Ergothioneine attenuates age-related declines in circadian rhythmicity

Aging is accompanied by the progressive deterioration of circadian clock function, characterized by reduced amplitude, increased period variability, and impaired metabolic coupling. Declining intracellular nicotinamide adenine dinucleotide (NAD⁺) levels and SIRT1 activity have been implicated as key mediators of age-associated circadian disruption. Ergothioneine (EGT), a diet-derived antioxidant with longevity-associated effects, has recently been reported to improve healthspan and modulate redox metabolism. However, its effects on the circadian clock remain unclear. Here, we examined whether EGT mitigates age-related decline in circadian rhythmicity using PER2::LUC mouse embryonic fibroblasts (MEFs). Chronic EGT treatment enhanced the amplitude of the PER2::LUC rhythm in a dose-dependent manner without markedly altering the baseline period length. Aging-associated NAD⁺ decline was modeled using FK866, a NAMPT inhibitor that depletes intracellular NAD⁺, which reduced rhythm amplitude, lengthened the period, and increased cycle-to-cycle variability. Notably, co-treatment with EGT significantly restored rhythm amplitude, attenuated FK866-induced period lengthening, and reduced period variability. Biochemical analyses revealed that EGT increased the NAD⁺/NADH ratio under basal conditions and significantly elevated both NAD⁺ levels and the NAD⁺/NADH ratio under FK866-induced NAD⁺ depletion. This effect was not attributable solely to cytoprotection. This study demonstrates that EGT enhances circadian rhythm robustness and counteracts NAD⁺-depletion-induced clock dysfunction. EGT may ameliorate age-related circadian decline by improving intracellular redox balance and NAD⁺ metabolism. Given that EGT crosses the blood-brain barrier, it may represent a novel nutritional strategy to preserve circadian function during aging.