Discriminatory effects of protein kinase inhibitors and calcium ionophore on endothelial ICAM-1 induction.
Intercellular adhesion molecule 1 (ICAM-1) is a proinflammatory adhesion glycoprotein induced by cytokines such as interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) as well as lipopolysaccharide (LPS). Little is known, however, concerning the intracellular regulatory mechanisms that modulate ICAM-1 expression in endothelial cells. We probed the involvement of protein kinase function and intracellular calcium ion upon ICAM-1 expression of human umbilical vein endothelial cells activated alternatively by TNF-alpha, IL-1 beta, LPS, or phorbol 12-myristate 13-acetate (PMA). Methodologies for the detection of ICAM-1 included both enzyme-linked immunosorbent assay and immunoprecipitation from biosynthetically labeled cells. The protein kinase inhibitor H-7 blocked induction of ICAM-1 by all of the activators; nonlinear regression analysis revealed 50% inhibitory concentration (IC50) values of 6-10 microM. Another kinase inhibitor, HA1004, did not block expression of the adhesion molecule at concentrations up to 50 microM. In contrast, the kinase inhibitor staurosporine dose dependently inhibited ICAM-1 expression triggered by PMA (IC50 67 +/- 4 nM) but, at similar concentrations, did not inhibit ICAM-1 expression induced by the other inflammatory stimuli. The divalent cation ionophore ionomycin (0.5 microM) interacted synergistically with PMA but not with cytokines or LPS in upregulating ICAM-1. We conclude from these data that although PMA-induced ICAM-1 expression may be triggered through activation of protein kinase C, ICAM-1 induction by IL-1 beta, TNF-alpha, or LPS may involve distinct regulatory pathway(s).
Boehringer Ingelheim Pharmaceuticals, Ridgefield, Connecticut 06877.
C L
CL
Myers
S N
SN
Desai
J
J
Schembri-King
G L
GL
Letts
R W
RW
Wallace
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000470:Alkaloids / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology*; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D016207:Cytokines / Q000494:pharmacology; D004357:Drug Synergism; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D018799:Intercellular Adhesion Molecule-1; D015759:Ionomycin / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D008070:Lipopolysaccharides; D010879:Piperazines / Q000494:pharmacology; D014176:Protein Biosynthesis; D047428:Protein Kinase Inhibitors; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000470:Alkaloids; D015815:Cell Adhesion Molecules; D016207:Cytokines; D007546:Isoquinolines; D008070:Lipopolysaccharides; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D018799:Intercellular Adhesion Molecule-1; D000001:Calcimycin; D015759:Ionomycin; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate