Metabolic disruptions induced by low concentrations of DMSO in RTgill-W1 fish cells: The importance of solvent controls in in vitro studies

Dimethyl sulfoxide (DMSO) is a widely used solvent in biological research due to its ability to enhance membrane permeability, facilitating drug delivery and molecular transport across cellular membranes. However, its effects on cellular metabolism, especially at low concentrations, remain insufficiently understood. This study investigated the metabolic disruptions induced by 0.1-10 % DMSO in the RTgill-W1 fish cell line, focusing on changes in cell viability, oxidative stress, and key metabolic pathways. Results revealed that DMSO exposure caused dose-dependent declines in cell viability at 0.5 % DMSO and increases in reactive oxygen species (ROS) at 4 % and higher, indicating elevated oxidative stress. Metabolomic profiling revealed altered levels of numerous metabolites and significant impacts on 41 metabolic pathways belonging to five major functional groups: amino acid metabolism, carbohydrate metabolism, lipid metabolism, vitamin and co-factor metabolism, and nucleotide metabolism. The effects were observed across all exposure concentrations (0.1, 0.5, 1, 4, and 8 %), with more pronounced impacts at higher concentrations. These findings highlight that DMSO, even at low concentrations (≤ 0.5 %), can have widespread effects on cellular metabolism, impacting experimental outcomes in in vitro studies. This study provides valuable insights into the biochemical impacts of DMSO on fish cell lines and emphasizes a caution in using DMSO in biological research to minimize unintended cellular effects. Additionally, it highlights the critical need to include solvent controls at matching concentrations to accurately distinguish solvent-induced effects from those caused by experimental treatments.