Inhibition of adhesion molecule expression on human venous endothelial cells by non‐viral siRNA transfection

• Published in: Journal of cellular and molecular medicine Vol. 11; no. 1; pp. 139 - 147
• Main Authors: Walker, Tobias, Wendel, Hans P., Tetzloff, Liane, Raabe, Claudia, Heidenreich, Olaf, Simon, Perikles, Scheule, Albertus M., Ziemer, Gerhard
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01.01.2007; John Wiley & Sons, Inc
• Subjects: adhesion molecules; Antibiotics; Antibodies; Atherosclerosis; Cardiovascular disease; Cell adhesion molecules; Cell Adhesion Molecules - antagonists & inhibitors; Cells, Cultured; Collagen; coronary bypass grafting; E-selectin; Endothelial cells; Endothelial Cells - drug effects; Endothelial Cells - metabolism; Endothelium; Endothelium, Vascular - cytology; Endothelium, Vascular - metabolism; Flow cytometry; Fluorescein-5-isothiocyanate; Fluorescent Dyes; Glycoproteins; Graft rejection; Humans; Hyperplasia; Immunoblotting; Inflammation; Intercellular adhesion molecule 1; Intercellular Adhesion Molecule-1 - pharmacology; Laboratories; Leukocytes; Phosphatase; Polymerase chain reaction; Proteins; RNA, Small Interfering - genetics; RNAi; Saphenous Vein - cytology; Saphenous Vein - metabolism; siRNA; Smooth muscle; Transfection; Tumor Necrosis Factor-alpha - pharmacology; Tumor necrosis factor-α; Veins - cytology; Veins - metabolism; venous graft disease; Germany
• ISSN: 1582-1838; 1582-4934
• DOI: 10.1111/j.1582-4934.2007.00006.x

Objective: Expression of adhesion molecule receptors on venous endothelial cells crucially influences the fate of venous grafts by mediating leukocyte‐endothelium interactions. These interactions include adhesion of leuko‐cytes to the endothelium, followed by transendothelial migration, leading to neointimal hyperplasia (NIH) and finally graft occlusion. Therefore, inhibition of adhesion molecule expression may be a promising strategy to improve the quality of venous grafts.We tested the efficiency of non‐viral transfection of human venous endothe‐lial cells (HVEC) with short interfering RNA (siRNA) to specifically down‐regulate adhesion molecule expression. Methods: Primary cultures of HVEC were examined for expression of the adhesion molecules ICAM1, VCAM1 and E‐selectin (SELE) after non viral siRNA transfection. Adhesion molecule expression was measured by flow cytom‐etry, real‐time polymerase chain reaction and immunoblotting after stimulation with TNF‐α, an inflammatory cytokine. Results: Non‐transfected cells showed a strong increase of adhesion molecule expression following cytokine stimulation (P < 0.01). Upon transfection with specific siRNAs a sixfold decrease in ICAM1 (P < 0.001) and SELE expression and cell positivity (P < 0.05) and a twofold decrease in VCAM1 expression and cell positiv‐ity (P < 0.01) Pcould be observed. SiRNA‐mediated gene suppression of adhesion molecules was also reflected by corresponding decreases in adhesion protein and transcript levels. Conclusions: The expression of adhesion molecules on HVECs can be effectively inhibited by specific siRNAs using a safe, non‐viral transfection approach. This is a promising tool to pre‐condition venous bypass grafts in order to interfere with endothelium‐leukocyte interactions and to prohibit neointima thickening or ath‐erosclerosis, which are regarded to be the most important causes of venous graft failure.