Background <p>Acute lung injury (ALI) represents a life-threatening condition triggered by diverse etiological factors. The expression and function of miR-769-5p within the context of ALI remain poorly characterized.</p> Purpose <p>This study aims to examine the expression levels of miR-769-5p in the serum of ALI patients and assess its synergistic diagnostic potential when combined with the lung ultrasound (LUS) score for ALI.</p> Methods <p>Serum miR-769-5p expression levels were quantified using reverse transcription quantitative polymerase chain reaction (RT-qPCR). The diagnostic accuracy of miR-769-5p and the LUS score for ALI was evaluated via receiver operating characteristic (ROC) curve analysis. Binary logistic regression was used to analyze their impact on ALI. Serum nuclear factor kappa B (NF-κB) and reactive oxygen species (ROS) levels were measured by enzyme-linked immunosorbent assay (ELISA). Pearson correlation assessed relationships between miR-769-5p and NF-κB, ROS, and LUS score. In lipopolysaccharide (LPS)-A549 cells, RT-qPCR and flow cytometry evaluated how miR-769-5p modulated NF-κB, ROS, and apoptosis.</p> Results <p>ALI patients exhibited significantly decreased serum miR-769-5p expression. Combining miR-769-5p with the LUS score enhanced diagnostic accuracy, with both correlating significantly with ALI risk. Serum NF-κB and ROS levels were elevated in ALI and inversely correlated with miR-769-5p. In LPS-A549 cells, miR-769-5p overexpression suppressed LPS-induced NF-κB activation, ROS generation, and cellular apoptosis.</p> Conclusions <p>The combined application of miR-769-5p and the LUS score demonstrated enhanced diagnostic efficacy. Furthermore, miR-769-5p overexpression effectively attenuated NF-κB activation, ROS production, and cellular apoptosis.</p>