Anlotinib ameliorates myositis-associated interstitial lung disease (MAILD) via suppression of the NETs-PI3K/Akt-driven epithelial-mesenchymal transition.

Myositis-associated interstitial lung disease (MAILD) is one of the most severe complications of idiopathic inflammatory myopathy, characterised by rapidly progressive pulmonary fibrosis and high mortality. Treatment options are limited, and mechanisms driving epithelial-mesenchymal transition (EMT) in MAILD are incompletely understood. Anlotinib, a multitarget tyrosine kinase inhibitor, shows potential in fibrotic diseases; however, its role and mechanism in MAILD need clarification. To assess anlotinib's therapeutic effects, we established a MAILD mouse model and a neutrophil extracellular trap (NET)-induced human alveolar epithelial cell (A549) model. H&E and Masson staining analysed lung pathological changes and collagen deposition. Immunohistochemistry, immunofluorescence and Western blot detected expressions of NETs markers (myeloperoxidase, citrullinated histone H3), phosphatidylinositol 3-kinase/protein kinase B (PI3K)/Akt components and EMT markers (E-cadherin, α-smooth muscle actin). RNA sequencing and gene set enrichment analysis identified differentially expressed genes and signalling pathways. Cell counting kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU) and wound healing assays assessed cellular repair and migration. MAILD mouse lungs showed structural damage, inflammatory infiltration, collagen deposition, increased NETs formation, PI3K/Akt activation and enhanced EMT. Anlotinib significantly ameliorated pulmonary fibrosis and reduced pro-inflammatory cytokines (tumour necrosis factor-α (TNF-α), interleukin (IL)-6, IL-1β). In vivo and in vitro, anlotinib suppressed NETs formation, PI3K/Akt activation and EMT, while enhancing alveolar epithelial cell repair and migration. Anlotinib alleviates MAILD progression by inhibiting the NETs-PI3K/Akt axis and subsequent EMT, providing a theoretical basis for drug repurposing and supporting its clinical translation potential in MAILD.