Effect of Pulse Width and Intensity on Cell Death in Reversible Electroporation of Cancerous Cells

Year 2024, , 530 - 536, 26.09.2024

Mehmet Eşref Alkış , Yusuf Alan , Erhan Eser

https://doi.org/10.17798/bitlisfen.1375850

Abstract

Electroporation (EP) is the process of increasing the permeability of a biological cell or tissue by applying a short-term and sufficient external electric field. The utilization of proper pulse settings is required for EP-based treatments to be successful. Our aim in this study is to examine the effect of different electrical pulse widths and strength on EP efficiency. Human osteosarcoma cells (U20S) were used in the study. Eight-square-pulses with a frequency of 1Hz at 10µs, 1ms, 5ms, 10ms, and 20ms widths with low electric fields (20-500V/cm) were applied to U20S cells. 10-15 minutes after the applications, the cells were incubated in 96-well plates with 10 thousand cells in each well for 24 hours. Efficiency of pulses of different intensity and width was evaluated by MTT analysis method. The percent inhibition of U20S cancer cells elevated as the pulse width increased in almost all electric field values. The highest cell inhibition (%) occurred in pulses with an electric field of 500 V/cm and a width of 20ms (inhibition ratio: 76.25%). No inhibition was observed in the cells at 10µs, 1ms, 5ms, 10ms width pulses with 20 V/cm electric field and 10µs, 1ms width pulses with 50V/cm electric field. In conclusion, our findings show that the electric field intensity and pulse width used in electroporation play an important role in U20S cancer cell death. According to our results, it may be more appropriate to use high-voltage short-width pulses or low-voltage long-width pulses in reversible EP studies.

Keywords

Electroporation, Pulse Width, Pulse Strength, Osteosarcoma, Toxicity

References

• [1] Y. Lv, Z. Feng, S. Chen, X. Cheng, J. Zhang, and C. Yao, “A fundamental theoretical study on the different effect of electroporation on tumor blood vessels and normal blood vessels,” Bioelectrochemistry, vol. 144, no. 108010, p. 108010, 2022.
• [2] A. Groselj, M. Bosnjak, P. Strojan, M. Krzan, M. Cemazar, and G. Sersa, “Efficiency of electrochemotherapy with reduced bleomycin dose in the treatment of nonmelanoma head and neck skin cancer: Preliminary results,” Head Neck, vol. 40, no. 1, pp. 120–125, 2018.
• [3] P. Gupta et al., “Efficacy and safety of irreversible electroporation for malignant liver tumors: a systematic review and meta-analysis,” Eur. Radiol., vol. 31, no. 9, pp. 6511–6521, 2021.
• [4] L. G. Campana et al., “Electrochemotherapy of superficial tumors - Current status: Basic principles, operating procedures, shared indications, and emerging applications,” Semin. Oncol., vol. 46, no. 2, pp. 173–191, 2019.
• [5] A. Zupanic, B. Kos, and D. Miklavcic, “Treatment planning of electroporation-based medical interventions: electrochemotherapy, gene electrotransfer and irreversible electroporation,” Phys. Med. Biol., vol. 57, no. 17, pp. 5425–5440, 2012.
• [6] M. E. Alkış, K. Buldurun, N. Turan, Y. Alan, Ü. K. Yılmaz, and A. Mantarcı, “Synthesis, characterization, antiproliferative of pyrimidine based ligand and its Ni(II) and Pd(II) complexes and effectiveness of electroporation,” J. Biomol. Struct. Dyn., vol. 40, no. 9, pp. 4073–4083, 2022.
• [7] V. Novickij et al., “Electrochemotherapy using doxorubicin and nanosecond electric field pulses: A pilot in vivo study,” Molecules, vol. 25, no. 20, p. 4601, 2020.
• [8] O. Kozak, S. Hać, J. Pieńkowska, and M. Studniarek, “Benefitial role of electrochemotherapy in locally advanced pancreatic cancer - radiological perspective,” Pol. J. Radiol., vol. 87, no. 1, pp. e30–e42, 2022.
• [9] G. Sersa, D. Miklavcic, M. Cemazar, Z. Rudolf, G. Pucihar, and M. Snoj, “Electrochemotherapy in treatment of tumours,” Eur. J. Surg. Oncol., vol. 34, no. 2, pp. 232–240, 2008.
• [10] Z. Shankayi, S. M. P. Firoozabadi, and Z. S. Hassan, “Optimization of electric pulse amplitude and frequency in vitro for low voltage and high frequency electrochemotherapy,” J. Membr. Biol., vol. 247, no. 2, pp. 147–154, 2014.
• [11] G. Pucihar, J. Krmelj, M. Reberšek, T. B. Napotnik, and D. Miklavčič, “Equivalent pulse parameters for electroporation,” IEEE Trans. Biomed. Eng., vol. 58, no. 11, pp. 3279–3288, 2011.
• [12] J. C. Weaver, K. C. Smith, A. T. Esser, R. S. Son, and T. R. Gowrishankar, “A brief overview of electroporation pulse strength-duration space: a region where additional intracellular effects are expected,” Bioelectrochemistry, vol. 87, pp. 236–243, 2012.
• [13] M. E. Alkış, Ü. Keleştemür, Y. Alan, N. Turan, and K. Buldurun, “Cobalt and ruthenium complexes with pyrimidine based schiff base: Synthesis, characterization, anticancer activities and electrochemotherapy efficiency,” J. Mol. Struct., vol. 1226, no. 129402, p. 129402, 2021.
• [14] M. E. Alkış, N. Turan, Y. Alan, S. Irtegun Kandemir, and K. Buldurun, “Effects of electroporation on anticancer activity of 5-FU and newly synthesized zinc(II) complex in chemotherapy-resistance human brain tumor cells,” Med. Oncol., vol. 38, no. 11, p. 129, 2021.
• [15] V. Novickij et al., “Effects of time delay between unipolar pulses in high frequency nano-electrochemotherapy,” IEEE Trans. Biomed. Eng., vol. 69, no. 5, pp. 1726–1732, 2022.
• [16] M. A. Esmekaya, H. Kayhan, M. Yagci, A. Coskun, and A. G. Canseven, “Effects of electroporation on tamoxifen delivery in estrogen receptor positive (ER+) human breast carcinoma cells,” Cell Biochem. Biophys., vol. 75, no. 1, pp. 103–109, 2017.
• [17] M. E. Alkis, K. Buldurun, Y. Alan, N. Turan, and A. Altun, “Electroporation enhances the anticancer effects of novel Cu(II) and Fe(II) complexes in chemotherapy-resistant glioblastoma cancer cells,” Chem. Biodivers., vol. 20, no. 2, p. e202200710, 2023.
• [18] M. Cemazar, I. Hreljac, G. Sersa, and M. Filipic, “Construction of EGFP expressing HepG2 cell line using electroporation,” in IFMBE Proceedings, Berlin, Heidelberg: Springer Berlin Heidelberg, 2009, pp. 128–131.
• [19] C. H. Lu, S. H. Lin, C. H. Hsieh, W. T. Chen, and C. Y. Chao, “Enhanced anticancer effects of low-dose curcumin with non-invasive pulsed electric field on PANC-1 cells,” Onco. Targets. Ther., vol. 11, pp. 4723–4732, 2018.
• [20] Y. Q. Chen, T. Q. Yang, B. Zhou, M. X. Yang, H. J. Feng, and Y. L. Wang, “HOXA5 overexpression promotes osteosarcoma cell apoptosis through the p53 and p38α MAPK pathway,” Gene, vol. 689, pp. 18-23, 2019.
• [21] Y. N. Hsu, H. W. Shyu, T. W. Hu, J. P. Yeh, Y. W. Lin, L. Y. Lee, and S. J. Su, “Anti-proliferative activity of biochanin A in human osteosarcoma cells via mitochondrial-involved apoptosis,” Food Chem. Toxicol., vol. 112, pp. 194-204, 2018.
• [22] T. Wang, X. Gong, R. Jiang, H. Li, W. Du, and G. Kuang, “Ferulic acid inhibits proliferation and promotes apoptosis via blockage of PI3K/Akt pathway in osteosarcoma cell,” Am. J. Transl. Res., vol. 8, no. 2, pp. 968-980, 2016.
• [23] B. Bute and M. E. Alkis, “Anticancer activity of methotrexate in electrochemotherapy and electrochemotherapy plus ionizing radiation treatments in human breast cancer cells,” Med. Oncol., vol. 40, no. 1, p. 28, 2022.
• [24] T. Polajžer, J. Dermol-Černe, M. Reberšek, R. O’Connor, and D. Miklavčič, “Cancellation effect is present in high-frequency reversible and irreversible electroporation,” Bioelectrochemistry, vol. 132, no. 107442, p. 107442, 2020.
• [25] D. E. Chafai, A. Mehle, A. Tilmatine, B. Maouche, and D. Miklavčič, “Assessment of the electrochemical effects of pulsed electric fields in a biological cell suspension,” Bioelectrochemistry, vol. 106, no. Pt B, pp. 249–257, 2015.
• [26] M. E. Alkış, “Investigation of treatment potential of O-vanillin containing Schiff base ligand and pd(II) complex in glioblastoma multiforme cells and efficiency of electroporation,” Van Med. J., vol. 29, no. 1, pp. 69–75, 2022.
• [27] A. Bicek, I. Turel, M. Kanduser, and D. Miklavcic, “Combined therapy of the antimetastatic compound NAMI-A and electroporation on B16F1 tumour cells in vitro,” Bioelectrochemistry, vol. 71, no. 2, pp. 113–117, 2007.
• [28] B. Mali, T. Jarm, M. Snoj, G. Sersa, and D. Miklavcic, “Antitumor effectiveness of electrochemotherapy: a systematic review and meta-analysis,” Eur. J. Surg. Oncol., vol. 39, no. 1, pp. 4–16, 2013.
• [29] T. Jarm, M. Cemazar, D. Miklavcic, and G. Sersa, “Antivascular effects of electrochemotherapy: implications in treatment of bleeding metastases,” Expert Rev. Anticancer Ther., vol. 10, no. 5, pp. 729–746, 2010.
• [30] J. Saczko et al., “Combination of therapy with 5-fluorouracil and cisplatin with electroporation in human ovarian carcinoma model in vitro,” Biomed. Pharmacother., vol. 68, no. 5, pp. 573–580, 2014.
• [31] V. De Giorgi, F. Scarfì, E. Saqer, A. Gori, G. M. Tomassini, and P. Covarelli, “The use of cisplatin electrochemotherapy in nonmelanoma skin cancers: A single-center study,” Dermatol. Ther., vol. 33, no. 4, p. e13547, 2020.
• [32] B. Agerholm-Larsen et al., “Preclinical validation of electrochemotherapy as an effective treatment for brain tumors,” Cancer Res., vol. 71, no. 11, pp. 3753–3762, 2011.
• [33] R. Fusco, E. Di Bernardo, V. D’Alessio, S. Salati, and M. Cadossi, “Reduction of muscle contraction and pain in electroporation-based treatments: An overview,” World J. Clin. Oncol., vol. 12, no. 5, pp. 367–381, 2021.
• [34] V. Stankevic et al., “Compact square-wave pulse electroporator with controlled electroporation efficiency and cell viability,” Symmetry (Basel), vol. 12, no. 3, p. 412, 2020.
• [35] M. Y. Sulaeman and R. Widita, “The effect of electric field intensity, pulse width, and pulse rise time on conventional and supra electroporation,” The 5th International Conference on Mathematics and Natural Sciences, 2–3 November 2014 Bandung, Indonesia. AIP Conf. Proc. 1677, 100008, 2015.
• [36] I. Hyder, S. Eghbalsaied, and W. A. Kues, “Systematic optimization of square-wave electroporation conditions for bovine primary fibroblasts,” BMC Mol. Cell Biol., vol. 21, no. 1, p. 9, 2020.
• [37] M. P. Rols and J. Teissié, “Electropermeabilization of mammalian cells to macromolecules: control by pulse duration,” Biophys. J., vol. 75, no. 3, pp. 1415–1423, 1998.