Targeting CD8⁺ T cell-derived granzyme K alleviates radiation-induced pulmonary fibrosis by attenuating senescence and SASP

BACKGROUND: Radiation-induced pulmonary fibrosis (RIPF) is a severe complication lacking effective treatment after thoracic radiotherapy. Cellular senescence and the senescence-associated secretory phenotype (SASP) are central drivers of this condition. Granzyme K (GZMK) is an inflammatory amplifier, yet its role in promoting RIPF by regulating cellular senescence remains unknown. METHODS: A time-course mouse model of radiation-induced lung injury was established using single-dose 15 Gy thoracic irradiation. Histopathology and molecular analyses were performed to assess injury and GZMK expression. Cellular origins of GZMK were identified via publicly available single-cell data and multiplex immunofluorescence. In vitro, mouse lung epithelial (MLE12) and alveolar macrophage (MH-S) cell lines were treated with recombinant GZMK. Senescence was evaluated by SA-β-gal staining, EdU assay, and expression of p53/p21/p16 and SASP factors via WB and RT-qPCR. From week 12 post-irradiation, the GZMK inhibitor PPACK was administered to evaluate its therapeutic effects. RESULTS: GZMK was significantly upregulated during the fibrotic phase and localized predominantly to infiltrating CD8⁺ T cells. In vitro, GZMK directly induced cellular senescence and activated the p53/p21/p16 pathway along with SASP factor expression. In vivo inhibition of GZMK markedly attenuated lung collagen deposition, myofibroblast activation, senescent cell burden, and levels of SASP factors (IL-6, TNF-α, TGF-β). CONCLUSION: This study demonstrates that CD8⁺ T cell-derived GZMK drives RIPF by promoting cellular senescence and SASP. Targeting GZMK alleviated fibrosis, highlighting its potential as a therapeutic target for radiation-induced lung injury.