Adenoviral gene transfer of a u-PA receptor-binding plasmin inhibitor and green fluorescent protein: Inhibition of migration and visualization of expression

Smooth muscle cell migration plays a role in the development of intimal hyp erplasia. Given the established role of the plasminogen activation system i n cell migration, an approach to therapy is to overexpress an inhibitor of plasmin. Therefore, an adenoviral vector was constructed encoding the hybri d protein ATF.BPTI, which contains the active domain of bovine pancreas try psin inhibitor (BPTI), fused to ATF, the amino terminal fragment or recepto r-binding domain of u-PA. Adenoviral vectors expressing ATF and BPTI indivi dually were also constructed, and a fourth vector was constructed encoding ATF.BPTI linked by an internal ribosomal entry sire to Green Fluorescent Pr otein (ABIG). Both the expression and functionality of the recombinant prot eins were established in human vascular smooth muscle cells. Adenoviral ene transfer of ATF.BPTI inhibited SMC migration more efficiently than the exp ression of ATF or BPTI individually. Expression of ABIG resulted in the co- expression of ATF.BPTI and Green Fluorescent Protein, thereby providing a t ool to monitor transfection efficiency and the behavior of the transfected cells.