Inflammatory injury is a hallmark of sepsis-induced acute respiratory distress syndrome (ARDS)/acute lung injury (ALI). However, the mechanisms underlying inflammatory injury remain obscure. Here, we developed the novel strategy to suppress lung inflammation through maintaining microvascular endothelial barrier integrity. VE-cadherin is the main adherens junction protein that interacts with beta-catenin and forms a complex. We found that lung inflammation was accompanied by decreased VE-cadherin expression and increased beta-catenin activity in animal models and human pulmonary microvascular endothelial cells (HPMECs), illuminating the relationship among VE-cadherin/beta-catenin complex, microvascular endothelial barrier integrity, and inflammation. Furthermore, we showed that the VE-cadherin/beta-catenin complex dissociated upon lung inflammation, while Sirt3 promoted the stability of such a complex. Sirt3 was decreased during lung inflammation in vivo and in vitro. Sirt3 deficiency not only led to the downregulation of VE-cadherin but also enhanced the transcriptional activity of beta-catenin that further increased beta-catenin target gene MMP-7 expression, thereby promoting inflammatory factor COX-2 expression. Sirt3 overexpression promoted VE-cadherin expression, inhibited beta-catenin transcriptional activity, strengthened the stability of the VE-cadherin/beta-catenin complex, and suppressed inflammation in HPMECs. Notably, Sirt3 deficiency significantly damaged microvascular endothelial barrier integrity and intensified lung inflammation in animal model. These results demonstrated the role of Sirt3 in modulating microvascular endothelial barrier integrity to inhibit inflammation. Therefore, strategies that aim at enhancing the stability of endothelial VE-cadherin/beta-catenin complex are potentially beneficial for preventing sepsis-induced lung inflammation.