Integration of single-cell RNA sequencing and spatial transcriptomics reveals neutrophil diversity and spatial heterogeneity in acute kidney injury.

Acute kidney injury (AKI) is a critical clinical condition, with limited effective pharmacological treatments. Neutrophil-mediated inflammatory cascades play a pivotal role in AKI pathogenesis. However, the neutrophil diversity in AKI remains inadequately elucidated, impeding progress in mechanistic studies. In this study, through integrative single-cell and spatial transcriptomics, we constructed a comprehensive landscape of AKI neutrophils, encompassing multiple time points, severities, and etiologies of AKI. Nine neutrophil subpopulations were defined, among which six were first characterized in AKI, and three were novel in the literature. Transcriptional profiling suggested significant functional heterogeneity, including potential roles in phagocytosis, antigen presentation, inflammation, vascular modulation, and fibrosis. Phagocytic subtypes were associated with crystalline-related AKI, while various polymorphonuclear neutrophils were primary effectors of inflammation and fibrosis in drug-induced, obstructive, and ischemic injuries. Spatial neighborhood analysis revealed a novel neutrophil-centered microenvironment unit in AKI, characterized by interactions between neutrophils and the thick ascending limb (TAL). Further analysis on human AKI revealed correlations of neutrophils and neutrophil-centered microenvironment with worsening long-term prognosis. A prognostic risk model based on neutrophil subsets demonstrated superior predictive accuracy compared to existing models. Our study provides a foundational resource for understanding neutrophil heterogeneity in AKI and proposes a framework for future research into prognostic biomarkers and neutrophil-targeted therapeutic strategies.