Circadian Phase Determines Tissue-Specific Adaptations to Long-Term Exercise in Obese Mice

Background: Exercise interacts closely with the circadian system; however, whether long-term training elicits time-of-day-dependent metabolic adaptations in the context of obesity remains unclear. Methods: Male C57BL/6 mice were fed a high-fat diet and trained on a treadmill for 8 weeks during either the early rest phase (ZT3, Zeitgeber time) or the early active phase (ZT15). Sedentary mice served as controls. After the last session, animals were fasted for 4 h and sampled 48–49 h later. Plasma triglycerides (TGs) and glucose, as well as liver and epididymal white adipose tissue (EPI), were analyzed. Results: Plasma TGs showed a significant phase × exercise interaction (F(1, 25) = 5.25, p = 0.0307), with the lowest levels in ZT15-exe (27.22 mg/dL) compared with ZT15-sed (39.47 mg/dL, p < 0.01) and ZT3-exe (41.80 mg/dL, p < 0.01). Hepatic TG content was markedly lower in ZT3 than in ZT15 mice (F(1, 25) = 15.49, p < 0.001), and Oil Red O staining was associated with reduced lipid accumulation in exercised groups (p < 0.05). In EPI, Fasn expression was robustly decreased by exercise (F(1, 25) = 16.43, p = 0.0004, q = 0.0059), indicating long-term suppression of lipogenesis. In the liver, Cpt1a showed significant main effects of both phase (F(1, 25) = 10.11, p = 0.0039, q = 0.0158) and exercise (F(1, 25) = 13.42, p = 0.0012, q = 0.0353), being higher in ZT3 and under sedentary conditions, suggesting a circadian-dominant oxidative advantage in hepatic metabolism. Conclusions: Long-term exercise induced phase-dependent adaptations in lipid metabolism. Active-phase exercise promoted adipose lipid mobilization and lowered plasma TGs, while rest-phase training enhanced hepatic oxidative capacity. These results suggest a “tissue × time” framework of circadian-specific exercise responses, providing hypothesis-generating evidence for optimizing exercise timing in metabolic disorders.