Mechanistic analysis of an IRF7-dependent pathway in virus-induced fibrosis in chronic lung allograft dysfunction

BACKGROUND: Chronic lung allograft dysfunction (CLAD) significantly limits long-term survival of lung transplant recipients, with viral infections acting as critical contributors to its pathogenesis. The mechanisms linking viral infections to CLAD-associated airway fibrosis remain incompletely understood. This study investigates the role of the type I interferon (IFN) master regulator IRF7 in virus-induced airway fibrogenesis. METHODS: CLAD (Bronchiolitis obliterans syndrome (BOS)), Stable LTx, and non-transplanted lung tissues were analysed by spatial transcriptomics (GeoMx; n = 5 CLAD (BOS), n = 3 Stable LTx, n = 3 controls), Western blotting, and immunostaining. Primary bronchial epithelial cells in air-liquid-interface (ALI) culture and a human precision-cut lung slice (PCLS) ex vivo model were exposed to Influenza A virus (IAV) with or without IRF7 silencing, IL-33 blockade, or MMP-9 inhibition. Group comparisons used Mann-Whitney tests and one-way ANOVA with Tukey's post hoc test; spatial differential expression used linear models with Benjamini-Hochberg correction (α = 0.05). FINDINGS: Spatial transcriptomics identified enrichment of IFN-stimulated genes including IRF7, STAT1, IFI44L, and GBP1 in the CLAD (BOS) epithelial compartment alongside antiviral effector genes (DDX58, TLR3) and pro-fibrotic programmes (TGFB1, SMAD2/3, ACTA2). Western blotting confirmed significantly increased IRF7 and phosphorylated IRF7 with airway-centric distribution in CLAD (BOS) (p < 0.01; n = 3-4). IAV exposure upregulated IRF7, α-SMA, SMAD2/3, and soluble collagen in ALI cultures (all p < 0.01; n = 3-6); IRF7 silencing attenuated these markers and reduced virus-induced IL-33 at mRNA and protein levels (p < 0.05). IL-33 blockade independently reduced α-SMA (p < 0.05), confirming IL-33 as a downstream IRF7 mediator. IAV increased MMP-9 secretion (p < 0.0001); IRF7 or IL-33 blockade attenuated MMP-9, while MMP-9 inhibition reduced α-SMA (p < 0.0001). All findings were replicated in human PCLS (p < 0.05; n = 5-6). INTERPRETATION: These findings delineate an IRF7-IL-33-MMP-9 axis linking viral infections to airway fibrogenesis, offering mechanistic insight into CLAD (BOS) pathogenesis. Therapeutic targeting of this pathway may mitigate fibrotic remodelling in lung transplant recipients. FUNDING: NIH 1R01HL161620 (N.S.S.). Caredx IIT (N.S.S).