Abstract:Objective: To construction of recombinant adenovirus containing KDR promoter-HSV-tk suicide gene by using a pAdeasy system and evaluate its specific killing effect on vascular endothelial cells. Methods: By using pAdeasy system, recombinant adenoviral plasmid containing KDR or cytomegalovirus (CMV) promoter-controlled HSV-tk gene (AdKDR-tk and AdCMV-tk) was constructed. After packaging and amplification in 293 cells, the virus was used to infect KDR-expressed human umbilical venous endothelial cells (HUVEC) and KDR-unexpressed HepG2. Following administration of ganciclovir(GCV), the survival rate of gene-transfected HUVEC and HepG2 was evaluated by using MTT method.
Results: The pAdeasy System produced a higher titer of the recombinant adenovirus (1×1010pfu/ml). Under infection index of 100, with increasing GCV concentration from 0 up to 50ug/ml, the survival rate of AdKDR-tk-transfected HUVEC and HepG2 decreased from 100% to (28.94±5.67)% and(75.45±2.91)%, respectively (P<0.01), while the survival rate of AdCMV-tk-transfected HUVEC and HepG2 declined from 100% to (17.56±2.48)% and (23.15±5.72)%, respectively (P>0.05). Conclusions: KDR promoter-HSV-tk gene possess a specific killing effect on vascular endothelial cells treated with GCV.