Ellen Lund Sagen

Soluble CD36 in Plasma Is Increased in Patients With Symptomatic Atherosclerotic Carotid Plaques and Is Related to Plaque Instability

Background and Purpose— The risk for cardiovascular events is related to the composition and stability of an atherosclerotic plaque driven by inflammation and deposition of lipids. Scavenger receptors are a family of cell surface receptors involved in lipid uptake and inflammation. Recently, we found that soluble CD36 is increased in plasma from patients with diabetes strongly correlated with insulin resistance. Methods— We tested whether soluble CD36 is a marker of plaque stability in patients with high-grade internal carotid stenoses (n=62). The patients were classified according to plaque symptomatology and plaque echogenicity on ultrasound examination. Results— When patients were divided into 3 groups according to the latest clinical symptoms from plaques (ie, symptoms within the last 2 months [n=16], symptoms within the last 2 to 6 months [n=15], or asymptomatic [n=31]), the former group had significantly raised plasma levels of soluble CD36 as compared with the other 2 groups. In contrast, we found no differences in plasma levels of C-reactive protein, β-thromboglobulin, lipid parameters, or HbA1C between these groups. The patients with echolucent carotid plaques (n=20) tended to have higher soluble CD36 levels in plasma compared with those with echogenic/heterogenic plaque (n=39; P=0.087). By immunohistochemistry, CD36 was localized to macrophages-rich area of intima within the atherosclerotic lesion. Conclusion— We propose that sCD36 may be a marker of plaque instability and symptomatic carotid atherosclerosis, possibly at least partly as a result of CD36 release to the circulation from the foam cells within the atherosclerotic lesion.

Matrix Metalloproteinase 7 Is Associated with Symptomatic Lesions and Adverse Events in Patients with Carotid Atherosclerosis

Background Atherosclerosis is a major cause of cerebrovascular disease. Matrix metalloproteinases (MMPs) play an important role in matrix degradation within the atherosclerotic lesion leading to plaque destabilization and ischemic stroke. We hypothesized that MMP-7 could be involved in this process. Methods Plasma levels of MMP-7 were measured in 182 consecutive patients with moderate (50–69%) or severe (≥70%) internal carotid artery stenosis, and in 23 healthy controls. The mRNA levels of MMP-7 were measured in atherosclerotic carotid plaques with different symptomatology, and based on its localization to macrophages, the in vitro regulation of MMP-7 in primary monocytes was examined. Results Our major findings were (i) Patients with carotid atherosclerosis had markedly increased plasma levels of MMP-7 compared to healthy controls, with particularly high levels in patients with recent symptoms (i.e., within the last 2 months). (ii) A similar pattern was found within carotid plaques with markedly higher mRNA levels of MMP-7 than in non-atherosclerotic vessels. Particularly high protein levels of MMP-7 levels were found in those with the most recent symptoms. (iii) Immunhistochemistry showed that MMP-7 was localized to macrophages, and in vitro studies in primary monocytes showed that the inflammatory cytokine tumor necrosis factor-α in combination with hypoxia and oxidized LDL markedly increased MMP-7 expression. (iv) During the follow-up of patients with carotid atherosclerosis, high plasma levels of MMP-7 were independently associated with total mortality. Conclusion Our findings suggest that MMP-7 could contribute to plaque instability in carotid atherosclerosis, potentially involving macrophage-related mechanisms.

8-isoprostane increases scavenger receptor A and matrix metalloproteinase activity in THP-1 macrophages, resulting in long-lived foam cells.

Background  Oxidative stress is a key factor in atherogenesis, in which it is closely associated with the inflammation and formation of bioactive lipids. Although 8‐isoprostane is regarded as a reliable marker of oxidative stress in vivo, the pathogenic role of this F2‐isoprostane in atherogenesis is far from clear. Based on the important role of foam cells in the initiation and progression of atherosclerosis we hereby examined the ability of 8‐isoprostane to modulate oxidized (ox)LDL‐induced foam cell formation and the function of these cells, particularly focusing on the effect on matrix degradation.

High Levels of S100A12 Are Associated With Recent Plaque Symptomatology in Patients With Carotid Atherosclerosis

Background and Purpose— Atherosclerosis is a progressive chronic disease, in which inflammation plays a key role. The calcium-binding proteins calgranulins including S100A8, S100A9, and S100A12 are involved in many cellular activities and pathological processes including inflammation. We therefore hypothesized that calgranulins may be markers of plaque instability in patients with carotid atherosclerosis. Methods— Plasma levels of S100A8/A9 and S100A10 were measured in 159 consecutive patients with high-grade carotid stenosis and in 22 healthy control subjects. The mRNA levels of calgranulins were also measured within the atherosclerotic carotid plaques, and their regulation was analyzed in vitro in monocytes. Results— Our main findings were: (1) plasma levels of S100A12 were significantly higher in patients with carotid atherosclerosis compared with healthy control subjects with the highest levels in patients with the most recent symptoms (ie, within 2 months); (2) plasma levels of S100A8/S100A9 showed a modest increase in patients with symptoms in the previous 2 to 6 months but not in the other patients; (3) mRNA levels of S100A8, S100A9, and S100A12 showed increased expression in atherosclerotic carotid plaques from patients with the most recent symptoms compared with the remaining patients; (4) in THP-1 monocytes, activation of Toll-like receptors 2 and 4 increased mRNA levels of S100A8, S100A9, and S10012 and interleukin-1&bgr;, interferon &ggr;, and releasate from thrombin-activated platelets significantly enhanced the expression of S100A12. Conclusions— Our findings support a link between calgranulins and atherogenesis and suggest that these mediators, and in particular S100A12, may be related to plaque instability.

Increased levels of the homeostatic chemokine CXCL13 in human atherosclerosis – Potential role in plaque stabilization

OBJECTIVES Based on the newly recognized role of the homeostatic chemokines in inflammation, we hypothesized that CXCL13 could modulate atherogenesis and plaque destabilization. METHODS The study included in vivo analyses in patients with carotid atherosclerosis and in vitro experiments in cells involved in atherogenesis (ie, monocytes/macrophages, vascular smooth muscle cells [SMC], and platelets). RESULTS Our main findings were: (i) Patients with carotid atherosclerosis (n = 130) had increased plasma levels of CXCL13 with particularly high levels in symptomatic disease. (ii) CXCL13 showed increased expression within atherosclerotic carotid plaques as compared with non-atherosclerotic vessels. (iii) Within the atherosclerotic lesions, CXCR5 and CXCL13 were expressed by macrophages and SMC in all stages of plaque progression. (iv) Releasate from activated platelets and toll-like receptor activation enhanced the expression of CXCL13 in THP-1 monocytes and primary monocytes. (v) In vitro, CXCL13 exerted anti-apoptotic effects in primary monocytes, THP-1 macrophages, and vascular SMC. (vi) CXCL13 increased arginase-1, transforming growth factor-β, and interleukin-10 expression in THP-1 cells and in samples from isolated carotid plaques. CONCLUSION Levels of CXCL13 are increased in carotid atherosclerosis both systemically and within the atherosclerotic lesion. Based on our in vitro findings, we hypothesize a potential plaque stabilizing effects of CXCL13-CXCR5 interaction.

Increased expression of NAMPT in PBMC from patients with acute coronary syndrome and in inflammatory M1 macrophages.

AIM The aim of the present study were to elucidate the role of NAMPT in atherosclerosis, by examine NAMPT expression in peripheral blood mononuclear cells (PBMC) in patients with coronary artery disease (CAD) and healthy controls and by examining the regulation and effect of NAMPT on macrophage polarization, hypothesizing that it could influence the polarization to inflammatory and resolving macrophages. METHOD AND RESULTS We analyzed RNA levels of NAMPT in PBMC from CAD and healthy controls and found NAMPT to be increased in PBMC from patients with acute coronary syndrome (n = 39) compared to healthy controls (n = 20) and patients with stable CAD (n = 22). Within the PBMC NAMPT was correlated to several inflammatory cytokines and the antioxidant enzyme superoxide dismutase 2. In vitro cell experiments revealed that NAMPT is increased both intracellular and extracellular in inflammatory M1 macrophages compared to in anti-inflammatory M2 macrophages. In addition, inhibiting NAMPT enzymatic activity inhibited M1 polarization in macrophages. CONCLUSION Based on our in vivo and in vitro findings we suggest that NAMPT could contribute to systemic and plaque inflammation in atherosclerotic disorders at least partly through effect on macrophages.

Activated platelets promote increased monocyte expression of CXCR5 through prostaglandin E2-related mechanisms and enhance the anti-inflammatory effects of CXCL13

BACKGROUND We have previously shown that the homeostatic chemokine CXCL13 is up-regulated in monocytes in atherosclerosis, mediating anti-apoptotic and anti-inflammatory effects. OBJECTIVE To investigate the regulation of CXCL13s receptor, CXCR5. METHODS/PATIENTS In vitro studies in THP-1 and primary monocytes and studies of CXCR5 expression in thrombus material obtained at the site of plaque rupture during myocardial infarction (MI). RESULTS Our major findings were: (i) toll-like receptor agonists and particularly β-adrenergic receptor activation and releasate from thrombin-activated platelets increased CXCR5 mRNA levels in monocytes. (ii) The platelet-mediated induction of CXCR5 involved prostaglandin E2/cAMP/protein kinase A-dependent as well as RANTES-dependent pathways with NFκB activation as a potential common down-stream mediator. (iii) Releasate from thrombin-activated platelets augmented the anti-inflammatory effects of CXCL13 in monocytes at least partly by enhancing the effects of CXCL13 on CXCR5 expression. (iv) We found strong immunostaining of CXCR5 in thrombus material obtained at the site of plaque rupture in patients with ST elevation MI (STEMI) and in unstable carotid lesions, co-localized with platelets. CONCLUSION Our findings suggest that platelet-mediated signaling through CXCR5 may be active in vivo during plaque destabilization, potentially representing a counteracting mechanism to inflammation.

Systemic biomarkers of inflammation and haemostasis in patients with chronic necrotizing pulmonary aspergillosis.

BackgroundThe purpose of this study was to investigate mediators of inflammation and haemostasis in patients with chronic necrotizing pulmonary aspergillosis (CNPA), a locally, destructive process of the lung due to invasion by Aspergillus species.MethodsMeasurements of selected biomarkers in 10 patients with CNPA and 19 healthy, matched controls were performed with enzyme-linked immunosorbent assay (ELISA) and multiplex methodology. The gene expressions of relevant biomarkers were analyzed with real-time quantitative RT-PCR.ResultsIncreased concentrations of circulating mediators of inflammation interleukin (IL)-6, IL-8, RANTES, TNF-α, ICAM-1 and mediators involved in endothelial activation and thrombosis (vWF, TF and PAI-1) were observed in patients with CNPA. The concentration of the anti-inflammatory cytokine IL-10 was increased both in plasma and in PBMC in the patient population. The gene expression of CD40L was decreased in PBMC from the patient group, accompanied by decreased concentrations of soluble (s) CD40L in the circulation.ConclusionsThe proinflammatory response against Aspergillus may be counteracted by reduced CD40L and sCD40L, as well as increased IL-10, which may compromise the immune response against Aspergillus in patients with CNPA.

Interleukin-10 increases reverse cholesterol transport in macrophages through its bidirectional interaction with liver X receptor α

Interleukin (IL)-10 is a prototypical anti-inflammatory cytokine that has been shown to attenuate atherosclerosis development. In addition to its anti-inflammatory properties, the anti-atherogenic effect of IL-10 has recently also been suggested to reflect a complex effect of IL-10 on lipid metabolism in macrophages. In the present study we examined the effects of IL-10 on cholesterol efflux mechanism in lipid-loaded THP-1 macrophages. Our main findings were: (i) IL-10 significantly enhanced cholesterol efflux induced by fetal-calf serum, high-density lipoprotein (HDL)2 and apolipoprotein A-1. (ii) The IL-10-mediated effects on cholesterol efflux were accompanied by an increased IL-10-mediated expression of the ATP-binding cassette transporters ABCA1 and ABCG1, that was further enhanced when the cells were co-activated with the liver X receptor (LXR)α agonist (22R)-hydroxycholesterol. (iii) The effect of LXRα activation on the IL-10-mediated effects on the ATP-binding cassette transporters seems to include enhancing effects on the IL-10 receptor 1 (IL10R1) expression and interaction with STAT-3 signaling. (iv) These enhancing effects on ABCA1 and ABCG1 was not seen when the cells were stimulated with the IL-10 family members IL-22 and IL-24. This study suggests that the anti-atherogenic properties of IL-10 may include enhancing effects on cholesterol efflux mechanism that involves cross-talk with LXRα activation.

Interleukin 27 is increased in carotid atherosclerosis and promotes NLRP3 inflammasome activation.

Aim Interleukin-27 (IL-27) is involved in different inflammatory diseases; however, its role in atherosclerosis is unclear. In this study we investigated the expression of IL-27 and its receptor in patients with carotid atherosclerosis and if IL-27 could modulate the inflammatory effects of the NLRP3 inflammasome in vitro. Methods Plasma IL-27 was measured by enzyme immunoassay in patients with carotid stenosis (n = 140) and in healthy controls (n = 19). Expression of IL-27 and IL-27R was analyzed by quantitative PCR and immunohistochemistry in plaques from patients and in non-atherosclerotic vessels. THP-1 monocytes, primary monocytes and peripheral blood mononuclear cells (PBMCs) were used to study effects of IL-27 in vitro. Results Our main findings were: (i) Plasma levels of IL-27 were significantly elevated in patients with carotid atherosclerotic disease compared to healthy controls. (ii) Gene expression of IL-27 and IL-27R was significantly elevated in plaques compared to control vessels, and co-localized to macrophages. (iii) In vitro, IL-27 increased NLRP3 inflammasome activation in monocytes with enhanced release of IL-1 β. Conclusions We demonstrate increased levels of IL-27 and IL-27R in patients with carotid atherosclerosis. Our in vitro findings suggest an inflammatory role for IL-27, which can possibly be linked to atherosclerotic disease development.