Endothelial cells upregulate eosinophil superoxide generation via VCAM-1 expression

Background In vitro eosinophil (EOS) adhesion to recombinant human (rh)-vas cular cell adhesion molecule (VCAM)-1 stimulates superoxide anion (O-2(-)) generation and enhances formyl-methionyl-leucyl phenylalanine (FMLP)-activa ted O-2(-) generation. Therefore, EOS adhesion via VLA-4 to VCAM-1 expresse d on endothelium may be instrumental in the selective recruitment and funct ion of EOS in airway inflammation. Objective We hypothesized that EOS interaction with endothelial cells expre ssing VCAM-1 will undergo an enhancement in inflammatory function. Methods To determine this possibility, human umbilical vein endothelial cel ls (HUVEC) were stimulated with either a combination of interleukin (IL)-4 and tumour necrosis factor (TNF)-alpha (100pM) or medium alone for 24h; the expression of adhesion proteins on HUVEC and their effect on EOS O-2(-) ge neration was subsequently determined. Results As determined by both enzyme-linked immunosorbent assay and flow cy tometry, IL-4 and TNF alpha acted synergistically to induce VCAM-1 expressi on on HUVEC. Treating HUVEC with IL-4/TNF alpha also increased EOS adhesion and primed subsequent FMLP (0.1 mu M) activated EOS O-2(-) generation. Alt hough EOS adhesion was partially inhibited by both anti alpha(4) and anti b eta(2) monoclonal antibodies (MoAbs), O-2(-) generation was completely inhi bited by either anti alpha(4) integrin MoAb (HP1/2) or anti-VCAM MoAb (BBIG -V1). Furthermore, enhanced O-2(-) generation, but not adhesion, associated with IL-4 + TNF alpha-treatment of HUVEC was inhibited when EOS were treat ed with the platelet activating factor (PAF)-antagonist WEB 2086 (20 mu M), thus suggesting an involvement of PAF in priming EOS. However, paraformald ehyde fixation of IL-4/TFN-alpha treated HUVEC did not significantly alter EOS function, Conclusions These results suggest EOS adhesion to endothelial cells via an VLA-4/VCAM-1 interaction may be important in the development of the functio n of this cell, Furthermore, our results suggest that modulation of EOS fun ction involves two priming factors: EOS adhesion to HUVEC expressing VCAM-1 and PAF.