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Thrombin-induced p65 homodimer binding to downstream NF-kappa B site of the promoter mediates endothelial ICAM-1 expression and neutrophil adhesion.
J Immunol. 1999 May 01; 162(9):5466-76.JI

Abstract

We investigated the mechanisms by which proinflammatory mediator, thrombin, released during intravascular coagulation and tissue injury, induces ICAM-1 (CD54) expression in endothelial cells. Stimulation of HUVEC with thrombin resulted in dose- and time-dependent increases in ICAM-1 mRNA and cell surface expression and in ICAM-1-dependent endothelial adhesivity toward polymorphonuclear leukocytes. Transient transfection of endothelial cells with ICAM-1 promoter luciferase reporter gene (ICAM-1LUC) constructs indicated that deletion of upstream NF-kappa B site (-533 bases from translation start site) had no effect on thrombin responsiveness, whereas mutation/deletion of downstream NF-kappa B site (-223 bases from the translation start site) prevented the activation of ICAM-1 promoter, indicating that the downstream NF-kappa B site is critical for thrombin inducibility. NF-kappa B-directed luciferase activity increased approximately 3-fold when cells transfected with the plasmid pNF-kappa BLUC containing five copies of consensus NF-kappa B site linked to a minimal adenovirus E1B promoter-luciferase gene were exposed to thrombin, indicating that activation of NF-kappa B was essential for thrombin response. Gel supershift assays demonstrated that thrombin induced binding of NF-kappa Bp65 (Rel A) to downstream NF-kappa B site of the ICAM-1 promoter. Thrombin receptor activation peptide, a 14-amino-acid peptide representing the new NH2 terminus of proteolytically activated receptor-1, mimicked thrombin's action in inducing ICAM-1 expression. These data indicate that thrombin activates endothelial ICAM-1 expression and polymorphonuclear leukocyte adhesion by NF-kappa Bp65 binding to the downstream NF-kappa B site of ICAM-1 promoter after proteolytically activated receptor-1 activation.

Authors+Show Affiliations

Department of Pharmacology, College of Medicine, University of Illinois, Chicago 60612, USA. ARahman@uic.eduNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

10228026

Citation

Rahman, A, et al. "Thrombin-induced P65 Homodimer Binding to Downstream NF-kappa B Site of the Promoter Mediates Endothelial ICAM-1 Expression and Neutrophil Adhesion." Journal of Immunology (Baltimore, Md. : 1950), vol. 162, no. 9, 1999, pp. 5466-76.
Rahman A, Anwar KN, True AL, et al. Thrombin-induced p65 homodimer binding to downstream NF-kappa B site of the promoter mediates endothelial ICAM-1 expression and neutrophil adhesion. J Immunol. 1999;162(9):5466-76.
Rahman, A., Anwar, K. N., True, A. L., & Malik, A. B. (1999). Thrombin-induced p65 homodimer binding to downstream NF-kappa B site of the promoter mediates endothelial ICAM-1 expression and neutrophil adhesion. Journal of Immunology (Baltimore, Md. : 1950), 162(9), 5466-76.
Rahman A, et al. Thrombin-induced P65 Homodimer Binding to Downstream NF-kappa B Site of the Promoter Mediates Endothelial ICAM-1 Expression and Neutrophil Adhesion. J Immunol. 1999 May 1;162(9):5466-76. PubMed PMID: 10228026.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Thrombin-induced p65 homodimer binding to downstream NF-kappa B site of the promoter mediates endothelial ICAM-1 expression and neutrophil adhesion. AU - Rahman,A, AU - Anwar,K N, AU - True,A L, AU - Malik,A B, PY - 1999/5/5/pubmed PY - 1999/5/5/medline PY - 1999/5/5/entrez SP - 5466 EP - 76 JF - Journal of immunology (Baltimore, Md. : 1950) JO - J Immunol VL - 162 IS - 9 N2 - We investigated the mechanisms by which proinflammatory mediator, thrombin, released during intravascular coagulation and tissue injury, induces ICAM-1 (CD54) expression in endothelial cells. Stimulation of HUVEC with thrombin resulted in dose- and time-dependent increases in ICAM-1 mRNA and cell surface expression and in ICAM-1-dependent endothelial adhesivity toward polymorphonuclear leukocytes. Transient transfection of endothelial cells with ICAM-1 promoter luciferase reporter gene (ICAM-1LUC) constructs indicated that deletion of upstream NF-kappa B site (-533 bases from translation start site) had no effect on thrombin responsiveness, whereas mutation/deletion of downstream NF-kappa B site (-223 bases from the translation start site) prevented the activation of ICAM-1 promoter, indicating that the downstream NF-kappa B site is critical for thrombin inducibility. NF-kappa B-directed luciferase activity increased approximately 3-fold when cells transfected with the plasmid pNF-kappa BLUC containing five copies of consensus NF-kappa B site linked to a minimal adenovirus E1B promoter-luciferase gene were exposed to thrombin, indicating that activation of NF-kappa B was essential for thrombin response. Gel supershift assays demonstrated that thrombin induced binding of NF-kappa Bp65 (Rel A) to downstream NF-kappa B site of the ICAM-1 promoter. Thrombin receptor activation peptide, a 14-amino-acid peptide representing the new NH2 terminus of proteolytically activated receptor-1, mimicked thrombin's action in inducing ICAM-1 expression. These data indicate that thrombin activates endothelial ICAM-1 expression and polymorphonuclear leukocyte adhesion by NF-kappa Bp65 binding to the downstream NF-kappa B site of ICAM-1 promoter after proteolytically activated receptor-1 activation. SN - 0022-1767 UR - https://www.unboundmedicine.com/medline/citation/10228026/Thrombin_induced_p65_homodimer_binding_to_downstream_NF_kappa_B_site_of_the_promoter_mediates_endothelial_ICAM_1_expression_and_neutrophil_adhesion_ L2 - http://www.jimmunol.org/cgi/pmidlookup?view=long&pmid=10228026 DB - PRIME DP - Unbound Medicine ER -