Single Cell and Spatial Transcriptomics Define a Proinflammatory and Profibrotic Niche After Kidney Injury

Abstract Kidney fibrosis is the common outcome of chronic kidney disease (CKD). It often instigates in the focal sites by forming the fibrogenic niche after injury. In this study, using single‐cell RNA sequencing (scRNA‐seq) and a spatial transcriptomic (ST) approach, the cellular heterogeneity, spatial organization, and molecular interactions are delineated in the fibrotic kidney. Through analyses of the scRNA‐seq and ST data from normal and fibrotic kidneys in mice subjected to unilateral ischemia‐reperfusion injury, a tenascin C (TNC)‐enriched, proinflammatory, and profibrotic microenvironment is identified that facilitated macrophage activation and promoted renal inflammation and fibrosis. Both TNC‐enriched decellularized kidney tissue scaffold and exogenous TNC protein promoted bone marrow‐derived macrophages activation though Toll‐like receptor 4 (TLR4)/NF‐κB signaling. Either pharmacological inhibition of TLR4 signaling or genetic knockout of its gene alleviated renal inflammation and fibrosis by inhibiting macrophage activation in vivo. Finally, chimeric mice that received bone marrow transplantation from TLR4‐deficient donors are protected against kidney inflammation and fibrosis. These results suggest that TNC plays a crucial role in orchestrating the formation of a proinflammatory and profibrotic niche that promotes renal inflammation and fibrosis by activating macrophages via TLR4/NF‐κB signaling. The findings underscore the complex interplay among fibroblasts, extracellular microenvironment, and macrophages that drive kidney fibrosis.