Folia Histochemica et Cytobiologica

open access

Vol 57, No 4 (2019)
Original paper
Submitted: 2019-04-24
Accepted: 2019-11-12
Published online: 2019-11-20
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In vitro evaluation of electroporated gold nanoparticles and extremely-low frequency electromagnetic field anticancer activity against Hep-2 laryngeal cancer cells

Mohammed A. Alshehri1, Piotr M. Wierzbicki2, Hassan F. Kaboo3, Mohamed S.M. Nasr3, Mohamed E. Amer4, Tamer M.M. Abuamara3, Doaa A. Badr5, Kamel A. Saleh1, Ahmed E. Fazary5, Aly F. Mohamed6
DOI: 10.5603/FHC.a2019.0018
·
Pubmed: 31746453
·
Folia Histochem Cytobiol 2019;57(4):159-167.
Affiliations
  1. Department of Biology, Science Collage, King Khalid University, Abha, Saudi Arabia
  2. Department of Histology, Medical University of Gdansk, Gdansk, Poland
  3. Histology Department, Faculty of Medicine, Al-Azhar University, Cairo, Egypt
  4. Histology Department, Faculty of Medicine, Al-Azhar University, Damietta, Egypt
  5. Applied Research Department, Research & Development Sector, Egyptian Organization for Biological Products and Vaccines (VACSERA Holding Company), Giza, Egypt
  6. The International Center for Advanced Researches (ICTAR-Egypt), Cairo, Egypt

open access

Vol 57, No 4 (2019)
ORIGINAL PAPERS
Submitted: 2019-04-24
Accepted: 2019-11-12
Published online: 2019-11-20

Abstract

Introduction. The extremely-low frequency electromagnetic field (ELFEMF) has been proposed for use in cancer therapy since it was found that magnetic waves interfere with many biological processes. Gold nanoparticles (Au-NPs) have been widely used for drug delivery during cancer in vitro studies due to their low cytotoxity and high biocompatibility. The electroporation of cancer cells in a presence of Au-NPs (EP Au-NPs) can induce cell apoptosis, alterations of cell cycle profile and morphological changes. The impact of ELFEMF and EP Au-NPs on morphology, cell cycle and activation of apoptosis-associated genes on Hep-2 laryngeal cancer cell line has not been studied yet.


Materials and methods. ELFEMF on Hep-2 cells were carried out using four different conditions: 25/50 mT at 15/30 min, while Au-NPs were used as direct contact (DC) or with electroporation (EP, 10 pulses at 200V, equal time intervals of 4 sec). MTT assay was used to check the toxicity of DC Au-NPs. Expression of CASP3, P53, BAX and BCL2 genes was quantified using qPCR. Cell cycle was analyzed by flow cytometry. Hematoxylin and eosin (HE) staining was used to observe cell morphology.


Results. Calculated IC50 of DC Au-NPs 24.36 μM (4.79 μg/ml) and such concentration was used for further DC and EP AuNPs experiments. The up-regulation of pro-apoptotic genes (CASP3, P53, BAX) and decreased expression of BCL2, respectively, was observed for all analyzed conditions with the highest differences for EP AuNPs and ELFEMF 50 mT/30 min in comparison to control cells. The highest content of cells arrested in G2/M phase was observed in ELFEMF-treated cells for 30 min both at 25 or 50 mT, while the cells treated with EP AuNPs or ELFEMF 50 mT/15 min showed highest ratios of apoptotic cells. HE staining of electroporated cells and cells exposed to ELFEMF’s low and higher frequencies for different times showed nuclear pleomorphic cells. Numerous apoptotic bodies were observed in the irregular cell membrane of neoplastic and necrotic cells with mixed euchromatin and heterochromatin.


Conclusions. Our observations indicate that treatment of Hep-2 laryngeal cancer cells with ELFEMF for 30 min at 25–50 mT and EP Au-NPs can cause cell damage inducing apoptosis and cell cycle arrest.

Abstract

Introduction. The extremely-low frequency electromagnetic field (ELFEMF) has been proposed for use in cancer therapy since it was found that magnetic waves interfere with many biological processes. Gold nanoparticles (Au-NPs) have been widely used for drug delivery during cancer in vitro studies due to their low cytotoxity and high biocompatibility. The electroporation of cancer cells in a presence of Au-NPs (EP Au-NPs) can induce cell apoptosis, alterations of cell cycle profile and morphological changes. The impact of ELFEMF and EP Au-NPs on morphology, cell cycle and activation of apoptosis-associated genes on Hep-2 laryngeal cancer cell line has not been studied yet.


Materials and methods. ELFEMF on Hep-2 cells were carried out using four different conditions: 25/50 mT at 15/30 min, while Au-NPs were used as direct contact (DC) or with electroporation (EP, 10 pulses at 200V, equal time intervals of 4 sec). MTT assay was used to check the toxicity of DC Au-NPs. Expression of CASP3, P53, BAX and BCL2 genes was quantified using qPCR. Cell cycle was analyzed by flow cytometry. Hematoxylin and eosin (HE) staining was used to observe cell morphology.


Results. Calculated IC50 of DC Au-NPs 24.36 μM (4.79 μg/ml) and such concentration was used for further DC and EP AuNPs experiments. The up-regulation of pro-apoptotic genes (CASP3, P53, BAX) and decreased expression of BCL2, respectively, was observed for all analyzed conditions with the highest differences for EP AuNPs and ELFEMF 50 mT/30 min in comparison to control cells. The highest content of cells arrested in G2/M phase was observed in ELFEMF-treated cells for 30 min both at 25 or 50 mT, while the cells treated with EP AuNPs or ELFEMF 50 mT/15 min showed highest ratios of apoptotic cells. HE staining of electroporated cells and cells exposed to ELFEMF’s low and higher frequencies for different times showed nuclear pleomorphic cells. Numerous apoptotic bodies were observed in the irregular cell membrane of neoplastic and necrotic cells with mixed euchromatin and heterochromatin.


Conclusions. Our observations indicate that treatment of Hep-2 laryngeal cancer cells with ELFEMF for 30 min at 25–50 mT and EP Au-NPs can cause cell damage inducing apoptosis and cell cycle arrest.

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Keywords

gold nanoparticles; electroporation; extremely-low frequency electromagnetic field; Hep-2 cells; apoptosis; qPCR

About this article
Title

In vitro evaluation of electroporated gold nanoparticles and extremely-low frequency electromagnetic field anticancer activity against Hep-2 laryngeal cancer cells

Journal

Folia Histochemica et Cytobiologica

Issue

Vol 57, No 4 (2019)

Article type

Original paper

Pages

159-167

Published online

2019-11-20

DOI

10.5603/FHC.a2019.0018

Pubmed

31746453

Bibliographic record

Folia Histochem Cytobiol 2019;57(4):159-167.

Keywords

gold nanoparticles
electroporation
extremely-low frequency electromagnetic field
Hep-2 cells
apoptosis
qPCR

Authors

Mohammed A. Alshehri
Piotr M. Wierzbicki
Hassan F. Kaboo
Mohamed S.M. Nasr
Mohamed E. Amer
Tamer M.M. Abuamara
Doaa A. Badr
Kamel A. Saleh
Ahmed E. Fazary
Aly F. Mohamed

References (36)
  1. Girgert R, Schimming H, Körner W, et al. Induction of tamoxifen resistance in breast cancer cells by ELF electromagnetic fields. Biochem Biophys Res Commun. 2005; 336(4): 1144–1149.
  2. Chen G, Upham BL, Sun W, et al. Effect of electromagnetic field exposure on chemically induced differentiation of friend erythroleukemia cells. Environ Health Perspect. 2000; 108(10): 967–972.
  3. Barbault A, Costa FP, Bottger B, et al. Amplitude-modulated electromagnetic fields for the treatment of cancer: discovery of tumor-specific frequencies and assessment of a novel therapeutic approach. J Exp Clin Cancer Res. 2009; 28: 51.
  4. Galloni P, Marino C. Effects of 50 Hz magnetic field exposure on tumor experimental models. Bioelectromagnetics. 2000; 21(8): 608–614, doi: 10.1002/1521-186x(200012)21:8<608::aid-bem7>3.0.co;2-z.
  5. Yasui M, Kikuchi T, Ogawa M, et al. Carcinogenicity test of 50 Hz sinusoidal magnetic fields in rats. Bioelectromagnetics. 1997; 18(8): 531–540, doi: 10.1002/(sici)1521-186x(1997)18:8<531::aid-bem1>3.0.co;2-3.
  6. Williams CD, Markov MS, Hardman WE, et al. Therapeutic electromagnetic field effects on angiogenesis and tumor growth. Anticancer Res. 2001; 21(6A): 3887–3891.
  7. Cameron IL, Sun LZ, Short N, et al. Therapeutic Electromagnetic Field (TEMF) and gamma irradiation on human breast cancer xenograft growth, angiogenesis and metastasis. Cancer Cell Int. 2005; 5: 23.
  8. Rannug A, Holmberg B, Ekström T, et al. Rat liver foci study on coexposure with 50 Hz magnetic fields and known carcinogens. Bioelectromagnetics. 1993; 14(1): 17–27.
  9. Sriram MI, Kanth SB, Kalishwaralal K, et al. Antitumor activity of silver nanoparticles in Dalton's lymphoma ascites tumor model. Int J Nanomedicine. 2010; 5: 753–762.
  10. Repacholi MH, Greenebaum B. Interaction of static and extremely low frequency electric and magnetic fields with living systems: health effects and research needs. Bioelectromagnetics. 1999; 20(3): 133–160, doi: 10.1002/(sici)1521-186x(1999)20:3<133::aid-bem1>3.0.co;2-o.
  11. Zhang Y, Xu D, Li W, et al. Effect of Size, Shape, and Surface Modification on Cytotoxicity of Gold Nanoparticles to Human HEp-2 and Canine MDCK Cells. Journal of Nanomaterials. 2012; 2012: 1–7.
  12. Rajeshkumar S. Anticancer activity of eco-friendly gold nanoparticles against lung and liver cancer cells. J Genet Eng Biotechnol. 2016; 14(1): 195–202.
  13. Lim HK, Asharani PV, Hande MP, et al. Cytotoxicity and genotoxicity of silver nanoparticles in human cells. ACS Nano. 2009; 3(2): 279–290.
  14. Artacho-Cordón F, Salinas-Asensio Md, Calvente I, et al. Could radiotherapy effectiveness be enhanced by electromagnetic field treatment? Int J Mol Sci. 2013; 14(7): 14974–14995.
  15. Kah JC, Wong KYi, Neoh KG, et al. Critical parameters in the pegylation of gold nanoshells for biomedical applications: an in vitro macrophage study. J Drug Target. 2009; 17(3): 181–193.
  16. Bailon P, Won CY. PEG-modified biopharmaceuticals. Expert Opin Drug Deliv. 2009; 6(1): 1–16.
  17. Behzadi S, Serpooshan V, Tao W, et al. Cellular uptake of nanoparticles: journey inside the cell. Chem Soc Rev. 2017; 46(14): 4218–4244.
  18. Brisdelli F, Bennato F, Bozzi A, et al. ELF-MF attenuates quercetin-induced apoptosis in K562 cells through modulating the expression of Bcl-2 family proteins. Mol Cell Biochem. 2014; 397(1-2): 33–43.
  19. Frederix F, Friedt JM, Choi KH, et al. Biosensing based on light absorption of nanoscaled gold and silver particles. Anal Chem. 2003; 75(24): 6894–6900.
  20. Hainfeld JF, Slatkin DN, Smilowitz HM. The use of gold nanoparticles to enhance radiotherapy in mice. Phys Med Biol. 2004; 49(18): N309–N315.
  21. Paciotti GF, Myer L, Weinreich D, et al. Colloidal gold: a novel nanoparticle vector for tumor directed drug delivery. Drug Deliv. 2004; 11(3): 169–183.
  22. Podsiadlo P, Sinani VA, Bahng JH, et al. Gold nanoparticles enhance the anti-leukemia action of a 6-mercaptopurine chemotherapeutic agent. Langmuir. 2008; 24(2): 568–574.
  23. Niidome T, Yamagata M, Okamoto Y, et al. PEG-modified gold nanorods with a stealth character for in vivo applications. J Control Release. 2006; 114(3): 343–347.
  24. Huang YC, Yang YC, Yang KC, et al. Pegylated gold nanoparticles induce apoptosis in human chronic myeloid leukemia cells. Biomed Res Int. 2014; 2014: 182353.
  25. Liu CJ, Wang CH, Chen ST, et al. Enhancement of cell radiation sensitivity by pegylated gold nanoparticles. Phys Med Biol. 2010; 55(4): 931–945.
  26. Chen HH, Chien CC, Petibois C, et al. Quantitative analysis of nanoparticle internalization in mammalian cells by high resolution X-ray microscopy. J Nanobiotechnology. 2011; 9: 14.
  27. Fruijtier-Pölloth C. Safety assessment on polyethylene glycols (PEGs) and their derivatives as used in cosmetic products. Toxicology. 2005; 214(1-2): 1–38.
  28. Liu CJ, Yang TY, Wang CH, et al. Enhanced photocatalysis, colloidal stability and cytotoxicity of synchrotron X-ray synthesized Au/TiO2 nanoparticles. Materials Chemistry and Physics. 2009; 117(1): 74–79.
  29. Liu CJ, Wang CH, Chien CC, et al. Enhanced x-ray irradiation-induced cancer cell damage by gold nanoparticles treated by a new synthesis method of polyethylene glycol modification. Nanotechnology. 2008; 19(29): 295104.
  30. Wang CH, Liu CJ, Wang CL, et al. Optimizing the size and surface properties of polyethylene glycol (PEG)–gold nanoparticles by intense x-ray irradiation. Journal of Physics D: Applied Physics. 2008; 41(19): 195301.
  31. Wang CH, Liu CJ, Chien CC, et al. X-ray synthesized PEGylated (polyethylene glycol coated) gold nanoparticles in mice strongly accumulate in tumors. Materials Chemistry and Physics. 2011; 126(1-2): 352–356.
  32. Nogueira-Pedro A, Cesário TA, Dias CC, et al. Hydrogen peroxide (H2O2) induces leukemic but not normal hematopoietic cell death in a dose-dependent manner. Cancer Cell Int. 2013; 13(1): 123.
  33. Liu Y, Liu Wb, Liu Kj, et al. Effect of 50 Hz Extremely Low-Frequency Electromagnetic Fields on the DNA Methylation and DNA Methyltransferases in Mouse Spermatocyte-Derived Cell Line GC-2. Biomed Res Int. 2015; 2015: 237183.
  34. Arellanes-Robledo J, Márquez-Rosado L, Pérez-Carreón JI, et al. Celecoxib induces regression of putative preneoplastic lesions in rat liver. Anticancer Res. 2006; 26(2A): 1271–1280.
  35. Carrasco-Legleu CE, Márquez-Rosado L, Fattel-Fazenda S, et al. Chemoprotective effect of caffeic acid phenethyl ester on promotion in a medium-term rat hepatocarcinogenesis assay. Int J Cancer. 2004; 108(4): 488–492.
  36. Márquez-Rosado L, Trejo-Solís MC, García-Cuéllar CM, et al. Celecoxib, a cyclooxygenase-2 inhibitor, prevents induction of liver preneoplastic lesions in rats. J Hepatol. 2005; 43(4): 653–660.

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