Background/Purpose: Neutrophils are important effector cells in systemic immune-mediated diseases. Neutrophil phenotypes vary depending on their age, maturity, activation state, and local environment; however, differences among neutrophils across different inflammatory conditions are poorly understood.

Methods: We analyzed mass cytometry data on blood neutrophils generated in the Accelerating Medicines Partnership RA/SLE Network, which included > 65 million neutrophils from 10 healthy controls, 13 patients with rheumatoid arthritis and 24 patients with biopsy-confirmed lupus nephritis. Neutrophil phenotypes were assessed by dimensional reduction and visualization with UMAP, and subclusters were partitioned by FlowSOM clustering and ConsensusClusterPlus metaclustering.

Results: Using unsupervised dimensionality reduction, we observed that neutrophils clustered by surface phenotype rather than by disease. Comparison of expression levels of surface proteins on neutrophils revealed a significant increase in the expression of Lysosomal-associated membrane protein 1 (LAMP1; CD107a) on neutrophils from SLE patients compared to controls (p=0.022). LAMP-1 expression largely segregated to a subcluster of neutrophils by UMAP visualization, suggesting an increase in a subset of neutrophils with recent de-granulation in SLE patients. SLE patients showed a range of expression of LAMP-1 on neutrophils, and LAMP-1 expression did not correlate with frequencies of PD-1hi CXCR5- T peripheral helper (Tph) cells, PD-1hi CXCR5+ T follicular helper (Tfh) cells, or CD11c+ B cells, which are highly expanded lymphocyte populations in lupus nephritis patients. Hierarchical clustering based solely on neutrophil protein expression distinguished two subgroups of SLE patients. Patients in the SLE1 group had an expanded population of LAMP1+ degranulated neutrophils, and this group also had higher chronicity scores in kidney biopsies. In contrast, patients in SLE2 group displayed an increased frequency of C5aR+ CXCR1+ neutrophils and had higher dsDNA titers and a higher SLEDAI composite score. These two subgroups did not differ in frequencies of Tph cells, Tfh cells, CD11c+ B cells, again suggesting that the neutrophil phenotypes may be independent of activated T/B cell pathways.

Conclusion: This work highlights an altered neutrophil phenotype in patients with lupus nephritis, in particular in a subset of patients associated with increased LAMP-1 expression. Neutrophil surface phenotypes may reveal a unique axis of immune activation in SLE patients.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/mass-cytometry-reveals-activation-heterogeneity-of-circulating-neutrophils-in-systemic-lupus-erythematosus/