In vitro study of cold atmospheric plasma-activated liquids inhibits malignant melanoma by affecting macrophage polarization through the ROS/JAK2/STAT1 pathway

Melanoma is a prevalent malignant skin tumor known for its high invasive ability and a high rate of metastasis, making clinical treatment exceptionally challenging. Tumor-associated macrophages (TAMs) are the most abundant immune cells in the tumor microenvironment and play a crucial role in tumor survival and development. Cold atmospheric plasma (CAP) is an emerging tool for tumor treatment that has garnered attention from scholars due to its interaction with non-tumor cells in the tumor microenvironment. Here, we used the macrophage lines THP-1 and RAW264.7, as well as the melanoma cell lines A375 and MV3, as research subjects to investigate the effect of plasma-activated liquid (PAL) on macrophage differentiation and its inhibitory effect on melanoma cell proliferation. We confirmed that the killing effect of PAL on melanoma cells was selective. Using flow cytometry and PCR, we discovered that PAL can influence macrophage differentiation. Through in vitro cell coculture, we demonstrated that PAL-treated macrophages can significantly impede tumor cell development and progression, and the effect is more potent than that of PAL directly targeting tumor cells. Furthermore, we have proposed the hypothesis that PAL promotes the differentiation of macrophages into the M1 type through the ROS/JAK2/STAT1 pathway. To test the hypothesis, we employed catalase and fludarabine to block different sites of the pathway. The results were then validated through Western Blot, qPCR and ELISA. This study illustrates that PAL therapy is an effective tumor immunotherapy and expands the scope of tumor immunotherapy. Furthermore, these findings establish a theoretical foundation for potential clinical applications of PAL.