Endothelial NEDD4L exacerbates acute lung injury by targeting A20 for ubiquitination degradation

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) are life-threatening critical diseases driven by uncontrolled inflammation and endothelial dysfunction, with limited effective targeted therapies available. NEDD4L, an E3 ubiquitin ligase, has been linked to ALI pathogenesis, but its role in endothelial cells remains undefined. Here, we explored the function and mechanism of endothelial NEDD4L in ALI. Single-cell RNA sequencing of a sepsis-induced ALI mouse model and in vitro assays revealed marked upregulation of NEDD4L in endothelial cells under ALI-related pathological stimuli (LPS, TNFα, H₂O₂). Endothelial cell-specific NEDD4L knockdown (NEDD4Lᴷᴰ) in mice significantly alleviated LPS-induced lung injury, as evidenced by reduced pathological damage, inflammatory infiltration, cytokine release and improved survival, and similar protective effects were observed in a CLP-induced ALI model. In vitro experiments confirmed that NEDD4L knockdown suppressed TNFα-induced endothelial activation and leukocyte adhesion. Mechanistically, NEDD4L directly bound to the anti-inflammatory protein A20, promoting its polyubiquitination and proteasomal degradation, which impaired A20-mediated inhibition of the JNK/p38/NF-κB pathway and upregulated the expression of pro-inflammatory cytokines and adhesion molecules. Rescue assays verified that A20 overexpression partially reversed NEDD4L induced pro-inflammatory effects. Collectively, our study identifies a novel NEDD4L-A20 axis in endothelial cells that contributes to ALI progression by modulating inflammatory signaling and endothelial dysfunction. Targeting this regulatory axis may provide a potential precise and cell-selective therapeutic strategy for alleviating ALI/ARDS.