Augmentation of pulmonary host defense against Pseudomonas by FcγRIIA cDNA transfer to the respiratory epithelium
Stefan Worgall, Petr Bezdicek, Moo-Kyung Kim, Jong-Gu Park, Ravi Singh, Melpo Christofidou-Solomidou, Alice Prince, Imre Kovesdi, Alan D. Schreiber, Ronald G. Crystal
Stefan Worgall, Petr Bezdicek, Moo-Kyung Kim, Jong-Gu Park, Ravi Singh, Melpo Christofidou-Solomidou, Alice Prince, Imre Kovesdi, Alan D. Schreiber, Ronald G. Crystal
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Augmentation of pulmonary host defense against Pseudomonas by FcγRIIA cDNA transfer to the respiratory epithelium

Abstract

Fcγ receptors on the surface of phagocytic cells bind the Fc region of IgG and mediate binding, phagocytosis, and destruction of particulate antigens opsonized by the antigen-specific IgG molecule. The present study evaluates the feasibility of converting lung epithelial cells into phagocytic cells using adenovirus (Ad) vector–mediated gene transfer of FcγRIIA cDNA to induce expression of the human FcγRIIA receptor. Binding and phagocytosis of opsonized sheep red blood cells (SRBCs) by the A549 human lung epithelial cell line after Ad-mediated FcγRIIA gene transfer was demonstrated using light and fluorescence microscopy and phagocytic assays with 51Cr-labeled SRBCs. When A549 cells were infected with an Ad vector expressing a FcγRIIA mutant in which 2 of 3 cytoplasmic tyrosines have been replaced with phenylalanine, only binding, but not phagocytosis, of opsonized SRBCs was observed. In vivo expression of FcγRIIA in the lung after intratracheal administration of the AdFcγRIIA enhanced clearance of opsonized Pseudomonas aeruginosa from the lung in normal rats and in mice deficient in Fcγ receptor expression. Similar results were observed with a chimeric FcγRIIA construct containing the extracellular domain of FcγRIIIA. Together, these data demonstrate that Ad-mediated FcγRIIA receptor cDNA expression can mediate the binding and phagocytosis of opsonized particulate antigens by normally nonphagocytic cells, suggesting that gene-transfer strategies might be used to utilize nonphagocytic cells to clear bacteria or other opsonized particulate antigens from the respiratory tract.

Authors

Stefan Worgall, Petr Bezdicek, Moo-Kyung Kim, Jong-Gu Park, Ravi Singh, Melpo Christofidou-Solomidou, Alice Prince, Imre Kovesdi, Alan D. Schreiber, Ronald G. Crystal

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Enhanced clearance of P. aeruginosa from the lung after Ad-mediated tran...
Enhanced clearance of P. aeruginosa from the lung after Ad-mediated transfer of FcγRIIA and FcγRIIIA cDNA to the respiratory epithelial surface of experimental animals. (a) Enhanced clearance in normal rats. Sprague-Dawley rats were infected intratracheally with AdFcγRIIA, AdFcγRIIIA, or AdNull (109 PFU). After 48 hours, the animals were challenged via the intratracheal route with 108 CFU of the PAO1 strain of P. aeruginosa opsonized with rabbit anti-PAO1 polyclonal antibody. After 20 hours, animals were sacrificed, the lungs were removed, and bacterial counts of the lysates were determined. Data are expressed as CFU of PAO1 per gram of lung tissue. Shown are means of CFU per gram lung tissue from naive controls and AdNull-, AdFcγRIIA-, and AdFcγRIIIA-infected animals (n = 5 animals per group). (b) Enhanced clearance in Fcγ receptor–knockout mice. Fcγ receptor–knockout mice were infected intratracheally with AdFcγRIIA or AdNull (109 PFU). After 48 hours, the mice were infected intranasally with 4 × 107 CFU of the PAO1 strain of P. aeruginosa opsonized with rabbit anti-PAO1 polyclonal antibody. After 20 hours, animals were sacrificed, the lungs were removed, and bacterial counts of the lysates were determined. Data are expressed as CFU of PAO1 per gram of lung tissue. Shown are means of CFU per gram of lung tissue from AdNull-infected animals (n = 7) and AdFcγRIIA-infected animals (n = 10). The data in a and b represent mean ± SEM.