In vivo retention of endothelial cells adenovirally transduced with tissue-type plasminogen activator and seeded onto expanded polytetrafluoroethylene

Abstract

Purpose: Seeding a prosthetic graft with genetically engineered vascular endothelial cells (ECs) has the potential to enhance the graft's antithrombotic properties. However, it has been reported that ECs transduced with tissue-type plasminogen activator (tPA) have very low levels of retention on grafts, probably because of increased proteolytic activity. We examined the retention of human tPA (htPA)-transduced ECs after the cells were seeded onto expanded polytetrafluoroethylene (ePTFE) grafts and implanted into dogs. We also examined the function of secreted htPA in this model. Methods and Results: Canine jugular venous ECs were transduced with adenoviral vectors encoding htPA (Adex1CAhtPA) and β-galactosidase (Adex1CALacZ). There was a positive relationship between the percentage of X-gal ECs staining and the multiplicity of infection (MOI) of Adex1CALacZ. The level of htPA production in vitro increased with the increasing MOI of Adex1CAhtPA, but decreased gradually 4 days after infection. ECs coinfected with Adex1CAhtPA and Adex1CALacZ (htPAEC) or ECs infected with Adex1CALacZ alone (LacZEC) were seeded onto ePTFE grafts at densities equivalent to confluence to visualize retained ECs in an in vivo flow study. The grafts were implanted into canine carotid arteries and harvested after 5 hours of exposure to blood flow. The harvested grafts showed patchy defects in ECs, most of which were covered with mural thrombi. There was no significant difference in retention between htPAEC (29.3% ± 8.7%) and LacZEC (19.5% ± 3.6%). There was a significant negative correlation between the in vivo EC retention on the grafts and the in vitro cellular passage level of ECs (P =.041; r = –.40). htPAEC produced 210.3 ± 22.2 ng htPA antigen/106 cells per 6 hours in vitro and continued to secrete htPA on the harvested graft. Conclusions: We demonstrated that a large amount of functional htPA was produced by adenovirally modified canine ECs. The results of the in vivo study may suggest that overexpression of tPA has little effect on the short-term retention of early passage ECs seeded onto ePTFE grafts. (J Vasc Surg 2000;32:353-63.)