Immature leukocyte and plasma-induced cell death reveal subclinical immune activation in EGPA patients in remission

Objective: ANCA-associated vasculitis (AAV) is a group of rare autoimmune diseases characterized by pauci-immune necrotizing inflammation of small to medium-sized blood vessels, in which ANCAs targeting neutrophil antigens promote neutrophil activation, endothelial injury and organ damage. Although AAV follows a relapsing-remitting course, the immune landscape during remission remains poorly defined. This study investigated leukocyte alterations across AAV subtypes and examined whether plasma from ANCA-positive and ANCA-negative eosinophilic granulomatosis with polyangiitis (EGPA) patients in remission modulates peripheral blood mononuclear cell (PBMC) responses in healthy donors (HDs). Methods: Peripheral blood was collected from 62 AAV patients in remission and 28 age- and sex-matched HDs. Leukocyte morphology was assessed via May-Grünwald Giemsa staining. Circulating cytokines and chemokines were quantified by ELISA. HD-derived PBMCs were exposed to plasma from EGPA patients, antiphospholipid-positive controls, or HDs. Cell death, metabolic activity, and cytokine production were evaluated using Trypan Blue, Annexin V/PI staining, MTT assays, and ELISA. Chemotaxis assays assessed cell migration in response to conditioned media, with or without Anakinra or CCR1 inhibitor J113863. Results: AAV patients showed increased immature neutrophils. Plasma from ANCA-positive EGPA patients induced PBMC death, inflammasome-related cytokine release, and secretion of chemotactic and proangiogenic factors. Conditioned media enhanced immune cell migration in a cytokine-dependent manner. Conclusion: These findings indicate persistent subclinical immune activation during AAV remission, particularly in ANCA-positive EGPA, suggesting a role for mononuclear cell-mediated inflammation in relapse risk and the potential utility of immune monitoring.