Search Results (43)

Epidermal melanin synthesis determines an individual’s skin color. In humans, melanin is formed by melanocytes within the epidermis. The process of melanin synthesis strongly depends on a range of cellular factors, including the fine-tuned interplay with reactive oxygen species (ROS). In this context, a role of cold atmospheric plasma (CAP) on melanin synthesis was proposed due to its tunable ROS generation. Herein, the argon-driven plasma jet kINPen^® MED was employed, and its impact on melanin synthesis was evaluated by comparison with known stimulants such as the phosphodiesterase inhibitor IBMX and UV radiation. Different available model systems were employed, and the melanin content of both cultured human melanocytes (in vitro) and full-thickness human skin biopsies (in situ) were analyzed. A histochemical method detected melanin in skin tissue. Cellular melanin was measured by NIR autofluorescence using flow cytometry, and a highly sensitive HPLC-MS method was applied, which enabled the differentiation of eu- and pheomelanin by their degradation products. The melanin content in full-thickness human skin biopsies increased after repeated CAP exposure, while there were only minor effects in cultured melanocytes compared to UV radiation and IBMX treatment. Based on these findings, CAP does not appear to be a useful option for treating skin pigmentation disorders. On the other hand, the risk of hyperpigmentation as an adverse effect of CAP application for wound healing or other dermatological diseases seems to be neglectable. Full article

Tissue inflammation drives the infiltration of innate immune cells that generate reactive species to kill bacteria and recruit adaptive immune cells. Neutrophil activation fosters the release of myeloperoxidase (MPO) enzyme, a heme-containing protein generating hypochlorous acid (HOCl) from hydrogen peroxide (H₂O₂) and chloride ions. MPO-dependent oxidant formation initiates bioactive oxidation and chlorination products and induces oxidative post-translational modifications (oxPTMs) on proteins and lipid oxidation. Besides HOCl and H₂O₂, further reactive species such as singlet oxygen and nitric oxide are generated in inflammation, leading to modified proteins, potentially resulting in their altered bioactivity. So far, knowledge about multiple free radical-induced modifications of MPO and its effects on HOCl generation is lacking. To mimic this multi-oxidant microenvironment, human MPO was exposed to several reactive species produced simultaneously via argon plasma operated at body temperature. Several molecular gas admixes were used to modify the reactive species type profiles generated. MPO was investigated by studying its oxPTMs, changes in protein structure, and enzymatic activity. MPO activity was significantly reduced after treatment with all five tested plasma gas conditions. Dynamic light scattering and CD-spectroscopy revealed altered MPO protein morphology indicative of oligomerization. Using mass spectrometry, various oxPTMs, such as +1O, +2O, and +3O, were determined on methionine and cysteine (Cys), and -1H-1N+1O was detected in asparagine (Asp). The modification types identified differed between argon-oxygen and argon-nitrogen plasmas. However, all plasma gas conditions led to the deamidation of Asp and oxidation of Cys residues, suggesting an inactivation of MPO due to oxPTM-mediated conformational changes. Full article

Background cold atmospheric plasma (CAP) is known to be a surface-friendly yet antimicrobial and activating process for surfaces such as titanium. The aim of the present study was to describe the decontaminating effects of CAP on contaminated collagen membranes and their influence on the properties of this biomaterial in vitro. Material and Methods: A total of n = 18 Bio-Gide^® (Geistlich Biomaterials, Baden-Baden, Germany) membranes were examined. The intervention group was divided as follows: n = 6 membranes were treated for one minute, and n = 6 membranes were treated for five minutes with CAP using kINPen^® MED (neoplas tools GmbH, Greifswald, Germany) with an output of 5 W, respectively. A non-CAP-treated group (n = 6) served as the control. The topographic alterations were evaluated via X-ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM). Afterward, the samples were contaminated with E. faecalis for 6 days, and colony-forming unit (CFU) counts and additional SEM analyses were performed. The CFUs increased with CAP treatment time in our analyses, but SEM showed that the surface of the membranes was essentially free from bacteria. However, the deeper layers showed remaining microbial conglomerates. Furthermore, we showed, via XPS analysis, that increasing the CAP time significantly enhances the carbon (carbonyl group) concentration, which also correlates negatively with the decontaminating effects of CAP. Conclusions: Reactive carbonyl groups offer a potential mechanism for inhibiting the growth of E. faecalis on collagen membranes after cold atmospheric plasma treatment. Full article

Cold physical plasma is a partially ionized gas operated at body temperature and utilized for heat-sensitive technical and medical purposes. Physical plasma is a multi-component system consisting of, e.g., reactive species, ions and electrons, electric fields, and UV light. Therefore, cold plasma technology is an interesting tool for introducing biomolecule oxidative modifications. This concept can be extended to anticancer drugs, including prodrugs, which could be activated in situ to enhance local anticancer effects. To this end, we performed a proof-of-concept study on the oxidative prodrug activation of a tailor-made boronic pinacol ester fenretinide treated with the atmospheric pressure argon plasma jet kINPen operated with either argon, argon–hydrogen, or argon–oxygen feed gas. Fenretinide release from the prodrug was triggered via Baeyer–Villiger-type oxidation of the boron–carbon bond based on hydrogen peroxide and peroxynitrite, which were generated by plasma processes and chemical addition using mass spectrometry. Fenretinide activation led to additive cytotoxic effects in three epithelial cell lines in vitro compared to the effects of cold plasma treatment alone regarding metabolic activity reduction and an increase in terminal cell death, suggesting that cold physical plasma-mediated prodrug activation is a new direction for combination cancer treatment studies. Full article

Several physico-chemical modifications have been developed to improve cell contact with prosthetic oral implant surfaces. The activation with non-thermal plasmas was one option. Previous studies found that gingiva fibroblasts on laser-microstructured ceramics were hindered in their migration into cavities. However, after argon (Ar) plasma activation, the cells concentrated in and around the niches. The change in surface properties of zirconia and, subsequently, the effect on cell behavior is unclear. In this study, polished zirconia discs were activated by atmospheric pressure Ar plasma using the kINPen^®09 jet for 1 min. Surfaces were characterized by scanning electron microscopy, X-ray photoelectron spectroscopy (XPS), and water contact angle. In vitro studies with human gingival fibroblasts (HGF-1) focused on spreading, actin cytoskeleton organization, and calcium ion signaling within 24 h. After Ar plasma activation, surfaces were more hydrophilic. XPS revealed decreased carbon and increased oxygen, zirconia, and yttrium content after Ar plasma. The Ar plasma activation boosted the spreading (2 h), and HGF-1 cells formed strong actin filaments with pronounced lamellipodia. Interestingly, the cells’ calcium ion signaling was also promoted. Therefore, argon plasma activation of zirconia seems to be a valuable tool to bioactivate the surface for optimal surface occupation by cells and active cell signaling. Full article

Gas plasma jet technology was recently identified as a potential adjuvant in the fight against cancer. Here, the partial ionization of gas yields the local formation of an exceptional variety of highly reactive oxygen (ROS) and nitrogen (RNS) species, which are considered the main actors of plasma-induced antitumor effects. Yet, fundamental knowledge in preclinical plasma research relies on the predominant use of two-dimensional cell culture systems, despite causing significant shifts in redox chemistries that largely limit translational relevance. So far, the intricacy of studying complex plasma–tissue interactions causes substantial knowledge gaps concerning the key mechanisms and therapeutical limitations of plasma treatment in a living organism. Identifying physiologically relevant yet simplified tissue models is vital to address such questions. In our study, a side-by-side comparison of conventional and pre-established hydrogel models emphasized this discrepancy, revealing a marked difference in plasma-induced toxicity related to species distribution dynamics. Chemically embedded, fluorescent reporters were further used to characterize reactive species’ fingerprints in hydrogels compared to liquids. In addition, a thirteen cell-line screening outlined the widespread applicability of the approach while indicating the need to optimize growth conditions dependent on the cell line investigated. Overall, our study presents important implications for the implementation of clinically relevant tissue culture models in preclinical plasma medicine in the future. Full article

Background: The use of cold atmospheric plasma (CAP) in oncology has been intensively investigated over the past 15 years as it inhibits the growth of many tumor cells. It is known that reactive oxidative species (ROS) produced in CAP are responsible for this effect. However, to translate the use of CAP into medical practice, it is essential to know how CAP treatment affects non-malignant cells. Thus, the current in vitro study deals with the effect of CAP on human bone cancer cells and human osteoblasts. Here, identical CAP treatment regimens were applied to the malignant and non-malignant bone cells and their impact was compared. Methods: Two different human bone cancer cell types, U2-OS (osteosarcoma) and A673 (Ewing’s sarcoma), and non-malignant primary osteoblasts (HOB) were used. The CAP treatment was performed with the clinically approved kINPen MED. After CAP treatment, growth kinetics and a viability assay were performed. For detecting apoptosis, a caspase-3/7 assay and a TUNEL assay were used. Accumulated ROS was measured in cell culture medium and intracellular. To investigate the influence of CAP on cell motility, a scratch assay was carried out. Results: The CAP treatment showed strong inhibition of cell growth and viability in bone cancer cells. Apoptotic processes were enhanced in the malignant cells. Osteoblasts showed a higher potential for ROS resistance in comparison to malignant cells. There was no difference in cell motility between benign and malignant cells following CAP treatment. Conclusions: Osteoblasts show better tolerance to CAP treatment, indicated by less affected viability compared to CAP-treated bone cancer cells. This points toward the selective effect of CAP on sarcoma cells and represents a further step toward the clinical application of CAP. Full article

Medical gas plasma is an experimental technology for anticancer therapy. Here, partial gas ionization yielded reactive oxygen and nitrogen species, placing the technique at the heart of applied redox biomedicine. Especially with the gas plasma jet kINPen, anti-tumor efficacy was demonstrated. This study aimed to examine the potential of using passive flow shaping to enhance the medical benefits of atmospheric plasma jets (APPJ). We used an in-house developed, proprietary Modular Nozzle System (MoNoS; patent-pending) to modify the flow properties of a kINPen. MoNoS increased the nominal plasma jet-derived reactive species deposition area and stabilized the air-plasma ratio within the active plasma zone while shielding it from external flow disturbances or gas impurities. At modest flow rates, dynamic pressure reduction (DPR) adapters did not augment reactive species deposition in liquids or tumor cell killing. However, MoNoS operated at kINPen standard argon fluxes significantly improved cancer organoid growth reduction and increased tumor immunogenicity, as seen by elevated calreticulin and heat-shock protein expression, along with a significantly spurred cytokine secretion profile. Moreover, the safe application of MoNoS gas plasma jet adapters was confirmed by their similar-to-superior safety profiles assessed in the hen’s egg chorioallantoic membrane (HET-CAM) coagulation and scar formation irritation assay. Full article

Gas plasma is a medical technology with antimicrobial properties. Its main mode of action is oxidative damage via reactive species production. The clinical efficacy of gas plasma-reduced bacterial burden has been shown to be hampered in some cases. Since the reactive species profile produced by gas plasma jets, such as the kINPen used in this study, are thought to determine antimicrobial efficacy, we screened an array of feed gas settings in different types of bacteria. Antimicrobial analysis was performed by single-cell analysis using flow cytometry. We identified humidified feed gas to mediate significantly greater toxicity compared to dry argon and many other gas plasma conditions. The results were confirmed by inhibition zone analysis on gas-plasma-treated microbial lawns grown on agar plates. Our results may have vital implications for clinical wound management and potentially enhance antimicrobial efficacy of medical gas plasma therapy in patient treatment. Full article

Survival rates among patients with pancreatic cancer, the most lethal gastrointestinal cancer, have not improved compared to other malignancies. Early tumor dissemination and a supportive, cancer-promoting tumor microenvironment (TME) limit therapeutic options and consequently impede tumor remission, outlining an acute need for effective treatments. Gas plasma-oxidized liquid treatment showed promising preclinical results in other gastrointestinal and gynecological tumors by targeting the tumor redox state. Here, carrier solutions are enriched with reactive oxygen (ROS) and nitrogen (RNS) species that can cause oxidative distress in tumor cells, leading to a broad range of anti-tumor effects. Unfortunately, clinical relevance is often limited, as many studies have forgone the use of medical-grade solutions. This study investigated the efficacy of gas plasma-oxidized Ringer’s lactate (oxRilac), a physiological solution often used in clinical practice, on two pancreatic cancer cell lines to induce tumor toxicity and provoke immunogenicity. Tumor toxicity of the oxRilac solutions was further confirmed in three-dimensional tumor spheroids monitored over 72 h and in ovo using stereomicroscope imaging of excised GFP-expressing tumors. We demonstrated that cell death signaling was induced in a dose-dependent fashion in both cell lines and was paralleled by the increased surface expression of key markers of immunogenic cell death (ICD). Nuclear magnetic resonance (NMR) spectroscopy analysis suggested putative reaction pathways that may cause the non-ROS related effects. In summary, our study suggests gas plasma-deposited ROS in clinically relevant liquids as an additive option for treating pancreatic cancers via immune-stimulating and cytotoxic effects. Full article

In modern oncology, therapies are based on combining monotherapies to overcome treatment resistance and increase therapy precision. The application of microsecond-pulsed electric fields (PEF) is approved to enhance local chemotherapeutic drug uptake within combination electrochemotherapy regimens. Reactive oxygen species (ROS) have been implicated in anticancer effects, and cold physical plasma produces vast amounts of ROS, which have recently been shown to benefit head and neck cancer patients. PEF and cold plasma technology have been linked to immunogenic cell death (ICD) induction, a regulated cell death accompanied by sterile inflammation that promotes antitumor immunity. To this end, we investigated the combined effect of both treatments regarding their intracellular ROS accumulation, toxicity, ICD-related marker expression, and optimal exposure sequence in a leukemia model cell line. The combination treatment substantially increased ROS and intracellular glutathione levels, leading to additive cytotoxic effects accompanied by a significantly increased expression of ICD markers, such as the eat-me signal calreticulin (CRT). Preconditioned treatment with cold plasma followed by PEF exposure was the most potent treatment sequence. The results indicate additive effects of cold plasma and PEF, motivating further studies in skin and breast tumor models for the future improvement of ECT in such patients. Full article

In cancer, antigen-presenting cells (APC), including dendritic cells (DCs), take up and process proteins to mount adaptive antitumor immune responses. This often happens in the context of inflamed cancer, where reactive oxygen species (ROS) are ubiquitous to modify proteins. However, the inflammatory consequences of oxidized protein uptake in DCs are understudied. To this end, we investigated human monocyte-derived cell surface marker expression and cytokine release profiles when exposed to oxidized and native proteins. Seventeen proteins were analyzed, including viral proteins (e.g., CMV and HBV), inflammation-related proteins (e.g., HO1 and HMGB1), matrix proteins (e.g., Vim and Coll), and vastly in the laboratory used proteins (e.g., BSA and Ova). The multifaceted nature of inflammation-associated ROS was mimicked using gas plasma technology, generating reactive species cocktails for protein oxidation. Fourteen oxidized proteins led to elevated surface marker expression levels of CD25, CD40, CD80, CD86, and MHC-II as well as strongly modified release of IL6, IL8, IL10, IL12, IL23, MCP-1, and TNFα compared to their native counterparts. Especially IL8, heme oxygenase 2, and vimentin oxidation gave pronounced effects. Furthermore, protein kinase phospho-array studies in monocyte-derived cells pulsed with native vs. oxidized IL8 and insulin showed enhanced AKT and RSK2 phosphorylation. In summary, our data provide for the first time an overview of the functional consequences of oxidized protein uptake by human monocyte-derived cells and could therefore be a starting point for exploiting such principle in anticancer therapy in the future. Full article

Protein vaccines rely on eliciting immune responses. Inflammation is a prerequisite for immune responses to control infection and cancer but is also associated with disease onset. Reactive oxygen species (ROSs) are central during inflammation and are capable of inducing non-enzymatic oxidative protein modifications (oxMods) associated with chronic disease, which alter the functionality or immunogenicity of proteins that are relevant in cancer immunotherapy. Specifically, antigen-presenting cells (APCs) take up and degrade extracellular native and oxidized proteins to induce adaptive immune responses. However, it is less clear how oxMods alter the protein’s immunogenicity, especially in inflammation-related short-lived reactive species. Gas plasma technology simultaneously generates a multitude of ROSs to modify protein antigens in a targeted and controlled manner to study the immunogenicity of oxMods. As model proteins relevant to chronic inflammation and cancer, we used gas plasma-treated insulin and CXCL8. We added those native or oxidized proteins to human THP-1 monocytes or primary monocyte-derived cells (moDCs). Both oxidized proteins caused concentration-independent maturation phenotype alterations in moDCs and THP-1 cells concerning surface marker expression and chemokine and cytokine secretion profiles. Interestingly, concentration-matched H₂O₂-treated proteins did not recapitulate the effects of gas plasma, suggesting sufficiently short diffusion distances for the short-lived reactive species to modify proteins. Our data provide evidence of dendric cell maturation and activation upon exposure to gas plasma- but not H₂O₂-modified model proteins. The biological consequences of these findings need to be elucidated in future inflammation and cancer disease models. Full article

The emerging use of low-temperature plasma in medicine, especially in wound treatment, calls for a better way of documenting the treatment parameters. This paper describes the development of a mobile sensory device (referred to as MSD) that can be used during the treatment to ease the documentation of important parameters in a streamlined process. These parameters include the patient’s general information, plasma source device used in the treatment, plasma treatment time, ambient humidity and temperature. MSD was developed as a standalone Raspberry Pi-based version and attachable module version for laptops and tablets. Both versions feature a user-friendly GUI, temperature–humidity sensor, microphone, treatment report generation and export. For the logging of plasma treatment time, a sound-based plasma detection system was developed, initially for three medically certified plasma source devices: kINPen^® MED, plasma care^®, and PlasmaDerm^® Flex. Experimental validation of the developed detection system shows accurate and reliable detection is achievable at 5 cm measurement distance in quiet and noisy environments for all devices. All in all, the developed tool is a first step to a more automated, integrated, and streamlined approach of plasma treatment documentation that can help prevent user variability. Full article

Reactive species generated by medical gas plasma technology can be enriched in liquids for use in oncology targeting disseminated malignancies, such as metastatic colorectal cancer. Notwithstanding, reactive species quantities depend on the treatment mode, and we recently showed gas plasma exposure in conductive modes to be superior for cancer tissue treatment. However, evidence is lacking that such a conductive mode also equips gas plasma-treated liquids to confer augmented intraperitoneal anticancer activity. To this end, employing atmospheric pressure argon plasma jet kINPen-treated Ringer’s lactate (oxRilac) in a CT26-model of colorectal peritoneal carcinomatosis, we tested repeated intraabdominal injection of such remotely or conductively oxidized liquid for antitumor control and immunomodulation. Enhanced reactive species formation in conductive mode correlated with reduced tumor burden in vivo, emphasizing the advantage of conduction over the free mode for plasma-conditioned liquids. Interestingly, the infiltration of lymphocytes into the tumors was equally enhanced by both treatments. However, significantly lower levels of interleukin (IL)4 and IL13 and increased levels of IL2 argue for a shift in intratumoral T-helper cell subpopulations correlating with disease control. In conclusion, our data argue for using conductively over remotely prepared plasma-treated liquids for anticancer treatment. Full article