Delivery and expression of fluid shear stress-inducible promoters to the vessel wall: applications for cardiovascular gene therapy.
In atherosclerosis, endothelial cells at sites of stenosis experience elevated levels of shear stress. We have constructed a series of shear stress-inducible transcription units (SITUs) expressing the luciferase reporter gene and determined their activation by fluid shear stress in transfected endothelial cells. Chimeric promoters were constructed that comprised basal transcription factor-binding sites coupled to a shear stress response element (SSRE). We have used consensus binding sites for transcription factors NF-kappaB, Ap1, Sp1, Oct1, and Egr-1/Sp1 in either the presence or absence of the previously defined "GAGACC" SSRE. The response of the promoters to shear stress was determined after transfection into human umbilical vein endothelial cells (HUVECs). After transient transfection into HUVECs, fluid shear stress activated the promoters by between two- and eightfold. The most responsive SITUs comprised an overlapping Sp1/Egr-1-binding site linked to a TATA box with (SP5) or without (SP7) the GAGACC SSRE. Instillation of SP5 DNA in vivo into the left carotid artery of rabbit and subsequent generation of a stenosis using a mechanical wire occluder caused a 10-fold upregulation of luciferase reporter gene expression at the site of vessel occlusion. These vectors show promise for therapeutic gene expression at sites of occlusive vascular disease.
Houston
P
P
Endothelial Cell Gene Expression Group, Vascular Diseases Unit, Glaxo Wellcome Medicines Research Centre, Stevenage, Herts, England.
White
B P
BP
Campbell
C J
CJ
Braddock
M
M
D000818:Animals; D001483:Base Sequence; D002318:Cardiovascular Diseases / Q000628:therapy*; D002478:Cells, Cultured; D017931:DNA Primers; D004730:Endothelium, Vascular / Q000378:metabolism*; D015316:Genetic Therapy; D006801:Humans; D011401:Promoter Regions, Genetic; D011817:Rabbits; D014158:Transcription, Genetic; D014162:Transfection
D017931:DNA Primers