高压纳秒脉冲电场对同源细胞的杀伤差异性研究

doi:10.19595/j.cnki.1000-6753.tces.231473

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Abstract

In terms of biological effects caused by the pulsed electric fields, the duration of the nanosecond pulse is less than the charging time constant of the cell membrane (~1μs), which has more profound effects on the intracellular organelles, known as the intracellular electron-manipulation. Based on this, the nanosecond pulsed electric fields (nsPEFs) could have unique bioelectromagnetic properties in the field of tumor ablation. Therefore, we studied the killing effects of homologous cells under the action of nsPEFs to investigate the difference between homologous cells to high-voltage nsPEFs.
The homologous cells, human immortal keratinocytes (HaCaT), human melanoma cells (A375), and human melanoma chemoresistant cells (A375/R), were chosen as research objects. The homemade pulse generator can output high-voltage nanosecond pulses with flexible and adjustable parameters. The selected pulse parameters were 5-15 kV/cm of field intensity, 300 ns of pulse width, 1 Hz of repetition frequency, and 80 of pulse number. Cell viability assay based on CCK-8 was used to study the survival rate of homologous cells after pulse treatment. The ablation effect of nanosecond pulses was further studied based on hydrogel-simulated tissue. The lethal threshold for a certain ablation area was calculated based on the ablation area. Furthermore, the electroporation response of the single cell to the nanosecond pulse is simulated by the finite element simulation model.
The survival rate of the three cells decreased with the increase of the electric field intensity of nsPEFs. The results of the cell viability assay showed that under the same parameters of nsPEFs, the survival rate of normal cells was the highest, followed by tumor cells, and tumor chemoresistant cells had the lowest survival rate. The results of the hydrogel simulated tissue ablation experiment showed that under the same pulse parameters, the ablation area of tumor chemoresistant cells was the largest, reaching 20.28±0.85 mm², while the ablation area of normal cells was the smallest, only 10.26±0.49 mm², which is significant difference. Meanwhile, the lethal threshold of normal cells and chemoresistant cells was calculated to be 7.15±0.65 kV/cm and 4.75±0.15 kV/cm, respectively, with a difference of approximately 2.4 kV/cm. These results indicated that there were significant differences in the sensitivity of normal cells, tumor cells, and tumor chemoresistant cells to the high-voltage nanosecond pulsed electric fields, and the nsPEFs had the least effect on normal cells and the strongest effect on tumor chemoresistant cells.
To analyze the potential reasons for the killing difference of cells by nsPEFs, the cell size and nucleus size of HaCaT, A375, and A375/R cells were measured and recorded by fluorescence microscopy. There was no significant difference in the cell size between HaCaT, A375, and A375/R, but there was a significant difference in the nucleus size. Based on the real cell geometry, the electroporation area and the pore number in the nuclear membrane were simulated. The results suggested that the difference in cell response to nsPEFs may be related to the nucleus size. Hence, tumor chemoresistant cells with the larger nucleus were more susceptible to nsPEFs, which indicated that high-voltage nanosecond pulsed electric fields have potential clinical application in the field of tumor ablation without affecting surrounding normal tissues.

Key words： Nanosecond pulsed electric fields homologous cells cell viability hydrogel scaffolds killing difference

PACS:	TM836
	Q64

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	Peng Wencheng
	Tang Xiao
	Liu Hongmei
	Dong Shoulong
	Yao Chenguo

Cite this article:

Peng Wencheng,Tang Xiao,Liu Hongmei等. Study on the Killing Difference of Homologous Cells by High-Voltage Nanosecond Pulsed Electric Fields[J]. Transactions of China Electrotechnical Society, 0, (): 9043-43.

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https://dgjsxb.ces-transaction.com/EN/10.19595/j.cnki.1000-6753.tces.231473 OR https://dgjsxb.ces-transaction.com/EN/Y0/V/I/9043

[1] Youlden D R, Khosrotehrani K, Green A C, et al.Diagnosis of an additional in situ melanoma does not influence survival for patients with a single invasive melanoma: A regis-try-based follow-up study[J]. Australasian Journal of Dermatology, 2016, 57(1): 57-60.
[2] Nornal M, Wright C Y.The epidemiology of cutaneous melanoma in the white and black African population groups in South Africa[M]. Brisbane (AU): Cutaneous Melanoma: Etiology and Therapy, 2017.
[3] 陈敏华,Nahum Goldberg S.肝癌射频消融-基础与临床[M].北京:人民卫生出版社,2009.
Chen Minhua, Nahum Goldberg S.Radiofrequency ablation of liver tumor - basic and clinical studies[M]. Beijing, China: People's Medical Publishing House, 2009.
[4] 安东均,郑晓燕,张成.358例肝肿瘤微波消融并发症的临床分析[J].肝胆外科杂志,2015,23(1):24-26.
An Dongjun, Zheng Xiaoyan, Zhang Cheng.A clinical study of complication arising from microwave ablation for 358 liver tu-mors[J]. Journal of Hepatobiliary Surgery, 2015, 23(1): 24-26.
[5] 张积仁. 氩氦刀冷冻消融治疗肿瘤[J].中国肿瘤,2007,16(5):335-337.
Zhang Jiren.Argon-helium cryoablation in the treatment for can-cer[J]. Bulletin of Chinese Cancer, 2007, 16(5): 335-337.
[6] 苏海兵,邹建中,王智彪.高强度聚焦超声肿瘤治疗技术原理[J].中华肝胆外科杂志,2011,17(3):271-272.
Su Haibing, Zou Jianzhong, Wang Zhibiao.Mechanism of high intensity focused ultrasound thermal ablation therapy[J]. Chinese Journal of Hepatobiliary Surgery, 2011, 17(3): 271-272.
[7] Hamid O, Robert C, Daud A, et al.Five-year survival outcomes for patients with advanced melanoma treated with pembrolizumab in KEYNOTE-001[J]. Annals of Oncology, 2019, 30(4): 582-588.
[8] Si L, Kong Y, Xu X, et al.Prevalence of BRAF V600E mutation in Chinese melanoma patients: large scale analysis of BRAF and NRAS mutations in a 432-case cohort[J]. European Journal of Cancer, 2012, 48(1): 94-100.
[9] Zaretsky J M, Garcia-diaz A, Shin D S, et al. Mutations associated with acquired resistance to PD-1 blockade in melanoma[J]. The New England Journal of Medicine, 2016, 375(9): 819-829.
[10] Shi W.Radiation therapy for melanoma[M]. Brisbane (AU): Cutaneous Melanoma: Etiology and Therapy, 2017.
[11] Dummer R, Ascierto P A, Gogas H, et al.Overall survival in patients with BRAF-mutant melanoma receiving encorafenib plus binimetinib versus vemurafenib or encorafenib (COLUMBUS): a multicentre, open-label, randomised, phase 3 trial[J]. Lancet Oncology, 2018, 19(10): 1315-1327.
[12] Sosman J A, Kim K B, Schuchter L M, et al.Survival in BRAF V600-mutant advanced melanoma treated with vemurafenib[J]. The New England Journal of Medicine, 2012, 366(8): 707-714.
[13] 齐忠慧,斯璐.恶性黑色素瘤治疗新进展[J].中国肿瘤临床,2019,46(5):213-217.
Qi Zhonghui, Si Lu.Advances in the treatment of malignant melanoma[J]. Chinese Journal of Clinical Oncology, 2019, 46(5): 213-217.
[14] Solyanik G I.Multifactorial nature of tumor drug resistance[J]. Experimental Oncology, 2010, 32(3): 181-185.
[15] Yu T, Luo L, Wang L.Ultrasound as a cancer chemotherapy sensitizer: the gap between laboratory and bedside[J]. Expert Opinion on Drug Delivery, 2016, 13(1): 37-47.
[16] Ren W, Beebe S J.An apoptosis targeted stimulus with nanosecond pulsed electric fields (nsPEFs) in E4 squamous cell carcinoma[J]. Apoptosis, 2011, 16(4): 382-393.
[17] Miklavcic D, Sersa G, Brecelj E, et al.Electrochemotherapy: technological advancements for efficient electroporation-based treatment of internal tumors[J]. Medical & Biological Engineering & Computing, 2012, 50(12): 1213-1225.
[18] Nuccitelli R, Berridge J C, Mallon Z, et al.Nanoelectro-ablation of murine tumors triggers a CD8-dependent inhibition of secondary tumor growth[J]. Plos One, 2015, 10(7): e0134364.
[19] 程显,陈硕,吕彦鹏,等.纳秒脉冲作用下核孔复合体影响细胞核膜电穿孔变化的仿真研究[J].电工技术学报,2021,36(18):3821-3828.
Chen Xian, Chen Shuo, Lv Yanpeng, et al.Simulation Study on the Effect of Nuclear Pore Complexes on Cell Electroporation Under Nanosecond Pulse[J]. Transactions of China Electrotechnical Society, 2021,36(18): 3821-3828.
[20] 李成祥,李新皓,柯强,等.纳/微秒复合脉冲提高细胞电融合效率的仿真与实验研究[J].电工技术学报,2022,37(6):1522-1532.
Li Chengxiang, Li Xinhao, Ke Qiang, et al.Simulation and Experimental Research on Improving the Efficiency of Cell Electrofusion by Nano-Microsecond Pulsed Electric Fields[J]. Transactions of China Electrotechnical Society, 2022,37(6): 1522-1532.
[21] Gianulis E C, Labib C, Saulis G, et al.Selective susceptibility to nanosecond pulsed electric field (nsPEF) across different human cell types[J]. Cellular and Molecular Life Sciences, 2017, 74(9): 1741-1754.
[22] 米彦,代璐健,刘权,等.靶向金纳米棒联合纳秒脉冲电场对体外A375黑色素瘤2D模拟组织的消融效果研究[J].电工技术学报,2021,36(18):3766-3775.
Mi Yan, Dai Lujian, Liu Quan, et al.Ablation Effects of A375 Melanoma Cells Treated by Targeted Gold Nanorods Combined with Nanosecond Pulsed Electric Fields[J]. Transactions of China Electrotechnical Society, 2021,36(18): 3766-3775.
[23] 米彦,吴晓,徐进,等.细胞膜力学性能对电穿孔影响的仿真研究[J].电工技术学报,2022,37(17):4326-4334+4400.
Mi Yan, Wu Xiao, Xu Jin, et al. Simulation Study on the Effect of Cell Membrane Mechanical Properties on Electroporation[J]. Transactions of China Electrotechnical Society, 2022,37(17): 4326-4334+4400.
[24] Gianulis E C, Labib C, Saulis G, et al.Selective susceptibility to nanosecond pulsed electric field (nsPEF) across different human cell types[J]. Cellular and Molecular Life Sciences, 2017, 74(9): 1741-1754.
[25] Ivey J W, Latouche E L, Sano M B, et al.Targeted cellular ablation based on the morphology of malignant cells[J]. Scientific Reports, 2015, 5: 17157.
[26] 吴晓,米彦,郑伟,等.脉冲电场对细胞膜电穿孔面积的影响研究[J].电工技术学报,2023,38(14):3779-3788.
Wu Xiao, Mi Yan, Zheng Wei, et al.Effect of Pulsed Electric Field on Electroporation Area of Cell Membrane[J]. Transactions of China Electrotechnical Society, 2023,38(14): 3779-3788.
[27] 姚陈果,宁郡怡,刘红梅,等.微/纳秒脉冲电场靶向不同尺寸肿瘤细胞内外膜电穿孔效应研究[J].电工技术学报,2020,35(1):115-124.
Yao Chenguo, Ning Junyi, Liu Hongmei, et al.Study of Electroporation Effect of Different Size Tumor Cells Targeted by Micro-Nanosecond Pulsed Electric Field[J]. Transactions of China Electrotechnical Society, 2020, 35(1): 115-124.
[28] 刘红梅,董守龙,宁郡怡,等.纳秒脉冲高频透膜效应优先杀伤化疗抗性肿瘤细胞的仿真与实验研究[J].电工技术学报,2019,34(22):4839-4848.
Liu Hongmei, Dong Shoulong, Ning Junyi, et al.Simulation and experimental study on preferential killing of chemoresistance tumor cells induced by the high-frequency permeation effect of nanosecond pulse field[J]. Transactions of China Electrotechnical Society, 2019, 34(22): 4839-4848.
[29] 王昌金,姚陈果,董守龙,等.基于Marx和LTD拓扑的全固态复合模式脉冲源的研制[J].电工技术学报,2018,33(13):3089-3097.
Wang Changjin, Yao Chenguo, Dong Shoulong, et al.The development of an all-solid-state mixed pulse generator based on marx and LTD topologies[J]. Transactions of China Electrotechnical Society, 2018, 33(13): 3089-3097.
[30] Yao C, Liu H, Zhao Y, et al.Analysis of dynamic processes in single-cell electroporation and their effects on parameter selection based on the finite-element model[J]. IEEE Transactions on Plasma Science, 2017, 45(5): 889-900.
[31] Wegner L H, Frey W, Schonwalder S.A critical evaluation of whole cell patch clamp studies on electroporation using the voltage sensitive dye ANNINE-6[J]. Bioelectrochemistry, 2013, 92(8): 42-46.
[32] Tolstykh G P, Beier H T, Roth C C, et al.Activation of intracellular phosphoinositide signaling after a single 600 nanosecond electric pulse[J]. Bioelectrochemistry, 2013, 94(12): 23-29.