Imaging-Based Analysis of Circadian Reprogramming and Cellular Remodeling Induced by Time-Restricted Feeding in Aging and Neurodegeneration Models

Time-restricted feeding (TRF) has emerged as a promising intervention to improve metabolic health and promote healthy aging, yet the cellular mechanisms underlying its effects remain incompletely understood. Here, we applied imaging-based and quantitative cellular analyses to investigate how TRF modulates aging-associated and neurodegeneration-related phenotypes in vitro. Human fibroblasts and AC16 cardiomyocytes were used as models of cellular aging, alongside fibroblast-based models of neurodegeneration. TRF was simulated through cyclic nutrient availability, and cellular responses were evaluated using microscopy-based assessment of cellular morphology, senescence-associated features, metabolic state, and circadian rhythm-associated gene expression dynamics. Imaging analyses demonstrated that TRF modulated key hallmarks of cellular senescence, including changes in cell morphology and intracellular organization, consistent with enhanced cellular resilience and altered metabolic adaptation. In AC16 cardiomyocytes, TRF influenced aging-associated cellular phenotypes, indicating that its effects extend beyond proliferative cell systems to cardiac-relevant models. In neurodegeneration-associated fibroblast models, TRF altered disease-related cellular signatures and stress-associated phenotypes, supporting a potential protective role in neurodegenerative conditions. Quantitative analyses further revealed significant TRF-induced changes in circadian rhythm characteristics across all models, including altered oscillatory amplitude, supporting a mechanistic link between nutrient timing and cellular timekeeping. Together, these findings demonstrate that TRF induces measurable changes in cellular architecture and circadian regulation associated with improved aging- and neurodegeneration-related phenotypes. This work highlights the utility of imaging-based approaches for investigating the spatiotemporal cellular effects of metabolic interventions and supports TRF as a potential therapeutic strategy for age-associated diseases.