RETROVIRAL-MEDIATED TRANSDUCTION OF ENDOTHELIAL-CELLS WITH THE LAC-Z GENE IMPAIRS CELLULAR PROLIFERATION IN-VITRO AND GRAFT ENDOTHELIALIZATION IN-VIVO

Purpose: An endothelialized lumen within a synthetic graft that expres ses recombinant proteins with anticoagulant or antiproliferative activ ity has the potential to improve graft function. However, preliminary data suggest that-genetic modification of endothelial cells (ECs) impa irs their proliferation. The purpose of this investigation was to test the hypothesis that retroviral transduction of cultured ECs with the lac Z gene encoding for beta-galactosidase would decrease EC prolifera tion in vitro and graft endothelialization in vivo. Methods: In vitro studies compared canine EC proliferation over a 14-day period among ea rly-passage ECs (two and three) and late-passage ECs (six and nine) tr ansduced with the BAG vector (containing the lac Z gene and the neomyc in resistance gene), ECs transduced with the neomycin resistance gene only, and nontransduced ECs. In vivo canine studies assessed endotheli alization of expanded polytetrafluoroethylene thoracoabdominal grafts seeded with autologous lac Z-transduced ECs (n = 7) or nontransduced E Cs (n = 3) compared with that of nonseeded grafts (n = 3). Histochemic al staining and DNA polymerase chain reaction was used 6 weeks after i mplantation to detect the presence of the lac Z gene in the grafts' ce llular Linings and perigraft tissues. Endothelialization was assessed by Light microscopy and electron microscopy. Results: Proliferation of late-passage lac Z-transduced ECs in vitro was significantly decrease d compared with neomycin resistance-transduced ECs or nontransduced EC s. Among early-passage ECs smaller but significant decreases in prolif eration were noted among (Rc Z-transduced cells compared with nontrans duced cells. Six of seven expanded polytetrafluoroethylene grafts seed ed with transduced ECs showed lac Z gene expression. Lac Z gene expres sion mas not found on grafts seeded with nontransduced ECs or nonseede d grafts. The endothelialized luminal surface area was significantly l ess in grafts seeded with lac Z-transduced ECs compared with grafts se eded with nontransduced ECs. Conclusions: Retroviral-mediated transduc tion of canine ECs with the lac Z gene encoding for beta-galactosidase impairs EC proliferation in vitro and the ability of transduced ECs t o form a confluent EC monolayer on the luminal surface of synthetic gr afts in vivo.