[Cold atmospheric pressure plasma for the treatment of acute and chronic wounds]. / Kaltes Atmosphärendruckplasma zur Behandlung akuter und chronischer Wunden.

BACKGROUND: Besides acute wounds (through trauma or surgical interventions), chronic wounds comprise a relatively large and heterogeneous group of diseases. These include leg ulcers with venous disease greatly prevailing arterial disease, diabetic foot syndrome, and pressure ulcers. Due to a considerable treatment resistance against such therapies, new and effective, additive treatment options especially for chronic wounds are needed. Wound treatment with cold atmospheric plasma (CAP) constitutes such an innovative option. OBJECTIVES: Current research regarding the efficacy of cold plasma for healing of acute and chronic wounds is summarized. MATERIALS AND METHODS: The literature on CAP applications in wound healing has been screened and reviewed. RESULTS: With CAP, several effects that promote wound healing can be simultaneously applied in one application. On the one hand, CAP exerts a strong and broad antimicrobial activity against biofilm. On the other hand, the plasma cocktail, which consists of reactive nitrogen and oxygen species, UV, and charged particles (electrical current), mediates tissue-stimulating, blood flow-promoting, and anti-inflammatory effects. Marked germ reduction on wounds and accelerated wound healing have already been convincingly demonstrated in controlled clinical studies. CONCLUSIONS: The comprehensive CAP study landscape with structured case report summaries and randomized case-control studies allows the conclusion that CAP is safe, effective, and easy to handle for wound treatment. The utilization of CAP in addition to standard wound treatments is starting to enter routine clinical practice.

Induction of proliferation of basal epidermal keratinocytes by cold atmospheric-pressure plasma.

BACKGROUND: Over the past few decades, new cold plasma sources have been developed that have the great advantage of operating at atmospheric pressure and at temperatures tolerable by biological material. New applications for these have emerged, especially in the field of dermatology. Recently it was demonstrated that cold atmospheric-pressure plasma positively influences healing of chronic wounds. The potential of cold plasma lies in its capacity to reduce bacterial load in the wound while at the same time stimulating skin cells and therefore promoting wound closure. In recent years, there have been great advances in the understanding of the molecular mechanisms triggered by cold plasma involving signalling pathways and gene regulation in cell culture. AIM: To investigate cold plasma-induced effects in ex vivo treated human skin biopsies. METHODS: Human skin tissue was exposed to cold plasma for different lengths of time, and analysed by immunofluorescence with respect to DNA damage, apoptosis, proliferation and differentiation markers. RESULTS: After cold plasma treatment, the epidermal integrity and keratin expression pattern remained unchanged. As expected, the results revealed an increase in apoptotic cells after 3 and 5 min of treatment. Strikingly, an induction of proliferating basal keratinocytes was detected after cold plasma exposure for 1 and 3 min. As these are the cells that regenerate the epidermis, this could indeed be beneficial for wound closure. CONCLUSION: We investigated the effect of cold plasma on human skin by detecting molecules for growth and apoptosis, and found that both processes are dependent on treatment time. Therefore, this approach offers promising results for further applications of cold plasma in clinical dermatology.

Diabetogenic glucose and insulin concentrations modulate transcriptome and protein levels involved in tumour cell migration, adhesion and proliferation.

BACKGROUND: During the last decade, epidemiological studies uncovered the tremendous impact of metabolic syndrome/diabetes mellitus type 2 (DM T2) as risk factors of the progression of cancer. Therefore, we studied the impact of diabetogenic glucose and insulin concentrations on the activities of tumour cells, because little is known about how high glucose and insulin levels are influencing gene activities causing changes in the signal cascade activities with respect to kinases involved in the proliferation and migration of cancer cells. METHODS: To address this question we analysed the activity of more than 400 gene signatures related to (i) cell cycle, (ii) cell movement as well as (iii) signal transduction. We examined transcriptomes of kinases (PKCα, PI3K), cadherins (E-, N- VE-), integrins and cyclins by comparing physiological (5.5 mM) vs diabetogenic (11 mM) glucose concentrations (without and with insulin). RESULTS: Proliferation assays revealed that high levels of glucose (11 mM) and insulin (100 ng ml(-1)) did promote the proliferation of the tumour cell lines HT29, SW480, MCF-7, MDA MB468, PC3 and T24. Using a 3D-migration assay, we have shown that high glucose concentrations caused increased motility rates of the tumour cells. The increase in migratory activity at high glucose and insulin concentrations was mediated by an activation of PI3K, PKCα and MLCK, as figured out by the pharmacological inhibitors wortmannin, Go6976 and ML-7. CONCLUSION: We present molecular and functional data, which could help to understand how hyperglycaemia and hyperinsulinemia might trigger tumour cell proliferation and motility in patients, too.

High PKC alpha and low E-cadherin expression contribute to high migratory activity of colon carcinoma cells.

The protein kinase C (PKC) is a family of serine/threonine kinases that are key regulatory enzymes involved in growth, differentiation, cytoskeletal reorganization, tumor promotion, and migration. We investigated the functional involvement of PKC isotypes and of E-cadherin in the regulation of the locomotion of six human colon-adenocarcinoma cell lines. The different levels of the PKC alpha and the E-cadherin expression have predictable implications in the spontaneous locomotory activity. With the use of PKC alpha--specific inhibitors (safingol, Go6976) as well as the PKC delta--specific inhibitor rottlerin, we showed that only PKC alpha plays a major role in the regulation of tumor cell migration. The results were verified by knocking out the translation of PKC isozymes with the use of an antisense oligonucleotide strategy. After stimulation with phorbol ester we observed a translocation and a colocalization of the activated PKC alpha at the plasma membrane to the surrounding extracellular matrix. Furthermore, we investigated the functional involvement of E-cadherin in the locomotion with the use of a blocking antibody. A high level of PKC alpha expression together with a low E-cadherin expression was strongly related to a high migratory activity of the colon carcinoma cells. This correlation was independent of the differentiation grade of the tumor cell lines.

DPPIV inhibitors extend GLP-2 mediated tumour promoting effects on intestinal cancer cells.

BACKGROUND: The glucagon-like peptides-1 and -2 (GLP-1 and -2) are co-secreted after food intake from intestinal L cells. Since both peptides are rapidly degraded by dipeptidyl peptidase-IV (DPPIV), research is focused on the development of DPPIV inhibitors or DPPIV resistant. AIMS: In this study we investigated, whether the inhibition of DPPIV activity and the resulting increased half-life of DPPIV substrates may influence cancer development and progression. METHODS: We examined proliferation and migratory activity of two human colon cancer cell lines (SW480, HT29) after stimulation with GLP-2 in combination with or without DPPIV inhibitors. RESULTS: Migratory activity was increased by 25% from 20% matrix induced activity to a maximum of 45% (100 nM GLP-2). In cells expressing CD26, migration was prolonged by addition of DPPIV inhibitors in a concentration dependent manner. After treatment with GLP-2 doubling time decreased from 2.4 to 1.5 days - and addition of DPPIV inhibitors enhanced the effect of GLP-2. CONCLUSIONS: The use of DPPIV inhibitors together with GLP-2 led to increased proliferation as well as elevated migratory activity. Therefore, the use of DPPIV inhibitors could increase the risk of promoting an already existing intestinal tumour and may support the potential of colon cancer cells to metastasize.

Norepinephrine-induced migration of SW 480 colon carcinoma cells is inhibited by beta-blockers.

Beta-adrenoceptors are highly expressed on SW 480 colon carcinoma cells as was assessed by flow cytometry. We investigated the influence of norepinephrine on the migration of these cells using time-lapse videomicroscopy. Norepinephrine-treatment increased the locomotor activity within the population from 25% spontaneously locomoting cells to 65% locomoting cells. The beta1/2-blocker propranolol but not the beta1-blocker atenolol inhibited this increase. The intracellular signaling solely of norepinephrine-induced locomotion involved protein tyrosine kinase activity, whereas both spontaneous and norepinephrine-induced migration were reduced by inhibiting phospholipase Cgamma and protein kinase Calpha activity. In summary, norepinephrine-induced locomotion of SW 480 cells is beta2-adrenoceptor mediated and distinct from spontaneous locomotion concerning the PTK involvement.

Risk assessment of a cold argon plasma jet in respect to its mutagenicity.

Cold atmospheric pressure plasmas represent a favorable option for the treatment of heat sensitive materials and human or animal tissue. Beneficial effects have been documented in a variety of medical conditions, e.g., in the treatment of chronic wounds. It is assumed that the main mechanism of the plasma's efficacy is mediated by a stimulating dissipation of energy via radiation and/or chemical energy. Although no evidence on undesired side effects of a plasma treatment has yet been presented, skepticism toward the safety of the exposure to plasma is present. However, only little data regarding the mutagenic potential of this new treatment option is available. Accordingly, we investigated the mutagenic potential of an argon plasma jet (kinpen) using different testing systems in accordance with ISO norms and multiple cell lines: a HPRT1 mutation assay, a micronucleus formation assay, and a colony formation assay. Moderate plasma treatment up to 180 s did not increase genotoxicity in any assay or cell type investigated. We conclude that treatment with the argon plasma jet kinpen did not display a mutagenic potential under the test conditions applied and may from this perspective be regarded as safe for the use in biomedical applications.

Hydrogen peroxide: A central player in physical plasma-induced oxidative stress in human blood cells.

Plasma medicine is an interdisciplinary field and recent clinical studies showed benefits of topical plasma application to chronic wounds. Whereas most investigations have focused on plasma-skin cell interaction, immune cells are omnipresent in most tissues as well. They not only elicit specific immune responses but also regulate inflammation, which is central in healing and regeneration. Plasma generates short-lived radicals and species in the gas phase. Mechanisms of plasma-cell interactions are not fully understood but it is hypothesized that reactive oxygen and nitrogen species (RONS) mediate effects of plasma on cells. In this study human blood cells were investigated after cold atmospheric plasma treatment with regard to oxidation and viability. Plasma generates hydrogen peroxide (H2O2) and the responses were similar in cells treated with concentration-matched H2O2. Both treatments gave an equivalent reduction in viability and this was completely abrogated if catalase was added prior to plasma exposure. Further, five oxidation probes were utilized and fluorescence increase was observed in plasma-treated cells. Dye-dependent addition of catalase diminished most but not all of the probe fluorescence, assigning H2O2 a dominant but not exclusive role in cellular oxidation by plasma. Investigations for other species revealed generation of nitrite and formation of 3-nitrotyrosine but not 3-chlorotyrosine after plasma treatment indicating presence of RNS which may contribute to cellular redox changes observed. Together, these results will help to clarify how oxidative stress associates with physical plasma treatment in wound relevant cells.