Rationale: Acute lung injury and Acute Respiratory Distress Syndrome are common clinical syndromes resulting largely from the accumulation of and inability to clear pulmonary edema, due to injury to the alveolar epithelium. Gene therapy may represent an important alternative for the treatment and prevention of these diseases by restoring alveolar epithelial function. We have recently developed an electroporation strategy to transfer genes to the lungs of mice with high efficiency and low inflammation. Objectives: We asked whether electroporation-mediated transfer of genes encoding subunits of the Na⁺,K⁺-ATPase could protect from LPS-induced lung injury or can be used to treat already injured lungs by upregulating mechanisms of pulmonary edema clearance. Methods: Plasmids were delivered to the lungs of mice using transthoracic electroporation. Lung injury was induced by intratracheal administration of LPS (4 mg/ kg BW). Biochemical, cellular, and physiologic measurements were taken to assess gene transfer and lung injury. Measurements and Main Results: Improvements in wet-to-dry ratios, pulmonary effusions, BAL protein levels and cellularity, alveolar fluid clearance, and respiratory mechanics were seen following delivery of plasmids expressing Na⁺,K⁺-ATPase subunits, but not control plasmids, in LPS-injured lungs. Delivery of plasmids expressing Na⁺,K⁺-ATPase subunits both protected from subsequent lung injury andpartially reversed existing lung injury by these measures. Conclusions: These results demonstrate that electroporation can be used effectively in healthy and injured lungs to facilitate gene delivery and expression. To our knowledge, this is the first successful use of gene delivery to treat existing lung injury and may have future clinical potential.