Evaluation of the anticancer effects induced by cold atmospheric plasma in 2D and 3D cell-culture models

The biological characteristics of three-dimensional (3D) tumors cannot be adequately represented through two-dimensional (2D) cell cultures. Here, to acquire more precise information about plasma–tumor interactions, the plasma-induced cytotoxicity was tested in both 2D cell culture and 3D tumor-spheroid culture. The results indicated that the toxic effects were markedly reduced in 3D cancer cells compared with 2D cells. We found that reactive oxygen and nitrogen species can penetrate through the entire depth of 3D tumor spheroid, but local apoptosis was observed only on the outermost layer. The permeation depth of plasma-generated reactive species was then tracked in 3D tumor spheroids. We hypothesized that plasma-induced apoptosis within a tumor depends on the “deep” delivery of H₂O₂, NO₂ ⁻, and NO₃ ⁻ rather than O₂ ⁻, OH•, and ONOOH/ONOO⁻.