Dose-Dependent Modulation of Tissue Factor Protein and Procoagulant Activity in Human Monocyte-Derived Macrophages by Oxidized Low Density Lipoprotein

Atherosclerosis Research Center, Division of Cardiology, Department of Medicine, The CSMC Burns & Allen Research Institute.
Journal of atherosclerosis and thrombosis (Impact Factor: 2.73). 04/2011; 18(7):596-603. DOI: 10.5551/jat.7179
Source: PubMed


Oxidized low-density lipoprotein (oxLDL) interacts with macrophages and is implicated in atherogenesis. Macrophages are also the major source within the atherosclerotic plaque of tissue factor (TF), the membrane-bound glycoprotein receptor that triggers the coagulation cascade in vivo and contributes to plaque thrombogenicity. In this study we tested the hypothesis that oxLDL modulates TF expression in human monocyte-derived macrophages (MDMs).
Mononuclear cells were isolated from human blood, allowed to differentiate into MDMs during 8 days in cell culture, and then exposed to varying concentrations of oxLDL in the presence or absence of lipopolysaccharide (LPS). TF procoagulant activity (TF-PCA) of MDMs was measured by one-stage recalcification clotting assay using human recombinant TF as standard. TF protein was evaluated by Western blotting, and TF mRNA was determined by Northern blot analysis.
OxLDL at 5-10 µg/mL increased TF-PCA, TF protein, and mRNA in MDMs, whereas 20-100 µg/mL oxLDL inhibited TF-PCA, protein expression, and mRNA expression in these cells even in the face of LPS stimulation.
Low concentrations of oxLDL enhance TF expression in MDMs, whereas higher concentrations attenuate TF expression both at baseline as well as following LPS stimulation. Both TF-PCA and TF protein follow this dose-response pattern that is preceded by concordant mRNA changes. Thus, we have demonstrated modulation by oxLDL of TF protein and bioactivity in MDMs.

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    • "We recently reported that BMP-7 contributes to the thrombogenicity of lipid-rich atherosclerotic plaques via stimulation of tissue factor (TF) expression in circulating monocytes, and atherosclerotic lesions [12]. TF, the main physiologic initiator of blood coagulation, is also a well-established modulator of plaque thrombogenicity [13] [14] [15] [16] [17]. TF becomes exposed to factor FVII/FVIIa, which activates FIX and FX, leading to a rapid thrombin generation, platelet activation, and thrombogenesis [18] [19]. "
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    ABSTRACT: Bone morphogenetic protein (BMP)-7, a major regulator of bone metabolism, inhibits ectopic calcification in atherosclerotic plaques. We have recently reported that BMP-7 is also a potent inducer of tissue factor (TF) in human mononuclear cells (MNCs). While nuclear factor kappa beta (NF-kB) and activation protein-1 (AP-1) are the transcription factors essential for inducible expression of human TF gene (F3), the mechanisms responsible for TF induction by BMP-7 are not known. To elucidate the molecular mechanisms governing BMP-7-triggered TF expression in human MNCs. Human blood monocytes were stimulated with BMP-7 and western blotting, qRT-PCR, and flow cytometry studies were carried out to assess F3 expression; promoter studies were also performed using a panel of reporter constructs. Procoagulant TF activity was measured using a validated FXa generation assay. The significance of NF-kB transcriptional activity was verified via pharmacological inhibition. BMP-7 increased TF protein levels, procoagulant activity, surface presentation, and TF mRNA expression. This increase was accompanied by activation of NF-kB as evidenced by reduced IkB-α levels and elevated transcriptional activity of an NF-kB-sensitive reporter in transfected MNCs. Although treatment with BMP-7 also led to a strong phosphorylation of c-Jun, activation of AP-1 alone was not sufficient to induce TF expression: JSH-23, a potent and specific NF-kB inhibitor, completely blocked BMP-7-induced TF expression. We report that BMP-7-dependent activation of TF in human MNCs is mediated via increased activity of NF-kB, leading to enhanced F3 transcription in human MNCs. Copyright © 2014 Elsevier Ltd. All rights reserved.
    Thrombosis Research 12/2014; 135(2). DOI:10.1016/j.thromres.2014.11.031 · 2.45 Impact Factor
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    • "There is a common agreement that stimulation of circulating monocytes with lipopolysaccharides induces TF expression in vitro and in vivo [81–83]. It has also been shown that the expression of monocyte and monocyte-derived macrophage TF could be induced by oxidized low density lipoprotein [84, 85]. As a consequence, an increased TF-related procoagulant activity was observed. "
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    ABSTRACT: Tissue factor (TF) is an integral membrane protein that is essential to life. It is a component of the factor VIIa-TF complex enzyme and plays a primary role in both normal hemostasis and thrombosis. With a vascular injury, TF becomes exposed to blood and binds plasma factor VIIa, and the resulting complex initiates a series of enzymatic reactions leading to clot formation and vascular sealing. Many cells, both healthy, and tumor cells, produce detectable amounts of TF, especially when they are stimulated by various agents. Despite the relative simplicity and small size of TF, there are numerous contradictory reports about the synthesis and presentation of TF on blood cells and circulation in normal blood either on microparticles or as a soluble protein. Another subject of controversy is related to the structure/function of TF. It has been almost commonly accepted that cell-surface-associated TF has low (if any) activity, that is, is "encrypted" and requires specific conditions/reagents to become active, that is, "decrypted." However there is a lack of agreement related to the mechanism and processes leading to alterations in TF function. In this paper TF structure, presentation, and function, and controversies concerning these features are discussed.
    12/2012; 2012(7):964862. DOI:10.6064/2012/964862
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    ABSTRACT: Dendritic cells (DCs) and oxLDL play an important role in the atherosclerotic process with DCs accumulating in the plaques during plaque progression. Our aim was to investigate the role of oxLDL in the modulation of the DC homing-receptor CCR7 and endothelial-ligand CCL21. The expression of the DC homing-receptor CCR7 and its endothelial-ligand CCL21 was examined on atherosclerotic carotic plaques of 47 patients via qRT-PCR and immunofluorescence. In vitro, we studied the expression of CCR7 on DCs and CCL21 on human microvascular endothelial cells (HMECs) in response to oxLDL. CCL21- and CCR7-mRNA levels were significantly downregulated in atherosclerotic plaques versus non-atherosclerotic controls [90% for CCL21 and 81% for CCR7 (P < 0.01)]. In vitro, oxLDL reduced CCR7 mRNA levels on DCs by 30% and protein levels by 46%. Furthermore, mRNA expression of CCL21 was significantly reduced by 50% (P < 0.05) and protein expression by 24% in HMECs by oxLDL (P < 0.05). The accumulation of DCs in atherosclerotic plaques appears to be related to a downregulation of chemokines and their ligands, which are known to regulate DC migration. oxLDL induces an in vitro downregulation of CCR7 and CCL21, which may play a role in the reduction of DC migration from the plaques.
    Mediators of Inflammation 04/2012; 2012(2):320953. DOI:10.1155/2012/320953 · 3.24 Impact Factor
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