Proteomic analysis identifies novel molecular signatures and immune-metabolic pathways in rheumatoid arthritis-associated interstitial lung disease

Rheumatoid arthritis (RA)-associated interstitial lung disease (RA-ILD) is a severe extra-articular manifestation of RA characterized by complex pathogenesis and limited therapeutic options. The present study aimed to identify circulating serum proteins that may reveal novel molecular mechanisms underlying RA-ILD and inform the development of disease-modifying strategies. A multi-center cohort study was conducted including patients with RA-ILD (n=40), patients with RA but without ILD (n=40) and healthy controls (n=7). Pooled serum samples were analyzed using a high-throughput antibody array targeting 440 proteins. Differentially expressed proteins were defined by statistical criteria (P<0.05) and a fold change >1.2 or <0.83. Functional enrichment and protein-protein interaction (PPI) analyses were performed to explore associated biological pathways. A total of 20 proteins that showed a stepwise increase in expression were identified: Levels were significantly higher in patients with RA compared with healthy controls and further elevated in patients with RA-ILD relative to RA alone. Hierarchical clustering and principal component analysis revealed distinct protein expression profiles across groups. Gene Ontology analysis indicated enrichment in pathways related to immune cell activation, proliferation and cytokine production. Kyoto Encyclopedia of Genes and Genomes pathway analysis highlighted cytokine-cytokine receptor interactions and PI3K-Akt signaling. PPI network analysis identified insulin as a central hub interacting with IGF-1R, IL-7, and other profibrotic mediators. Several proteins (for example, CA9, EDA-A2, Gas1, CRTAM, IL-2Rb and IL-31) emerged as novel candidates not previously linked to RA-ILD. The present study identified a panel of 20 dysregulated serum proteins in RA-ILD, implicating immune dysregulation, fibrotic processes and metabolic signaling-particularly the insulin/IGF-1R-PI3K-Akt axis-in disease pathogenesis. These findings provide potential therapeutic targets for RA-ILD that warrant further validation.