Increasing endothelial cell permeability improves the efficiency of myocyte adenoviral vector infection

Background Gene delivery to the myocardium using blood-borne adenoviral vec tors is hindered by the endothelium, which represents a barrier limiting th e infection rate of underlying myocytes. However, endothelial permeability may be modulated by pharmacological agents. Methods In the present study, we modeled the endothelial barrier in vitro u sing a human umbilical vein endothelial cell (HUVEC) monolayer seeded on a Transwell membrane as a support and diffusion of fluorescent dextrans as a permeability index. We used alpha -thrombin (100 nM) as a pharmacological a gent known to increase endothelial permeability and tested the barrier func tion of the endothelial cell monolayer on adenovector-mediated luciferase g ene transfer to underlying isolated cardiac myocytes. Results A confluent HUVEC monolayer represented a considerable physical bar rier to dextran diffusion; it reduced the permeability of the micropore mem brane alone to fluorescein isothiocyanate (FITC)-labeled dextrans of molecu lar weights 4, 70, 150 and 2000 kDa by approximately 54, 78, 88 and 98%, re spectively. alpha -Thrombin (100 nM) increased the permeability coefficient s (P-EC) by 276, 264, 562 and 4166% for the same dextrans, respectively. A confluent HUVEC monolayer represented a major impediment to adenovector-med iated luciferase gene transfer to cardiac myocytes, largely reducing gene t ransfer efficiency. However thrombin induced a nine-fold increase in myocyt e infection. Conclusion In our model, the endothelial cell monolayer represents a major impediment to myocyte adenovector-mediated gene transfer which can be parti ally improved by pharmacologically increasing endothelial permeability. The Transwell model is therefore particularly useful for testing the efficienc y of pharmacological agents in modulating adenovector passage through the e ndothelial barrier. Copyright lj 2000 John Wiley & Sons, Ltd.