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Plasmin-induced expression of cytokines and tissue factor in human monocytes involves AP-1 and IKKbeta-mediated
T Syrovets1, M Jendrach, A Rohwedder
1Department of Pharmacology of Natural Products and Clinical Pharmacology, University of Ulm, Helmholtzstrasse 20, D-89081 Ulm, Germany.
Abstract:
It was previously shown that plasmin activates human peripheral monocytes in terms of lipid mediator release and chemotactic migration. Here it is demonstrated that plasmin induces proinflammatory cytokine release and tissue factor (TF) expression by monocytes. Plasmin 0.043 to 1.43 CTA U/mL, but not active site-blocked plasmin, triggered concentration-dependent expression of mRNA for interleukin-1alpha (IL-1alpha), IL-1beta, tumor necrosis factor-alpha (TNF-alpha), and TF with maximum responses after 4 hours. Plasmin-mediated mRNA expression was inhibited in a concentration-dependent manner by the lysine analogue trans-4-(aminomethyl)cyclohexane-1-carboxylic acid (t-AMCA). Increases in mRNA levels were followed by concentration- and time-dependent release of IL-1alpha, IL-1beta and TNF-alpha and by TF expression on monocyte surfaces. Neither cytokines nor TF could be detected when monocytes were preincubated with actinomycin D or cycloheximide. Electrophoretic mobility shift assays indicated plasmin-induced activation of NF-kappaB; DNA-binding complexes were composed of p50, p65, and c-Rel, as shown by supershift experiments. Nuclear translocation of NF-kappaB/Rel proteins coincided with IkappaBalpha degradation. At variance with endotoxic lipopolysaccharide, plasmin elicited the rapid degradation of another cytoplasmic NF-kappaB inhibitor, p105. Proteolysis of NF-kappaB inhibitors was apparently due to transient activation of IkappaB kinase (IKK) beta that reached maximum activity at 1 hour after plasmin stimulation. In addition, AP-1 binding was increased in plasmin-treated monocytes, with most complexes composed of JunD, c-Fos, and FosB. These findings further substantiate the role of plasmin as a proinflammatory activator of human monocytes and reveal an important new link between the plasminogen-plasmin system and inflammation. (Blood. 2001;97:3941-3950)
Insights
Plasmin, a key protein, activates human monocytes, inducing the release of inflammatory cytokines and expression of tissue factor. This highlights a significant connection between the plasminogen-plasmin system and inflammatory processes.
Area of Science:
- Biochemistry
- Immunology
- Molecular Biology
Background:
- Previous research established plasmin's role in activating human peripheral monocytes for lipid mediator release and chemotactic migration.
- This study investigates plasmin's impact on proinflammatory cytokine release and tissue factor expression in monocytes.
Purpose of the Study:
- To demonstrate that plasmin induces proinflammatory cytokine release and tissue factor (TF) expression in human monocytes.
- To elucidate the molecular mechanisms underlying plasmin-mediated monocyte activation, including transcription factor involvement.
Main Methods:
- Monocytes were stimulated with varying concentrations of plasmin, and mRNA expression for IL-1alpha, IL-1beta, TNF-alpha, and TF was analyzed.
- Inhibition studies using trans-4-(aminomethyl)cyclohexane-1-carboxylic acid (t-AMCA), actinomycin D, and cycloheximide were performed.
- Electrophoretic mobility shift assays (EMSAs) and supershift experiments were used to assess NF-kappaB and AP-1 activation.
- IkappaB kinase (IKK) beta activity and degradation of NF-kappaB inhibitors were measured.
Main Results:
- Plasmin triggered concentration-dependent mRNA expression of IL-1alpha, IL-1beta, TNF-alpha, and TF, which was inhibited by t-AMCA.
- Monocyte preincubation with actinomycin D or cycloheximide blocked cytokine and TF release.
- Plasmin induced NF-kappaB activation, characterized by nuclear translocation of p50, p65, and c-Rel, and degradation of IkappaBalpha and p105.
- Transient activation of IKK beta was observed, and AP-1 binding increased, with complexes containing JunD, c-Fos, and FosB.
Conclusions:
- Plasmin acts as a proinflammatory activator of human monocytes.
- The study reveals a novel link between the plasminogen-plasmin system and inflammatory responses.
- Plasmin-induced monocyte activation involves the NF-kappaB and AP-1 signaling pathways.