Integrated single-cell TCR analysis across tissue identifies GZMK+CD8+ T cells promoting fibrosis via TGF-β signalling in IPF.

Abstract

Idiopathic pulmonary fibrosis (IPF) is a fatal interstitial lung disease with progressive scarring and an unknown aetiology, but the cross-tissue immune network remains poorly understood. Here, we performed paired single-cell RNA sequencing and T cell receptor profiling of lung, mediastinal lymph node, and peripheral blood samples from patients with severe IPF. We identified a GZMK⁺CD8⁺ T cell subset enriched in fibrotic lung tissue with an inflammatory, low-cytotoxic phenotype, and a clonally related CD8_HSPA1A population enriched in lymph nodes, suggesting a cross-tissue clonal connection. Higher GZMK⁺CD8⁺ T cell signatures were associated with worse survival and impaired lung function. Functionally, GZMK overexpressing CD8⁺ T cells promoted fibroblast-to-myofibroblast differentiation and proliferation through TGF-β1/Activin A signalling, whereas GZMK knockdown attenuated these responses. Importantly, genetic ablation of GZMK markedly reduced collagen deposition and lung fibrosis in bleomycin-induced mice. Furthermore, inhibition of TGFβR1/ALK4 using the inhibitor TEW-7197 effectively suppressed fibroblast activation in vitro and significantly attenuated pulmonary fibrosis in bleomycin-induced mice. Together, these findings suggest a potential profibrotic immune-stromal circuit in severe IPF in which GZMK⁺CD8⁺ T cells promote fibroblast activation through TGF-β1/Activin A signalling, highlighting this pathway as a potential therapeutic target.