Multi-Omics and Experimental Validation Reveal the Targets and Mechanisms of Urolithin A in diabetic kidney disease

Background: Urolithin A (UA) has therapeutic potential in diabetic kidney disease (DKD), but its molecular targets and mechanisms remain unclear. Methods: Transcriptomic and single-cell datasets were integrated with UA target prediction. Machine learning, ROC analysis, GSEA, immune infiltration, and pseudotime trajectory inference were applied to identify biomarkers and proximal convoluted tubule (PCT) subtypes. Findings were validated in db/db mice and high-glucose-exposed HK-2 cells. Results: Four UA-responsive biomarkers—HSPA1A, ESR2, NUAK1, and PLK1—demonstrated strong diagnostic performance (AUC > 0.7). GSEA implicated these biomarkers in ascorbate/aldarate and butanoate metabolism pathways, while immune profiling revealed elevated CD8+ T cells and reduced naive B cells in DKD. Single-cell analysis identified PCT as a key disease-relevant population comprising three functional subtypes. Pseudotime trajectory revealed PLK1 enrichment at early differentiation stages, whereas NUAK1 was enriched at late stages. In vivo and in vitro experiments confirmed that UA treatment significantly attenuated the aberrant upregulation of NUAK1 and PLK1 in db/db mouse kidneys and HG-exposed HK-2 cells, accompanied by restoration of renal architecture and improved metabolic parameters. Conclusions: UA exerts renoprotective effects in DKD by modulating stage-specific expression of NUAK1 and PLK1 along PCT differentiation, offering insights into its mechanism of action.