.1038/sj.onc.1207116.CrossRefPubMedGoogle Scholar
  39. 39.
    She Q, Bode AM, Ma W. Resveratrol-induced activation of p53 and apoptosis is mediated by extracellular-signal-regulated protein kinases and p38 kinase. 2001;1604–1610.Google Scholar
  40. 40.
    Duarte RLDM, Paschoal MEM. Marcadores moleculares no câncer de pulmão: papel prognóstico e sua relação com o tabagismo. J Bras Pneumol. 2006;32:56–65. doi: 10.1590/S1806-37132006000100012.CrossRefPubMedGoogle Scholar
  41. 41.
    Abrantes A. Hipóxia Tumoral - Metabonómica e Imagem, Estudo Experimental. Coimbra: University of Coimbra; 2013.Google Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Fernando Mendes
    • 1
    • 2
    • 3
  • Tiago Sales
    • 1
  • Cátia Domingues
    • 3
    • 4
  • Susann Schugk
    • 1
  • Ana Margarida Abrantes
    • 1
    • 3
  • Ana Cristina Gonçalves
    • 3
    • 4
  • Ricardo Teixo
    • 1
  • Rita Silva
    • 1
  • João Casalta-Lopes
    • 1
    • 5
  • Clara Rocha
    • 6
    • 7
  • Mafalda Laranjo
    • 1
  • Paulo César Simões
    • 5
  • Ana Bela Sarmento Ribeiro
    • 3
    • 4
    • 8
    • 9
  • Maria Filomena Botelho
    • 1
    • 3
  • Manuel Santos Rosa
    • 10
  1. 1.Biophysics Unit-CNC.IBILI, Faculty of MedicineUniversity of CoimbraCoimbraPortugal
  2. 2.Department Biomedical Laboratory Sciences, ESTESC-Coimbra Health SchoolPolytechnic Institute of CoimbraCoimbraPortugal
  3. 3.Faculty of Medicine, Center of Investigation in Environment, Genetics and Oncobiology (CIMAGO)University of CoimbraCoimbraPortugal
  4. 4.Faculty of Medicine, Applied Molecular Biology and Clinical University of HematologyUniversity of CoimbraCoimbraPortugal
  5. 5.Radiation Oncology DepartmentHospital and University Center of CoimbraCoimbraPortugal
  6. 6.Department Complementary Sciences, ESTESC-Coimbra Health SchoolPolytechnic Institute of CoimbraCoimbraPortugal
  7. 7.Institute for Systems Engineering and Computers at CoimbraCoimbraPortugal
  8. 8.Cento Hospitalar Universitário de CoimbraCoimbraPortugal
  9. 9.CNC.IBILI, Faculty of MedicineUniversity of CoimbraCoimbraPortugal
  10. 10.Faculty of Medicine, Immunology InstituteUniversity of CoimbraCoimbraPortugal

Personalised recommendations