Retroviral mediated gene transduction alters integrin expression on vascular endothelial cells.
Genetically recombinant endothelial cells (rEC) may improve the patency of small diameter vascular grafts by preventing thrombosis or limiting neointimal hyperplasia. Previous work has shown that rEC have reduced adhesion to vascular bypass grafts in vivo. Poor adhesion may be due to altered adhesion (integrin) receptors. This study evaluated the expression of the alpha 5 beta 1 (fibronectin), alpha 2 beta 1 (collagen IV), and alpha v beta 3 (vitronectin) integrin subunits on rEC. Human umbilical vein EC or canine jugular vein EC were transduced with neoR, neoR and human tPA or hygromycin resistance genes using retroviral vectors. Naive EC and EC exposed to empty viral particles (mEC) were controls. Naive EC, mEC, and all rEC's were evaluated for alpha and beta subunits for each integrin receptor studied using immunoblotting. Blotting for alpha 2, alpha 5, and alpha v exhibited expression of the alpha integrin subunits in all cells. The beta 1 and beta 3 subunits were present in mEC and nEC but were absent or truncated in all rEC. The decreased adhesion of rEC's to synthetic vascular grafts may be accounted for by their altered beta 1 and beta 3 integrin subunit expression. The beta subunit is critical for organization of the cytoskeleton and cellular signal transduction. Diminished beta subunit expression in rEC is neither vector specific nor related to retroviral exposure alone. Alteration of beta integrin expression may be to associated with the over-expression of phosphotransferase genes such as neoR or hygromycin B used as selectable markers in gene transfer protocols.
Sackman
J E
JE
Department of Surgery, University of Tennessee Medical Center, Knoxville 37920, USA.
Wymore
A M
AM
Reddick
T T
TT
Freeman
M B
MB
Stevens
S L
SL
Goldman
M H
MH
D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D004285:Dogs; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D018014:Gene Transfer Techniques; D006801:Humans; D015151:Immunoblotting; D016023:Integrins / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics*; D014161:Transduction, Genetic; D019096:Vitronectin / Q000378:metabolism
D005353:Fibronectins; D016023:Integrins; D019096:Vitronectin; D003094:Collagen