Unstable Atherosclerotic Plaques Research Articles

Effects of dyslipidaemia on monocyte production and function in cardiovascular disease.

Monocytes are heterogeneous effector cells involved in the maintenance and restoration of tissue integrity. Monocytes and macrophages are involved in cardiovascular disease progression, and are associated with the development of unstable atherosclerotic plaques. Hyperlipidaemia can accelerate cardiovascular disease progression. However, monocyte responses to hyperlipidaemia are poorly understood. In the past decade, accumulating data describe the relationship between the dynamic blood lipid environment and the heterogeneous circulating monocyte pool, which might have profound consequences for cardiovascular disease. In this Review, we explore the updated view of monocytes in cardiovascular disease and their relationship with macrophages in promoting the homeostatic and inflammatory responses related to atherosclerosis. We describe the different definitions of dyslipidaemia, highlight current theories on the ontogeny of monocyte heterogeneity, discuss how dyslipidaemia might alter monocyte production, and explore the mechanistic interface linking dyslipidaemia with monocyte effector functions, such as migration and the inflammatory response. Finally, we discuss the role of dietary and endogenous lipid species in mediating dyslipidaemic responses, and the role of these lipids in promoting the risk of cardiovascular disease through modulation of monocyte behaviour.

SURGICAL TREATMENT OF MULTIFOCAL ATHEROSCLEROSIS: CORONARY AND BRACHIOCEPHALIC PATHOLOGY AND PREDICTORS OF EARLY ADVERSE EVENTS DEVELOPMENT

Aim. Analysis of clinical and demographic, anatomic and angiographic, instrumental and perioperational factors of adverse prognosis in inpatient period for various surgical managing strategies in patients with multifocal atherosclerosis. Material and methods . The analysis is done, of surgical treatment results in 391 patient with combination lesion of coronary and brachocephalic arteries under the frame work of single center prospective registry. According to the presence of in-hospital complications (HC) all patients were selected to 2 groups: I — patients with HC (n=84); II — patients with no HC (n=307). In this selection, 4 revasculariztion tactics were realized: staged surgery within the procedures of coronary bypass and carotid endarterectomy (CEE) with various sequence (n=222, 56,8%); combinational operation of coronary bypass and CEE (n=141; 36%); hybrid revascularization within percutaneous coronary intervention and CEE (n=28; 7,2%). Results . The wide spectrum of prognostically significant factors was analized in details, including clinical and instrumental, anatomic and angiographic, perioperational factors. Adverse prognosis factors were found, and protecting parameters defined that decrease the risk of adverse cardiovascular events occurence. Predictors of adverse prognosis in post-operation period were found: clinical and demographic — older age; high surgical risk by EuroScore II ≥3; chronic renal failure; cerebrovascular — chronic cerebral ischemia III grade; the unclosed Willis circle; unstable atherosclerotic plaque or subtotal stenosis of the internal carotid arteria unilateral with operation; subtotal stenosis or occlusion of internal carotid artery contralateral. Conclusion . The results can be applied in algorithms development for selection of surgical tactics in multifocal atherosclerosis patients.

The usefulness of Duplex Doppler ultrasound in the angiological and dermatological diagnosis of patients with blue toe syndrome.

IntroductionPeripheral microembolism is one of the most frequent causes of acute limb ischemia. In order to effectively prevent relapses it is essential to localize and eliminate the source of embolism.AimTo evaluate the role of Duplex Doppler ultrasound examination in identifying the causes of blue toe syndrome (BTS).Material and methodsThe group of 165 patients with clinical symptoms of BTS on their upper limbs (n = 16) and lower limbs (n = 149) was investigated. They all underwent Duplex Doppler ultrasound of the major arteries of the extremities, where ischemic changes occurred.ResultsMorphological and functional changes which might be potential sources of microembolism were identified in 146 patients. These changes included significant short-length stenoses or unstable atherosclerotic plaque (n = 73), true aneurysms (n = 42) and pseudoaneurysms (n = 17). In 11 cases, pathology of vascular prostheses in the form of anastomotic aneurysms, infection and residual thrombi after fibrinolysis was detected. In all cases, Duplex diagnosis was confirmed by other imaging and intraoperative tests.ConclusionsDuplex Doppler ultrasound of the arteries in the affected limb with a full length view should be the first-line examination in diagnosing patients with BTS. In the absence of hemodynamic blood flow disturbances in the major arteries in patients with symptoms of BTS, it is advisable to start haematological tests to identify/exclude congenital or acquired thrombophilia.

Pseudomonas aeruginosa Microcolonies in Coronary Thrombi from Patients with ST-Segment Elevation Myocardial Infarction.

Chronic infection is associated with an increased risk of atherothrombotic disease and direct bacterial infection of arteries has been suggested to contribute to the development of unstable atherosclerotic plaques. In this study, we examined coronary thrombi obtained in vivo from patients with ST-segment elevation myocardial infarction (STEMI) for the presence of bacterial DNA and bacteria. Aspirated coronary thrombi from 22 patients with STEMI were collected during primary percutaneous coronary intervention and arterial blood control samples were drawn from radial or femoral artery sheaths. Analyses were performed using 16S polymerase chain reaction and with next-generation sequencing to determine bacterial taxonomic classification. In selected thrombi with the highest relative abundance of Pseudomonas aeruginosa DNA, peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) with universal and species specific probes was performed to visualize bacteria within thrombi. From the taxonomic analysis we identified a total of 55 different bacterial species. DNA from Pseudomonas aeruginosa represented the only species that was significantly associated with either thrombi or blood and was >30 times more abundant in thrombi than in arterial blood (p<0.0001). Whole and intact bacteria present as biofilm microcolonies were detected in selected thrombi using universal and P. aeruginosa-specific PNA-FISH probes. P. aeruginosa and vascular biofilm infection in culprit lesions may play a role in STEMI, but causal relationships remain to be determined.

Increased levels of legumain in plasma and plaques from patients with carotid atherosclerosis

Background and aimsThe cysteine protease legumain has been shown to be up-regulated in unstable atherosclerotic plaques. This study aims to further elucidate legumain in atherosclerosis, by examining legumain in plasma and carotid plaques from patients with carotid stenosis. Furthermore, legumain secretion from monocyte-derived macrophages treated with atherogenic lipids during macrophage polarization was studied. MethodsPlasma levels of legumain from patients with carotid stenosis (n = 254), healthy controls (n = 91), and secreted from monocyte-derived macrophages were assessed by enzyme-linked-immunosorbent assay. Quantitative PCR and immunoblotting of legumain were performed on isolated plaques and legumain localization was visualized by immunohistochemistry and fluorescence microscopy. Monocyte-derived macrophages polarized to M1 or M2 macrophages were treated with VLDL, oxLDL or cholesterol crystals (CC) and the level of legumain analysed. ResultsPatients with carotid stenosis had significantly higher levels of plasma legumain compared with healthy controls (median 2.0 versus 1.5 ng/ml, respectively; p = 0.003), although there was no correlation between the level of legumain and the degree of stenosis, and legumain was not an independent factor to identify patients with carotid plaques. Moreover, patients with symptoms the last 2 months had higher expressions of mature legumain, cystatin C and E/M, and the macrophage markers CD80 (M1) and CD163 (M2). Legumain co-localized with both M1 and M2 macrophages within plaques, whereas legumain mRNA expression was significantly higher (p < 0.0001) in plaques compared to non-atherosclerotic arteries (controls). Furthermore, in vitro studies showed significantly increased secretion of legumain from pro-inflammatory M1 compared to pro-resolving M2 macrophages (p = 0.014), and particularly in M1 treated with CC. In plaques, legumain was localized to structures resembling foam cells. ConclusionsLegumain is increased in both plasma and plaques of patients with carotid stenosis and might be a new and early biomarker of atherosclerosis.

Effects of berberine on serum inflammatory factors and carotid atherosclerotic plaques in patients with acute cerebral ischemic stroke

This study aims to analyze the effect of berberine on serum inflammatory factors and carotid atherosclerotic plaques in ppatients with acute cerebral ischemic stroke(AIS). In the study, 120 patients with AIS were randomly divided into berberine group(n=60) and general group (n=60). The 60 cases in the general group were provided with general therapy according to the latest guidelines of diagnosis and treatment of AIS. The berberine group received berberine 300 mg(tid) in addition to the therapy of the general group. The levels of serum inflammatory factors, the nerve function defect grades and the indexes of carotid atherosclerosis plaques [including the total plaque area(TPA), intima-media thickness(IMT) and the number of unstable carotid atherosclerotic plaques] were measured and compared. The results indicated that the levels of serum inflammatory factors, the NIHSS(national institute of health stroke scales) cores and the indexes of carotid atherosclerosis plaques were not significantly different between the berberine groups of general group, with positive correlation between serum inflammatory factors and NIHSS scores(P<0.05). The levels of serum inflammatory factors and NIHSS scores of the berberine groups on 14 d were significantly lower than those on 1 d(P<0.05). The levels of serum inflammatory factors and NIHSS scores of the berberine group on 14 d were significantly lower than those of the general group(P<0.05). The TPA and the number of unstable carotid atherosclerotic plaques of the berberine groups on 90 d were significantly lower than those of general group, with significant differences(P<0.05). The IMT showed a downward trend, but with significant difference.The mRS(modified rankin scale) scores of the berberine group on 90 d were significantly lower, with a higher rate of short-term favorable prognosis (P<0.05). There was no significant difference in the incidence of adverse reactions between the two groups. This study showed that berberine in addition to the general therapy can significantly lower the levels of serum MIF and IL-6, reduce the degree of carotid atherosclerosis to some extent and improve neurological impairment and the prognosis of patients with AIS.

Talin and vinculin are downregulated in atherosclerotic plaque; Tampere Vascular Study

Background and aimsFocal adhesions (FA) play an important role in the tissue remodeling and in the maintenance of tissue integrity and homeostasis. Talin and vinculin proteins are among the major constituents of FAs contributing to cellular well-being and intercellular communication. MethodsMicroarray analysis (MA) and qRT-PCR low-density array were implemented to analyze talin-1, talin-2, meta-vinculin and vinculin gene expression in circulating blood and arterial plaque. ResultsAll analyzed genes were significantly and consistently downregulated in plaques (carotid, abdominal aortic and femoral regions) compared to left internal thoracic artery (LITA) control. The use of LITA samples as controls for arterial plaque samples was validated using immunohistochemistry by comparing LITA samples with healthy arterial samples from a cadaver. Even though the differences in expression levels between stable and unstable plaques were not statistically significant, we observed further negative tendency in the expression in unstable atherosclerotic plaques. The confocal tissue imaging revealed gradient of talin-1 expression in plaque with reduction close to the vessel lumen. Similar gradient was observed for talin-2 expression in LITA controls but was not detected in plaques. This suggests that impaired tissue mechanostability affects the tissue remodeling and healing capabilities leading to development of unstable plaques. ConclusionsThe central role of talin and vinculin in cell adhesions suggests that the disintegration of the tissue in atherosclerosis could be partially driven by downregulation of these genes, leading to loosening of cell-ECM interactions and remodeling of the tissue.

A role for autophagy in carotid atherosclerosis.

Autophagy has emerged in recent years as a critical cellular survival mechanism for cell homeostasis and may play a protective role in atherosclerosis. We aimed to review here the role autophagy plays in different cell types present in carotid atherosclerotic plaques and that may be associated with the development of unstable carotid atheroma plaque. We performed a thorough literature exploration in this area of research covering the three main cell types present in carotid atheroma plaques. Reviewed reports indicate that the role of autophagy in stable or unstable carotid atherosclerotic plaques depends on the different cell types and phenotypes, the stage and morphology of the plaque and the specific autophagy factor/s involved. Although defective autophagy could be one of the causes for carotid atheroma plaques to become unstable, it is important to take into account that autophagic players can act differentially in different cell types and different stages of the developed plaque. This review provides an overview of the role of autophagy in the main cell types in carotid atherosclerosis (i.e. macrophages, endothelial cells and smooth muscle cells).

GW27-e0841 Hyperhomocysteinemia promotes unstable plaque formation in ApoE-/- mice: A role of Endoplasmic Reticulum Stress induced macrophage apoptosis

Hyperhomocysteinemia(Hhcy) promotes unstable atherosclerotic plaque formation. Macrophage apoptosis and plaque necrosis are the two hallmarks of advanced plaque. Endoplasmic reticulum stress plays a critical role in macrophage apoptosis and plaque instability induced by homocysteine. The present

Gene and Protein Expression of Chemokine (C-C-Motif) Ligand 19 is Upregulated in Unstable Carotid Atherosclerotic Plaques

The aim was to investigate the expression of genes associated with carotid plaque instability and their protein products at a local and systemic level. Carotid plaques from 24 patients undergoing carotid endarterectomy (CEA) were classified as stable or unstable using clinical, histological, ultrasound, and transcranial Doppler criteria, and compared using whole genome microarray chips. Initial results of differentially expressed genes were validated by quantitative reverse transcriptase polymerase chain reaction in an independent group of 96 patients undergoing CEA. The protein product of genes significantly differentially expressed between patients with stable and unstable plaques were analysed by plaque immunohistochemistry and serum protein quantification by enzyme-linked immunosorbent assay on a further independent cohort. Expression of chemokine (c-c-motif) ligand 19 (CCL19) was significantly upregulated in plaques from patients with clinically unstable disease (p<.001). Cathepsin G expression was upregulated in histologically unstable plaques (p=.04). Serum concentration of CCL19 was significantly higher in patients with clinically unstable plaques (p=.02). Immunohistochemical staining for CCL19 demonstrated positive staining in histologically and clinically unstable plaques (p=.03). CCL19 also co-localised with CD3+ T-cell lymphocytes in the core region, around where CCL19 was expressed. CCL19 is significantly overexpressed in patients with unstable carotid atherosclerotic plaques and may be a possible novel biomarker for identifying high-risk patients in whom more urgent intervention may be indicated.

Adipo/cytokines in atherosclerotic secretomes: increased visfatin levels in unstable carotid plaque.

BackgroundNovel pro-inflammatory and anti-inflammatory derivatives from adipose tissue, known as adipokines, act as metabolic factors. The aim of this study was to analyse the secreted expression of different adipo/cytokines in secretomes of unstable carotid atherosclerotic plaque versus non-atherosclerotic mammary artery.MethodsWe evaluated the secretion levels of adiponectin, visfatin, lipocalin-2, resistin, IL-6 and TNFR2 by ELISA in human secretomes from cultured unstable carotid atherosclerotic plaque (n = 18) and non-atherosclerotic mammary artery (n = 13). We also measured visfatin serum levels in patients suffering from atherosclerosis and in a serum cohort of healthy subjects (n = 16).ResultsWe found that visfatin levels were significantly increased in unstable carotid atherosclerotic plaque secretome than in non-atherosclerotic mammary artery secretome. No differences were found with regard the other adipo/cytokines studied. Regarding visfatin circulating levels, there were no differences between unstable carotid atherosclerotic plaque and non-atherosclerotic mammary artery group. However, these visfatin levels were increased in comparison to serum cohort of healthy subjects.ConclusionsOf all the adipo/cytokines analysed, only visfatin showed increased levels in secretomes of unstable carotid atherosclerotic plaque. Additional human studies are needed to clarify the possible role of visfatin as prognostic factor of unstable carotid atherosclerotic plaque.

MiR-106b-5p Inhibits Tumor Necrosis Factor-α-induced Apoptosis by Targeting Phosphatase and Tensin Homolog Deleted on Chromosome 10 in Vascular Endothelial Cells.

Background:Apoptosis of endothelial cells (ECs) plays a key role in the development of atherosclerosis and there are also evidence indicated that phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a viable target in therapeutic approaches to prevent vascular ECs apoptosis. Aberrant miR-106b-5p expression has been reported in the plasma of patients with unstable atherosclerotic plaques. However, the role and underlying mechanism of miR-106-5p in the genesis of atherosclerosis have not been addressed. In this study, we explored the anti-apoptotic role of miR-106-5p by regulating PTEN expression in vascular ECs.Methods:Real-time reverse transcription polymerase chain reaction (RT-PCR) was performed to detect the expression levels of miR-106b-5p in human atherosclerotic plaques and normal vascular tissues. Human umbilical vein endothelial cells (HUVEC) were transfected with miR-106b-5p mimic or negative control mimic, and apoptosis was induced by serum starvation and tumor necrosis factor-α (TNF-α) treat. Western blotting and real-time RT-PCR experiments were used to detect PTEN expression levels and TNF-α-induced apoptosis was evaluated by the activation of caspase-3 and cell DNA fragmentation levels in HUVEC.Results:The expression of miR-106b-5p was significantly downregulated in plaques than in normal vascular tissues. TNF-α significantly downregulated miR-106b-5p expression levels and upregulated activation of caspase-3 and cell DNA fragmentation levels in HUVEC. Overexpression of miR-106b-5p with miR-106b-5p mimic inhibited PTEN expression and TNF-α-induced apoptosis in HUVEC. Luciferase reporter assays confirmed that miR-106b-5p binds to PTEN mRNA 3’ untranslated region site.Conclusion:MiR-106b-5p could inhibit the expression of PTEN in vascular ECs, which could block TNF-α-induced activation of caspase-3, thus prevent ECs apoptosis in atherosclerosis diseases.

Circulating Chemerin Is Associated With Carotid Plaque Instability, Whereas Resistin Is Related to Cerebrovascular Symptomatology.

The rupture of unstable carotid atherosclerotic plaques is one of the main causes of cerebrovascular ischemic events. There is need for circulating markers that can predict plaque instability and risk of stroke. Proinflammatory chemerin, leptin, and resistin, along with anti-inflammatory adiponectin, are adipokines with direct influence on vascular function. We investigated the association of circulating adipokines with carotid plaque instability and cerebrovascular symptomatology. Neurologically symptomatic and asymptomatic patients (n=165) scheduled for carotid endarterectomy were recruited. Fasting blood samples were collected preoperatively; adiponectin and leptin levels were determined by radioimmunoassay; and chemerin and resistin levels were measured by enzyme-linked immunosorbent assays. The instability of plaque specimens was assessed using gold-standard histological classifications. Chemerin was significantly associated with plaque instability. The fully adjusted model, accounting for age, sex, body mass index, high-sensitivity C-reactive protein, type 2 diabetes mellitus, and circulating adiponectin, leptin, and resistin, yielded an odds ratio of 0.991 (95% confidence interval 0.985-0.998) for plaque instability per unit increase in chemerin. High leptin levels were significantly associated with presence of specific features of plaque instability. In subjects with type 2 diabetes mellitus, resistin levels were significantly elevated in symptomatic when compared with asymptomatic subjects (P=0.001) and increased the risk of cerebrovascular symptomatology (adjusted odds ratio 1.264, 95% confidence interval 1.004-1.594). Low chemerin and high resistin levels were associated with carotid disease severity, suggesting that these adipokines may act as potential markers for plaque instability and stroke risk. Future studies are needed to assess causation between circulating adipokines and plaque instability.

Mast cells are increased in the media of coronary lesions in patients with myocardial infarction and may favor atherosclerotic plaque instability

ObjectivesMast cells (MCs) may play an important role in plaque destabilization and atherosclerotic coronary complications. Here, we have studied the presence of MCs in the intima and media of unstable and stable coronary lesions at different time points after myocardial infarction (MI). MethodsCoronary arteries were obtained at autopsy from patients with acute MI (up to 5 days old; n=27) and with chronic MI (5–14 days old; n=18), as well as sections from controls without cardiac disease (n=10). Herein, tryptase-positive MCs were quantified in the intima and media of both unstable and stable atherosclerotic plaques in infarct-related and non-infarct-related coronary arteries. ResultsIn the media of both acute and chronic MI patients, the number of MCs was significantly higher than in controls. This was also found when evaluating unstable and stable plaques separately. In patients with chronic MI, the number of MCs in unstable lesions was significantly higher than in stable lesions. This coincided with a significant increase in the relative number of unstable plaques in patients with chronic MI compared with control and acute MI. No differences in MC density were found between infarct-related and non-infarct-related coronary arteries in patients with MI. ConclusionThe presence of MCs in the media of both stable and unstable atherosclerotic coronary lesions after MI suggests that MCs may be involved in the onset of MI and, on the other hand, that MI triggers intra-plaque infiltration of MCs especially in unstable plaques, possibly increasing the risk of re-infarction.

Imaging of hypoxia in mouse atherosclerotic plaques with 64Cu-ATSM

IntroductionCardiovascular disease is the leading cause of death in the United States. The identification of vulnerable plaque at risk of rupture has been a major focus of research. Hypoxia has been identified as a potential factor in the formation of vulnerable plaque, and it is clear that decreased oxygen plays a role in the development of plaque angiogenesis leading to plaque destabilization. The purpose of this study is to demonstrate the feasibility of copper-64 labeled diacetyl-bis (N4-methylthiosemicarbazone) (64Cu-ATSM), a positron-emitting radiopharmaceutical taken up in low-oxygen-tension cells, for the identification of hypoxic and potentially unstable atherosclerotic plaque in a mouse model. Methods64Cu-ATSM PET was performed in 21 atherosclerotic apolipoprotein E knockout (ApoE−/−) mice, 6 of which were fed high-fat diet (HFD) while the others received standard-chow diet (SCD), and 13 control wild type mice fed SCD. 4 SCD ApoE−/− mice and 4 SCD wild type mice also underwent 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography (PET) imaging one day prior to 64Cu-ATSM PET. Results64Cu-ATSM uptake was increased in the aortic arch in SCD ApoE−/− mice (average aortic arch/muscle (A/M) standardized uptake value ratio 7.5–30min post injection: (5.66±0.23) compared to control mice (A/M SUV ratio 7.5–30min post injection (3.87±0.22), p<0.0001). HFD ApoE−/− mice also showed similarly increased aortic arch uptake on PET imaging in comparison to control mice. Immunohistochemistry in both HFD and SCD ApoE−/− mice revealed noticeable hypoxia by pimonidazole stain in atherosclerosis which was co-localized to macrophage by CD68 staining. Autoradiography assessment demonstrated the presence of hypoxia by 64Cu-ATSM uptake correlated with pimonidazole uptake within the ex vivo atherosclerotic aortic arch specimens. A significant increase in 18F-FDG uptake in the SCD ApoE−/− mice in comparison to controls was also observed at delayed time points. ConclusionThis pre-clinical study suggests that 64Cu-ATSM is a potential PET tracer for hypoxia imaging in atherosclerosis. Advances in Knowledge and Implications for Patient CareWhile studies in humans are necessary for conclusive data, in the long term, a 64Cu-ATSM PET imaging strategy could help facilitate the study of plaque biology in human patients.

Smooth Muscle Enriched Long Noncoding RNA (SMILR) Regulates Cell Proliferation.

Phenotypic switching of vascular smooth muscle cells from a contractile to a synthetic state is implicated in diverse vascular pathologies, including atherogenesis, plaque stabilization, and neointimal hyperplasia. However, very little is known about the role of long noncoding RNA (lncRNA) during this process. Here, we investigated a role for lncRNAs in vascular smooth muscle cell biology and pathology. Using RNA sequencing, we identified >300 lncRNAs whose expression was altered in human saphenous vein vascular smooth muscle cells following stimulation with interleukin-1α and platelet-derived growth factor. We focused on a novel lncRNA (Ensembl: RP11-94A24.1), which we termed smooth muscle-induced lncRNA enhances replication (SMILR). Following stimulation, SMILR expression was increased in both the nucleus and cytoplasm, and was detected in conditioned media. Furthermore, knockdown of SMILR markedly reduced cell proliferation. Mechanistically, we noted that expression of genes proximal to SMILR was also altered by interleukin-1α/platelet-derived growth factor treatment, and HAS2 expression was reduced by SMILR knockdown. In human samples, we observed increased expression of SMILR in unstable atherosclerotic plaques and detected increased levels in plasma from patients with high plasma C-reactive protein. These results identify SMILR as a driver of vascular smooth muscle cell proliferation and suggest that modulation of SMILR may be a novel therapeutic strategy to reduce vascular pathologies.

The curative effect comparison of two kinds of therapeutic regimens on decreasing the relative intensity of microembolic signal in CLAIR trial

BackgroundMicroembolic signals (MESs) are direct markers of unstable large artery atherosclerotic plaques. In a previous study, we found that the number of MESs is associated with stroke recurrence and that clopidogrel plus aspirin more effectively reduce the number of MESs than does aspirin alone. Stroke recurrence is associated with not only the number of MESs but also the size of the MES, which can theoretically be estimated by monitoring the MES intensity via transcranial doppler (TCD). Thus, we compared the effects of clopidogrel and aspirin with aspirin alone on MES intensity using TCD. MethodsWe recruited 100 patients who experienced acute ischemic stroke or transient ischemic attack (TIA) within 7days of symptom onset. All patients also had large artery stenosis in the cerebral or carotid arteries and the presence of MES as revealed by TCD. The patients were randomized to receive either aspirin or clopidogrel and aspirin for 7days. MES monitoring was performed on days 2 and 7. ResultsIntent-to-treat (ITT) analysis (46 patients in the dual therapy group, 52 patients in the monotherapy group) and per–protocol (PP) analysis (25 patients in the dual therapy group, 31 patients in the monotherapy group) were performed on 98 patients. The primary finding was that the MES intensity was dramatically reduced in the dual therapy group. ITT analysis of the dual therapy group revealed that the MES intensity was 8.04 (0–16) dB before treatment, 0.00 (0–17) dB on day 2, and 0.00 (0−12) dB on day 7 (P=0.000). In the monotherapy group, the MES intensity was 9.00 (0−20) dB before treatment, 8.25 (0–17) dB on day 2, and 7.0 (0–18) dB on day 7 (P=0.577). PP analysis revealed similar results. No severe hemorrhagic complications were detected. The two patients in this study who experienced stroke recurrence were in the monotherapy group. ConclusionsClopidogrel and aspirin more effectively decrease the MES intensity than aspirin alone in patients with large artery stenotic minor stroke or TIA.

Frequency Analysis of the Photoacoustic Signal Generated by Coronary Atherosclerotic Plaque

The identification of unstable atherosclerotic plaques in the coronary arteries is emerging as an important tool for guiding percutaneous coronary interventions and may enable preventive treatment of such plaques in the future. Assessment of plaque stability requires imaging of both structure and composition. Spectroscopic photoacoustic (sPA) imaging can visualize atherosclerotic plaque composition on the basis of the optical absorption contrast. It is an established fact that the frequency content of the photoacoustic (PA) signal is correlated with structural tissue properties. As PA signals can be weak, it is important to match the transducer bandwidth to the signal frequency content for in vivo imaging. In this ex vivo study on human coronary arteries, we combined sPA imaging and analysis of frequency content of the PA signals. Using a broadband transducer (−3-dB one-way bandwidth of 10–35 MHz) and a 1-mm needle hydrophone (calibrated for 1–20 MHz), we covered a large frequency range of 1–35 MHz for receiving the PA signals. Spectroscopic PA imaging was performed at wavelengths ranging from 1125 to 1275 nm with a step of 2 nm, allowing discrimination between plaque lipids and adventitial tissue. Under sPA imaging guidance, the frequency content of the PA signals from the plaque lipids was quantified. Our data indicate that more than 80% of the PA energy of the coronary plaque lipids lies in the frequency band below 8 MHz. This frequency information can guide the choice of the transducer element used for PA catheter fabrication.

0265 : Tumour necrosis factor- carried by microparticles from apoptotic RAW 264.7 macrophage cells triggers deleterious effects on cardiomyocytes from adult mice

After ischaemic injury and in patients with atherosclerosis, the pool of inflammatory macrophages is enlarged in the heart and in atherosclerotic plaques. Monocyte/macrophage-derived microparticles (MPs) are part of the pathological process of unstable atherosclerotic plaques. In the present study, we have focused on MPs released by apoptotic murine RAW 264.7 macrophage cell line. The hypothesis that these MPs could induce deleterious effects in adult myocardial cells was evaluated. Flow cytometry and western blot analysis showed that these MPs contained the soluble form of tumour necrosis factor alpha (TNF-α). Furthermore, cardiomyocyte sarcomere shortening amplitudes and kinetics were reduced within 5 min of exposure to MPs. Conversely, Ca 2+ transient amplitudes and kinetics were not modified. The contractile effects of MPs were completely prevented after pre-treatment with nitric oxide synthase, guanylate cyclase or TNF-α inhibitors as well as blocking TNF-α receptor 1 with neutralizing antibody. Moreover, confocal microscopy showed that, after 1 h, MPs were clearly surrounding rod-shaped cardiomyocytes, and after 2 h they were internalized into cardiomyocytes undergoing apoptosis. Indeed, after 4 h of treatment with MPs, cardiomyocytes expressed increased apoptotic markers expression (caspase-3, caspase-8, Bax and cytochrome C) as demonstrated by Western Blot. The present study showed that these MPs carry functional TNF-α and activate TNFR1 in isolated adult murine ventricular cardiomyocytes, leading to cell shortening without affecting calcium signalling via a mechanism sensitive to NO synthase inhibitor and that the mitochondrial intrinsic pathway was implicated in their proapoptotic effects. Thus, the present study allows supporting the hypothesis that MPs from human carotid atherosclerotic plaques may contain active TNF-α, which could contribute to MP-induced inflammatory signals in human atherosclerotic lesions leading to myocardial infarction. The author hereby declares no conflict of interest

Assessment of vulnerable and unstable carotid atherosclerotic plaques on endarterectomy specimens.

The types of lesion instability responsible for the majority of acute coronary events frequently include plaque disruption and plaque erosion with superimposed thrombosis. The term 'vulnerable plaque is used to describe atherosclerotic (ATS) plaques that are particularly prone to rupture and susceptible to thrombus formation, such as the thin-cap fibroatheroma (TCFA). The aim of the present study was to assess the morphological and histological differences between plaques that are unstable and those that are vulnerable to instability. Carotid artery endarterectomy specimens were obtained from 26 patients with carotid artery stenosis, consisting of 20 men and 6 women (age range, 35-80 years). Histological and morphometric methods were used to visualize and characterize the ATS plaques. Among the 26 carotid ATS plaques, 23% were stable, 23% were unstable and 54% were vulnerable. With regard to morphometric characteristics, the following mean values were obtained for the TCFA and unstable plaques, respectively: Fibrous cap thickness, 21.91 and 11.66 µM; proportion of necrotic core area in the total plaque area, 25.90 and 22.03%; and the proportion of inflammatory area in the total plaque area, 8.41 and 3.04%. No plaque calcification was observed in any of them. Since ATS coronary artery disease is considerably widespread and fatal, it is crucial to further study ATS lesions to obtain an improved understanding of the nature of vulnerable and unstable plaques. The methods used to detect plaque size, necrotic core area and fibrous cap thickness are considered to be particularly useful for identifying vulnerable and unstable plaques.