LAUSR

Introduction Considering the increased need for alternatives to current treatments, a new therapy based on plasma, the fourth state of matter, has recently raised the medical community’s attention. The aim of this work was to evaluate the effect, selectivity and mechanisms of action of cold atmospheric plasma (CAP) in a human retinoblastoma cell line. Material and methods An electronic device was designed to generate CAP, in open air above multiwell plates where Y79 cell cultures were seeded. Plasma emission spectrum was captured by a spectrometer. In order to evaluate the cytotoxicity and selectivity of CAP, metabolic activity of similarly treated Y79 and human fibroblasts HFF1 cells was measured. Apoptosis detection, analysis of mitochondrial membrane potential (MMP) and cell morphology were studied to determine the type of cell death. Propidium-iodide/RNAse staining was used to study the cell cycle and genotoxic effects were assessed by comet assay. Oxygen and nitrogen reactive species (RS) and oxidative defenses were measured. In order to explore the interaction of the electric field with voltage-gated calcium channels, blockade with verapamil was applied. Clonogenic assay screened for long term survival. Results and discussions After 60 s of CAP treatment, the metabolic activity of Y79 cells decreased more than 50%, mostly due to apoptosis, while HFF1 endured viable. Cell survival was shortened. Accumulation of Y79 cells in S and G2/M phases was recorded, nevertheless, no DNA strand breaks were detected. Plasma emission spectrum displayed several peaks in ultraviolet domain. Concerning RS, the concentration of intracellular peroxides and nitric oxide was increased. However, antioxidative defenses were not triggered and reactive oxygen species inhibitors were not capable of abrogating cytotoxic effects of CAP. Similarly, verapamil did not protect cells from death. Conclusion This study suggests a potential novel therapy based on plasma able to selectively target tumour cells while preserving the non-inflammatory environment. Funding FCT, Portugal (UID/NEU/04539/2013), FEDER-COMPETE (FCOMP-01–0124-FEDER-028417, POCI-01–0145-FEDER-007440)