Sepsis-associated acute kidney injury (SA-AKI) is closely linked to profound metabolic reprogramming and mitochondrial dysfunction, yet effective metabolic targeted therapies remain limited. In this study, we elucidated the molecular mechanism by which ethyl pyruvate (EP), an exogenous metabolic substrate, alleviates SA-AKI by regulating mitochondrial bioenergetics through the AMPK-mTOR-SIRT3 signaling pathway. Using a cecal ligation and puncture-induced SA-AKI mouse model and LPS-stimulated HK-2 cells, we demonstrated that EP synergized with the AMPK activator AICAR to significantly improve renal function, reduce injury markers and inflammation, and enhance survival. Single-cell RNA sequencing (scRNA-seq) identified extensive metabolic reprogramming in renal epithelial cells during sepsis, characterized by mitochondrial dysfunction and suppression of oxidative phosphorylation (OXPHOS). Mechanistically, combined EP and AICAR treatment, rather than EP alone, alleviated mitochondrial dysfunction by restoring NAD/NADH balance, increasing mitochondrial DNA (mtDNA) content, improving mitochondrial respiratory chain complex activity, and rebalancing mitochondrial fission-fusion dynamics. Combined EP and AICAR treatment also enhanced autophagy and reduced apoptosis, partly through a SIRT3-dependent mechanism downstream of AMPK-mTOR signaling. This effect was enhanced or diminished by pretreatment with rapamycin (Rapa) or interferon-γ (IFN-γ). Pharmacological inhibition of SIRT3 partially abolished the mitochondrial and cytoprotective benefits of EP, confirming SIRT3 as a key downstream effector of AMPK-mTOR signaling. Collectively, our data identified EP as a metabolic modulator that synergizes with AMPK activation to alleviate SA-AKI. This effect is achieved by restoring mitochondrial function and OXPHOS through the AMPK-mTOR-SIRT3 signaling pathway, providing a mechanistic basis for metabolism-oriented therapeutic strategies in SA-AKI. +
