Immunoenzymometric analysis for expression and shedding of intercellular adhesion molecule-1 on human endothelial cells stimulated with cytokines or lipopolysaccharide.
Unstimulated endothelial cell (EC) cultures express low levels of intercellular adhesion molecule-1 (ICAM-1) and their expression can be enhanced by inflammatory cytokines such as tumor necrosis factor alpha (TNF). Three monoclonal antibodies (MoAbs) highly reactive with TNF-stimulated human ECs were established and defined to recognize a 95 kDa cell surface protein specifically expressed on cytokine-activated ECs, which was immunochemically identified as ICAM-1. The quantitative immunoassay of soluble and insoluble ICAM-1 could be performed with two different MoAbs. Secretion of fibronectin or the von Willebrand factor, was not significantly enhanced with TNF stimulation. Cellular expression of ICAM-1 was drastically induced by TNF or interleukin-1 stimulation, and the moderate expression with delayed-action was observed only by lipopolysaccharide stimulation. A maximal amount of soluble ICAM-1 was released from ECs stimulated only by TNF, apparently in a dose dependent manner, but no significant release of ICAM-1 was induced by thrombin interleukin-2, or lipopolysaccharides. Released levels of soluble ICAM-1 from interleukin-1-stimulated ECs were apparently diminished as compared with those from TNF-stimulated cells. These results suggest that release of soluble ICAM-1 from EC surfaces can be most significantly enhanced by TNF-specific signaling, and prospectively, should be a sensitive indicator of intravascular inflammation in acute endothelium injury.
Sano
Y
Y
Cell Technology Reagent Section, Takara Shuzo Company Limited, Shiga, Japan.
Hirai
S
S
Katayama
M
M
Kato
I
I
D000911:Antibodies, Monoclonal; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D005434:Flow Cytometry; D006801:Humans; D015151:Immunoblotting; D018799:Intercellular Adhesion Molecule-1 / Q000276:immunology / Q000378:metabolism*; D007375:Interleukin-1 / Q000494:pharmacology; D007376:Interleukin-2 / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D000911:Antibodies, Monoclonal; D016207:Cytokines; D000939:Epitopes; D007375:Interleukin-1; D007376:Interleukin-2; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1