Disparate periphery and lung immune microenvironments induced monocyte-macrophage activation as a key factor in anti-synthetase syndrome-associated interstitial lung disease

BACKGROUND: Anti-synthetase syndrome (ASS) is an autoimmune disease characterized by serum anti-aminoacyl-tRNA synthetase (ARS) antibodies and frequently associated with recurrent and refractory interstitial lung disease (ASS-ILD). Its pathogenesis remains poorly understood. METHOD: Paired peripheral blood mononuclear cells (PBMCs) and bronchoalveolar lavage fluid (BALF) cells from 4 patients with new-onset ASS-ILD were analyzed by single-cell RNA sequencing (scRNA-seq), whereas paired plasma and BALF samples from 11 new-onset ASS-ILD patients underwent Pro-DIA proteomics. Ten patients with idiopathic pulmonary fibrosis (IPF) and 10 healthy donors served as disease and healthy controls (HC) for the proteomics. In vitro experiments were performed to support the multi-omics findings. RESULT: Integrated scRNA-seq (117,960 PBMCs and 91,230 BALF cells) and Pro-DIA proteomics (4,629 plasma and 3,919 BALF proteins) analyses were performed. In ASS-ILD BALF, monocyte-macrophages showed significantly higher cytokine and inflammatory gene module scores compared with IPF and HCs (all P < 2e-16). The proportion of alveolus pro-inflammatory monocyte-macrophages was markedly increased (52.7% vs. 6.2% in IPF and 3.6% in HC). These subsets exhibited upregulated tRNA aminoacylation pathway and elevated multiple ARS gene expressions (adj.P < 1e-5). BALF proteomics also revealed increased ARS-related proteins and inflammatory mediators in ASS-ILD (adj.P < 0.05). In vitro, human histidyl ARS-containing immune complexes (IC) induced higher expression of MHC-I/II, CD80/CD86, pro-inflammatory cytokines, and ARS genes in primary human macrophages compared with control IC. CONCLUSION: Our findings provide novel insights into ASS-ILD pathogenesis and suggest the underlying pathogenicity of ARS containing IC. These results underscore the role of monocyte-macrophage activation in ASS-ILD related immunopathogenesis and pave the way for targeted therapies directed at IC formation and aberrant monocyte-macrophage dynamics.