Non-equilibrium atmospheric-pressure plasma (APP) is used medically for plasma-induced cell permeabilization. However, how plasma irradiation specifically triggers permeabilization remains unclear. In an attempt to identify the dominant factor( s ), the distribution of plasma-produced reactive species was investigated, primarily focusing on OH radicals. A stronger plasma discharge, which produced more OH radicals in the gas phase, also produced more OH radicals in the liquid phase (OH _aq ), enhancing the cell membrane permeability. In addition, plasma irradiation-induced enhancement of cell membrane permeability decreased markedly with increased solution thickness (<1 mm), and the plasma-produced OH _aq decayed in solution (diffusion length on the order of several hundred micrometers). Furthermore, the horizontally center-localized distribution of OH _aq corresponded with the distribution of the permeabilized cells by plasma irradiation, while the overall plasma-produced oxidizing species in solution (detected by iodine-starch reaction) exhibited a doughnut-shaped horizontal distribution. These results suggest that OH _aq, among the plasma-produced oxidizing species, represents the dominant factor in plasma-induced cell permeabilization. These results enhance the current understanding of the mechanism of APP as a cell-permeabilization tool.

The cell membrane permeability, which is strongly related to gene transfection, is improved using a cell-solution electrode for generating atmospheric-pressure plasma (APP) just above the solution. In the case of the floating cells, the cell membrane permeability is significantly improved by the cell-solution electrode APP compared with the conventional diffusion type APP, because the distance between the plasma generation area and the cell solution surface becomes short, which could reduce the radial diffusion loss of the plasma irradiated to the cell suspended solution. In the case of the adherent cells, cell membrane permeability is found to be enhanced by the shorter distance between the solution surface and the adherent cells as well as using the cell-solution electrode, which means that the short-lived reactive oxygen species generated at the solution surface are essential for the improvement of cell membrane permeability.

This study was designed to assess the effects of cold atmospheric plasma on osteoblastic differentiation in pre-osteoblastic MC3T3-E1 cells. Plasma was irradiated directly to a culture medium containing plated cells for 5 s or 10 s. Alkaline phosphatase (ALP) activity assay and alizarin red staining were applied to assess osteoblastic differentiation. The plasma-generated radicals were detected directly using an electron spin resonance-spin trapping technique. Results show that plasma irradiation under specific conditions increased ALP activity and enhanced mineralization, and demonstrated that the yield of radicals was increased in an irradiation-time-dependent manner. Appropriate plasma irradiation stimulated the osteoblastic differentiation of the cells. This process offers the potential of promoting bone regeneration.

Gas-liquid interfacial atmospheric-pressure plasma jets (GLI-APPJ) are used medically for plasma-induced cell-membrane permeabilization. In an attempt to identify the dominant factors induced by GLI-APPJ responsible for enhancing cell-membrane permeability, the concentration and distribution of plasma-produced reactive species in the gas and liquid phase regions are measured. These reactive species are classified in terms of their life-span: long-lived (e.g., H2O2), short-lived (e.g., O2•-), and extremely-short-lived (e.g., •OH). The concentration of plasma-produced •OHaq in the liquid phase region decreases with an increase in solution thickness (<1 mm), and plasma-induced cell-membrane permeabilization is found to decay markedly as the thickness of the solution increases. Furthermore, the horizontally center-localized distribution of •OHaq, resulting from the center-peaked distribution of •OH in the gas phase region, corresponds with the distribution of the permeabilized cells upon APPJ irradiation, whereas the overall plasma-produced oxidizing species such as H2O2aq in solution exhibit a doughnut-shaped horizontal distribution. These results suggest that •OHaq is likely one of the dominant factors responsible for plasma-induced cell-membrane permeabilization.

Non-equilibrium helium atmospheric-pressure plasma (He-APP), which allows for a strong non-equilibrium chemical reaction of O₂ and N₂ in ambient air, uniquely produces multiple extremely reactive products, such as reactive oxygen species (ROS), in plasma-irradiated solution. We herein show that relatively short-lived unclassified reactive species (i.e., deactivated within approximately 10 min) generated by the He-APP irradiation can trigger physiologically relevant Ca²⁺ influx through ruthenium red- and SKF 96365-sensitive Ca²⁺-permeable channel(s), possibly transient receptor potential channel family member(s). Our results provide novel insight into understanding of the interactions between cells and plasmas and the mechanism by which cells detect plasma-induced chemically reactive species, in addition to facilitating development of plasma applications in medicine.