Low-temperature plasma induced melanoma apoptosis by triggering a p53/PIGs/caspase-dependent pathway in vivo and in vitro

Abstract

The potential antitumor application of low-temperature plasma (LTP) has attracted wide attention from researchers. The mechanism of plasma antitumor action may be related to LTP-induced reactive oxygen and nitrogen species. The aim of this study was to identify the apoptotic effects of plasma on melanoma cells (B16) in vivo and in vitro. For this purpose, a helium atmospheric-pressure plasma jet (APPJ) was used to generate the plasma-activated saline and medium (PAS and PAM). The PAS was injected subcutaneously to treat B16-tumor bearing mice in vivo and the PAM was used to treat B16 cells in vitro. Catalase (CAT) was added into the PAM to degrade H2O2 as the intervention group. The tumor nodules’ weight, cryosections, B16 cell viability, apoptosis, DNA damage, p53 pathway-related protein and the concentration of H2O2 in PAM were detected. The in vivo results showed that PAS could inhibit tumor growth and induce apoptosis. The in vitro results revealed the concentration of H2O2 in the PAM increased as LTP treatment time was longer, while the cell viability decreased as APPJ irradiation time was increased. Flow cytometry results showed that, compared with the control group, all PAM treatment groups showed a higher apoptotic rate. A single cell gel assay and an 8-OHdG assay showed that the PAM could induce DNA damage. A western blotting assay indicated that the PAM could increase the expressions of p53, p53-induced gene 8 protein, PIG3, cytoplasmic cytochrome c, cleaved-caspase-9 and -3. However, CAT could alleviate all these changes effectively. These findings suggested that plasma exerted an apoptotic effect on B16 cells mainly by activating the p53/PIGs/caspase pathway.