Cold Atmospheric Plasma Triggers Apoptosis via the Unfolded Protein Response in Melanoma Cells.

Cold atmospheric plasma (CAP) describes a partially ionized gas carrying large amounts of reactive oxygen (ROS) and nitrogen species (RNS). Numerous studies reported strong antitumor activity of CAP, thus rendering it a promising approach for tumor therapy. Although several cellular mechanisms of its cytotoxicity were identified in recent years, the exact molecular effects and contributing signaling pathways are yet to be discovered. We discovered a strong activation of unfolded protein response (UPR) after CAP treatment with increased C/EBP homologous protein (CHOP) expression, which was mainly caused by protein misfolding and calcium loss in the endoplasmic reticulum. In addition, both ceramide level and ceramide metabolism were reduced after CAP treatment, which was then linked to the UPR activation. Pharmacological inhibition of ceramide metabolism resulted in sensitization of melanoma cells for CAP both in vitro and ex vivo. This study identified a novel mechanism of CAP-induced apoptosis in melanoma cells and thereby contributes to its potential application in tumor therapy.

From the Defense Health Board: Military Accessions and the Continuum of Mental Health Research.

The Defense Health Board conducted a year-long examination of mental health accession screening and related issues. In its August 2020 report, Examination of Mental Health Accession Screening: Predictive Value of Current Measures and Processes, the Board recommends a paradigm shift in how mental health impacts on readiness are understood and addressed. This shift can only occur with the development and implementation of a research plan that follows cohorts of military personnel from recruitment through their military career. The following article describes this research plan as an excerpt of the larger report.

Cold Atmospheric Plasma Changes the Amino Acid Composition of Solutions and Influences the Anti-Tumor Effect on Melanoma Cells.

Cold Atmospheric Plasma (CAP) is an ionized gas near room temperature. Its anti-tumor effect can be transmitted either by direct treatment or mediated by a plasma-treated solution (PTS), such as treated standard cell culture medium, which contains different amino acids, inorganic salts, vitamins and other substances. Despite extensive research, the active components in PTS and its molecular or cellular mechanisms are not yet fully understood. The purpose of this study was the measurement of the reactive species in PTS and their effect on tumor cells using different plasma modes and treatment durations. The PTS analysis yielded mode- and dose-dependent differences in the production of reactive oxygen and nitrogen species (RONS), and in the decomposition and modification of the amino acids Tyrosine (Tyr) and Tryptophan (Trp). The Trp metabolites Formylkynurenine (FKyn) and Kynurenine (Kyn) were produced in PTS with the 4 kHz (oxygen) mode, inducing apoptosis in Mel Im melanoma cells. Nitrated derivatives of Trp and Tyr were formed in the 8 kHz (nitrogen) mode, elevating the p16 mRNA expression and senescence-associated ß-Galactosidase staining. In conclusion, the plasma mode has a strong impact on the composition of the active components in PTS and affects its anti-tumor mechanism. These findings are of decisive importance for the development of plasma devices and the effectiveness of tumor treatment.

Acidified Nitrite Contributes to the Antitumor Effect of Cold Atmospheric Plasma on Melanoma Cells.

Cold atmospheric plasma (CAP) is partially ionized gas near room temperature with previously reported antitumor effects. Despite extensive research and growing interest in this technology, active components and molecular mechanisms of CAP are not fully understood to date. We used Raman spectroscopy and colorimetric assays to determine elevated nitrite and nitrate levels after treatment with a MiniFlatPlaster CAP device. Previously, we demonstrated CAP-induced acidification. Cellular effects of nitrite and strong extracellular acidification were assessed using live-cell imaging of intracellular Ca2+ levels, cell viability analysis as well as quantification of p21 and DNA damage. We further characterized these observations by analyzing established molecular effects of CAP treatment. A synergistic effect of nitrite and acidification was found, leading to strong cytotoxicity in melanoma cells. Interestingly, protein nitration and membrane damage were absent after treatment with acidified nitrite, thereby challenging their contribution to CAP-induced cytotoxicity. Further, phosphorylation of ERK1/2 was increased after treatment with both acidified nitrite and indirect CAP. This study characterizes the impact of acidified nitrite on melanoma cells and supports the importance of RNS during CAP treatment. Further, it defines and evaluates important molecular mechanisms that are involved in the cancer cell response to CAP.

Biological effects of a new ultraviolet A1 prototype based on light-emitting diodes on the treatment of localized scleroderma.

Ultraviolet A1 (UVA1 ) phototherapy (spectral range 340-400 nm) is a well-established treatment option for various skin diseases such as localized scleroderma. Recent improvements of conventional UVA1 light sources (metal-halide or fluorescent lamps) have brought attention to a new light-emitting diode (LED) technology with remarkable advantages in handling and clinical routine. This study provides a preclinical histological and molecular evaluation of an LED-based UVA1 prototype with a narrower spectral range (360-400 nm) for treating localized scleroderma. Scleroderma mouse models and fibroblasts in vitro were exposed to LED-based UVA1 phototherapy or to irradiation with a commercially available metal-halide lamp emitting low-dose (20, 40 J/cm2 ), medium-dose (60 J/cm2 ) and high-dose (80, 100 J/cm2 ) UVA1 light. Both UVA1 light sources affected inflammatory genes (IL-1α and IL-6) and growth factors (TGFß-1 and TGFß-2). Increased collagen type 1 was reduced after UVA1 phototherapy. Matrix metalloproteinase-1 was more enhanced after a medium dose of LED-based UVA1 phototherapy than after conventional treatment. In vivo, dermal thickness and the amount of collagen were reduced after both treatment methods. Remarkably, myofibroblasts were more effectively reduced by a medium dose of LED-based UVA1 phototherapy. The study indicates that LED-based UVA1 phototherapy yields similar or even better results than conventional treatment. In terms of biosafety and patient comfort, LED-based UVA1 phototherapy offers clear advantages over conventional treatment because of the use of a narrower and less harmful UVA1 spectrum, less heat generation and shorter treatment times at the same irradiation intensity. Clinical studies are required to confirm these results in patients with localized scleroderma.

A fully human anti-BMP6 antibody reduces the need for erythropoietin in rodent models of the anemia of chronic disease.

Recombinant erythropoietin (EPO) and iron substitution are a standard of care for treatment of anemias associated with chronic inflammation, including anemia of chronic kidney disease. A black box warning for EPO therapy and concerns about negative side effects related to high-dose iron supplementation as well as the significant proportion of patients becoming EPO resistant over time explains the medical need to define novel strategies to ameliorate anemia of chronic disease (ACD). As hepcidin is central to the iron-restrictive phenotype in ACD, therapeutic approaches targeting hepcidin were recently developed. We herein report the therapeutic effects of a fully human anti-BMP6 antibody (KY1070) either as monotherapy or in combination with Darbepoetin alfa on iron metabolism and anemia resolution in 2 different, well-established, and clinically relevant rodent models of ACD. In addition to counteracting hepcidin-driven iron limitation for erythropoiesis, we found that the combination of KY1070 and recombinant human EPO improved the erythroid response compared with either monotherapy in a qualitative and quantitative manner. Consequently, the combination of KY1070 and Darbepoetin alfa resulted in an EPO-sparing effect. Moreover, we found that suppression of hepcidin via KY1070 modulates ferroportin expression on erythroid precursor cells, thereby lowering potentially toxic-free intracellular iron levels and by accelerating erythroid output as reflected by increased maturation of erythrocyte progenitors. In summary, we conclude that treatment of ACD, as a highly complex disease, becomes more effective by a multifactorial therapeutic approach upon mobilization of endogenous iron deposits and stimulation of erythropoiesis.

Acidification is an Essential Process of Cold Atmospheric Plasma and Promotes the Anti-Cancer Effect on Malignant Melanoma Cells.

(1) Background: Cold atmospheric plasma (CAP) is ionized gas near room temperature. The anti-cancer effects of CAP were confirmed for several cancer types and were attributed to CAP-induced reactive species. However, the mode of action of CAP is still not well understood. (2) Methods: Changes in cytoplasmic Ca2+ level after CAP treatment of malignant melanoma cells were analyzed via the intracellular Ca2+ indicator fura-2 AM. CAP-produced reactive species were determined by fluorescence spectroscopic and protein nitration by Western Blot analysis. (3) Results: CAP caused a strong acidification of water and solutions that were buffered with the so-called Good buffers, while phosphate-buffered solutions with higher buffer capacity showed minor pH reductions. The CAP-induced Ca2+ influx in melanoma cells was stronger in acidic pH than in physiological conditions. NO formation that is induced by CAP was dose- and pH-dependent and CAP-treated solutions only caused protein nitration in cells under acidic conditions. (4) Conclusions: We describe the impact of CAP-induced acidification on the anti-cancer effects of CAP. A synergistic effect of CAP-induced ROS, RNS, and acidic conditions affected the intracellular Ca2+ level of melanoma cells. As the microenvironment of tumors is often acidic, further acidification might be one reason for the specific anti-cancer effects of CAP.

BMP6-induced modulation of the tumor micro-milieu.

Melanoma is the deadliest form of skin cancer with rising incidence, creating a significant health problem. We discovered increased expression of bone morphogenetic protein 6 (BMP6) in melanoma cells and tissues, and observed that BMP6 deficiency caused significantly delayed tumor onset and decelerated tumor progression in a melanoma mouse model. Moreover, we determined that BMP6 inhibits dermal mast cell recruitment and found that mast cell-derived mediators significantly reduced melanoma growth in vitro. In line with this, mast cell deficiency accelerated tumor onset and progression in a melanoma mouse model. Analysis of human melanoma tissues revealed a strong negative correlation between melanoma proliferation and mast cell infiltration. This study elucidates a novel role of BMP6-induced modulation of the tumor microenvironment.

Cold atmospheric plasma causes a calcium influx in melanoma cells triggering CAP-induced senescence.

Cold atmospheric plasma (CAP) is a promising approach in anti-cancer therapy, eliminating cancer cells with high selectivity. However, the molecular mechanisms of CAP action are poorly understood. In this study, we investigated CAP effects on calcium homeostasis in melanoma cells. We observed increased cytoplasmic calcium after CAP treatment, which also occurred in the absence of extracellular calcium, indicating the majority of the calcium increase originates from intracellular stores. Application of previously CAP-exposed extracellular solutions also induced cytoplasmic calcium elevations. A substantial fraction of this effect remained when the application was delayed for one hour, indicating the chemical stability of the activating agent(s). Addition of ryanodine and cyclosporin A indicate the involvement of the endoplasmatic reticulum and the mitochondria. Inhibition of the cytoplasmic calcium elevation by the intracellular chelator BAPTA blocked CAP-induced senescence. This finding helps to understand the molecular influence and the mode of action of CAP on tumor cells.

Cold atmospheric plasma treatment inhibits growth in colorectal cancer cells.

Plasma oncology is a relatively new field of research. Recent developments have indicated that cold atmospheric plasma (CAP) technology is an interesting new therapeutic approach to cancer treatment. In this study, p53 wildtype (LoVo) and human p53 mutated (HT29 and SW480) colorectal cancer cells were treated with the miniFlatPlaSter - a device particularly developed for the treatment of tumor cells - that uses the Surface Micro Discharge (SMD) technology for plasma production in air. The present study analyzed the effects of plasma on colorectal cancer cells in vitro and on normal colon tissue ex vivo. Plasma treatment had strong effects on colon cancer cells, such as inhibition of cell proliferation, induction of cell death and modulation of p21 expression. In contrast, CAP treatment of murine colon tissue ex vivo for up to 2 min did not show any toxic effect on normal colon cells compared to H2O2 positive control. In summary, these results suggest that the miniFlatPlaSter plasma device is able to kill colorectal cancer cells independent of their p53 mutation status. Thus, this device presents a promising new approach in colon cancer therapy.

Cold atmospheric plasma (CAP) activates angiogenesis-related molecules in skin keratinocytes, fibroblasts and endothelial cells and improves wound angiogenesis in an autocrine and paracrine mode.

BACKGROUND: Cold atmospheric plasma (CAP) emerged as a novel therapeutic field with applications developed for bacterial sterilization, wound healing and cancer treatment. For clinical implementation it is important to know how CAP works and which molecular changes occur after the CAP treatment. Vascularization is an important step during wound healing, however, the effects of CAP on wound angiogenesis are not well examined so far. Furthermore, it has not been investigated, whether CAP primarily affects endothelial cells directly or via paracrine mechanisms to modulate the vasculature. OBJECTIVE: This study concentrates on the influence of CAP on angiogenesis-related molecules in human epidermal keratinocytes, dermal fibroblasts and endothelial cells. METHODS: CAP was generated by the MicroPlaSter ß® plasma torch system and CAP effects on angiogenesis were determined in vitro and in vivo. RESULTS: We observed that CAP significantly induces the expression of Artemin, EGF, EG-VEGF (PK1), Endothelin-1 (ET-1), FGF-2 (FGF basic), IL-8 (CXCL8) and uPA in keratinocytes and Angiogenin (ANG), Endostatin (Col18A1), MCP-1 (CCL2), MMP-9, TIMP-1, uPA and VEGF in fibroblasts. In addition, CAP activates the expression of Angiopoietin-2 (Ang-2), Angiostatin (PLG), Amphiregulin (AR), Endostatin, FGF-2 and angiogenic-involved receptor expression of FGF R1 and VEGF R1 in HUVEC endothelial cells. It was also demonstrated that supernatants collected from CAP activated fibroblasts and keratinocytes elevate tube formation by endothelial cells and FGF-2 appears to be an important pro-angiogenic factor that controls vascularization via paracrine mechanisms. Mouse experiments supplement that CAP promotes angiogenesis during wound healing in vivo. CONCLUSIONS: Taken together, these results suggest that CAP modulates angiogenesis-involved factors via autocrine and paracrine mechanisms and may be used to affect angiogenesis during wound healing.

Broadening the cancer and cognition landscape: the role of self-regulatory challenges.

BACKGROUND: The potentially detrimental effects of cancer and related treatments on cognitive functioning have emerged as one of the key foci of cancer survivorship research, but little is known about how psychological variables other than depression influence these relationships. To illustrate the potential of social psychological perspectives, we examine how a self-regulatory analysis and specific self-regulatory challenges of contending with cancer-related expectancies and stereotypes provide conceptual frameworks for understanding some of the potential causes and consequences of cancer-related cognitive deficits. METHODS: Literatures on cancer-related cognitive deficits, self-regulatory ego depletion, expectancy stereotypes, and their points of convergence are briefly reviewed. RESULTS: A review and conceptual integration of relevant literatures suggest that coping with cancer can impair self-regulatory capacity. There is an overlap between cognitive deficits associated with self-regulatory challenge and with cancer and its treatment, and restoring self-regulatory resources can attenuate cancer-related cognitive deficits. Examination of specific regulatory challenges of contending with expectancies and stereotypes related to treatment suggests insights that can inform when and among whom cognitive deficits may most likely emerge. CONCLUSIONS: Integrating social psychological ideas with a substantial knowledge base can illustrate novel research trajectories that can deepen our understanding of cancer-related cognitive deficits and their impact on psychosocial well-being.

Cold atmospheric plasma, a new strategy to induce senescence in melanoma cells.

Over the past few years, the application of cold atmospheric plasma (CAP) in medicine has developed into an innovative field of research of rapidly growing importance. One promising new medical application of CAP is cancer treatment. Different studies revealed that CAP may potentially affect the cell cycle and cause cell apoptosis or necrosis in tumor cells dependent on the CAP device and doses. In this study, we used a novel hand-held and battery-operated CAP device utilizing the surface micro discharge (SMD) technology for plasma production in air and consequently analysed dose-dependent CAP treatment effects on melanoma cells. After 2 min of CAP treatment, we observed irreversible cell inactivation. Phospho-H2AX immunofluorescence staining and Flow cytometric analysis demonstrated that 2 min of CAP treatment induces DNA damage, promotes induction of Sub-G1 phase and strongly increases apoptosis. Further, protein array technology revealed induction of pro-apoptotic events like p53 and Rad17 phosphorylation of Cytochrome c release and activation of Caspase-3. Interestingly, using lower CAP doses with 1 min of treatment, almost no apoptosis was observed but long-term inhibition of proliferation. H3K9 immunofluorescence, SA-ß-Gal staining and p21 expression revealed that especially these low CAP doses induce senescence in melanoma cells. In summary, we observed differences in induction of apoptosis or senescence of tumor cells in respond to different CAP doses using a new CAP device. The mechanism of senescence with regard to plasma therapy was so far not described previously and is of great importance for therapeutic application of CAP.

fussel (fuss)--A negative regulator of BMP signaling in Drosophila melanogaster.

The TGF-ß/BMP signaling cascades control a wide range of developmental and physiological functions in vertebrates and invertebrates. In Drosophila melanogaster, members of this pathway can be divided into a Bone Morphogenic Protein (BMP) and an Activin-ß (Act-ß) branch, where Decapentaplegic (Dpp), a member of the BMP family has been most intensively studied. They differ in ligands, receptors and transmitting proteins, but also share some components, such as the Co-Smad Medea (Med). The essential role of Med is to form a complex with one of the two activating Smads, mothers against decapentaplegic (Mad) or dSmad, and to translocate together to the nucleus where they can function as transcriptional regulators of downstream target genes. This signaling cascade underlies different mechanisms of negative regulation, which can be exerted by inhibitory Smads, such as daughters against decapentaplegic (dad), but also by the Ski-Sno family. In this work we identified and functionally analyzed a new member of the Ski/Sno-family, fussel (fuss), the Drosophila homolog of the human functional suppressing element 15 (fussel-15). fuss codes for two differentially spliced transcripts with a neuronal expression pattern. The proteins are characterized by a Ski-Sno and a SAND homology domain. Overexpression studies and genetic interaction experiments clearly reveal an interaction of fuss with members of the BMP pathway, leading to a strong repression of BMP-signaling. The protein interacts directly with Medea and seems to reprogram the Smad pathway through its influence upon the formation of functional Mad/Medea complexes. This leads amongst others to a repression of downstream target genes of the Dpp pathway, such as optomotor blind (omb). Taken together we could show that fuss exerts a pivotal role as an antagonist of BMP signaling in Drosophila melanogaster.

Stress, coping and cognitive deficits in women after surgery for breast cancer.

Research on neuropsychological difficulties among cancer patients has focused on chemotherapy as a primary cause, yet several studies have now shown that some patients evidence cognitive weaknesses prior to chemotherapy. As an alternative to the 'chemo-brain' theory, this study examined the hypothesis that stress and coping style may be associated with observed neuropsychological difficulties among female cancer patients. Thirty-six women completed neuropsychological testing and psychological questionnaires following surgery for breast cancer and prior to any subsequent treatments. Twenty-seven percent of participants evidenced deficits on at least one measure of verbal fluency, and 14% of participants were impaired on at least one memory measure. Self-reported stress was correlated with deficits in memory, verbal fluency, and attention. Subsequent mediational analyses indicated that use of passive coping styles may underlie this relationship between stress and neuropsychological deficits. These findings highlight the potential relevance of psychological mechanisms, such as coping style, in cancer patients' experience of neuropsychological deficits.

Considering the unspoken: the role of death cognition in quality of life among women with and without breast cancer.

This study examined how breast cancer diagnosis influences underlying cognitions and explicit worries about death, and their roles in health-related quality of life (QOL). Forty-two women who underwent surgery for the removal of either a cancerous or benign breast mass indicated their worries about dying and completed measures of death-thought accessibility and QOL. Women with cancer reported lowered physical, emotional, and functional well-being. Further, although they did not differ in explicit worry about death, women with cancer (compared to those with a benign mass) evidenced greater death-thought accessibility, which in turn mediated the effect of cancer diagnosis on well-being.

Tai Chi effects on neuropsychological, emotional, and physical functioning following cancer treatment: a pilot study.

OBJECTIVE: To examine the effects of a 10-week Tai Chi (TC) program on neuropsychological, psychological, and physical health of female cancer survivors. DESIGN: Twenty-three women with a history of cancer participated in 60-min TC classes two times/week for 10-weeks. MAIN OUTCOME MEASURES: Before and after the intervention, participants completed neuropsychological tests (memory, executive functioning, language, and attention); 5 tests of balance; and self-report questionnaires of neuropsychological complaints, stress and mood, and fatigue. RESULTS: After the 10-week session, participants evidenced fewer neuropsychological complaints and enhanced neuropsychological functioning. They also demonstrated improved balance and reported better psychological functioning. CONCLUSIONS: Results suggest that TC may promote gains in neuropsychological functioning, in addition to previously demonstrated improvements in physical and psychological health. These findings support the need for controlled trials examining the potential benefits of TC on neuropsychological functioning after cancer.

Validation of WAIS-III four-subtest short forms in patients with traumatic brain injury.

In an effort to identify four-subtest Wechsler Adult Intelligence Scale-Third Edition (WAIS-III) short forms valid for estimating Full-Scale IQ (FSIQ) among individuals with traumatic brain injury (TBI), seven tetrad versions of the WAIS-III were evaluated in a convenience sample of patients referred for neuropsychological assessment (n = 176). Estimated FSIQ scores were compared to actual FSIQ scores via correlation analyses, repeated-measures analyses of variance (ANOVAs), and frequency analyses. All short form-estimated FSIQ scores correlated highly with actual scores (all rs > .91, ps < .001). Repeated-measures ANOVAs identified no significant differences between actual and short form-estimated FSIQ scores for two of the seven short forms. These same two short forms had the highest percentage of scores within ±5 points of actual FSIQ scores (75.6% and 71.6%). Thus, two tetrad versions were consistently superior to others in accuracy of estimating FSIQ; these may be helpful when time constraints or other issues necessitate use of an abbreviated battery for estimating FSIQ among individuals with TBI.

Downregulation of hemojuvelin prevents inhibitory effects of bone morphogenetic proteins on iron metabolism in hepatocellular carcinoma.

Recently, we revealed that bone morphogenetic protein (BMP) 4 is increased in hepatocellular carcinoma (HCC). Furthermore, latest reports described BMPs, in particular BMP6, as important regulators of hepcidin expression in iron homeostasis. Therefore, we aimed to unravel why enhanced BMP expression in HCC patients does not lead to severe changes in iron metabolism. Initial analysis of the BMP4 and BMP6 expression patterns revealed enhanced expression on mRNA and protein level in HCC cell lines and tissue samples compared with primary human hepatocytes (PHHs) and normal liver tissues. However and interestingly, hepcidin expression was reduced in HCC cell lines and tissues. Analysis of BMP6 receptor expression revealed loss of BMP6-specific receptor subunit in HCC. To identify a possible regulatory mechanism causing lack of reaction to BMP4 we analyzed the expression of hemojuvelin (HJV), which is involved in iron metabolism as BMP co-receptor. HJV expression was markedly decreased in HCC cell lines and tissues. HJV promoter analysis revealed potential HNF-1α and snail-binding sites, but functional analysis ruled out that these transcriptional regulators or promoter methylation are the cause of HJV downregulation in HCC. However, we identified AU-rich elements in the HJV 3'-untranslated region and revealed significantly faster decay of HJV mRNA in HCC cells as compared with PHH indicating decreased mRNA-stability as the reason for the loss of HJV expression in HCC.

Does rurality affect quality of life following treatment for breast cancer?

PURPOSE: The present research examined the extent to which rural residence and social support seeking are associated with quality of life (QOL) among breast cancer patients following chemotherapy. METHODS: Female breast cancer patients (n = 46) from communities of varying degrees of rurality in a Midwestern state completed psychological and QOL measures at 1-month postchemotherapy. Analyses assessed the relationships between QOL outcomes, rurality, and social support seeking. FINDINGS: Using age and education as covariates, regression analyses were conducted to determine the extent to which QOL was related to social support seeking and rural/urban residence. Analyses revealed that social support seeking was associated with lower scores on multiple indices of QOL, and it was associated with higher self-reported symptoms of depression. Several significant associations with rural/urban residence were noted as well. Specifically, increasing rurality, as defined by USDA Rural-Urban continuum codes, was associated with lower overall QOL, lower functional well-being, and increased complaints of breast cancer specific symptoms. CONCLUSIONS: These findings highlight the relevance of continued efforts to address social support needs among women with a history of breast cancer living in rural and urban communities. They also suggest that individuals in more rural communities may be at risk for lowered QOL in the early period following cancer treatment. Future research is needed to replicate these results with larger and more diverse samples of rural and urban dwelling individuals, and to determine whether these effects may be attributed to identifiable characteristics of rural communities (eg, fewer cancer-related resources).