A Chimeric Airway Model Enables Evaluation of Essential Genes In Vivo

A pseudostratified epithelium lines the upper airways and is maintained by stem cells that express a basal cell phenotype. These airway stem cells share molecular and functional similarities with other tissue stem cells, particularly those that are involved in development or postnatal maintenance of the esophagus and epidermis. Thus, analysis of gene function in the airway is complicated by the potential for lethal non-respiratory phenotypes. Conditional genetic approaches employing cell type-specific and/or temporally controlled expression systems provide some options, but limitations remain. To overcome these concerns, we tested a novel microsurgical approach in which a chimeric airway is created by orthotopic transplantation of transgenic mouse tissue into the trachea. We hypothesized that precise temporal and spatial control of gene expression in the pseudostratified tracheal epithelium would prevent life-threatening collateral tissue injury and allow analysis of essential genes. First, we demonstrated that the graft was revascularized allowing parenteral administration of tamoxifen. Second, we transplanted tracheal tissue from Krt5CreERT2; Rosa26mTmG or Col1a2CreERT2; Rosa26mTmG donors into wild type mice and established that tamoxifen-dependent recombination in basal cells or fibroblasts was highly efficient and restricted to the graft. Finally, we assessed the feasibility of knocking out an essential gene, Itgb1, by transplanting tracheal segments from Krt5CreERT2; Rosa26mTmG; Itgb1flox/flox transgenic mice into wild type mice. We demonstrated highly selective recombination in the graft and long-term survival. We conclude that the chimeric airway model allows analysis of essential gene function in the airway and has the potential to be a versatile tool for preclinical testing of targeted gene therapies.