Cold atmospheric-pressure nitrogen plasma induces the production of reactive nitrogen species and cell death by increasing intracellular calcium in HEK293T cells

Cold atmospheric-pressure plasma (CAP) has been emerging as a promising tool for cancer therapy in recent times. In this study, we used a CAP device with nitrogen gas (N(2)CAP) and investigated the effect of the N(2)CAP on the viability of cultured cells. Moreover, we investigated whether N(2)CAP-produced hydrogen peroxide (H2O2) in the medium is involved in N(2)CAP-induced cell death. Here, we found that the N(2)CAP irradiation inhibited cell proliferation in the human embryonic kidney cell line HEK293T and that the N(2)CAP induced cell death in an irradiation time- and distance-dependent manner. Furthermore, the N(2)CAP and H2O2 increased intracellular calcium levels and induced caspase-3/7 activation in HEK293T cells. The N(2)CAP irradiation induced a time dependent production of H2O2 and nitrite/nitrate in PBS or culture medium. However, the amount of H2O2 in the solution after N(2)CAP irradiation was too low to induce cell death. Interestingly, carboxy-PTIO, a nitric oxide scavenger, or BAPTA-AM, a cell-permeable calcium chelator, inhibited N(2)CAP-induced morphological change and cell death. These results suggest that the production of reactive nitrogen species and the increase in intracellular calcium were involved in the N(2)CAP-induced cell death.