Background/Purpose: Ankylosing spondylitis (AS) patients have irregular neutrophil responses, as indicated clinically by neutrophilia and increased neutrophil to lymphocyte ratios that positively associate with disease activity. CD4⁺ T helper cells that produce IL-17 (Th17 cells) drive the pathogenesis of AS, and IL-17 blockade is an effective therapy for this disease. Yet, a causative connection between neutrophils and induction of pathogenic Th17 responses in AS remains unknown.

Methods: Arthritis was induced by intraperitoneal injection of 1.5mg zymosan to female SKG mice pre-treated with neutrophil-depleting mAb 1A8 or an isotype control, and onset and severity of peripheral arthritis and axial disease was measured clinically. Neutrophils and CD4+ T cells were isolated from the bone marrow and spleen, respectively, of naïve SKG and WT mice. Zymosan-stimulated neutrophils and TCR-activated CD4+ T cells were co-cultured, together or separated by a 0.4-um pore transwell. IL-17A in supernatants was quantified by ELISA after 72h of co-culture. N=2-4 mice/experiment/genotype, experiments were repeated 3 times each. Statistical significance was calculated by multiple unpaired parametric student T tests.

Results: SKG mice developed Th17 responses and clinical arthritis as early as 5d post-zymosan exposure (pze). SKG mice-deplete of neutrophils showed decreased Th17 responses and undetectable arthritis 5d pze; thereby revealing neutrophils as a requirement for the onset of AS. SKG T cells co-cultured with stimulated neutrophils at a 1:2 (T cell:neutrophil) ratio produced 4.5-fold more IL-17A than SKG T cells cultured in the absence of neutrophils. While SKG neutrophils increased IL-17A production in a dose-dependent manner, WT neutrophils did not. Collectively, these data indicate that neutrophils are alone sufficient to expand Th17 cells. Intriguingly, the capacity of co-cultured neutrophils to potentiate Th17 cells was mitigated when physically separated in transwell plates, indicating that neutrophil-induced Th17 expansion works through a contact-dependent mechanism.

Conclusion: Our study suggests that activated SKG neutrophils interact physically with CD4+ T cells to induce potent arthritogenic Th17 responses. Understanding the intersection between innate and adaptive cellular mechanisms in AS pathogenesis will elucidate novel therapeutic targets for treatment of disease.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/neutrophils-induce-contact-dependent-expansion-of-arthritogenic-th17-cells-and-are-necessary-for-disease-in-experimental-ankylosing-spondylitis/