Eric Steinmetz

Multiplexed flow cytometric analyses of pro- and anti-inflammatory cytokines in the culture media of oxysterol-treated human monocytic cells and in the sera of atherosclerotic patients

Some oxysterols are identified in atheromatous plaques and in plasma of atherosclerotic patients. We asked whether they might modulate cytokine secretion on human monocytic cells. In healthy and atherosclerotic subjects, we also investigated the relationships between circulating levels of C‐reactive protein (CRP), conventional markers of hyperlipidemia, some oxysterols (7β‐hydroxycholesterol, 7‐ketocholesterol, and 25‐hydroxycholesterol), and various cytokines.

Impairment of the cytotoxic and oxidative activities of 7β-hydroxycholesterol and 7-ketocholesterol by esterification with oleate

Atherosclerosis involves inflammatory processes, as well as cytotoxic and oxidative reactions. In atherosclerotic plaques, these phenomena are revealed by the presence of dead cells, oxidized lipids, and oxidative DNA damage, but the molecules triggering these events are still unknown. As 7 beta-hydroxycholesterol and 7-ketocholesterol, which are present at elevated concentrations in atherosclerotic lesions, are strongly cytotoxic and pro-oxidative, their effects were determined on cell death, superoxide anion and nitric oxide production, lipid peroxidation, and oxidative DNA damage. 7-Ketocholesterol- and 7 beta-hydroxycholesterol-induced cell death leads to a loss of mitochondrial potential, to increased permeability to propidium iodide, and to morphological nuclear changes (swelling, fragmentation, and/or condensation of nuclei). These effects are preceded by the formation of cytoplasmic monodansylcadaverine-positive structures and are associated with a rapid enhancement of cells overproducing superoxide anions, a decrease in cells producing nitric oxide, lipid peroxidation (formation of malondialdehyde and 4-hydroxynonenal adducts, low ratio of [unsaturated fatty acids]/[saturated fatty acids]) as well as oxidative DNA damage (8-oxoguanine formation). Noteworthy, none of the cytotoxic features previously observed with 7 beta-hydroxycholesterol and 7-ketocholesterol were noted with cholesterol, 7 beta-hydroxycholesteryl-3-oleate and 7-ketocholesteryl-3-oleate, with the exception of a slight increase in superoxide anion production with 7 beta-hydroxycholesteryl-3-oleate. This finding supports the theory that 7 beta-hydroxycholesterol and 7-ketocholesterol could induce cytotoxic and oxidative processes observed in atherosclerotic lesions and that esterification of these compounds may contribute to reducing atherosclerosis progression.

Induction of Transglutaminase 2 by a Liver X Receptor/Retinoic Acid Receptor α Pathway Increases the Clearance of Apoptotic Cells by Human Macrophages

Rationale: Liver X receptors (LXRs) are oxysterol-activated nuclear receptors that are involved in the control of cholesterol homeostasis and inflammatory response. Human monocytes and macrophages express high levels of these receptors and are appropriate cells to study the response to LXR agonists. Objective: The purpose of this study was to identify new LXR targets in human primary monocytes and macrophages and the consequences of their activation. Methods and Results: We show that LXR agonists significantly increase the mRNA and protein levels of the retinoic acid receptor (RAR)&agr; in primary monocytes and macrophages. LXR agonists promote RAR&agr; gene transcription through binding to a specific LXR response element on RAR&agr; gene promoter. Preincubation of monocytes or macrophages with LXR agonists before RAR&agr; agonist treatment enhances synergistically the expression of several RAR&agr; target genes. One of these genes encodes transglutaminase (TGM)2, a key factor required for macrophage phagocytosis. Accordingly, the combination of LXR and RAR&agr; agonists at concentrations found in human atherosclerotic plaques markedly enhances the capabilities of macrophages to engulf apoptotic cells in a TGM2-dependent manner. Conclusions: These results indicate an important role for LXRs in the control of phagocytosis through an RAR&agr;-TGM2–dependent mechanism. A combination of LXR/RAR&agr; agonists that may operate in atherosclerosis could also constitute a promising strategy to improve the clearance of apoptotic cells by macrophages in other pathological situations.

High incidence of venous thrombosis after surgery for abdominal aortic aneurysm

OBJECTIVEnThe incidence of venous thromboembolism (VT) after aortic abdominal aneurysm (AAA) surgery is imprecisely reported. On one hand, thromboprophylaxis has improved, on the other hand, AAA patients have become older and/or present worse comorbidities. Herein, we prospectively analyzed the incidence of VT in a continuous series of patients operated on for AAA repair and looked for predictive factors.nnnMATERIALS AND METHODSnBetween January 1, 2005, and December, 31, 2006, 193 consecutive patients (177 men and 16 women), mean age 73 (range, 47-93) underwent elective AAA repair, 137 open (71%) and 56 endovascular (29%), in our institution. Thromboprophylaxis consisted of thigh-length compression bandages or stockings, early mobilization, and a daily subcutaneous injection of low-molecular-weight heparin (enoxaparin 40 mg per day). Patients with renal insufficiency or aged over 80 were given unfractionated heparin (5000 IU twice a day). Heparin was started between day 1 and day 5 (median = day 1) after surgery, according to the prescription of the surgeon. A bilateral lower limb duplex venous compression ultrasonography scan using 3 to 7.5 MHz transducers was systematically done before and after surgery in each patient. Two groups were considered: group 1 with postoperative VT (n = 17) and group 2 without (n = 176). The 17 patients with VT were compared with 51 patients randomly chosen among the 176 patients without VT. Different characteristics such as venous risk factors, preoperative antithrombotic treatment, anatomical features of the AAA, and perioperative data were studied.nnnRESULTSnSeventeen patients (8.1%) were diagnosed with postoperative VT (15 asymptomatic deep vein thrombosis [DVT] and 2 symptomatic pulmonary embolisms [PE]). VTE tended to be more frequent in open (10.2%) than endovascular (5.3%) repair (P = .28). Among perioperative data, delay to thromboprophylaxis was related to bleeding complication (P = .05) and blood transfusion (P = .02), and tended to be longer in VT patients (1.7 +/- 1.4 vs 0.9 +/- 0.9 day; P = .09).nnnCONCLUSIONnDespite systematic prevention with heparin, surgery for AAA repair induces a high incidence of postoperative VT. This series mandates for vigilance about VT, with particular attention to the patients who received transfusion with fresh frozen plasma. Even though this series is one of the largest ever published on this topic, the rarity of the events calls for confirmation with a larger prospective study.

Development of Abdominal Aortic Aneurysm Is Decreased in Mice with Plasma Phospholipid Transfer Protein Deficiency

Plasma phospholipid transfer protein (PLTP) increases the circulating levels of proatherogenic lipoproteins, accelerates blood coagulation, and modulates inflammation. The role of PLTP in the development of abdominal aortic aneurysm (AAA) was investigated by using either a combination of mechanical and elastase injury at one site of mouse aorta (elastase model) or continuous infusion of angiotensin II in hyperlipidemic ApoE-knockout mice (Ang II model). With the elastase model, complete PLTP deficiency was associated with a significantly lower incidence and a lesser degree of AAA expansion. With the Ang II model, findings were consistent with those in the elastase model, with a lower severity grade in PLTP-deficient mice, an intermediate phenotype in PLTP-deficient heterozygotes, and a blunted effect of the PLTP-deficient trait when restricted to bone marrow-derived immune cells. The protective effect of whole-body PLTP deficiency in AAA was illustrated further by a lesser degree of adventitia expansion, reduced elastin degradation, fewer recruited macrophages, and less smooth muscle cell depletion in PLTP-deficient than in wild-type mice, as evident from comparative microscopic analysis of aorta sections. Finally, cumulative evidence supports the association of PLTP deficiency with reduced expression and activity levels of matrix metalloproteinases, known to degrade elastin and collagen. We conclude that PLTP can play a significant role in the pathophysiology of AAA.

Long-Term Results of Stenting of the Aortic Bifurcation

BACKGROUNDnTo evaluate the long-term results in a multicentric continuous series of narrowing lesions of the aortic bifurcation treated with a kissing stent.nnnMETHODSnFrom January, 1st 1999 to the December, 31st 2001, all of the patients (n = 80) presenting with stenosis of the aortic bifurcation (n = 15) and/or the 2 common iliac arteries (n = 65), treated with a kissing stent, in 8 teaching hospitals were collected retrospectively. The risk factors were smoking (91%), dyslipidemia (60%), arterial hypertension (42%) and diabetes (27%). In 84% of cases, the indication for treatment was claudication. The lesions were stenotic < 70% (n = 76) and/or thrombotic (n = 18). The associated lesions were external iliac stenoses (n = 21), common femoral stenoses (n = 19), femoro-popliteal stenoses (n = 42), arteriopathy in the leg (n = 35). Follow-up was clinical examination and Doppler US scan.nnnRESULTSnThe success rate of the technique was 89%. There were 4 cases (5.3%) of residual stenosis and 4 cases (5.3%) of dissection. The length of the lesions treated in the aorta and the iliac arteries was respectively 17.1 ± 7 and 17.3 ± 9 mm. The stents were all placed as kissing stents, and had a mean diameter and a mean length of 13.75 mm and 56 mm in the aorta and 9 mm and 48 mm in the iliac arteries, respectively. At 5 years, 19 patients had required repeat angioplasty in the treated area, and 13 had undergone open surgery. Primary and assisted primary patency at 5 years were 64.5% and 81.8%, respectively.nnnCONCLUSIONnLong-term follow-up of endovascular treatment with kissing stents for stenosis of the aortic bifurcation shows that this technique gives good results, though it does not justify doing away with classical revascularisation surgery, in a population with major cardiovascular risk factors.

Specific enrichment of 2-arachidonoyl-lysophosphatidylcholine in carotid atheroma plaque from type 2 diabetic patients

BACKGROUND AND AIMSnDiabetic patients are at high risk of stroke and coronary artery disease. Recent data suggest that arachidonic acid metabolism is altered in diabetic conditions and that these alterations contribute to accelerated atherosclerosis. Little is known about how these alterations affect the metabolism and the proportions of different lipid species within the atherosclerotic plaque. The aim of our study was to perform a targeted lipidomic analysis of human atherosclerotic lesions, with a specific focus on PUFA-containing lipid species, to reveal differences in the fatty-acid composition of plaque in diabetic patients compared with non-diabetic controls.nnnMETHODSnCarotid atheroma plaque samples were obtained from 31 diabetic and 48 non-diabetic patients undergoing carotid endarterectomy. Targeted lipidomic analysis was then performed to determine the fatty acid composition of major glycerophospholipids and cholesteryl ester species by liquid chromatography-tandem mass spectrometry.nnnRESULTSnAtheroma plaques from diabetic patients were significantly enriched with 2-arachidonoyl-lysophosphatidylcholine (2-AA-LPC) (2.3xa0±xa00.8% Vs. 1.8xa0±xa00.6% pxa0=xa00.0002). Multivariable logistic regression showed that an increased 2-AA-LPC level was independently associated with diabetes. Finally, a positive relationship was found between 2-AA-LPC and HbA1c levels. Interestingly, endothelial lipase and calcium independent PLA2 gamma which could account for the production of 2-AA-LPC were detected in carotid plaques by immunohistochemistry.nnnCONCLUSIONSn2-AA-LPC stands at the crossroads of major metabolic pathways that lead to the synthesis of bioactive molecules such as AA-derived eicosanoids, 2-AA-lysophosphatidic acid and 2-AA-glycerol. 2-AA-LPC therefore appears to be a promising molecule to investigate in the context of diabetes.

Dynamic 4D blood flow representation in the aorta and analysis from cine-MRI in patients

Natural evolution of aortic disease is characterized by a diameter increase that can result in aortic dissection or rupture. Currently the evaluation of risk of rupture or dissection is based on the size of the aorta. However, this parameter is not always relevant and it appears necessary to define new parameters.

Markovian method for 2D, 3D and 4D segmentation of MRI

Magnetic resonance imaging (MRI) is well adapted for early detection of diseases such as aortic aneuryms or dissections. In this paper, we present a new Markovian method which evolves an active contour for 2D, 3D and 4D (3D + time) segmentation. As opposed to other Markovian contour-based methods, our approach considers an implicit contour as the boundary of a 2D region. The regions are modeled via a Markov random field (MRF) and their computation is based on the maximum a posteriori probability criterion solved using an ICM algorithm. Our method depends on only one parameter that controls region boundary smoothness, is fast, easy to implement and can accommodate different likelihood functions to handle images with very different characteristics. Results on real and synthetic MRI are presented.

Automatic detection of vessel wall contours from cine-MRI for aortic compliance determination

The aortic compliance is defined as the relative change of aortic cross-sectional area divided by the change in arterial pressure. Magnetic resonance imaging (MRI) can be used to evaluate aortic compliance. A knowledge of the aortic contour is essential to determine the aortic area. To prevent important intra-and inter-observer variability, the aortic contours are detected automatically. This work consists in extracting automatically the aortic contour at different phases of the cardiac cycle. An automatic edge detection making use of a Haralick filter and graph searching allows the estimation of aortic cross-sectional area on MR images, from which the aortic compliance is derived