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Targeting Tumor Necrosis Factor-? with Adalimumab: Effects on Endothelial Activation and Monocyte Adhesion.

ABSTRACT: It is well known that atherosclerotic inflammatory vascular disease is critically driven by oxidized lipids and cytokines. In this regard, tumor necrosis factor (TNF)-? is known as a crucial mediator of early pro-atherosclerotic events. Epidemiologic data suggest that blockade of TNF-? has beneficial effects on vascular outcomes in patients with rheumatoid arthritis, however, detailed mechanistic studies are still lacking. This study aims to elucidate effects of TNF-? blockade by adalimumab-which is approved for several inflammatory disorders-on endothelial activation and monocyte adhesion under pro-atherosclerotic conditions.Phorbol myristate acetate (PMA) differentiated THP-1 macrophages were stimulated with oxidized low density lipoprotein and subsequent analysis of this conditioned media (oxLDL CM) revealed a strong release of TNF-?. The TNF-? rich supernatant led to activation of human umbilical vein endothelial cells (HUVEC) as shown by enhanced expression of major adhesion molecules, such as vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1) and E-selectin which was suppressed by the TNF-? inhibitor adalimumab. Accordingly, adalimumab effectively prevented THP-1 monocyte adhesion to endothelial cells under static as well as under flow conditions. Furthermore, adalimumab suppressed endothelial leakage as shown by Evan's blue diffusion across a confluent endothelial monolayer. Of note, after intraperitoneal injection we detected abundant deposition of fluorophore-labelled adalimumab in atherosclerotic plaques of hypercholesterolemic mice.Our results show that adalimumab prevents major inflammatory effects of TNF-? on endothelial activation, endothelial monocyte adhesion, endothelial leakage and therefore extends the therapeutic options of adalimumab to limit vascular inflammation.

SUBMITTER: Oberoi R

PROVIDER: S-EPMC4965117 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Targeting Tumor Necrosis Factor-α with Adalimumab: Effects on Endothelial Activation and Monocyte Adhesion.

Oberoi Raghav R Schuett Jutta J Schuett Harald H Koch Ann-Kathrin AK Luchtefeld Maren M Grote Karsten K Schieffer Bernhard B

PloS one 20160728 7

<h4>Objective</h4>It is well known that atherosclerotic inflammatory vascular disease is critically driven by oxidized lipids and cytokines. In this regard, tumor necrosis factor (TNF)-α is known as a crucial mediator of early pro-atherosclerotic events. Epidemiologic data suggest that blockade of TNF-α has beneficial effects on vascular outcomes in patients with rheumatoid arthritis, however, detailed mechanistic studies are still lacking. This study aims to elucidate effects of TNF-α blockade ...[more]

PMID: 27467817

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Project description:BackgroundThe blood-spinal cord barrier (BSCB) is composed of a monolayer of endothelium linked with tight junctions and extracellular matrix (ECM)-rich basement membranes and is surrounded by astrocyte foot processes. Endothelial permeability is regulated by interaction between endothelial cells and ECM proteins. Fibronectin (FN) is a principal ECM component of microvessels. Excessive FN deposition disrupts cell-cell adhesion in fibroblasts through β1 integrin ligation. To determine whether excessive FN deposition contributes to the disruption of endothelial integrity, we used an in vitro model of the endothelial monolayer to investigate whether the FN inhibitor pUR4 prevents FN deposition into the subendothelial matrix and attenuates endothelial leakage.MethodsTo correlate the effects of excessive FN accumulation in microvessels on BSCB disruption, spinal nerve ligation-which induces BSCB leakage-was applied, and FN expression in the spinal cord was evaluated through immunohistochemistry and immunoblotting. To elucidate the effects by which pUR4 modulates endothelial permeability, brain-derived endothelial (bEND.3) cells treated with tumor necrosis factor (TNF)-α were used to mimic a leaky BSCB. A bEND.3 monolayer was preincubated with pUR4 before TNF-α treatment. The transendothelial electrical resistance (TEER) measurement and transendothelial permeability assay were applied to assess the endothelial integrity of the bEND.3 monolayer. Immunofluorescence analysis and immunoblotting were performed to evaluate the inhibitory effects of pUR4 on TNF-α-induced FN deposition. To determine the mechanisms underlying pUR4-mediated endothelial permeability, cell morphology, stress fiber formation, myosin light chain (MLC) phosphorylation, and β1 integrin-mediated signaling were evaluated through immunofluorescence analysis and immunoblotting.ResultsExcessive FN was accumulated in the microvessels of the spinal cord after spinal nerve ligation; moreover, pUR4 inhibited TNF-α-induced FN deposition in the bEND.3 monolayer and maintained intact TEER and endothelial permeability. Furthermore, pUR4 reduced cell morphology alteration, actin stress fiber formation, and MLC phosphorylation, thereby attenuating paracellular gap formation. Moreover, pUR4 reduced β1 integrin activation and downstream signaling.ConclusionspUR4 reduces TNF-α-induced β1 integrin activation by depleting ECM FN, leading to a decrease in endothelial hyperpermeability and maintenance of monolayer integrity. These findings suggest therapeutic benefits of pUR4 in pathological vascular leakage treatment.

Dataset Information

Targeting Tumor Necrosis Factor-? with Adalimumab: Effects on Endothelial Activation and Monocyte Adhesion.

Publications

Targeting Tumor Necrosis Factor-α with Adalimumab: Effects on Endothelial Activation and Monocyte Adhesion.

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