Development of a Single-Cell Spatial MetabolomicsMethod for the Characterization of Cell–Cell Metabolic Interactions

Tumor microenvironment (TME) is characterized by complex cellular composition and high molecular heterogeneity. Characterizing the metabolic interactions between different cells in the TME is important for understanding the molecular signatures of tumors and identifying potential metabolic vulnerabilities for tumor treatment. In this research, we develop a single-cell spatial metabolomics method to profile cell-specific metabolic signatures and cell–cell metabolic interactions using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI). Different low-molecular-weight metabolites and lipids including glutamate, aspartate, glutamine, taurine, phenylalanine, glutathione, fatty acids, phospholipids, etc. were successfully detected and imaged after optimizing cell culture conductive slides, cell washing, and fixation procedures. Subsequently, we carried out single-cell spatial metabolomics on H460 large-cell lung cancer cells, HT-29 colorectal cancer cells, A549 lung cancer cells, HUH-7 liver cancer cells, and cancer-fibroblasts coculture system. We revealed that the metabolic profiles of both cancer cells and fibroblasts were altered after cell coculture. Glutamate and aspartate significantly increased in fibroblasts after coculture with cancer cells, corresponding to their indispensable roles in the creation of pro-cancer microenvironment. In addition, we discovered that the expressions of fatty acids and phospholipids in tumor cells and fibroblasts were also changed after cell coculture, which is closely related to the competition for energy and nutrient metabolites between different cells. We anticipate this single-cell analysis method to be broadly used in the investigations of diverse cellular models and cell–cell metabolic interactions.