B19-06 Mgst2Hi PMNS Mediate Nets and Drive Pericyte-to-Myofibroblast Transition in RA-ILD

Abstract Objective Neutrophil extracellular trap (NET) formation is implicated in the pathogenesis of rheumatoid arthritis-associated interstitial lung disease (RA-ILD), yet the specific polymorphonuclear leukocyte (PMN) subsets involved and the mechanisms remain incompletely understood. Methods We established a zymosan A (ZYM)-induced interstitial pneumonia arthritis model and quantified NET-releasing polymorphonuclear leukocytes (PMNs) in lung, bronchoalveolar lavage fluid (BALF), and peripheral blood (PB). Microsomal glutathione S-transferase 2 (Mgst2) was selectively silenced in PMNs in vivo using a Cd11b-driven adeno-associated virus 9 (AAV9) shRNA vector. Additionally, Mgst2 interference was performed in HL-60 cells to assess its impact on NET formation and the transformation pericytes into myofibroblasts. Clinical relevance was examined using least absolute shrinkage and selection operator (LASSO) regression and receiver operating characteristic (ROC) curve analysis. Results We identified a significantly increased population of NET-releasing PMNs expressing high levels of Mgst2 (Mgst2hi PMNs) in the lungs, BALF, and PB of interstitial pneumonia arthritis mouse models. Targeted knockdown of Mgst2 in PMNs via an AAV9-shRNA-Mgst2 effectively mitigated pulmonary progression by inhibiting NET formation. Mechanistically, Mgst2 promoted NET release in neutrophil-differentiated HL-60 cells via a NADPH oxidases 2 (NOX2)-dependent pathway. Furthermore, NETs from these cells induced transdifferentiation of primary human pulmonary microvascular pericytes into myofibroblasts by activating the transforming growth factor-beta (TGF-β) pathway. Clinically relevant elevations of Mgst2hi NET-releasing PMNs were also confirmed in the lungs, BALF, and PB of RA-ILD patients. Employing LASSO regression analysis and ROC curves, we identified key diagnostic parameters, including high-resolution computed tomography (HRCT) metrics (lung vessel surface Std HU, 6mm, lung vessel diameter mean 9mm, lung vessel diameter Std 9mm), pulmonary function test (PFT) parameters (FVC% predicted), and the proportion of Mgst2+CD66b+ PMNs in PB, significantly enhancing diagnostic accuracy for RA-ILD. Conclusions these findings demonstrate a critical pathogenic role for Mgst2hi PMNs in RA-ILD and suggest their utility as both a diagnostic biomarker and potential therapeutic target through modulation of NET formation. This abstract is funded by: National Natural Science Foundation of China (No. 82460200),(No. 82500103)