Spatial transcriptomics identifies IL-32 as a lipid droplet-associated cytokine linked to tubular injury in human diabetic kidney disease

BACKGROUND: Diabetic kidney disease (DKD) is a severe complication of diabetes mellitus and the leading cause of chronic kidney disease worldwide. Among the many drivers of tubular injury, lipid accumulation and inflammation are emerging as major contributors to kidney disease progression, but the molecular link between lipid metabolism and inflammatory signaling remains to be determined. METHODS: Kidney biopsies from patients with DKD across pathologic classes were labelled for lipid droplets and analyzed by Nile Red spectroscopy. Digital spatial profiling and single-cell spatial transcriptomics were performed on samples from 14 patients representing different DKD classes. RNA scope and immunofluorescence microscopy were used for data validation and characterization. RESULTS: Lipid droplets (LD) were increasingly abundant in advanced stages of DKD, primarily accumulating in the proximal tubules. Single-cell spatial transcriptomics identified several genes-DUSP5, AZU1, COL9A1, HSPB1, and IGFBP7-as highly upregulated in DKD. Remarkably, IL32, which encodes a LD-associated cytokine, was highly enriched in injured proximal tubules. Immunofluorescence confirmed IL-32 localization to LDs predominantly within KIM1 positive tubules in moderate to advanced DKD. Furthermore, injured IL-32 expressing tubules were in close proximity to infiltrating neutrophils and macrophages, immune effectors of non-resolving inflammation and kidney disease progression. CONCLUSION: IL-32 is a LD-associated cytokine upregulated during tubular injury that represents a potential link between lipid dysregulation, inflammation and progression in human DKD.