The cold atmospheric pressure plasma jet interaction with the bio-target is studied in the plasma experiment, 2D fluid model simulations, and with MTT and iCELLigence assays of the viability of cancer cells. It is shown, for the first time, that the use of the grounded substrate under the media with cells considerably amplifies the effect of plasma cancer cell treatment in vitro. Plasma devices with cylindrical and plane geometries generating cold atmospheric plasma jets are developed and tested. The sequence of the streamers which forms the plasma jet is initiated with a voltage of 2.5–6.5 kV applied with the frequency 40 kHz. We suggest using the grounded substrate under the bio-target during the plasma jet treatment of cancer cells. The analysis of the measured plasma spectra and comparison of OH-line intensity for different voltages and gas flow rates allows us to find a range of optimal plasma parameters for the enhanced OH generation. The time-dependent viability is measured for human cell lines, A431 (skin carcinoma), HEK 293 (kidney embryonic cells), and A549 (human lung adenocarcinoma cells) after the plasma jet treatment. The results with cell-based experiments (direct treatment) performed with various plasma jet parameters confirm the maximum efficiency of the treatment with the optimal plasma parameters.

Cold atmospheric plasma (CAP) is an intensively-studied approach for the treatment of malignant neoplasms. Various active oxygen and nitrogen compounds are believed to be the main cytotoxic effectors on biotargets; however, the comprehensive mechanism of CAP interaction with living cells and tissues remains elusive. In this study, we experimentally determined the optimal discharge regime (or semi-selective regime) for the direct CAP jet treatment of cancer cells, under which lung adenocarcinoma A549, A427 and NCI-H23 cells demonstrated substantial suppression of viability, coupled with a weak viability decrease of healthy lung fibroblasts Wi-38 and MRC-5. The death of CAP-exposed cancer and healthy cells under semi-selective conditions was caspase-dependent. We showed that there was an accumulation of lysosomes in the treated cells. The increased activity of lysosomal protease Cathepsin D, the transcriptional upregulation of autophagy-related MAPLC3B gene in cancer cells and the changes in autophagy-related proteins may have indicated the activation of autophagy. The addition of the autophagy inhibitor chloroquine (CQ) after the CAP jet treatment increased the death of A549 cancer cells in a synergistic manner and showed a low effect on the viability of CAP-treated Wi-38 cells. Downregulation of Drp1 mitochondrial protein and upregulation of PINK1 protein in CAP + CQ treated cells indicated that CQ increased the CAP-dependent destabilization of mitochondria. We concluded that CAP weakly activated pro-survival autophagy in irradiated cells, and CQ promoted CAP-dependent cell death due to the destabilization of autophagosomes formation and mitochondria homeostasis. To summarize, the combination of CAP treatment with CQ could be useful for the development of cold plasma-based antitumor approaches for clinical application.