BackgroundThe rate of microscopic incomplete resections of gastrointestinal cancers including pancreatic cancer has not changed considerably over the past years. Future intra-operative applications of tissue tolerable plasmas (TTP) could help to address this problem. Plasma is generated by feeding energy, like electrical discharges, to gases. The development of non-thermal atmospheric plasmas displaying spectra of temperature within or just above physiological ranges allows biological or medical applications of plasmas.MethodsWe have investigated the effects of tissue tolerable plasmas (TTP) on the human pancreatic cancer cell line Colo-357 and PaTu8988T and the murine cell line 6606PDA in vitro (Annexin-V-FITC/DAPI-Assay and propidium iodide DNA staining assay) as well as in the in vivo tumour chorio-allantoic membrane (TUM-CAM) assay using Colo-357.ResultsTTP of 20 seconds (s) induced a mild elevation of an experimental surface temperature of 23.7 degree Celsius up to 26.63+/−0.40 degree Celsius. In vitro TTP significantly (p=0.0003) decreased cell viability showing the strongest effects after 20s TTP. Also, TTP effects increased over time levelling off after 72 hours (30.1+/−4.4% of dead cells (untreated control) versus 78.0+/−9.6% (20s TTP)). However, analyzing these cells for apoptosis 10s TTP revealed the largest proportion of apoptotic cells (34.8+/−7.2%, p=0.0009 versus 12.3+/−6.6%, 20s TTP) suggesting non-apoptotic cell death in the majority of cells after 20s TTP. Using solid Colo-357 tumours in the TUM-CAM model TUNEL-staining showed TTP-induced apoptosis up to a depth of tissue penetration (DETiP) of 48.8+/−12.3μm (20s TTP, p<0.0001). This was mirrored by a significant (p<0.0001) reduction of Ki-67+ proliferating cells (80.9+/−13.2% versus 37.7+/−14.6%, p<0.0001) in the top cell layers as well as typical changes on HE specimens. The bottom cell layers were not affected by TTP.ConclusionsOur data suggest possible future intra-operative applications of TTP to reduce microscopic residual disease in pancreatic cancer resections. Further promising applications include other malignancies (central liver/lung tumours) as well as synergistic effects combining TTP with chemotherapies. Yet, adaptations of plasma sources as well as of the composition of effective components of TTP are required to optimize their synergistic apoptotic actions.

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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.

<i>Background:</i> The formation of biofilms is crucial in the pathogenesis of many acute and subacute microbial infections, including chronic wounds and foreign-body-related infections. Topical antimicrobial therapy with chemical antiseptics or physical treatment with tissue-tolerable plasma (TTP) may be promising to control bacterial infection. <i>Methods:</i> We assessed the efficacy of 0.1% chlorhexidine digluconate (CHX), 0.02 and 0.04% polihexanide (polyhexamethylene biguanide, PHMB) and of TTP against <i>Pseudomonas aeruginosa</i> SG81 biofilm grown in microtitre plates (polystyrene) and on silicone materials in an artificial wound fluid. <i>Results:</i> Overall, PHMB was as effective as CHX in reducing the total amount of biofilm (gentian violet assay) and in reducing the bacterial metabolism in biofilms (XTT assay). TTP also led to a significant reduction in colony-forming units. <i>Conclusion:</i> The antimicrobial activity of PHMB in biofilms is comparable to that of CHX. TTP could become an interesting physical alternative to chemical antisepsis in the future.

The treatment of infected wounds is one possible therapeutic aspect of plasma medicine. Chronic wounds are often associated with microbial biofilms which limit the efficacy of antiseptics. The present study investigates two different surface barrier discharges with air plasma to compare their efficacy against microbial biofilms with chlorhexidine digluconate solution (CHX) as representative of an important antibiofilm antiseptic. Pseudomonas aeruginosa SG81 and Staphylococcus epidermidis RP62A were cultivated on polycarbonate discs. The biofilms were treated for 30, 60, 150, 300 or 600 s with plasma or for 600 s with 0.1% CHX, respectively. After treatment, biofilms were dispensed by ultrasound and the antimicrobial effects were determined as difference in the number of the colony forming units by microbial culture. A high antimicrobial efficacy on biofilms of both plasma sources in comparison to CHX treatment was shown. The efficacy differs between the used strains and plasma sources. For illustration, the biofilms were examined under a scanning electron microscope before and after treatment. Additionally, cytotoxicity was determined by the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay with L929 mouse fibroblast cell line. The cell toxicity of the used plasma limits its applicability on human tissue to maximally 150 s. The emitted UV irradiance was measured to estimate whether UV could limit the application on human tissue at the given parameters. It was found that the UV emission is negligibly low. In conclusion, the results support the assumption that air plasma could be an option for therapy of chronic wounds.

Aim: To compare the tissue tolerance and efficacy of two wound antiseptics with tissue-tolerable plasma (TTP) on enucleated contaminated eyes from slaughtered pigs in order to draw consequences for the use of TTP on wounds. Method: The corneas of extracted eyes were contaminated with Staphylococcus aureus or Pseudomonas aeruginosa. One and 10 min after application of 10% povidone (PVP)-iodine and 0.04% polyhexanide, respectively, the eyes were rinsed with inactivating solution. To test TTP, the plasma pen meandered over the eyes at a speed of 30 mm/s and a distance of 5 mm; the eyes were then rinsed with balanced salt solution. The reduction factor was calculated by the difference between the logarithm of colony-forming units in the rinse before and after antisepsis or TTP application. Results: The efficacy of TTP (reduction factor 2.4–2.9) was significantly higher (p < 0.001) than that of PVP-iodine and polyhexanide (reduction factor 1.7–2.1). Conclusion: TTP is more effective than the tested wound antiseptics. The lack of histological damage to the eyes of slaughtered pigs would seem to make its use as a wound antiseptic a viable alternative. In contrast to antiseptics, it supplies additional energy in the form of heat, electric fields and radicals by TTP.

Cold physical plasmas ignited a technological spark in industry, biotechnology, and medicine. Especially the field of hygiene benefited of the plasma's exceptional activity against pathogenic microorganisms. Together with plasma-based surface functionalization, these qualities are highly relevant in a variety of processes in health care, such as the decontamination or sterilization of medical devices, food, packaging materials, waste water, or indoor air. In medicine, plasma has proven to show promising antiseptic results on skin and mucosal membranes in infection-related diseases in dermatology and dentistry. This comprehensive review will discuss the current applications of cold plasma in the fields of hygiene, and will provide a promising outlook on many applications yet to come.

Microbial biofilms impair wound healing and the efficacy of antiseptics. This study used the plasma source kinpen 09 to investigate the efficacy of argon and an argon/oxygen‐mixture plasma against in vitro biofilms of Pseudomonas aeruginosa and Staphylococcus epidermidis compared to the antiseptic chlorhexidine digluconate (CHX). Biofilms were treated for 30, 60, 150, or 300 s with plasma and for 600 s with a 0.1% CHX solution. Thereafter, the biofilms were dispersed and their inactivation was determined as the reduction of colony forming units. Plasma treatment led to a significant antimicrobial effect compared to the untreated control and was at least as effective as the treatment with CHX, or even better, depending on the gas admixture, the bacterial strain used, and the exposure time. magnified image