Synergistic effect of cold gas plasma and experimental drug exposure exhibits skin cancer toxicity in vitro and in vivo.

INTRODUCTION: Skin cancer is often fatal, which motivates new therapy avenues. Recent advances in cancer treatment are indicative of the importance of combination treatments in oncology. Previous studies have identified small molecule-based therapies and redox-based technologies, including photodynamic therapy or medical gas plasma, as promising candidates to target skin cancer. OBJECTIVE: We aimed to identify effective combinations of experimental small molecules with cold gas plasma for therapy in dermato-oncology. METHODS: Promising drug candidates were identified after screening an in-house 155-compound library using 3D skin cancer spheroids and high content imaging. Combination effects of selected drugs and cold gas plasma were investigated with respect to oxidative stress, invasion, and viability. Drugs that had combined well with cold gas plasma were further investigated in vascularized tumor organoids in ovo and a xenograft mouse melanoma model in vivo. RESULTS: The two chromone derivatives Sm837 and IS112 enhanced cold gas plasma-induced oxidative stress, including histone 2A.X phosphorylation, and further reduced proliferation and skin cancer cell viability. Combination treatments of tumor organoids grown in ovo confirmed the principal anti-cancer effect of the selected drugs. While one of the two compounds exerted severe toxicity in vivo, the other (Sm837) resulted in a significant synergistic anti-tumor toxicity at good tolerability. Principal component analysis of protein phosphorylation profiles confirmed profound combination treatment effects in contrast to the monotherapies. CONCLUSION: We identified a novel compound that, combined with topical cold gas plasma-induced oxidative stress, represents a novel and promising treatment approach to target skin cancer.

Decreased inflammatory profile in oral leukoplakia tissue exposed to cold physical plasma ex vivo.

BACKGROUND: Oral leukoplakia (OL) is an unfavorable oral disease often resistant to therapy. To this end, cold physical plasma technology was explored as a novel therapeutic agent in an experimental setup. METHODS: Biopsies with a diameter of 3 mm were obtained from non-diseased and OL tissues. Subsequently, cold atmospheric pressure plasma (CAP) exposure was performed ex vivo in the laboratory. After 20 h of incubation, biopsies were cryo-conserved, and tissue sections were quantified for lymphocyte infiltrates, discriminating between naïve and memory cytotoxic and T-helper cells. In addition, the secretion pattern related to inflammation was investigated in the tissue culture supernatants by quantifying 10 chemokines and cytokines. RESULTS: In CAP-treated OL tissue, significantly decreased overall lymphocyte numbers were observed. In addition, reduced levels were observed when discriminating for the T-cell subpopulations but did not reach statistical significance. Moreover, CAP treatment significantly reduced levels of C-X-C motif chemokine 10 (CXCL10) and granulocyte-macrophage colony-stimulating factor in the OL biopsies' supernatants. In idiopathically inflamed tissues, ex vivo CAP exposure reduced T-cells and CXCL10 as well but also led to markedly increased interleukin-1ß secretion. CONCLUSION: Our findings suggest CAP to have immuno-modulatory properties, which could be of therapeutic significance in the therapy of OL. Future studies should investigate the efficacy of CAP therapy in vivo in a larger cohort.

Mechanical and Plasma Electrolytic Polishing of Dental Alloys.

(1) Background: In dentistry, a reduction in surface roughness is established mostly by conventional mechanical polishing to hinder biofilm adhesion. This is time- and labor-intensive. Plasma electrolytic polishing is believed to be an effective finishing method due to the reduced treatment time and materials used for applications in dentistry. (2) Methods: Co-Cr-Mo dental alloy samples were sandblasted and prepared with either plasma electrolytic or conventional mechanical polishing. Evaluation of the polishing methods was obtained by atomic force microscopy, scanning electron microscopy and energy-dispersive X-ray spectroscopy. (3) Results: The sandblasted samples showed the highest surface roughness (Heraenium® Sun 991 ± 288 nm; Wironit® 1187 ± 331 nm). Our results show that with plasma electrolytic polishing, Co-Cr-Mo surfaces can be polished with a surface roughness in the nanometer range, comparable to those achieved by conventional mechanical polishing. Conventional mechanical polishing (Heraenium® Sun 134 ± 23 nm; Wironit® 114 ± 11 nm) provided lower surface roughness values compared to plasma electrolytic polishing (Heraenium® Sun 288 ± 94 nm; Wironit® 261 ± 49 nm). We anticipate our pilot study as a starting point for future studies to refine process parameters and quantitative microbiological assays. (4) Conclusions: Plasma electrolytic polishing might have a promising future for polishing dental alloys.

Chronic oxidative stress adaptation in head and neck cancer cells generates slow-cyclers with decreased tumour growth in vivo.

BACKGROUND: Reactive oxygen species (ROS) are implicated in cancer therapy and as drivers of microenvironmental tumour cell adaptations. Medical gas plasma is a multi-ROS generating technology that has been shown effective for palliative tumour control in head and neck cancer (HNC) patients before tumour cells adapted to the oxidative stress and growth regressed fatally. METHODS: In a bedside-to-bench approach, we sought to explore the oxidative stress adaptation in two human squamous cell carcinoma cell lines. Gas plasma was utilised as a putative therapeutic agent and chronic oxidative stress inducer. RESULTS: Cellular responses of single and multiple treated cells were compared regarding sensitivity, cellular senescence, redox state and cytokine release. Whole transcriptome analysis revealed a strong correlation of cancer cell adaption with increased interleukin 1 receptor type 2 (IL1R2) expression. Using magnetic resonance imaging, tumour growth and gas plasma treatment responses of wild-type (WT) and repeatedly exposed (RE) A431 cells were further investigated in a xenograft model in vivo. RE cells generated significantly smaller tumours with suppressed inflammatory secretion profiles and increased epidermal growth factor receptor (EGFR) activity showing significantly lower gas plasma sensitivity until day 8. CONCLUSIONS: Clinically, combination treatments together with cetuximab, an EGFR inhibitor, may overcome acquired oxidative stress resistance in HNC.

Impact of dental restorations and removable prostheses on potentially malignant oral mucosal disorders in the general population.

STATEMENT OF PROBLEM: Dental restorations and removable dental prostheses have been considered as risk factors for potentially malignant disorders of the oral mucosa. It remains unclear whether amalgam, composite resins, and prosthesis materials can induce potentially malignant disorders. PURPOSE: The purpose of this clinical study was to determine the relationship between the presence of amalgam and composite resin restorations, crowns and fixed partial dentures, and removable prostheses in potentially malignant disorders. MATERIAL AND METHODS: The data of 6041 participants in the population-based Studies of Health in Pomerania (SHIP) were accessed. Potentially malignant disorders had been clinically diagnosed by calibrated dentists and documented with photographs. Dental treatment was subdivided into restored and replaced teeth. Dental restorations were subclassified as buccal composite resin or amalgam restorations. Prosthetic treatment was subclassified into removable partial or complete prostheses and definitive restorations with crowns and fixed partial dentures. RESULTS: In the maxilla, participants with removable prostheses had a higher incidence of potentially malignant disorders than participants not undergoing treatment with removable prostheses (OR 2.12; 95% CI: 1.08-4.18), but not in the mandible (OR 1.30; 95% CI: 0.67-2.53). The surfaces with composite resin restorations were associated with a slightly higher risk of mucosal lesions than those without the restorations (OR 1.04; 95% CI: 1.01-1.07). No significant association was found between amalgam restorations and mucosal lesions. CONCLUSIONS: Participants with removable prostheses have a higher risk of potentially malignant disorders. Composite resin restorations are associated with a higher risk of mucosal lesions, whereas no significant association was found between amalgam restorations and mucosal lesions.

Conductivity augments ROS and RNS delivery and tumor toxicity of an argon plasma jet.

Gas plasma jet technology was recently identified as a potential adjuvant in the palliation of cancer patients. However, a practical point raised is if higher therapeutic efficacy is achieved with the gas plasma applied in direct contact to the tumor tissue (conducting) or during treatment with the remote cloud of reactive oxygen and nitrogen species (ROS/RNS) being expelled. In a bedside-to-bench study, this clinical question was translated into studying these two distinct treatment modalities using a three-dimensional tumor cell-matrix-hydrogel assay with subsequent quantitative confocal imaging. Z-resolved fluorescence analysis of two cancer cell lines revealed greater toxicity of the conducting mode. This result was re-iterated in the growth analysis of vascularized tumor tissue cultured on chicken embryos' CAM using in ovo bioluminescence imaging. Furthermore, for conducting compared to free mode, optical emission spectroscopy revealed stronger RNS signal lines in the gas phase, while both ROS/RNS deposition in the liquid was drastically exacerbated in the conducting mode. Altogether, our results are vital in understanding the importance of standardized treatment distances on the therapeutic efficacy of gas plasma exposure in clinical oncology and will help to give critical implications for clinicians involved in plasma onco-therapy in the future.

Prevalence and risk factors of potentially malignant disorders of the mucosa in the general population: Mucosa lesions a general health problem?

AIM: Oral cancer mostly develops from oral mucosa regions with morphological alterations transforming malignant. These visible precancerous mucosa lesions are named potentially malignant disorders (PMD). We aimed to analyze the prevalence of PMD and its risk factors for PMD in a population-based sample in Northern Germany. MATERIAL AND METHODS: Data of 6078 individuals from the population-based Study of Health in Pomerania (SHIP) was used. PMD were photographically documented and periodontal health was assessed in a standardized procedure. RESULTS: PMD were observed in 54 individuals (0.9%). The most prevalent PMD was homogenous leukoplakia (n = 37) followed by Lichen ruber (n = 9). Smoking (Odds Ratio (OR) 2.70; 95% confidence interval (CI): 1.24-5.87), male sex (OR 3.32; 95%-CI: 1.77-6.21), type 2 diabetes mellitus (OR 2.07; 95%-CI: 1.08-3.98) and body mass index (OR 1.09; CI 1.04-1.14) were significantly associated with PMD, with the corresponding area under the curve (AUC) being 0.696 (CI: 0.655; 0.737). CONCLUSION: Our results suggest a clinically relevant prevalence of PMD in the population. We identified male sex, type 2 diabetes mellitus, current smoking, and obesity as risk factors. We recommend photographic documentation and intensified training of medical and dental staff to detect and monitor PMD.

Tumor cell metabolism correlates with resistance to gas plasma treatment: The evaluation of three dogmas.

Gas plasma is a partially ionized gas increasingly recognized for targeting cancer. Several hypotheses attempt to explain the link between plasma treatment and cytotoxicity in cancer cells, all focusing on cellular membranes that are the first to be exposed to plasma-generated reactive oxygen species (ROS). One proposes high levels of aquaporins, membrane transporters of water and hydrogen peroxide, to mark tumor cell line sensitivity to plasma treatment. A second focuses on membrane-expression of redox-related enzymes such as NADPH oxidases (NOX) that may modify or amplify the effects of plasma-derived ROS, fueling plasma-induced cancer cell death. Another hypothesis is that the decreased cholesterol content of tumor cell membranes sensitizes these to plasma-mediated oxidation and subsequently, cytotoxicity. Screening 33 surface molecules in 36 tumor cell lines in correlation to their sensitivity to plasma treatment, the expression of aquaporins or NOX members could not explain the sensitivity but were rather associated with treatment resistance. Correlation with transporter or enzyme activity was not tested. Analysis of cholesterol content confirmed the proposed positive correlation with treatment resistance. Strikingly, the strongest correlation was found for baseline metabolic activity (Spearman r = 0.76). Altogether, these data suggest tumor cell metabolism as a novel testable hypothesis to explain cancer cell resistance to gas plasma treatment for further elucidating this innovative field's chances and limitations in oncology.

Effects of cold physical plasma on oral lichen planus: An in vitro study (Effects of CAP on OLP).

OBJECTIVES: In the search for more effective and safe treatment avenues, we investigated cold physical plasma as a new treatment modality for therapy of oral lichen planus (OLP). MATERIAL AND METHODS: Healthy and diseased human mucosal tissue samples with a size of 3 mm in diameter obtained from OLP patients were subjected to plasma treatment ex vivo or were left untreated. Tissue sections were quantified for immune-infiltration of CD4+ , CD8+ , CD45RA+ , and CD45R0+ T cells. Moreover, the tissues' inflammatory profile was assessed by analyzing 12 different cytokines in the surrounding media. RESULTS: A significantly increased infiltrate of CD8+ and CD45-R0+ T cells was detected in OLP tissue samples when compared to healthy tissue. A higher concentration of interleukin (IL) 1ß, IL6, IL8, and granulocyte macrophage-colony stimulating factor (GM-CMF) was detected in OLP samples compared to healthy mucosal tissue. For all cytokines and chemokines investigated, 23 out of 24 comparisons showed a decrease in tendency (significant for IL1ß, IL2, IL10, and GM-CSF) in response to plasma treatment. In ex vivo-treated tissue, a decrease of T-cell infiltrate in OLP lesions compared with healthy tissue was observed. CONCLUSION: Our findings suggest cold physical plasma can be a promising therapeutic option for OLP that requires further validation in vivo.

Are third molars associated with orofacial pain? Findings from the SHIP study.

OBJECTIVES: To examine the association between third molars and orofacial pain. We hypothesized that impacted third molars are a cause of orofacial pain. METHODS: Magnetic resonance images of 1808 participants from two population-based cohorts from Northeastern Germany were analysed to define the status of third molars according to the Pell and Gregory classification. A self-reported questionnaire and a clinical dental examination were used to detect chronic and acute complaints of orofacial pain, masticatory muscle pain, migraine and other types of headache. Logistic regression models were used to analyse the associations between third molar status and orofacial pain. RESULTS: Individuals with impacted third molars in the maxilla had a higher chance of chronic orofacial pain than those with erupted third molars (odds ratio 2.19; 95% CI 1.19-4.02). No such association was detected for third molars in the lower jaw. Third molars were not associated with masticatory muscle pain, migraine or other types of headache. CONCLUSIONS: Impacted maxillary third molars might be a cause of chronic orofacial pain. Thus, physicians should consider the eruption/impaction status of third molars in their decision-making process when treating patients who complain of orofacial pain.

Long-term Risk Assessment for Medical Application of Cold Atmospheric Pressure Plasma.

Despite increasing knowledge gained based on multidisciplinary research, plasma medicine still raises various questions regarding specific effects as well as potential risks. With regard to significant statements about in vivo applicability that cannot be prognosticated exclusively based on in vitro data, there is still a deficit of clinical data. This study included a clinical follow-up of five probands who had participated five years previously in a study on the influence of cold atmospheric pressure plasma (CAP) on the wound healing of CO2 laser-induced skin lesions. The follow-up included a complex imaging diagnostic involving dermatoscopy, confocal laser scanning microscopy (CLSM) and hyperspectral imaging (HSI). Hyperspectral analysis showed no relevant microcirculatory differences between plasma-treated and non-plasma-treated areas. In summary of all the findings, no malignant changes, inflammatory reactions or pathological changes in cell architecture could be detected in the plasma-treated areas. These unique in vivo long-term data contribute to a further increase in knowledge about important safety aspects in regenerative plasma medicine. However, to confirm these findings and secure indication-specific dose recommendations, further clinical studies are required.

Molecular Mechanisms of the Efficacy of Cold Atmospheric Pressure Plasma (CAP) in Cancer Treatment.

Recently, the potential use of cold atmospheric pressure plasma (CAP) in cancer treatment has gained increasing interest. Especially the enhanced selective killing of tumor cells compared to normal cells has prompted researchers to elucidate the molecular mechanisms for the efficacy of CAP in cancer treatment. This review summarizes the current understanding of how CAP triggers intracellular pathways that induce growth inhibition or cell death. We discuss what factors may contribute to the potential selectivity of CAP towards cancer cells compared to their non-malignant counterparts. Furthermore, the potential of CAP to trigger an immune response is briefly discussed. Finally, this overview demonstrates how these concepts bear first fruits in clinical applications applying CAP treatment in head and neck squamous cell cancer as well as actinic keratosis. Although significant progress towards understanding the underlying mechanisms regarding the efficacy of CAP in cancer treatment has been made, much still needs to be done with respect to different treatment conditions and comparison of malignant and non-malignant cells of the same cell type and same donor. Furthermore, clinical pilot studies and the assessment of systemic effects will be of tremendous importance towards bringing this innovative technology into clinical practice.

Does craniofacial morphology affect third molars impaction? Results from a population-based study in northeastern Germany.

OBJECTIVES: It is still not clear why impaction of third molars occurs. Craniofacial morphology and facial parameters have been discussed to be strong predictors for third molar impaction. Thus, this study aimed to investigate the effect of craniofacial morphology on erupted or impacted third molars in a German population sample. MATERIALS AND METHODS: Erupted and impacted third molars in 2,484 participants from the Study of Health in Pomerania were assessed by whole-body magnetic resonance imaging. Markers of facial morphology were determined in 619 individuals of those participants in whose 421 participants (16.7%) had at least one impacted third molar. Craniofacial morphology was estimated as linear measurements and was associated in a cross-sectional study design with impacted and erupted third molars by multinomial logistic regression models. Erupted third molars were used as reference outcome category and regression models were adjusted for age and sex. RESULTS: Maximum Cranial Width (Eurion-Eurion distance) was significantly associated with impacted third molars (RR: 1.079; 95% confidence interval 1.028-1.132). This association was even more pronounced in the mandible. Individuals with a lower total anterior facial height (Nasion-Menton distance) and a lower facial index also have an increased risk for impacted third molars in the mandible (RR 0.953; 95% confidence interval 0.913-0.996 and RR: 0.943; 95% confidence interval 0.894-0.995). No significant associations of third molar status with facial width (Zygion-Zygion distance), and sagittal cranial dimension (Nasion-Sella distance; Sella-Basion distance) were observed. CONCLUSION: Individuals with an increased maximal cranial width have a higher risk for impaction of third molars in the mandible and in the maxilla. Individuals with a lower anterior total anterior facial height and lower facial index also have an increased risk for third molars impaction in the mandible. These findings could help orthodontic dentists, oral surgeons and oral and maxillofacial surgeons in decision-making for third molars removal in their treatment. These findings highlight the necessity of an additional analysis of the maximal cranial width by the Eurion- Eurion distance.

Physical plasma-treated saline promotes an immunogenic phenotype in CT26 colon cancer cells in vitro and in vivo.

Metastatic colorectal cancer is the fourth most common cause of cancer death. Current options in palliation such as hyperthermic intraperitoneal chemotherapy (HIPEC) present severe side effects. Recent research efforts suggested the therapeutic use of oxidant-enriched liquid using cold physical plasma. To investigate a clinically accepted treatment regimen, we assessed the antitumor capacity of plasma-treated saline solution. In response to such liquid, CT26 murine colon cancer cells were readily oxidized and showed cell growth with subsequent apoptosis, cell cycle arrest, and upregulation of immunogenic cell death (ICD) markers in vitro. This was accompanied by marked morphological changes with re-arrangement of actin fibers and reduced motility. Induction of an epithelial-to-mesenchymal transition phenotype was not observed. Key results were confirmed in MC38 colon and PDA6606 pancreatic cancer cells. Compared to plasma-treated saline, hydrogen peroxide was inferiorly toxic in 3D tumor spheroids but of similar efficacy in 2D models. In vivo, plasma-treated saline decreased tumor burden in Balb/C mice. This was concomitant with elevated numbers of intratumoral macrophages and increased T cell activation following incubation with CT26 cells ex vivo. Being a potential adjuvant for HIPEC therapy, our results suggest oxidizing saline solutions to inactivate colon cancer cells while potentially stimulating antitumor immune responses.

Association Between Symptoms of Posttraumatic Stress Disorder and Signs of Temporomandibular Disorders in the General Population.

AIMS: To estimate the association between signs of temporomandibular disorders (TMD) and symptoms of posttraumatic stress disorder (PTSD) in a representative sample from the general population of northeastern Germany. METHODS: Signs of TMD were assessed with a clinical functional analysis that included palpation of the temporomandibular joints (TMJs) and masticatory muscles. PTSD was assessed with the PTSD module of the Structured Clinical Interview for the Diagnostic and Statistical Manual of Mental Disorders, ed 4. The change-in-estimate method for binary logistic regression models was used to determine the final model and control for confounders. RESULTS: After the exclusion of subjects without prior traumatic events, the sample for joint pain consisted of 1,673 participants with a median age of 58.9 years (interquartile range 24.8), and the sample for muscle pain consisted of 1,689 participants with a median age of 59.1 years (interquartile range 24.8). Of these samples, 84 participants had pain on palpation of the TMJ, and 42 participants had pain on palpation of the masticatory muscles. Subjects having clinical PTSD (n = 62) had a 2.56-fold increase in joint pain (odds ratio [OR] = 2.56; 95% confidence interval [CI]: 1.14 to 5.71, P = .022) and a 3.86-fold increase (OR = 3.86; 95% CI: 1.51 to 9.85, P = .005) in muscle pain compared to subjects having no clinical PTSD. CONCLUSION: These results should encourage general practitioners and dentists to acknowledge the role of PTSD and traumatic events in the diagnosis and therapy of TMD, especially in a period of international migration and military foreign assignments.

Alexithymia and temporomandibular joint and facial pain in the general population.

BACKGROUND: Associations of alexithymia with temporomandibular pain disorders (TMD), facial pain, head pain and migraine have been described, but the role of the different dimensions of alexithymia in pain development remained incompletely understood. OBJECTIVES: We sought to investigate the associations of alexithymia and its subfactors with signs of TMD and with facial pain, head pain and migraine in the general population. METHODS: A total of 1494 subjects from the general population completed the Toronto Alexithymia Scale-20 (TAS-20) and underwent a clinical functional examination with palpation of the temporomandibular joint and masticatory muscles. Facial pain, migraine and head pain were defined by questionnaire. A set of logistic regression analyses was applied with adjustment for age, sex, education, number of traumatic events, depressive symptoms and anxiety. RESULTS: Alexithymia was associated with TMD joint pain (Odds Ratio 2.63; 95% confidence interval 1.60-4.32 for 61 TAS-20 points vs the median of the TAS-20 score) and with facial pain severity (Odds Ratio 3.22; 95% confidence interval 1.79-5.79). Differential effects of the subfactors were discovered with difficulties in identifying feelings as main predictor for joint, facial, and head pain, and externally oriented thinking (EOT) as U-shaped and strongest predictor for migraine. CONCLUSION: Alexithymia was moderately to strongly associated with signs and symptoms of TMD. These results should encourage dental practioners using the TAS-20 in clinical practice, to screen TMD, facial or head pain patients for alexithymia and could also help treating alexithymic TMD, facial or head pain patients.

General Precautions and Safety Aspects of Facial Laser Treatment.

The current overview of laser radiation therapy options and the legal aspects of the head and neck region provides insight into the available laser hardware, discusses the indications and treatment options and risks, as well as the necessary precautions to be taken when operating a laser device. Further handling recommendations and requirements for the laser devices are drawn and the question of transferring tasks to nonmedical personnel is discussed. The reader should get an idea about the benefits and risks of using lasers in treating patients and be able to set the precise indication on the background of possibilities, adverse effects, and legal aspects.