Killing Bacteria Using Acetic Acid and Nanosecond Pulsed Electric Fields—An In Vivo Superficial Infection Model Study and Immune Response

Invasive infections caused by drug-resistant bacteria are a problem responsible for many fatal cases, especially in burn wound care centers, while bacterial resistance to antibiotics is growing dramatically worldwide. In this work, we utilize pulsed electric fields (up to 25 kV/cm × 750 ns) in combination with low-concentration (1%) acetic acid for the inactivation of P. aeruginosa. An in vivo superficial infection model is developed in BALB/C mice using a luminescent strain of P. aeruginosa. We show that an up to 25 kV/cm electric field (3 kV, 1.2 mm gap), when combined with acetic acid, induces a bacteriostatic effect, preventing further infection for up to 7 days after treatment. Additionally, we evaluate antibodies against surface and intracellular P. aeruginosa bacteria antigens following the treatment. It is shown that the levels of surface IgG and IgG1 antibodies are significantly lower in the murine serum of electric-field-treated mice compared to the bacterial-infection-bearing group of mice treated with acetic acid alone. The results of this work are useful as a proof of concept for the development of novel clinical procedures to fight drug-resistive microorganisms responsible for wound contamination and chronic wounds

Antitumor Response and Immunomodulatory Effects of Sub-Microsecond Irreversible Electroporation and Its Combination with Calcium Electroporation

In this work, we have investigated the feasibility of sub-microsecond range irreversible electroporation (IRE) with and without calcium electroporation in vivo. As a model, BALB/C mice were used and bioluminescent SP2/0 myeloma tumor models were developed. Tumors were treated with two separate pulsed electric field (PEF) pulsing protocols PEF1: 12 kV/cm × 200 ns × 500 (0.006 J/pulse) and PEF2: 12 kV/cm × 500 ns × 500 (0.015 J/pulse), which were delivered with and without Ca2+ (168 mM) using parallel plate electrodes at a repetition frequency of 100 Hz. Both PEF1 and PEF2 treatments reduced tumor growth and prolonged the life span of the mice, however, the PEF2 protocol was more efficient. The delay in tumor renewal was the biggest when a combination of IRE with calcium electroporation was used, however, we did not obtain significant differences in the final mouse survival compared to PEF2 alone. Anti-tumor immune responses were also investigated after treatment with PEF2 and PEF2+Ca. In both cases the treated mice had enlarged spleens and increased spleen T cell numbers, lower percentages of suppressor cell subsets (conventional CD4+CD25+ Treg, CD4+CD25−DX5+ Tr1, CD8+DX5+, CD4+CD28−, CD8+CD28−), changed proportions of Tcm and Tef/Tem T cells in the spleen and increased amount of tumor cell specific antibodies in the sera. The treatment based on IRE was effective against primary tumors, destroyed the tumor microenvironment and induced an anti-tumor immune response, however, it was not sufficient for complete control of tumor metastasis.This article belongs to the Special Issue Electric Field Based Therapies in Cancer Treatment: a Selection of Studies Presented at the 3rd World Congress on ElectroporationThis research was funded by Research Council of Lithuania, grant number S-MIP-19-22

Antitumor response and immunomodulatory effects of sub-microsecond irreversible electroporation and its combination with calcium electroporation

In this work, we have investigated the feasibility of sub-microsecond range irreversible electroporation (IRE) with and without calcium electroporation in vivo. As a model, BALB/C mice were used and bioluminescent SP2/0 myeloma tumor models were developed. Tumors were treated with two separate pulsed electric field (PEF) pulsing protocols PEF1: 12 kV/cm 200 ns 500 (0.006 J/pulse) and PEF2: 12 kV/cm 500 ns 500 (0.015 J/pulse), which were delivered with and without Ca2+ (168 mM) using parallel plate electrodes at a repetition frequency of 100 Hz. Both PEF1 and PEF2 treatments reduced tumor growth and prolonged the life span of the mice, however, the PEF2 protocol was more e cient. The delay in tumor renewal was the biggest when a combination of IRE with calcium electroporation was used, however, we did not obtain significant di erences in the final mouse survival compared to PEF2 alone. Anti-tumor immune responses were also investigated after treatment with PEF2 and PEF2+Ca. In both cases the treated mice had enlarged spleens and increased spleen T cell numbers, lower percentages of suppressor cell subsets (conventional CD4+CD25+ Treg, CD4+CD25 DX5+ Tr1, CD8+DX5+, CD4+CD28, CD8+CD28), changed proportions of Tcm and Tef/Tem T cells in the spleen and increased amount of tumor cell specific antibodies in the sera. The treatment based on IRE was e ective against primary tumors, destroyed the tumor microenvironment and induced an anti-tumor immune response, however, it was not sufficient for complete control of tumor metastasisAplinkos tyrimų centrasValstybinis mokslinių tyrimų institutas, Inovatyvios medicinos centrasVilniaus Gedimino technikos universitetasVytauto Didžiojo universiteta

Non-invasive nanosecond electroporation for biocontrol of surface infections: an in vivo study

Invasive infections caused by drug-resistant bacteria are frequently responsible for fatal sepsis, morbidity and mortality rates. In this work, we propose a new methodology based on nanosecond high frequency electric feld bursts, which enables successful eradication of bacteria in vivo. High frequency (15kHz) 15–25kV/cm 300–900ns pulsing bursts were used separately and in combination with acetic acid (0.1–1%) to treat Pseudomonas aeruginosa in a murine model. Acetic acid 1% alone was efective resulting in almost 10-fold reduction of bacteria viability, however combination of nanosecond electric feld and acetic acid 1% treatment was the most successful showing almost full eradication (0.01% survival compared to control) of the bacteria in the contaminated area. The short duration of the pulses (sub-microsecond) and high frequency (kHz range) of the burst enabled reduction of the muscle contractions to barely detectable level while the proposed applicators ensured predominantly topical treatment, without electroporation of deeper tissues. The results of our study have direct application for treatment of wounds and ulcers when chemical treatment is no longer efective