Pulmonary Surfactant Inhibits Cationic Liposome-Mediated Gene Delivery to Respiratory Epithelial CellsIn Vitro

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Abstract
Cationic lipid-mediated transfection of the alveolar epithelium in vivo will require exposure of plasmid DNA and cationic lipids to endogenous surfactant lipids and proteins in the alveolar space. Effects of pulmonary surfactant and of surfactant constituents on transfection in vitro of two respiratory epithelial cell lines (MLE-15 and H441) with a plasmid encoding the luciferase reporter gene were studied using two cationic lipid formulations: 1,2-dimyristyloxypropyl-3-dimethyl-hydroxyethyl ammonium bromide/cholesterol (DMRIE/C) and 1,2-dioleoyl-3-trimethylammonium propane/dioleoyl phosphatidylethanolamine (DOTAP/DOPE). Gene expression, as assessed by luciferase activity, decreased as increasing concentrations of natural surfactant were added to cationic lipid–DNA complexes. Incorporation of phospholipids DOPC/DOPG or surfactant proteins SP-B or SP-C in the cationic lipid formulation inhibited transfection. A fluorescent lipid mixing assay was used to determine the effects of surfactant proteins SP-B and SP-C on mixing between cationic lipid–DNA complexes and surfactant lipid vesicles. Mixing between DOPC/DOPG vesicles and cationic lipid–DNA complexes in the absence of added proteins amounted to 10–20%. Addition of SP-B or SP-C increased the mixing of DOPC/DOPG vesicles with DOTAP/DOPE–DNA complexes, but not DMRIEC–DNA complexes. These results demonstrate that pulmonary surfactant lipids and proteins inhibit transfection with cationic lipid–DNA complexes in vitro, and may therefore represent a barrier to gene transfer in the lung. Pulmonary surfactant lining the alveoli and respiratory airways may pose a barrier to successful gene therapy in the lung. In the present work, the effects of pulmonary surfactant and its constituent lipids and proteins on cationic lipid-mediated gene transfer to pulmonary epithelial cells was studied in vitro. Pulmonary surfactant lipids and proteins SP-B and SP-C inhibited transfection of two respiratory epithelial cell lines. Lipid mixing occurred in vitro between pulmonary surfactant lipids and cationic lipid–DNA complexes. Incorporation of components of pulmonary surfactant into cationic lipid–DNA complexes reduced transfection efficiency. These results suggest that the efficacy of cationic lipid–DNA complexes may be limited by pulmonary surfactant lining the human airway epithelium.