1843: Tissue Neutrophils in The Pathogenesis of Lupus Nephritis

Background/Purpose: Systemic lupus erythematosus (SLE) is a multiorgan autoimmune disease that frequently affects the kidneys, with lupus nephritis (LN) being a major contributor to morbidity and mortality. A neutrophil gene signature in peripheral blood has been associated with LN, and neutrophil infiltration has been detected in the lupus kidney parenchyma. A putative pathogenic contribution of neutrophils to SLE is the externalization of modified autoantigens and proteins that are detrimental to the host, through dysregulated neutrophil death by the formation of neutrophil extracellular traps (NETs). However, the specific pathogenic impact of neutrophils/NETs in LN has not been systematically studied.

Methods: Lupus-prone NZB/W F₁ and control C57BL/6 (B6) mice were analyzed. The number and maturity of neutrophils were assessed in PBS-perfused kidneys by flow cytometry. The morphology and localization of neutrophils were evaluated by different light microscopy approaches such as, brightfield, namely, hematoxylin-eosin and immunohistochemical (IHC) staining, as well as by immunofluorescence (IF) staining. Kidney biopsies from patients with LN were also analyzed by IHC and IF to corroborate findings obtained from mice. Furthermore, human mesangial cells were stimulated with NETs, and their interactions were assessed by IF and RNA sequencing.

Results: NZB/W F₁ mice exhibited increased renal neutrophil infiltration in comparison to the control B6 mice, particularly following the onset of proteinuria, which correlated with its severity. Kidney-infiltrating neutrophils exhibited a predominantly mature phenotype and localized primarily in the glomerular region. Fragmented neutrophil remnants, suggesting cell death and enhanced debris deposition, accumulated in mesangial areas post-proteinuria onset and in association with mesangial matrix damage score. IF analysis confirmed that NETs deposited in the mesangial regions. Similar findings were observed in biopsies from SLE patients with a diagnosis of LN. In vitro, human mesangial cells internalized NETs via endocytosis and phagocytosis, leading to phenotypic changes detectable by RNA sequencing and functional studies.

Conclusion: These results suggest that neutrophils contribute to the pathogenesis of LN through infiltration, NET formation, and interaction with mesangial cells. These observations further implicate aberrant neutrophils in the pathogenesis of lupus-associated organ damage.