Objective To study the effect of Honokiol (HKL) on pulmonary microvascular endothelial cells in lipopolysaccharide (LPS)-induced acute respiratory distress syndrome (ARDS) and its potential mechanism. Methods Mouse lung microvascular endothelial cells (PMVECs) were cultured with DMEM+10%FBS in a six-well plate and divided into control (Con) group 1, Honokiol (HKL) group 1, LPS treated (LPS) group 1, and LPS+HKL treatment (HKL+LPS) group 1. The levels of malondialdehyde (MDA) and reactive oxygen species (ROS) in cell lysates were determined by lipid peroxidation assay kit and H2DCF-DA, respectively. TUNEL/DAPI double staining was used to detect apoptosis. Cell junctions were visualized via VE-cadherin/DAPI and Claudin-5/DAPI double staining. Western blot was used to detect caspase-3, cleaved caspase-3, Sirt3, SOD2, and acetylated SOD2 (Ac-SOD2) expression. Thirty-two mice were randomly divided into control (Con) group 2, Honokiol (HKL) group 2, LPS treated (LPS) group 2, and LPS+HKL treatment (HKL+LPS) group 2. Hematoxylin and eosin (HE) staining was used to observe pathological changes to the lung tissue. Results HKL pretreatment significantly reversed the LPS-induced increase in ROS and MDA levels(P<0.05), SOD2 acetylation and Sirt3 down-regulation(P<0.05). TUNEL and caspase analysis showed that HKL protected against the apoptosis of PMVECs induced by LPS. VE-cadherin fluorescence staining demonstrated that HKL pretreatment prevented LPS from disrupting cell adhesion junctions. Claudin-5 fluorescence staining showed that HKL pretreatment prevented LPS from disrupting the tight junctions between cells. In the animal experiments, HE staining showed that HKL significantly inhibited the typical pathological changes of ARDS in the lung tissue of mice in the LPS group. Conclusions HKL can significantly inhibit the LPS-induced oxidative stress, apoptosis, and cell-connection breakdown of PMVECs, thereby alleviating ARDS symptoms.