Inhibition of monocyte adhesion on brain‐derived endothelial cells by NF‐kappaB decoy/polyethylenimine complexes

Abstract

The nuclear factor (NF)-kappaB plays a key role in inflammatory reactions of the endothelium by controlling the expression of surface-adhesion molecules and other inflammatory mediators, which facilitate the attachment of monocytes and lymphocytes to the endothelial surface. We investigated the inhibition of monocyte adhesion by NF-kappaB transcription factor decoys complexed with polyethylenimines (PEIs) of different molecular weights and structures (800, 25, and 2.7 kDa PEI). Formation, size and stability of the PEI/decoy complexes were investigated by polyacrylamide gel electrophoresis and photon correlation spectroscopy. The efficiency of the complexes was studied in a cell adhesion assay using the murine brain-derived endothelial cell line bEnd5, activated with lipopolysaccharide as inflammatory model. U-937 monocytes were fluorescently labeled with BCECF-AM to permit quantitative measurement of adhesion. Expression of endothelial cell adhesion molecules was determined at the mRNA level by RT-PCR and at the protein level by ELISA. Depending on the N/P ratio, decoys formed complexes of <200 nm in size with all PEIs, which were stable against degradation by nucleases and dissociation by albumin. Treatment of bEnd5 and U-937 cells with NF-kappaB decoys complexed with 25 and 2.7 kDa PEI reduced the number of adherent U-937 cells and decreased the levels of ICAM-1 and VCAM-1 mRNA and protein. The effects were specific, time-dependent and increased with higher N/P ratios of complexes and lower cytotoxicity of polymers. In contrast, the efficiency of the 800 kDa PEI was much lower compared to the other polymers. Complexes of NF-kappaB decoy and PEIs effectively inhibited the adherence of monocytes on endothelial cells, which could be a promising strategy for the treatment of inflammatory diseases.