Davies, L. A., Varathalingam, A., Cheng, S. H., Hyde, S. C. & Gill, D. R.
Journal of Cystic Fibrosis, 3, S29Download Back
The murine lung has served as a model for the evaluation of a variety of gene therapy vectors for cystic fibrosis lung disease. Many studies have focussed on enhancing overall gene expression in the lung but few have examined if expression can be demonstrated in the ciliated epithelial cells believed to be the major target for cystic fibrosis gene therapy. We evaluated transgene expression in the murine lung using vectors incorporating the red-shifted variant of green fluorescent protein (EGFP). BALB/c mice were dosed intranasally with 300µg of the plasmid pEGFP-N1 or 80µg of pEGFP-N1 complexed to the cationic lipid GL67 (Genzyme, Framingham, USA). 24 hours after vector administration, 7µm cryosections were prepared from lung tissues and examined for GFP expression. GFP positive cells were identified in all treated mice with a mean expression of 347±280 positive cells in sections from animals dosed with pEGFP-N1. The majority (79%) of these cells were located in the epithelium of conducting airways and the remainder (21%) were located in the lung parenchyma. Subsequent immunohistochemical analysis revealed that most of these GFP positive cells were ciliated epithelial cells. In mice treated with DNA/GL67, 2,020 ±435 GFP positive cells per mouse were observed. In contrast to naked DNA the majority (99.4%) of GFP positive cells were located in the lung parenchyma and only a very small number (0.6%) were observed in the conducting airway epithelium. Antibody staining demonstrated that the majority of GFP positive cells in mice treated with DNA/GL67 were Type I pneumocytes. These data reflect that GFP expression is a robust first step in the detection and identification of cell types targeted by gene transfer vectors in the mouse lung.