A precision medicine approach uncovers a unique signature of neutrophils in patients with brushite kidney stones

Background: We have previously found that papillary histopathology differs greatly between calcium oxalate and brushite stone formers (SF); the latter have much more papillary mineral deposition and tissue fibrosis. Methods: In this study, we applied unbiased orthogonal omics approaches on biopsied renal papillae and extracted stones from patients with brushite or calcium oxalate (CaOx) stones. Our goal was to discover stone type-specific molecular signatures to advance our understanding of the underlying pathogenesis. Results: Brushite SF did not differ from CaOx SF with respect to metabolic risk factors for stones, but did exhibit increased tubule plugging in their papillae. Brushite SF had upregulation of inflammatory pathways in papillary tissue, and increased neutrophil markers in stone matrix compared to those with CaOx stones. Large-scale 3D tissue cytometry on renal papillary biopsies showed an increase in the number and density of neutrophils in the papillae of brushite vs. CaOx patients, thereby linking the observed inflammatory signatures to the neutrophils in the tissue. To explain how neutrophil proteins appear in the stone matrix, we measured neutrophil extracellular trap (NET) formation, NETosis, and found it significantly increased in the papillae of brushite compared to CaOx patients. Conclusions: We show that increased neutrophil infiltration and NETosis is an unrecognized factor that differentiates brushite and CaOx SF, and may explain the markedly increased scarring and inflammation seen in the papillae of brushite patients. Given the increasing prevalence of brushite stones, the role of neutrophil activation in brushite stone formation requires further study.