Acute lung injury (ALI), which can progress to the highly lethal acute respiratory distress syndrome (ARDS), remains a condition with limited pharmacological interventions, highlighting the urgent need for mechanism-informed repurposing strategies. Given the reported inhibitory effects of digoxin on nuclear factor kappa B (NF-κB)-mediated inflammation, we assessed its prophylactic and therapeutic efficacy in a murine model of lipopolysaccharide (LPS)-induced ALI. This evidence concerns the gene NFKB1 and acute lung injury.