LAUSR

Natural airway defences currently impede the efficacy of viral vector-mediated airway gene therapy. Conditioning airways prior to vector delivery can disrupt these barriers, improving viral vector access to target receptors and airway stem cells. This study aimed to assess and quantify the in vivo histological and gene transfer effects of physical perturbation devices to identify effective conditioning approaches. A range of flexible wire baskets with varying configurations, a Brush, biopsy forceps, and a balloon catheter were examined. We first evaluated the histological effects of physical perturbation devices in rat tracheas that were excised 10 minutes after conditioning. Based on the histological findings, a selection of devices were used to condition rat tracheas in vivo before delivering a lentiviral vector containing the LacZ reporter gene. After 7 days, excised tracheas were X-gal processed and examined en face to quantify the area of LacZ staining. Histological observations 10 minutes after conditioning found that physical perturbation dislodged cells from the basement membrane to varying degrees, with some producing significant levels of epithelial cell removal. When a subset of devices were assessed for their ability to enhance gene transfer, only the NGage® wire basket (Cook Medical) produced a significant increase in the proportion of X-gal-stained area when compared to unconditioned tracheas (8-fold, p = 0.00025). These results suggest that a range of factors contribute to perturbation-enhanced gene transfer. Overall, this study supports existing evidence that physical perturbation can assist airway gene transfer, and will help to identify the characteristics of an effective device for airway gene therapy.