USP11 exacerbates radiation induced pneumonitis by activating endothelial cell inflammatory response via OTUD5-STING signaling.

PURPOSE: Radiation induced pneumonitis (RIP) seriously limits the application of radiotherapy in the treatment of thoracic tumors, and its etiology and pathogenesis remain elusive. This study aims to elucidate the role of Ubiquitin-specific peptidase 11 (USP11) in the progression of RIP and the associated underlying mechanisms.

METHODS AND MATERIALS: The changes of cytokines and infiltrated immune cells were detected by ELISA and IHC after exposed to 20 Gy X-ray with whole thorax irradiation. USP11 expression on endothelial cell proliferation and apoptosis were analyzed by co-staining of CD31/Ki67 and CD31/Caspase-3 in vivo and the production of cytokines and ROS were confirmed by RT-PCR and flow cytometry in vitro. Comprehensive proteome and ubiquitinome were employed for USP11 substrate screening post-radiation. Results were verified by Western blot and Co-IP experiments. rAAV-lung vectors expressing OTUD5 were utilized for localized overexpression of OTUD5 in mouse pulmonary, and IHC were conducted to analyze cytokines expression.

RESULTS: The progression of RIP was significantly alleviated by reduced the expression of proinflammatory cytokines in both USP11 knockout (Usp11-/- ) and the USP11 inhibitor mitoxantrone treated mice. Likewise, absence of USP11 resulted in decreased permeability of pulmonary vessels and neutrophils and macrophages infiltration. The proliferation rates of endothelial cells were prominently increased in the Usp11-/- lung, while the apoptosis decreased compared to the Usp11+/+ following irradiation. Conversely, USP11 overexpression increased proinflammatory cytokines and ROS production in endothelial cells post-radiation. Comprehensive proteome and ubiquitinome analyses indicated that USP11 overexpression upregulated several deubiquitinating enzymes, including USP22, USP33 and OTUD5. We clarified that USP11 deubiquitinating OTUD5 and implicate OTUD5-STING signaling pathway in the progression of inflammatory response in endothelial cells.

CONCLUSIONS: USP11 exacerbated RIP by triggering an inflammatory response in endothelial cell both in vitro and in vivo, and OTUD5-STING pathway was involved in USP11 promotes RIP. This study provided experimental support for the development of precision intervention strategies targeting USP11 to mitigate RIP.