A Sustained Reduction in IκB-β May Contribute to Persistent NF-κB Activation in Human Endothelial Cells

David R Johnson,Iris Douglas,Andreas Jahnke,Sankar Ghosh,Jordan S Pober

doi:10.1074/jbc.271.27.16317

Abstract

The responses of vascular endothelial cells (EC) to tumor necrosis factor-alpha (TNF), interleukin-1alpha (IL-1), and phorbol myristate acetate (PMA) were compared with respect to the kinetics of (i) NF-kappaB activation, (ii) IkappaB-alpha and IkappaB-beta degradation, and (iii) NF-kappaB-dependent cell surface molecule expression. TNF rapidly (</=20 min) and persistently (>20 h) activates NF-kappaB; IL-1 rapidly activates NF-kappaB, but activity declines by 3 h and further by 20 h; PMA slowly and transiently activates NF-kappaB. Untreated EC contain the inhibitory proteins IkappaB-alpha and IkappaB-beta. The onset of NF-kappaB activation correlates with degradation of IkappaB-alpha, but IkappaB-alpha reappears by 4 h without resequestration of NF-kappaB. TNF causes a rapid but partial (50%) reduction in IkappaB-beta, which does not recover by 22 h; IL-1 and PMA cause slower and less sustained reductions in IkappaB-beta. All three agonists induce de novo expression of E-selectin (CD62E) and vascular cell adhesion molecule-1 (CD106) and increase expression of intercellular adhesion molecule-1 (CD54) at 4 h. TNF induces sustained increases in vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 and increases human leukocyte antigen class I molecules at 24 h. We conclude that TNF causes persistent activation of NF-kappaB in human EC and that this may result from sustained reductions in IkappaB-beta levels.

Highlights

The responses of vascular endothelial cells (EC) to tumor necrosis factor-␣ (TNF), interleukin-1␣ (IL-1), and phorbol myristate acetate (PMA) were compared with respect to the kinetics of (i) NF-␬B activation, (ii) I␬B-␣ and I␬B-␤ degradation, and (iii) NF-␬B-dependent cell surface molecule expression
In the present study we have examined the effects of TNF, IL-1, and PMA on NF-␬B activation and I␬B degradation in human EC
The time courses of NF-␬B activation by the agonists TNF, IL-1␣, or PMA in endothelial cells were determined by testing nuclear extracts prepared from replicate cultures of EC treated for 20 min, 3 h, or 20 h

Summary

Introduction

The responses of vascular endothelial cells (EC) to tumor necrosis factor-␣ (TNF), interleukin-1␣ (IL-1), and phorbol myristate acetate (PMA) were compared with respect to the kinetics of (i) NF-␬B activation, (ii) I␬B-␣ and I␬B-␤ degradation, and (iii) NF-␬B-dependent cell surface molecule expression. Activation of the transcription factor NF-␬B is required for the full expression of many TNF-inducible1 genes in vascular endothelial cells (EC), including E-selectin (CD62E, known as endothelial-leukocyte adhesion molecule-1) [1], vascular cell adhesion molecule-1 (VCAM-1, CD106) [2], intercellular adhesion molecule-1 (ICAM-1, CD54) [3, 4], and human leukocyte antigen (HLA) class I [5].

Results

Conclusion