Applied Sciences

Research

11 pages, 2257 KiB

Open AccessArticle

Dielectrophoretic Manipulation of Cell Transfection Efficiency during Electroporation Using a Center Needle Electrode

by Eivina Radzevičiūtė, Arūnas Murauskas, Paulius Ruzgys, Saulius Šatkauskas, Irutė Girkontaitė, Jurij Novickij and Vitalij Novickij

Appl. Sci. 2021, 11(15), 7015; https://doi.org/10.3390/app11157015 - 29 Jul 2021

Cited by 2 | Viewed by 1764

Abstract

Long duration electric pulses are frequently used to facilitate DNA electrotransfer into cells and tissues, while electroporation pulses can be combined with electrophoresis to maximize the transfection efficiency. In this work, we present the dielectrophoresis (DEP)-assisted methodology for electrotransfer of plasmid DNA (3.5 [...] Read more.

Long duration electric pulses are frequently used to facilitate DNA electrotransfer into cells and tissues, while electroporation pulses can be combined with electrophoresis to maximize the transfection efficiency. In this work, we present the dielectrophoresis (DEP)-assisted methodology for electrotransfer of plasmid DNA (3.5 kbp pmaxGFP) into mammalian cells (CHO-K1). A prototype of an electroporation cuvette with center needle electrode for DEP-assisted transfection is presented resulting in a 1.4-fold of transfection efficiency increase compared to the electroporation-only procedure (1.4 kV/cm × 100 µs × 8). The efficiency of transfection has been compared between three DEP frequencies of 1, 100, and 1 MHz. Lastly, the effects of exposure time (1, 3, and 5 min) during the DEP application step have been determined. It is concluded that the proposed methodology and exposure setup allow a significant improvement of transfection efficiency and could be used as an alternative to the currently popular electrotransfection techniques. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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10 pages, 2581 KiB

Open AccessCommunication

Induction of Apoptosis of Cancer Cells Using the Cisplatin Delivery Based Electrospray (CDES) System

by Myung Chul Lee, Shambhavi Pandey, Jae Woon Lim, Sangbae Park, Jae Eun Kim, Hyunmok Son, Jinsub Han, Hoon Seonwoo, Pankaj Garg and Jong Hoon Chung

Appl. Sci. 2021, 11(7), 3203; https://doi.org/10.3390/app11073203 - 02 Apr 2021

Cited by 2 | Viewed by 1756

Abstract

Cisplatin, a representative anticancer drug used to treat cancer, has many adverse effects. In particular, it causes significant damage to the kidneys. Thus, many researchers have studied the delivery of drugs, such as cisplatin, to cancer areas using targeted drug-delivery systems. Here, we [...] Read more.

Cisplatin, a representative anticancer drug used to treat cancer, has many adverse effects. In particular, it causes significant damage to the kidneys. Thus, many researchers have studied the delivery of drugs, such as cisplatin, to cancer areas using targeted drug-delivery systems. Here, we propose a new way to treat cancer by delivering anticancer drugs directly to the tumor site using the electrospray (ES) technique. We determined the optimal conditions for ES to promote the introduction of cisplatin into cancer cells. In our results, the group with cisplatin delivery-based electrospray (CDES) at 6 kV had an apoptosis of cancer cells approximately two times more than the group treated with cisplatin alone. We also confirmed that ES alone did not affect the survival of cells up to a voltage of 6 kV, but increased the permeability of the cell plasma membrane instantaneously, facilitating the influx of the drug. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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12 pages, 8893 KiB

Open AccessArticle

Antimicrobial Activity of L-Lysine and Poly-L-Lysine with Pulsed Electric Fields

by Jurgita Švedienė, Vitalij Novickij, Rokas Žalnėravičius, Vita Raudonienė, Svetlana Markovskaja, Jurij Novickij and Algimantas Paškevičius

Appl. Sci. 2021, 11(6), 2708; https://doi.org/10.3390/app11062708 - 17 Mar 2021

Cited by 6 | Viewed by 2352

Abstract

For the first time, the possibility to use L-lysine (Lys) and poly-L-lysine (PLL) as additives with pulsed electric fields (PEF) for antimicrobial treatment is reported. The antimicrobial efficacy of Lys and PLL for Escherichia coli, Staphylococcus aureus, Trichophyton rubrum and Candida [...] Read more.

For the first time, the possibility to use L-lysine (Lys) and poly-L-lysine (PLL) as additives with pulsed electric fields (PEF) for antimicrobial treatment is reported. The antimicrobial efficacy of Lys and PLL for Escherichia coli, Staphylococcus aureus, Trichophyton rubrum and Candida albicans was determined. Inactivation of microorganisms was also studied by combining Lys and PLL with PEF of 15 and 30 kV/cm. For PEF treatment, pulses of 0.5, 1, 10 or 100 μs were applied in a sequence of 10 to 5000 at 1 kHz frequency. The obtained results showed that 100 μs pulses were the most effective in combination with Lys and PLL for all microorganisms. Equivalent energy PEF bursts with a shorter duration of the pulse were less effective independently on PEF amplitude. Additionally, various treatment susceptibility patterns of microorganisms were determined and reported. In this study, the Gram-negative E. coli was the most treatment-resistant microorganism. Nevertheless, inactivation rates exceeding 2 log viability reduction were achieved for all analyzed yeast, fungi, and bacteria. This methodology could be used for drug-resistant microorganism’s new treatment development. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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10 pages, 2057 KiB

Open AccessArticle

Effects of Pulsed Electric Fields on Yeast with Prions and the Structure of Amyloid Fibrils

by Justina Jurgelevičiūtė, Nedas Bičkovas, Andrius Sakalauskas, Vitalij Novickij, Vytautas Smirnovas and Eglė Lastauskienė

Appl. Sci. 2021, 11(6), 2684; https://doi.org/10.3390/app11062684 - 17 Mar 2021

Cited by 3 | Viewed by 1878

Abstract

Prions are misfolded, self-replicating, and transmissible proteins capable of causing different conditions that affect the brain and nervous system in humans and animals. Yeasts are the perfect model to study prion formation, dissemination, and the structure of protein aggregates. Yeast prions are related [...] Read more.

Prions are misfolded, self-replicating, and transmissible proteins capable of causing different conditions that affect the brain and nervous system in humans and animals. Yeasts are the perfect model to study prion formation, dissemination, and the structure of protein aggregates. Yeast prions are related to stress resistance, cell fitness, and viability. Applying a pulsed electric field (PEF) as a factor capable of disintegrating the amyloid aggregates arises from the fact that the amyloid aggregates form via noncovalent bonds and stabilize via electrostatic interactions. In this research, we applied 2–26 kV/cm PEF delivered in sequences of 5 pulses of 1 ms duration to the Saccharomyces cerevisiae cell without prions and containing strong and weak variants of the [PSI+] prion (prion form of Sup35 translation termination factor). We determined that prions significantly increase cell survivability and resistance to PEF treatment. The application of PEF to the purified Sup35NM fibrils showed that the electric field causes significant reductions in the length of fibrils and the full disintegration of fibrils to Sup35 oligomers can be achieved in higher fields. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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11 pages, 2304 KiB

Open AccessArticle

Investigation of Plasmid DNA Delivery and Cell Viability Dynamics for Optimal Cell Electrotransfection In Vitro

by Sonam Chopra, Paulius Ruzgys, Martynas Maciulevičius, Milda Jakutavičiūtė and Saulius Šatkauskas

Appl. Sci. 2020, 10(17), 6070; https://doi.org/10.3390/app10176070 - 02 Sep 2020

Cited by 15 | Viewed by 3754

Abstract

Electroporation is an effective method for delivering plasmid DNA molecules into cells. The efficiency of gene electrotransfer depends on several factors. To achieve high transfection efficiency while maintaining cell viability is a tedious task in electroporation. Here, we present a combined study in [...] Read more.

Electroporation is an effective method for delivering plasmid DNA molecules into cells. The efficiency of gene electrotransfer depends on several factors. To achieve high transfection efficiency while maintaining cell viability is a tedious task in electroporation. Here, we present a combined study in which the dynamics of both evaluation types of transfection efficiency and the cell viability were evaluated in dependence of plasmid concentration as well as at the different number of high voltage (HV) electric pulses. The results of this study reveal a quantitative sigmoidal (R² > 0.95) dependence of the transfection efficiency and cell viability on the distance between the cell membrane and the nearest plasmid. We propose this distance value as a new, more accurate output parameter that could be used in further optimization studies as a predictor and a measure of electrotransfection efficiency. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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15 pages, 3374 KiB

Open AccessArticle

Electronic Emulator of Biological Tissue as an Electrical Load during Electroporation

by Eva Pirc, Bertrand Balosetti, Damijan Miklavčič and Matej Reberšek

Appl. Sci. 2020, 10(9), 3103; https://doi.org/10.3390/app10093103 - 29 Apr 2020

Cited by 3 | Viewed by 2585

Abstract

Electroporation is an emerging technology, with great potential in many different medical and biotechnological applications, food engineering and biomass processing. Large variations of biological load characteristics, however, represent a great challenge in electroporator design, which results in different solutions. Because a clinical electroporator [...] Read more.

Electroporation is an emerging technology, with great potential in many different medical and biotechnological applications, food engineering and biomass processing. Large variations of biological load characteristics, however, represent a great challenge in electroporator design, which results in different solutions. Because a clinical electroporator is a medical device, it must comply with medical device regulative and standards. However, none of the existing standards directly address the operation or electroporator’s performance requirements. In order to evaluate clinical, laboratory and prototype electroporation devices during the development process, or to evaluate their final performance considering at least from the perspective of output pulse parameters, we present a case study on the design of an electronic emulator of biological tissue as an electrical load during electroporation. The proposed electronic load emulator is a proof of concept, which enables constant and sustainable testing and unbiased comparison of different electroporators’ operations. We developed an analog electrical circuit that has equivalent impedance to the beef liver tissue in combination with needle electrodes, during high voltage pulse delivery and/or electroporation. Current and voltage measurements during electroporation of beef liver tissue ex vivo, were analyzed and parametrized to define the analog circuit equation. An equivalent circuit was simulated, built and validated. The proposed concept of an electronic load emulator can be used for “classical” electroporator (i.e., not nanosecond) performance evaluation and comparison of their operation. Additionally, it facilitates standard implementation regarding the testing protocol and enables quality assurance. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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17 pages, 9664 KiB

Open AccessFeature PaperArticle

Doxorubicin Assisted by Microsecond Electroporation Promotes Irreparable Morphological Alternations in Sensitive and Resistant Human Breast Adenocarcinoma Cells

by Nina Rembiałkowska, Magda Dubińska-Magiera, Andrzej Sikora, Wojciech Szlasa, Anna Szewczyk, Hanna Czapor-Irzabek, Małgorzata Daczewska, Jolanta Saczko and Julita Kulbacka

Appl. Sci. 2020, 10(8), 2765; https://doi.org/10.3390/app10082765 - 16 Apr 2020

Cited by 16 | Viewed by 3536

Abstract

Electroporation increases the transmembrane transport of molecules. The combination of electric pulses with cytostatic compounds is beneficial for cancer treatment. Doxorubicin (DOX) is a commonly used chemotherapeutic anticancer drug. Its fluorescence properties enable the investigation of drug distribution and metabolism. In this study, [...] Read more.

Electroporation increases the transmembrane transport of molecules. The combination of electric pulses with cytostatic compounds is beneficial for cancer treatment. Doxorubicin (DOX) is a commonly used chemotherapeutic anticancer drug. Its fluorescence properties enable the investigation of drug distribution and metabolism. In this study, doxorubicin was enhanced by electroporation to eliminate cancer cells more effectively. The influence of electroporation on the drug uptake was evaluated in two cell lines: MCF-7/WT and MCF-7/DOX. The intracellular localization of doxorubicin and its impact on the intracellular structure organization were examined under a confocal microscope. Cellular effects were examined with the 3(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test that estimates the rate of metabolism in viable cells. The ultrastructure (TEM) of tumor cells subjected to the electric field was analyzed. An enhanced doxorubicin efficacy was observed in MCF-7/DOX cells after combination with electroporation. The response of the resistant cell line was revealed to be more sensitive to electric pulses. Electroporation-based methods may be attractive for cancer treatment in human breast adenocarcinoma, especially with acquired resistance. Electroporation enables a reduction of the effective dose of the drugs and the exposure time in this type of cancer, diminishing side effects of the systemic therapy. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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Review

Jump to: Research, Other

20 pages, 4277 KiB

Open AccessReview

Concepts and Capabilities of In-House Built Nanosecond Pulsed Electric Field (nsPEF) Generators for Electroporation: State of Art

by Paulius Butkus, Arūnas Murauskas, Sonata Tolvaišienė and Vitalij Novickij

Appl. Sci. 2020, 10(12), 4244; https://doi.org/10.3390/app10124244 - 20 Jun 2020

Cited by 19 | Viewed by 4557

Abstract

Electroporation is a pulsed electric field triggered phenomenon of cell permeabilization, which is extensively used in biomedical and biotechnological context. There is a growing scientific demand for high-voltage and/or high-frequency pulse generators for electropermeabilization of cells (electroporators). In the scope of this article [...] Read more.

Electroporation is a pulsed electric field triggered phenomenon of cell permeabilization, which is extensively used in biomedical and biotechnological context. There is a growing scientific demand for high-voltage and/or high-frequency pulse generators for electropermeabilization of cells (electroporators). In the scope of this article we have reviewed the basic topologies of nanosecond pulsed electric field (nsPEF) generators for electroporation and the parametric capabilities of various in-house built devices, which were introduced in the last two decades. Classification of more than 60 various nsPEF generators was performed and pulse forming characteristics (pulse shape, voltage, duration and repetition frequency) were listed and compared. Lastly, the trends in the development of the electroporation technology were discussed. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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Other

Jump to: Research, Review

12 pages, 980 KiB

Open AccessBrief Report

The First Study of Irreversible Electroporation with Calcium Ions and Chemotherapy in Patients with Locally Advanced Pancreatic Adenocarcinoma

by Julia Rudno-Rudzińska, Wojciech Kielan, Maciej Guziński, Maciej Płochocki and Julita Kulbacka

Appl. Sci. 2020, 10(15), 5163; https://doi.org/10.3390/app10155163 - 27 Jul 2020

Cited by 13 | Viewed by 2721

Abstract

(1) Background: In the last two decades, anticancer treatment has been extensively developed based on various physical methods, including electroporation (EP). On the basis of many in vitro and in vivo studies, electroporation and further electrochemotherapy (ECT) have been established as a treatment [...] Read more.

(1) Background: In the last two decades, anticancer treatment has been extensively developed based on various physical methods, including electroporation (EP). On the basis of many in vitro and in vivo studies, electroporation and further electrochemotherapy (ECT) have been established as a treatment method for cutaneous and subcutaneous lesions. In this procedure, after placing electrodes in the tumor mass and the generation of electrical pulses, a reversible or irreversible rearrangement of the cell membrane occurs. Calcium electroporation has already been applied to treat skin tumors and subcutaneous tissue tumors. Here, we demonstrate the first application of irreversible electroporation (IRE) in combination with calcium ions and chemotherapy for patients with cancer. (2) Methods: This study aimed to present and compare the findings and outcomes of patients with locally advanced pancreatic adenocarcinoma who were qualified for the procedure. Two patients were treated with IRE with calcium ions after chemotherapy (paclitaxel + Gemcitabine (GCB) or FOLOX) or only FOLFOX (folinic acid calcium folinate, fluorouracil, and oxaliplatin). The clinicopathological data, overall survival, and the safety of the procedure were analyzed. (3) Results: Two patients were treated with calcium electroporation. One of the patients developed pancreatitis and the second developed pancreatic fistula, but both of them continued standard systemic treatment. Overall survival was 9 months in the first case and 21 months in the second case (and the patient is still alive). Calcium electroporation had a good impact on QOL (Quality of Life). (4) Conclusions: IRE accompanied chemotherapy, and intratumoral calcium ions administration might represent an additional therapy to surgery and chemotherapy in patients with locally advanced pancreatic cancer (LAPC), particularly in unresectable cases. However, further studies of randomized trials should be undertaken to elucidate the role of chemotherapy in IRE protocols. Full article

(This article belongs to the Special Issue Electroporation Systems and Applications)

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