Investigation of the mutagenic potential of cold atmospheric plasma at bactericidal dosages

Boxhammer, Veronika; Li, Yang‐Fang; Köritzer, Julia; Shimizu, Tetsuji; Maisch, Tim; Thomas, Hubertus M.; Schlegel, Jürgen; Morfill, G. E.; Zimmermann, Julia L.

doi:10.1016/j.mrgentox.2012.12.015

Cited by 84 publications

(60 citation statements)

References 37 publications

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“…Although cell culture experiments are commonly used to investigate the extent of damage of substances, animal experiments are more meaningful studies with regard to long-term effects. Plasma application was shown to lack genotoxic effects in vitro [30,32] and using an egg HET-MN model [31], which takes an intermediate position between cell culture and rodent models. Here, we have further quantitatively characterized the long-term side effects using an immunocompetent mouse model.…”

Section: Discussionmentioning

confidence: 99%

“…In contrast to low ROS and RNS quantities, higher concentrations are known to be responsible for apoptotic signaling [26,27,28] or finally for DNA damage [29]. Previous studies assessed the mutagenic risks of plasma in vitro [30,31,32]. The data provided in these studies demonstrated the absence of mutagenic or genotoxic effects in plasma-treated cells or in a hen’s egg test model for micronuclei induction (HET-MN), suggesting that a clinical application of the argon plasma jet does not pose mutagenic risks.…”

Section: Introductionmentioning

confidence: 99%

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One Year Follow-Up Risk Assessment in SKH-1 Mice and Wounds Treated with an Argon Plasma Jet

Schmidt

Woedtke

Stenzel³

et al. 2017

IJMS

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Multiple evidence in animal models and in humans suggest a beneficial role of cold physical plasma in wound treatment. Yet, risk assessment studies are important to further foster therapeutic advancement and acceptance of cold plasma in clinics. Accordingly, we investigated the long-term side effects of repetitive plasma treatment over 14 consecutive days in a rodent full-thickness ear wound model. Subsequently, animals were housed for 350 days and sacrificed thereafter. In blood, systemic changes of the pro-inflammatory cytokines interleukin 1β and tumor necrosis factor α were absent. Similarly, tumor marker levels of α-fetoprotein and calcitonin remained unchanged. Using quantitative PCR, the expression levels of several cytokines and tumor markers in liver, lung, and skin were found to be similar in the control and treatment group as well. Likewise, histological and immunohistochemical analysis failed to detect abnormal morphological changes and the presence of tumor markers such as carcinoembryonic antigen, α-fetoprotein, or the neighbor of Punc 11. Absence of neoplastic lesions was confirmed by non-invasive imaging methods such as anatomical magnetic resonance imaging and positron emission tomography-computed tomography. Our results suggest that the beneficial effects of cold plasma in wound healing come without apparent side effects including tumor formation or chronic inflammation.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

One Year Follow-Up Risk Assessment in SKH-1 Mice and Wounds Treated with an Argon Plasma Jet

Schmidt

Woedtke

Stenzel³

et al. 2017

IJMS

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“…Second, they utilized read-out system such as cytochrome C release, Comet-assay, or the ATM/ATR system that are not exclusively linked only to DNA-damage but also to general oxidative stress and apoptosis [30–32]. By contrast, mutagenicity studies carried out and according to OECD guidelines concluded a lack of permanent DNA damage following exposure to cold plasma [33, 34], also for the plasma jet used in this study [35]. Nonetheless, all studies mentioned were in vitro work, and such test models do not allow for biologically relevant conclusions in vertebrate organisms.…”

Section: Introductionmentioning

confidence: 99%

Investigating the Mutagenicity of a Cold Argon-Plasma Jet in an HET-MN Model

et al. 2016

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ObjectiveSo-called cold physical plasmas for biomedical applications generate reactive oxygen and nitrogen species and the latter can trigger DNA damage at high concentrations. Therefore, the mutagenic risks of a certified atmospheric pressure argon plasma jet (kINPen MED) and its predecessor model (kINPen 09) were assessed.MethodsInner egg membranes of fertilized chicken eggs received a single treatment with either the kINPen 09 (1.5, 2.0, or 2.5 min) or the kINPen MED (3, 4, 5, or 10 min). After three days of incubation, blood smears (panoptic May-Grünwald-Giemsa stain) were performed, and 1000 erythrocytes per egg were evaluated for the presence of polychromatic and normochromic nuclear staining as well as nuclear aberrations and binucleated cells (hen’s egg test for micronuclei induction, HET-MN). At the same time, the embryo mortality was documented. For each experiment, positive controls (cyclophosphamide and methotrexate) and negative controls (NaCl-solution, argon gas) were included. Additionally, the antioxidant potential of the blood plasma was assessed by ascorbic acid oxidation assay after treatment.ResultsFor both plasma sources, there was no evidence of genotoxicity, although at the longest plasma exposure time of 10 min the mortality of the embryos exceeded 40%. The antioxidant potential in the egg’s blood plasma was not significantly reduced immediately (p = 0.32) or 1 h (p = 0.19) post exposure to cold plasma.ConclusionThe longest plasma treatment time with the kINPen MED was 5–10 fold above the recommended limit for treatment of chronic wounds in clinics. We did not find mutagenic effects for any plasma treatment time using the either kINPen 09 or kINPen MED. The data provided with the current study seem to confirm the lack of a genotoxic potential suggesting that a veterinary or clinical application of these argon plasma jets does not pose mutagenic risks.

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“…Reactive nitrogen species are highly toxic and can lead to cell death by increasing DNA damage (15). One of the potential benefits of ACP as a sterilization or pasteurization technology is the reported low mutation level associated with it, which may be attributed to the "cocktail" of reactive species generated (16,17). However, different patterns of cellular damage between Gram-negative and Gram-positive bacteria were observed in previous studies (18,19).…”

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confidence: 97%

Mechanisms of Inactivation by High-Voltage Atmospheric Cold Plasma Differ for Escherichia coli and Staphylococcus aureus

Han¹,

Patil²,

Boehm³

et al. 2016

Appl Environ Microbiol

341

200

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bAtmospheric cold plasma (ACP) is a promising nonthermal technology effective against a wide range of pathogenic microorganisms. Reactive oxygen species (ROS) play a crucial inactivation role when air or other oxygen-containing gases are used. With strong oxidative stress, cells can be damaged by lipid peroxidation, enzyme inactivation, and DNA cleavage. Identification of ROS and an understanding of their role are important for advancing ACP applications for a range of complex microbiological issues. In this study, the inactivation efficacy of in-package high-voltage (80 kV [root mean square]) ACP (HVACP) and the role of intracellular ROS were investigated. Two mechanisms of inactivation were observed in which reactive species were found to either react primarily with the cell envelope or damage intracellular components. Escherichia coli was inactivated mainly by cell leakage and low-level DNA damage. Conversely, Staphylococcus aureus was mainly inactivated by intracellular damage, with significantly higher levels of intracellular ROS observed and little envelope damage. However, for both bacteria studied, increasing treatment time had a positive effect on the intracellular ROS levels generated.A tmospheric cold plasma (ACP) refers to nonequilibrated plasma generated at near-ambient temperatures and pressure. ACP is composed of particles, including free electrons, radicals, and positive and negative ions, but it is low in collision frequency of gas discharge compared to that with equilibrated plasma (1, 2). ACP technologies have widely been applied for many surface treatments and environmental processes. Recently, they have been studied for food sterilization and plasma medicine (2-5).ACP provides inactivation effects against a wide range of microbes, mainly by the generation of cell-lethal reactive species (6-8). By discharging in air, groups of reactive species are generated, such as reactive oxygen species (ROS), reactive nitrogen species (RNS), UV radiation, energetic ions, and charged particles (5). However, the inactivation efficacy can be varied by changing the working gases, which results in different types or amounts of reactive species generated (9-11). ROS are often identified as the principal effecting species, with a relatively long half-life and strong antimicrobial effects, which are generated in oxygen-containing gases (12).ROS generated during plasma discharge in air or oxygen-containing mixtures are assemblies of ozone, hydrogen peroxide, and singlet and atomic oxygen, while ozone is considered the most microbicidal species (13). With strong oxidative stress, cells are damaged by lipid peroxidation, enzyme inactivation, and DNA cleavage. The generation of plasma in air or a nitrogen-containing gas mixture can also generate NO x species. However, higher inactivation efficacy has been reported with the combined application of NO and H 2 O 2 on Escherichia coli than that with a treatment with NO or H 2 O 2 alone (14). Reactive nitrogen species are highly toxic and can lead to cell death by increas...

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Investigation of the mutagenic potential of cold atmospheric plasma at bactericidal dosages

Cited by 84 publications

References 37 publications

One Year Follow-Up Risk Assessment in SKH-1 Mice and Wounds Treated with an Argon Plasma Jet

One Year Follow-Up Risk Assessment in SKH-1 Mice and Wounds Treated with an Argon Plasma Jet

Investigating the Mutagenicity of a Cold Argon-Plasma Jet in an HET-MN Model

Mechanisms of Inactivation by High-Voltage Atmospheric Cold Plasma Differ for Escherichia coli and Staphylococcus aureus

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