Elucidation of Kidney Ischemia Tolerance Mechanisms in Fasting State with Energy Metabolism Visualization Techniques

The kidneys constantly require large amounts of ATP to meet the demands of their intricate functions, such as tubular reabsorption. Furthermore, abnormalities in kidney energy metabolism are closely associated with various types of kidney disease. We previously established an experimental technique that visualizes spatiotemporal ATP dynamics at single-cell resolution using two-photon microscopy and a novel mouse line systemically expressing a Förster resonance energy transfer–based ATP biosensor. Using this system, we reported that proximal tubule (PT) ATP recovery during ischemia reperfusion injury (IRI) in the acute phase was correlated with kidney prognosis. Meanwhile, caloric restriction and intermittent fasting have been reported to exert protective effects against metabolic disorders in multiple organs and even to extend lifespan. Moreover, fasting has been demonstrated to attenuate ischemic injury, particularly in the brains, hearts, and kidneys. While inhibited mTOR signaling, activated autophagy, and reduced oxidative stress have been suggested to contribute to this protective effect, the underlying mechanism and energy metabolic alteration in the acute phase remain unclear. Here, we analyzed the ATP dynamics and β-oxidation activity in the kidneys during IRI in fasted mice.

Using ATP visualizing mice and multiphoton microscopy, we analyzed ATP dynamics during unilateral 30-minute IRI in the 1-day fasting group and control group, evaluating the correlation between ATP recovery in the acute phase and tubular damage in the chronic phase. Next, RNA-seq analysis was performed on PT cells isolated from the kidneys of fasted mice. Furthermore, β-oxidation activity in the tubules during fasting was evaluated using quinone methide (QM)-releasing probes capable of in vivo visualization of fatty acid β-oxidation activity.

There was no significant difference in ATP levels in the PTs before ischemia between the control group and fasting group. In contrast, the ATP recovery rates in PTs at 10 min after IRI was 59.2% in the control group and 77.8% in the fasting group, and the percentages of impaired tubules at 14 days after IRI were 73.2% in the control group and 55.6% in the fasting group. ATP recovery rates in the acute phase were inversely correlated with tubular damage in the chronic phase. Gene ontology enrichment analysis showed upregulation of fatty acid metabolic pathways in PT cells of fasted mice in comparison with the control group. In addition, the administration of a QM release probe successfully visualized the activation of β-oxidation in the kidney at the single-cell level in vivo. Consequently, β-oxidation was predominantly activated in PTs and was further upregulated under fasting conditions.

The ATP recovery rate in PTs in the acute phase was improved in fasted mice, which was associated with reduced tubular injury in the chronic phase. Gene expression related to fatty acid metabolism was also elevated in PT cells of fasted mice. Moreover, QM-releasing probe administration demonstrated the activated β-oxidation in PTs after fasting. These data suggest that enhanced β-oxidation in the PT of fasting mice may be associated with improved ATP recovery rate and resistance to injury after IRI. This abstract was also submitted for JSN/ERA Symposium and ASN Kidney Week 2025 congress, and re-submission is permitted by JSN and ASN.