No abstract

Non-thermal atmospheric-pressure plasmas have been developed that will be used in future for several purposes, e.g. medicine. Living tissues and cells are at the focus of plasma treatment, e.g. to improve wound healing, or induce apoptosis and growth arrest in tumour cells. Detailed investigations of plasma-cell interactions are needed. Cell surface adhesion molecules as integrins, cadherins or the EGFR (epidermal growth factor receptor) are of importance in wound healing and also for development of cancer metastasis. This study has focused on measurement of cell surface molecules on human HaCaT keratinocytes (human adult low calcium temperature keratinocytes) promoting adhesion, migration and proliferation as one important feature of plasma-cell interactions. HaCaT keratinocytes were treated with plasma by a surface dielectric barrier discharge in air. Cell surface molecules and induction of intracellular ROS (reactive oxygen species) were analysed by flow cytometry 24 h after plasma treatment. Besides a reduction of cell viability a significant down-regulation of E-cadherin and the EGFR expression occurred. The influence on α2- and β1-integrins was less pronounced, and expression of ICAM-1 (intercellular adhesion molecule 1) was unaffected. The extent of effects depended on the exposure time of cells to the plasma and the treatment regimen. Intracellular level of ROS detected by the fluorescent dye H2DCFDA (2',7'-dichlorodihydrofluorescein diacetate) increased by plasma treatment, but it was neither dependent on the treatment time nor related to the different treatment regimens. Two-dimensional cultures of HaCaT keratinocytes appear to be a suitable method of investigating plasma-cell interactions.

The application of non-thermal atmospheric pressure plasma in medicine, especially in dermatology, is a very promising approach to improve the treatment of chronic wounds. This application has to be effective as well as safe. That is why it is important to understand the effects of plasma on living cells and on the genetic information. Subject of this research was the investigation of the influence of non-thermal atmospheric surface DBD plasma (working gas: air and argon) on the DNA of human keratinocytes. Adherent HaCaT cells were plasma-treated for 5, 10 and 20 min. DNA damage was measured subsequently as well as 24h after treatment using Alkaline Single-Cell Gel Electrophoresis. Further, cell number and induction of reactive oxygen species (ROS) were determined. Immediately after plasma treatment a correlation between duration of plasma treatment and DNA damage level was observed. 24h after plasma exposition (5 and 10 min), the DNA damage showed the same level as that of control cells although the number of adherent cells decreased. Plasma treatment of 20 min resulted in a permanent DNA damage. Probably one of the more important causes of DNA damage is the presence of ROS. Therefore, the H2DCFDA assay was used to indicate intracellular oxidative stress. Plasma treatment caused a time dependent increase of ROS. In conclusion, directly after plasma treatment DNA damage was observed in HaCaT cells, which was not longer evident 24h later. Whether this observation reflects DNA repair in damaged single cells or a recovery of the cell population remains to be clarified. Treatment times of 20 min induced irreparable damage. In addition to ROS it is also believed that other factors (e.g. RNS, UV light) are responsible for DNA damage. So there is still a need for further research into the cause of DNA damage and recovery after plasma treatment.*