Senescence Cell Induction Methods Display Diverse Metabolic Reprogramming and Reveal an Underpinning Serine/Taurine Reductive Metabolic Phenotype

The relationship between in vitro senescence cell induction and intracellular biomolecular dysregulation is still poorly understood. In this study, we have found that a range of metabolic subphenotypes exists and is dependent on the induction method that is used. To develop understanding of these subphenotypes, we developed and employed a novel bioanalytical pipeline integrating untargeted metabolomics, label-free proteomics, and stable isotope tracing alongside cellular deformability measurements and established senescence biomarkers. Initially, standard senescent markers indicated all induction methods were consistent by showing elevated SA-β-Gal expression, p21 levels, and γH2AX DNA damage markers alongside a decrease in Ki67 and an increase in shape, volume, and deformability. However, when probed at the metabolic and protein levels, all senescence models indicated both shared and unique biomolecular responses. A metabolic shift toward reductive pathways (driven by serine and taurine rewiring) and impaired proteostasis was an observed shared response. These findings suggest that targeting metabolic redox circuits, alongside serine and taurine metabolic processes, presents novel therapeutic strategies for addressing senescence and aging. But importantly, alongside this general shift, we found that significant metabolic and proteomic heterogeneity also exists across different senescence induction methods. This demonstrates that the method of senescence induction significantly influences cell metabolic and proteomic profiles. Critically, methods of senescence induction are not interchangeable, and careful consideration is needed when choosing between different induction methods and when comparing cellular phenotypes across different in vitro senescence experiments.