Carnosine ameliorates renal ischaemia–reperfusion injury by modulating MFN2/OPA1 to restore mitochondrial homeostasis in UPB1 deficiency

Renal ischaemia–reperfusion injury (IRI) is a major cause of acute kidney injury (AKI) and a critical determinant of transplant outcomes. β-ureidopropionase (UPB1) is the terminal enzyme of the pyrimidine degradation pathway, catalysing the production of β-alanine — the rate-limiting precursor for carnosine (β-alanyl-L-histidine) synthesis. However, the role of this metabolic axis in IRI-AKI remains unknown. This study investigated the expression and functional significance of UPB1 in IRI-AKI and evaluated whether carnosine, the dipeptide product downstream of UPB1-mediated pyrimidine catabolism, exerts therapeutic effects against renal IRI. UPB1 expression was examined in a mouse IRI-AKI model and publicly available transcriptomic datasets. Quantitative proteomic analysis was performed on UPB1-knockout HK-2 cells. Mitochondrial function was assessed by TMRM staining, MitoSOX Red detection and CCK-8 viability assays. The therapeutic effect of exogenous carnosine was evaluated both in vitro and in vivo . Molecular docking and dynamics simulations were used to predict carnosine binding to mitochondrial dynamics proteins. Upb1 expression was significantly downregulated after IRI, reaching a nadir at 24 h. Single-cell analysis revealed that Upb1 was almost exclusively expressed in proximal tubular cells. MitoCarta3.0 scoring demonstrated that Upb1 -positive proximal tubular cells retained significantly higher mitochondrial activity after IRI, and virtual knockout of Upb1 resulted in broad downregulation of oxidative phosphorylation genes. Proteomic analysis revealed that UPB1 knockout disrupted mitochondrial fatty acid β-oxidation and mitochondrial envelope components. UPB1-deficient cells exhibited decreased mitochondrial membrane potential, elevated mitochondrial reactive oxygen species (mtROS) and reduced viability, along with downregulation of fusion proteins Mitofusin-2(MFN2), Optic Atrophy 1(OPA1), and upregulation of the fission protein Dynamin-related protein 1(DRP1). Computational modeling predicted that carnosine may interact with the mitochondrial fusion proteins MFN2 and OPA1, with a much stronger predicted interaction for OPA1 (−19.49 kcal mol⁻ 1 ) than for MFN2 (−4.66 kcal mol⁻ 1 ), suggesting OPA1 as the primary effector target. Carnosine supplementation rescued mitochondrial membrane potential, reduced mitochondrial reactive oxygen species, and attenuated IRI-induced renal injury in mice. UPB1 deficiency disrupts mitochondrial homeostasis in renal tubular cells by impairing mitochondrial dynamics. Exogenous carnosine supplementation restores mitochondrial function, likely through interaction with the mitochondrial fusion proteins MFN2 and OPA1, positioning the UPB1–carnosine axis as a potential therapeutic target for IRI-AKI.