Recombinant adeno-associated virus vector-transduced vascular endothelial cells express the thrombomodulin transgene under the regulation of enhanced plasminogen activator inhibitor-1 promoter.
We were able to facilitate plasminogen activator inhibitor 1 (PAI-1) promoter activity approximately by 14-fold using an enhancer element. This enhanced PAI-1 promoter has a strong basal activity, comparable to CAG promoter activity, and has a response similar to the PAI-1 promoter with respect to TGFbeta 1 and TNFalpha stimulation. The characteristics of the enhanced PAI-1 promoter are thought to be suited to timely and tissue-specific expression of anticoagulant molecules in the vascular cells. Thus, we developed recombinant adeno-associated virus (rAAV) vectors using the enhanced PAI-1 promoter and were successful in transducing vascular endothelial cells to express the thrombomodulin transgene under the regulation of the enhanced PAI-1 promoter in vitro. Thromobomodulin transgene expression driven by the enhanced PAI-1 promoter in rAAV vector-transduced cultured endothelial cells was between 600- and 1000-fold higher than constitutive thrombomodulin gene expression in cultured human umbilical vein endothelial cells and was up-regulated by TGFbeta1 and TNFalpha stimulation which may down-regulate endogenous thrombomodulin gene expression in endothelial cells. The brain vascular endothelial cells of Mongolian gerbils could also be transduced by the same rAAV vector in vivo. Transduction of endothelial cells by rAAV vectors to express enhanced PAI-1 promoter-driven transgenes may be a useful gene therapy approach for vascular diseases.
Mimuro
J
J
Division of Cell and Molecular Medicine, Center for Molecular Medicine, Jichi Medical School, Tochigi-ken Japan.
Muramatsu
S
S
Hakamada
Y
Y
Mori
K
K
Kikuchi
J
J
Urabe
M
M
Madoiwa
S
S
Ozawa
K
K
Sakata
Y
Y
D000818:Animals; D017536:Carotid Artery, Common; D002417:Cattle; D002478:Cells, Cultured; D000229:Dependovirus / Q000235:genetics*; D004730:Endothelium, Vascular / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D005822:Genetic Vectors / Q000008:administration & dosage*; D005849:Gerbillinae; D006801:Humans; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics*; D011401:Promoter Regions, Genetic; D018180:Thrombomodulin / Q000235:genetics*; D014161:Transduction, Genetic; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D053773:Transforming Growth Factor beta1; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D017395:Plasminogen Activator Inhibitor 1; C508960:TGFB1 protein, human; D018180:Thrombomodulin; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D014409:Tumor Necrosis Factor-alpha