Objective: Micro-RNA (miR)-155 is a novel gene regulator with important roles in inflammation. Herein, our study was aimed at exploring the role of miR-155 in lipopolysaccharide (LPS)-induced acute lung injury(ALI). Measurements and Results: ALI in mice was induced by intratracheally delivered LPS. Loss-of-function experiments performed on miR-155 knockout mice showed that miR-155 gene inactivation protected mice from LPS-induced ALI as manifested preserved lung permeability and reduced lung inflammation compared with wild-type controls. Bone marrow transplantation experiments identified leukocytes, but not lung parenchymal derived miR-155 promoted acute lung inflammation. Real-time PCR analysis showed that the expression of miR-155 in lung tissue was greatly elevated in wild-type mice after LPS stimulation. In situ hybridization showed that miR-155 was mainly expressed in alveolar macrophages. In vitro experiments performed in isolated alveolar macrophages and polarized bone marrow derived macrophages confirmed that miR-155 expression in macrophages was increased in response to LPS stimulation. Conversely, miR-155 gain-of-function in alveolar macrophages remarkably exaggerated LPS-induced acute lung injury. Molecular studies identified the inflammation repressor SOCS-1 as the downstream target of miR-155. By binding to the 3'-UTR of the SOCS-1 mRNA, miR-155 downregulated SOCS-1 expression thus permitting the inflammatory response during lung injury. Finally, we generated a novel miR-155 knockout rat strain and showed that the pro-inflammatory role of miR-155 was conserved in rats. Conclusion Our study identified miR-155 is a pro-inflammatory factor after LPS stimulation, and AM-derived miR-155 has an important role in LPS-induced ALI.