Background/Purpose: Neutrophils are present in the early phases of spondyloarthritis (SpA)-associated uveitis, skin and intestinal disease, but their role in enthesitis remains unknown. We investigated the role of neutrophils in the experimental SKG mouse model and in human axial enthesis tissue. 

Methods: SKG mice have been shown to manifest multiple features of SpA. Arthritis in 9-week old female SKG mice was synchronized via intraperitoneal injections of curdlan and peripheral arthritis was scored weekly. Histology of ankles and spine were performed at serial time points. Formalin-fixed, paraffin-embedded (FFPE) tissue samples from entheseal sites around ankles and proximal tail vertebrae were obtained 2-3 weeks after curdlan injection (representing early inflammation) and subjected to laser capture microscopy. Whole transcriptome analysis was carried out using Affymetrix gene array MTA 1.0 and data was analyzed via IPA. Immunohistochemical staining for S100A8, a neutrophil product, and for MPO, a neutrophil marker, were performed at both sites. In conjunction with murine experimental work, normal human spinous process entheses were obtained from patients undergoing spinal decompression or thoracic or lumbar scoliosis surgery. IHC for MPO was performed on human FFPE enthesis to confirm the presence of neutrophils. Isolated neutrophils from peri-entheseal bone were stimulated with the fungal adjuvant zymosan and IFNγ and supernatants were probed for IL-23 secretion by ELISA.

Results: SKG mice developed early inflammation at enthesial sites around ankles and proximal tail vertebrae that included many neutrophils. The highest expression of transcripts arising from neutrophils included the alarmins S100A8 and S100A9 (calprotectin) and IL-23-associated genes, identified in both peripheral and axial enthesitis. In normal human axial enthesis, occasional neutrophils were evident. Upon fungal stimulation in vitro, these produced IL-23 protein, while isolated human entheseal fibroblasts stimulated with the fungal adjuvant mannan produced chemokines including IL-8, an important in the recruitment of neutrophils.

Conclusion: Neutrophil genes and pathways (including S100A8/9, cathepsin S, clec4d, clec7a) are upregulated early in the inflammatory infiltrate at entheseal sites. Neutrophils obtained from human entheses can be induced to produce IL-23 and may be an important early source of this cytokine that is critical for early SpA pathogenesis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-role-for-neutrophils-in-the-early-phases-of-enthesitis-in-spondyloarthritis/