Atherosclerotic Subjects Research Articles

Decreased atherosclerosis and abdominal aortic aneurysm in mice deficient in polymeric immunoglobulin receptor

Abstract Funding Acknowledgements Type of funding sources: Foundation. Main funding source(s): La Caixa Banking Foundation and Comunidad de Madrid Background Polymeric immunoglobulin receptor (PIGR) is a transmembrane protein involved in the transcytosis of polymeric immunoglobulins that is widely expressed in mucosal epithelial cells. Recent data from our group described increased PIGR levels in plasma and tissue of atherosclerotic subjects. However, the role for PIGR in the mechanisms involved in vascular remodeling has not been addressed. Purpose To analyse the role for PIGR in experimental mouse models of atherosclerosis and abdominal aortic aneurysm (AAA). Methods We analyzed PIGR expression, secretion and distribution in early atherosclerotic plaques and AAA tissues compared to healthy aortic samples, as well as in macrophages derived from the human monocytic THP-1 cell line. Next, we studied the effect of global PIGR deficiency in experimental atherosclerosis (Ldlr-/- Pigr-/- mice fed with atherogenic diet for 10 weeks). In addition, the contribution of hematopoietic PIGR was assessed in both experimental atherosclerosis and AAA (Ldlr-/- mice fed with atherogenic diet and 1 µg/Kg/min angiotensin II infusion for 28 days) by bone marrow transplantation experiments. Finally, bone marrow-derived macrophages (BMDMs) from Pigr-/- and control mice were isolated to assess their inflammatory expression profile and modulation of polarization in response to oxidized LDL (ox-LDL), as well as their ability to promote foam cell formation. Results PIGR expression was increased in the intima of early human atherosclerotic lesions and showed colocalization with CD68+ expression. PIGR expression was also increased during the in vitro differentiation of THP-1 monocytes to macrophages with phorbol myristate acetate. Both global and hematopoietic-specific PIGR deficient mice displayed reduced plaque size in the aortic sinuses as well as a strong decrease in macrophage infiltration (CD68+ cells), while no changes in smooth muscle cells (α-SMA+ cells) and collagen content were observed. Regarding AAA, increased PIGR levels were found in human AAA tissues, associated with infiltrating CD68+ cells. PIGR deficiency in hematopoietic cells also showed significantly reduced AAA dilatation and reduced macrophage infiltration. In vitro experiments with BMDMs obtained from Pigr-/- mice and stimulated with ox-LDL showed reduced proinflammatory gene expression (TNF-a and IL-6) and M1 markers (iNOS and CD11c) as well as diminished foam cell formation. Conclusions PIGR deficiency in hematopoietic cells decreases atherosclerosis and AAA progression by modulating macrophage inflammatory responses, suggesting a novel proinflammatory role for PIGR in vascular pathologies. Future experiments will test the potential therapeutic effect of modulating PIGR to dampen the inflammatory profile in vascular diseases.Results in human and mouse studies

A Machine Learning Approach for Atherosclerosis Detection in Murine Models

The early detection of atherosclerosis has been the interest of researchers in order to prevent diseases progression. And, most of cases diagnosed with atherosclerosis in late stages resulted of subjective ways of diagnosis in the healthcare sector. Consequently, in order to help cardiologists diagnose atherosclerosis in early stages with an objective, fast and accurate way, we are proposing a machine learning model based on autoencoders that enables atherosclerosis detection from MRI images of murine subjects. This novel way of automated system consists of applying various image processing techniques on the input MRI images. Then, after training, testing and validation it will be capable of classifying the images as atherosclerotic or not based on a specific threshold for the reconstruction error calculated from the autoencoders' output. An autoencoder is a feed-forward neural network that has its input neurons equal to the output neuron. The classification works by comparing the reconstructed images to the original input images and evaluating loss between them. Since the autoencoder is trained on healthy images, reconstruction error of the healthy images would be low while that of atherosclerotic subjects would be higher. By setting a threshold for the loss, we can classify the images as healthy or atherosclerotic. The pre-processing of these images was made using a Block Matching 3D (BM3D) filter to remove the noise in the images prior to application of a Contrast Limited Adaptive Histogram Equalization (CLAHE) to enhance the contrast. The next step included introducing the dataset into the autoencoder to start training on the healthy images, after increasing the images number with augmentation. The results showed a reconstruction loss of 0.018 while using the stacked architecture of the autoencoder and 0.0366 when using the convolutional autoencoder architecture.

Outcomes of Mechanical Thrombectomy of Acute Basilar Artery Occlusion Due to Underlying Intracranial Atherosclerotic Disease

Background Intracranial atherosclerotic disease (ICAD) is a common cause of posterior circulation acute ischemic stroke. We aimed to compare baseline characteristics, recanalization rates, and clinical outcomes in patients with acute basilar occlusion due to underlying ICAD to patients with other causes of occlusion. Methods The Trevo Registry (ClinicalTrials.gov Identifier: NCT02040259) was a prospective open‐label mechanical thrombectomy registry that included 2008 patients from 76 sites across 12 countries. For a secondary analysis, we selected patients with isolated acute basilar occlusion. We then classified patients into 2 groups: patients with acute basilar artery occlusion due to ICAD (ICAD group) and patients with acute basilar occlusion due to another cause (non‐ICAD group). Results We identified 90 subjects with acute basilar occlusion. According to stroke mechanism, there were 9 (10.0%) patients in the ICAD group and 81 (90.0%) in the non‐ICAD group. There was a significant difference ( P =0.001) in the lesion location between the 2 groups, with 60.5% of non‐ICAD occlusions located in the distal region and no ICAD occlusions there. The 2 groups showed a significant difference ( P =0.003) for rescue therapy with balloon angioplasty, with 33.3% in the ICAD group and 1.2% in the non‐ICAD group, respectively. In a multivariable model adjusted for age, baseline National Institutes of Health Stroke Scale, and intravenous tissue‐type plasminogen activator (tPA), the odds of 90‐day death (odds ratio=4.6; P =0.10) were higher for atherosclerotic subjects. Conclusion Acute basilar occlusions related to ICAD showed a similar good clinical outcome (modified Rankin scale (mRS) 0–2) and a tendency for a higher rate of 90‐day mortality compared with non‐ICAD occlusions.

The Relationship between Atherosclerosis and Gut Microbiome in Patients with Obstructive Sleep Apnoea

Background: Obstructive sleep apnoea (OSA) and gut dysbiosis are known risk factors for atherosclerosis. However, only very few studies have been focused on the relationship between OSA, atherosclerosis, and the intestinal microbiome, all in animal models. Methods: Twenty-two patients with OSA, 16 with and 6 without carotid atherosclerosis were involved in the study. After a diagnostic sleep examination, the intima media thickness (IMT) was measured and plaques were found using carotid ultrasound. Blood was also drawn for metabolic profile, and a stool sample was provided for 16S ribosomal RNA microbiome investigation. Results: An increased maximal common carotid artery (CCA) IMT was significantly associated with decreased phylum-level diversity. The level of Peptostreptococcaceae was significantly lower in atherosclerotic subjects. Some other candidate microbes appeared in the two groups at the genus level as well: Bilophila, Romboutsia, Slackia, and Veillonella in the non-atherosclerotic group; and Escherichia-Shigella, Prevotella, and Ruminococcaceae in the atherosclerotic group. Conclusions: This is the first pilot research to analyze the association between the gut microbiome and atherosclerosis in adult patients with OSA with and without carotid atherosclerosis. Dysbiosis and individual bacteria may contribute to the development of carotid atherosclerosis in patients with OSA. Further investigations are necessary to reveal a more precise background in a larger sample.

Abdominal arterial lesions associated with antiphospholipid antibodies: a comparative cross-sectional magnetic resonance angiography study.

Case reports and small case series suggest that stenotic lesions of the renal, coeliac and mesenteric arteries may occur in the antiphospholipid syndrome (APS) resulting in clinical consequences such as hypertension and abdominal angina. The objective was to determine the prevalence of stenotic lesions in arteries arising from the middle aorta in patients with antiphospholipid antibodies (aPL) compared with healthy, hypertensive and atherosclerotic controls. In a cross-sectional comparative radiological study using magnetic resonance angiography (MRA), we assessed five groups of subjects for the prevalence of stenotic lesions in arteries arising from the middle aorta: APS/aPL positive, healthy renal donors, patients with hypertension, patients with atherosclerosis defined radiologically and patients with systemic lupus erythematosus and vasculitis who were negative for aPL. All subjects underwent MRA in suspended respiration and images were assessed by two senior radiologists blinded to the clinical details. In the atherosclerosis group, vascular stenotic lesions were more prevalent (71%) than in any other group (P ≤0.000002). The prevalence of all stenotic lesions in aPL positive patients (33%) was significantly higher than in the renal donors (18%) and hypertensive patients (19%) (P ≤0.009). Renal artery stenosis was significantly more prevalent in aPL positive patients than in renal donors (P ≤0.0006) but similar to the prevalence in hypertensive patients. Coeliac and/or mesenteric lesions were significantly more common in aPL positive patients vs hypertensive patients (P ≤0.001). Stenoses did not correlate with traditional risk factors. Arterial stenotic lesions in arteries arising from the middle aorta were highly prevalent in atherosclerotic subjects and were more common in aPL-positive patients than in hypertensive patients and healthy renal donors.

P4442miR-200c is up-regulated in lesional skin and plasma of patients affected by psoriasis and correlates with disease severity and cardiovascular risk

Abstract Background Psoriasis is a common, chronic inflammatory disease involving skin. Psoriatic patients mostly show the plaque-type clinical form and several comorbidities, including cardiovascular (CV) diseases. We previously showed that miR-200 family members (miR-200s) is induced by reactive oxygen species (ROS). miR-200c is the most up-regulated member and is responsible for apoptosis, senescence, endothelial dysfunction, ROS increase and nitric oxide decrease. Circulating miR-200c is upregulated in Familial Hypercholesterolemia in children, a pathology associated with ROS increase and atherosclerosis. miR-200c increases also in carotid plaques and plasma of atherosclerotic pts vs healthy subjects and positively correlates with plaque instability biomarkers. Purpose Given the role of miR-200s in ROS modulation, endothelial dysfunction, cardiac remodelling and inflammation, all features associated with psoriasis, we wondered whether miR-200s were modulated in lesional skin (LS) and plasma of psoriatic patients (Pso) and whether miR-200 levels correlated with CV risk. Methods 29 Pso were compared to 29 control subjects (Ctrl) age- and sex-matched. All Pso had a severe psoriasis, defined as Psoriasis Area and Severity Index (PASI) >10 and one of the following: at least two systemic psoriasis treatments, psoriasis onset <40 years of age, disease duration >10 years. Exclusion criteria were: diabetes, cerebrovascular events, myocardial infarction, and/or myocardial revascularization, psoriatic arthritis. Total RNA was extracted from plasma and miR-200 levels assayed by quantitative real-time PCR. The clinical parameters were similar between the two groups except for Total Cholesterol (mg/dl) (Ctrl 193.3±6.2; Pso 213.2±6.9; P<0.05). Blood pressure measurement, wave reflection analysis and pulse wave velocity (PWV) were similar between groups, echocardiographic parameters were different for left ventricular (LV) mass index (g/m2) (Ctrl 84.2±5.2; Pso 102.5±4.7; P<0.05) and relative wall thickness (RWT) (Ctrl 0.4±0.0; Pso 0.48±0.0; P<0.01). Total RNA was extracted from biopsies of nonlesional (NLS) and LS of 6 Pso and 6 healthy subject skin (HS). Results miR-200s were increased in LS vs NLS samples. miR-200c was the most expressed and was upregulated also in LS vs HS (2.0+0.2 fold-increase; P<0.01). Circulating miR-200c (2.5+0.5 fold-increase; P<0.05) and miR-200a (4.8+0.7 fold; P<0.0001) were up-regulated in Pso vs Ctr. Circulating miR-200c positively correlated with LV mass (Rs=0.32; P<0.05), RWT (Rs=0.32; P<0.05) and diastolic dysfunction assessed with E/e' parameters (Rs=0.34; P<0.05). Circulating miR-200a correlated only with LV mass, although not significantly (Rs=0.30; P=0.06). miR-200c exhibited a significant positive correlation with PASI (Rs=0.43; P<0.05) and with disease duration of (Rs=0.40; P<0.05). Conclusion miR-200c is upregulated in skin plaques and plasma of Pso, and might be involved in inflammatory and CV risk increase in these patients. Acknowledgement/Funding Ministero della Salute RF-2016-02362708 to AM and MCC

P4425The exercise-induced troponin I elevation is highly correlated with power output during exercise in recreational cyclists with coronary atherosclerosis

Abstract Background Following strenuous exercise there is an increase cardiac Troponins (cTn) elevation considered being a physiological response. During prolonged strenuous physical activity, high work-loads may induce demand myocardial ischemia due to an oxygen demand/supply mismatch in susceptible subjects, causing an excessive cTn elevation. Purpose This study aimed to assess the relationship between exercise-induced cTnI elevation and direct measurement of work performed during prolonged strenuous exercise in subjects with and without atherosclerotic CAD. Methods Work during a 91 km mountain bike race was quantified by Stages™ power meters. Power (Watt) and heart rate data were stored in Garmin™ Forerunner 935 monitors. Coronary computed tomography angiography was performed after the race. Blood pressure was measured 4 times during the race. Blood samples (hs-cTnI from Abbot Diagnostics) were obtained one day prior to the race and at 3 and 24 h after the race. Data are presented as mean±SD or median (25th and 75th percentile). Results 40 subjects (10 women) were included in the final analysis. 15 Participants (4 women) had atherosclerosis, none had obstructive CAD. These participants were significantly older (55±8 years vs. 46±8 years p=0.007) and had higher training volumes (METS: 69 (64–102) hrs/week) compared with normal subjects (METS: 51 (33–88) hrs/week) (p=0.03). Baseline cTnI was higher (p=0.04) in the atherosclerotic group (4.5 (3.4–8.8) ng/L) compared with normals (2.6 (1.6–4.8) ng/L). There were no differences in baseline blood pressure, peak VO2 max, heart rate or BMI. There was no significant difference in race duration between normals (3.9 (3.5–4.5) hrs) and subjects with atherosclerosis (4.1 (3.6–4.5) hrs). During the race there were no differences in peak power or peak Watt/kg. cTnI increased after the race in all participants, but there were no differences between groups: 3h: atherosclerosis: 89 (27–131) ng/L vs. normal 77 (36–104) ng/L, 24h: atherosclerosis: 13 (6.3–23.7) ng/L vs. normal: 17 (12–37) ng/L. There were no significant difference between the groups in average power during the race: atherosclerosis: 167±50 Watt vs. 174 (±50) Watt or ratio: 2.0±0.49 Watt/kg vs 2.2±0.58 Watt/kg during the race. Maximal systolic and diastolic blood pressures during the race were higher (p=0.002) in the atherosclerotic group: SBP: 241±14 mmHg vs. 219±26 mmHg, DBP: 107±8 vs 95±8 mmHg. In atherosclerotic subjects cTnI both at 3h and 24 h were highly correlated (p<0.001) with Watt/kg ratio during the race in contrast to no correlations in the normal group (Figure). Conclusions Our findings suggest that the presence of coronary atherosclerosis, even in the absences of significant stenosis, alters the relationship between workload and the troponin response. This indicates different release kinetics in exercise-induced cTn in participants with and without CAD, with prolonged elevation in cTnI in CAD subjects exceeding the highest work-intensities. Acknowledgement/Funding Grant Western Norway Health service, Grant ConocoPhillips, Grant Simon Fougner Hartmanns Familyfund

Metabolic activity in bone metastases of breast and prostate cancer were similar as studied by 18F-FDG PET/CT. The role of 99mTc-MDP.

The aim of this study was to compare the metabolic activity of metastatic foci from breast and prostate cancer patients as scanned by fluorine-18-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) and by technetium-99m methyl diphosphonate (99mTc-MDP) bone scan (BS). Forty one patients were studied, divided into 2 groups based on histologically confirmed diagnosis: a) Breast cancer group, 23 women, mean age: 61±12 years, range: 37-79 years and, b) Prostate cancer group, 18 men, mean age 68±8 years, range: 52-82 years. Another group of 17 non cancer atherosclerotic subjects 9 women and 8 men, of mean age and age range similar to the above were also studied for comparison. The R index (the total count rate in bone metastases divided by the total count rate in a contralateral area), the maximum semi-quantitative standardized uptake value (SUVmax) of BS lesions and the mean number of metastases were evaluated. For the metastatic findings in the PET/CT scans the automatic method of contouring with 50% background cut-off was used, while for the 99mTc-MDP BS metastases were delineated manually. The mean R index of the bone metastatic foci studied by 18F-FDG PET/CT was 1.89±0.69 for Groups I and II patients. There was no significant difference of the R index between prostate cancer and breast cancer metastases (1.95±0.86 vs 1.83±0.52). The average SUVmax value was significantly higher in breast cancer patients than in prostate cancer patients (5.15±2.54 vs 4.01±1.71; P<0.05). There was no significant correlation in both cancer groups between R index and SUVmax values. The number of metastatic foci diagnosed by the 99mTc-MDP BS scan was much less than by the 18F-FDG PET/CT. No significant correlation was noticed in the metabolic activity-glucose utilization of metastatic bone foci between breast and prostate cancer cases. This observation validates the independent value of analyzed diagnostic methods and suggests negligible influence of glucose utilization in bone re-modeling in the above metastatic cancer cells. The 18F-FDG PET/CT bone scan was much better in diagnosing metastases compared to the 99mTc-MDP scan.

ATHEROSCLEROSIS;

Objectives: To compare the adiponectin/leptin ratio and intima media thicknessbetween normal and atherosclerotic individuals. To find out the association of A/L ratio withIMT in atherosclerotic patients. Study Design: Comparative cross sectional study. Placeand Duration of Study: This study was conducted at department of Physiology, BMSI incollaboration with Radiology department, Jinnah Postgraduate Medical Centre Karachi, from16th December 2014 to 15th December 2015. Methodology: 160 subjects having Dopplerultrasound of neck were selected on the basis of pre-determined criteria. They were grouped incases i.e. atherosclerotic group (80 subjects) and control i.e. healthy group (80 subjects). Age&gt; 20 years, both genders, patients for ultrasound Doppler for neck region irrespective of theirprovisional diagnosis, patients suffering from co-morbid conditions like diabetes, hypertension,chronic kidney disease, chronic liver disease, asthma and COPD and Healthy individuals(Controls) were included in this study. Patients who fail to gave consent and patients whoseultrasound findings were not up to the mark required for diagnosis were excluded from thisstudy. Results: Out of 160 patients in this study in control Male: Female ratio was 1:1.1 whilein cases, Male: Female ratio = 1:1. Majority (53.1%) of cases had age &gt;50 years {41(51.25%)in controls and 44(55%) in cases}. Mean ages in two groups (controls and cases) were49.39+12.30 years and 53.31+10.09 years respectively (p = 0.029).44 (22.5%) patients weresmoker and 116 (72.5%) were non-smoker. In control group, 13 (16.3%) were smokers whereasin cases 31 (38.8%) were smokers (p = 0.002). Mean triglyceride levels were statistically highin atherosclerotic subjects as compared to healthy individuals (138.31+62.65 mg/dl and125.73+46.17 mg/dl respectively; p=0.024). Mean IMT of both right and left carotid arteriesexamined via carotid Doppler ultrasound were statistically high in atherosclerotic individuals ascompared to healthy subjects {(Right carotid; 0.83+0.23 mm and 0.63+0.16 mm respectively;p=0.022) (Left carotid; 0.85+0.25 mm and 0.64+0.16 mm respectively; p=0.004)}. MeanLeptin levels were statistically high in atherosclerotic individuals as compared to healthysubjects (332.32+555.31 ng/ml and 254.55+224.66 ng/ml respectively; p=0.001). However,the difference in Adiponectin levels was insignificant (7.93+4.80 μg/ml and 9.45+4.73 μg/mlrespectively; p=0.343). Conclusion: We conclude that plasma levels of Adiponectin and Leptinare significantly higher in atherosclerotic than normal individuals.

English

Objectives: To compare the adiponectin/leptin ratio and intima media thicknessbetween normal and atherosclerotic individuals. To find out the association of A/L ratio withIMT in atherosclerotic patients. Study Design: Comparative cross sectional study. Placeand Duration of Study: This study was conducted at department of Physiology, BMSI incollaboration with Radiology department, Jinnah Postgraduate Medical Centre Karachi, from16th December 2014 to 15th December 2015. Methodology: 160 subjects having Dopplerultrasound of neck were selected on the basis of pre-determined criteria. They were grouped incases i.e. atherosclerotic group (80 subjects) and control i.e. healthy group (80 subjects). Age&gt; 20 years, both genders, patients for ultrasound Doppler for neck region irrespective of theirprovisional diagnosis, patients suffering from co-morbid conditions like diabetes, hypertension,chronic kidney disease, chronic liver disease, asthma and COPD and Healthy individuals(Controls) were included in this study. Patients who fail to gave consent and patients whoseultrasound findings were not up to the mark required for diagnosis were excluded from thisstudy. Results: Out of 160 patients in this study in control Male: Female ratio was 1:1.1 whilein cases, Male: Female ratio = 1:1. Majority (53.1%) of cases had age &gt;50 years {41(51.25%)in controls and 44(55%) in cases}. Mean ages in two groups (controls and cases) were49.39+12.30 years and 53.31+10.09 years respectively (p = 0.029).44 (22.5%) patients weresmoker and 116 (72.5%) were non-smoker. In control group, 13 (16.3%) were smokers whereasin cases 31 (38.8%) were smokers (p = 0.002). Mean triglyceride levels were statistically highin atherosclerotic subjects as compared to healthy individuals (138.31+62.65 mg/dl and125.73+46.17 mg/dl respectively; p=0.024). Mean IMT of both right and left carotid arteriesexamined via carotid Doppler ultrasound were statistically high in atherosclerotic individuals ascompared to healthy subjects {(Right carotid; 0.83+0.23 mm and 0.63+0.16 mm respectively;p=0.022) (Left carotid; 0.85+0.25 mm and 0.64+0.16 mm respectively; p=0.004)}. MeanLeptin levels were statistically high in atherosclerotic individuals as compared to healthysubjects (332.32+555.31 ng/ml and 254.55+224.66 ng/ml respectively; p=0.001). However,the difference in Adiponectin levels was insignificant (7.93+4.80 μg/ml and 9.45+4.73 μg/mlrespectively; p=0.343). Conclusion: We conclude that plasma levels of Adiponectin and Leptinare significantly higher in atherosclerotic than normal individuals.

Macrophage migration inhibitory factor promoter polymorphisms (\u2212794 CATT5\u20138): Relationship with soluble MIF levels in coronary atherosclerotic disease subjects

BackgroundWe analyzed the relationship of −794 CATT5–8 MIF polymorphisms with soluble MIF in Coronary Atherosclerotic Disease (CAD) patients.MethodsA total of 256 patients selected, on which 186 normal-coronary and 70 Coronary artery disease subjects, were recruited in the study (Retrospectively registered). Genotyping of −794 CATT5–8 polymorphisms were performed by PCR and DNA sequencing. Serum MIF levels were measured using an ELISA kit. Patients were classified by coronary angiogram, and CAD based on Gensini’s integral degree (angiographic scoring system).ResultsThe allele frequency and genotype frequency of −794 CATT5–8 did not show any differences in normal-coronary subjects and CAD subjects. In CAD patients, serum MIF levels was lower in CATT (5) subjects than in CATT (7) subjects, while the genotype of −794 CATT5–8 did not show differences in serum MIF levels. In addition, we found a decrease in serum MIF levels in carriers of the (5/5) genotypes the −794 CATT5–8 MIF polymorphisms, although it was not significant. There was no relationship of CAD class and the allele frequency of −794 CATT5–8.ConclusionsThis study found no association between CAD class and −794 CATT5–8 MIF polymorphisms with soluble MIF levels in CAD Subjects.Trial registrationNCT01750502 (November 2012, Retrospectively registered).

Abstract WP141: Genetic Polymorphisms Influence Carotid Atherosclerosis

Introduction: Genetic susceptibility affects atherosclerosis in humans. Polymorphisms of genes of angiotensin-converting enzyme (ACE), lipoprotein APOE (APOE), IL1 receptor antagonist (IL1Ra) and myeloperoxidase (MPO) are associated with several components of atherosclerotic disease. We evaluated allelic and genotypic frequencies and their association with age at presentation, vascular risk factors, and presence of symptoms in subjects with carotid atherosclerosis. Methods: We studied Argentine patients with severe carotid atherosclerosis and controls from the general population. Age, vascular risk factors and presence of neurological symptoms were recorded. DNA was obtained from peripheral blood and PCR or PCR-RFLP were used to typify ACE, APOE, IL1Ra, and MPO genes. Allelic and genotypic frequencies were compared and genotypic susceptibility variants were established. Chi-square and good fit test were applied for differences between expected and observed frequencies. Results: There were 137 patients, 36 women and 101 men, aged 67±8 years. Symptomatic subjects younger than 60 years had higher frequency of the alleles ACE-DD, associated to vasoconstriction, endothelial proliferation, oxidation, and apoptosis (p&lt;0.01); IL1RN-12/22, associated to inflammation and apoptosis (p&lt;0.01); and MPO*GA/AA, associated to less oxidative response and proatherogenic (p&lt;0.05). Subjects older than 60 years had a genetic profile similar to the general population without atherosclerosis, with similar prevalence of ACE-ID/II, IL1RN-11, and MPO-GG and a higher frequency of APOE23, 24 and 34m. Independent associations of ACE*D and IL1RN*2 with dyslipidemia and of MPO-GA and APOE-34 with hypertension were observed. Conclusions: Subjects with carotid atherosclerosis are genetically different from the general population. Carriers of certain gene variants were predominant among atherosclerotic subjects, suggesting susceptibility, and others were more prevalent in controls, suggesting protection. Some polymorphisms and their combinations are associated with occurrence of symptomatic disease at an earlier age. The genetic profile of older patients does not substantially differ from the general population.

Simvastatin Efficiently Lowers Small LDL-IgG Immune Complex Levels: A Therapeutic Quality beyond the Lipid-Lowering Effect.

We investigated a polyethylene glycol non-precipitable low-density lipoprotein (LDL) subfraction targeted by IgG and the influence of statin therapy on plasma levels of these small LDL-IgG-immune complexes (LDL-IgG-IC). LDL-subfractions were isolated from 6 atherosclerotic subjects and 3 healthy individuals utilizing iodixanol density gradient ultracentrifugation. Cholesterol, apoB and malondialdehyde (MDA) levels were determined in each fraction by enzymatic testing, dissociation-enhanced lanthanide fluorescence immunoassay and high-performance liquid chromatography, respectively. The levels of LDL-IgG-IC were quantified densitometrically following lipid electrophoresis, particle size distribution was assessed with dynamic light scattering and size exclusion chromatography. The influence of simvastatin (40 mg/day for three months) on small LDL-IgG-IC levels and their distribution among LDL-subfractions (salt gradient separation) were investigated in 11 patients with confirmed coronary artery disease (CAD). We demonstrate that the investigated LDL-IgG-IC are small particles present in atherosclerotic patients and healthy subjects. In vitro assembly of LDL-IgG-IC resulted in particle density shifts indicating a composition of one single molecule of IgG per LDL particle. Normalization on cholesterol levels revealed MDA values twice as high for LDL-subfractions rich in small LDL-IgG-IC if compared to dominant LDL-subfractions. Reactivity of affinity purified small LDL-IgG-IC to monoclonal antibody OB/04 indicates a high degree of modified apoB and oxidative modification. Simvastatin therapy studied in the CAD patients significantly lowered LDL levels and to an even higher extent, small LDL-IgG-IC levels without affecting their distribution. In conclusion simvastatin lowers levels of small LDL-IgG-IC more effectively than LDL-cholesterol and LDL-apoB levels in atherosclerotic patients. This antiatherogenic effect may additionally contribute to the known beneficial effects of this drug in the treatment of atherosclerosis.

PP40 - Total Antioxidant Capacity (TEAC) is decreased despite high levels of uric acid in patient with atherosclerosis

Introduction The atherosclerosis development and plasma antioxidant protection are strongly linked. The intima damage is reflected through the difference between the antioxidants and oxidants in patients with atheromatous plaques. A high level of oxidative stress in atherosclerotic subjects lead to an imbalance in the serum antioxidant capacity. The purpose of the study was to evaluated the plasma Trolox Antioxidant Capacity (TEAC) and residual antioxidant activity (GAP) in the atherosclerotic patients compared with healthy humans. Materials and Methods The study included 33 subjects: 19 patients newly diagnosed with monovascular disease and 14 subjects apparently healthy. The atherosclerotic patients were selected in The Cardiology Department of University and Emergency Hospital, Bucharest. Inclusion criteria for atherosclerotic patients were: recently diagnosed monovascular disease by coronary angiography or high intima media thickness in patients with cardiovascular risk factors. None of the patients were taken statins for more than one week. The lipid profile: total cholesterol (TC), triglycerides (TG), LDLc, HDLc was determined in EDTA plasma blood after an overnight fast. Uric acid, albumin, TEAC and GAP were also evaluated. Results The LDLc, TC, HDLc and TG plasma levels were not statistically significant different between the both groups of study. The uric acid increase was statistically significant in the atherosclerosis group (p Conclusion A statistically significant difference was found for TEAC, GAP and uric acid in atherosclerotic patients compared to the controls. The decreased antioxidant capacity may signify a reduced antioxidant protection for this type of patients. Acknowledgement Ms. Elena Torac was supported by the POSDRU/159/1.5/S/137390.

Calcipressin plasma level difference between atherosclerotic and non-atherosclerotic subjects

Calcipressin plasma level difference between atherosclerotic and non-atherosclerotic subjects

A novel non-invasive ultrasonic method to assess total axial stress of the common carotid artery wall in healthy and atherosclerotic men

In the present study, developing a new non-invasive method independent from blood flow, we estimated and compared the total axial stress of the common carotid artery wall in healthy and atherosclerotic subjects. Consecutive ultrasonic images of the common carotid artery of 48 male subjects including healthy, with less and more than 50% stenosis in carotid artery were recorded. Longitudinal displacement and acceleration was extracted from ultrasonic image processing using a block matching algorithm. Furthermore, images were examined using a maximum gradient algorithm and time rate changes of the internal diameter and intima-media thickness were extracted. Finally, axial stress was estimated using an appropriate constitutive equation. Statistical analysis results showed that with stenosis initiation and its progression, axial acceleration and stress increase significantly. According to the results of the present study, maximum axial stress of the arterial wall is 1.713±0.546, 1.993±0.731 and 2.610±0.603 (kPa) in normal, with less and more than 50% stenosis in carotid artery respectively. Whereas minimum axial stress is −1.714±0.676, −1.982±0.663 and −2.593±0.661 (kPa) in normal, with less and more than 50% stenosis in carotid artery respectively. Moreover, internal diameter and intima-media thickness of the artery also increase significantly with stenosis initiation and its progression. In this study, the feasibility of axial wall stress computation for human common carotid arteries based on non-invasive in vivo clinical data is concluded. We found a strong and graded association between axial stress and severity of carotid stenosis, which might be used to discriminate healthy from atherosclerotic arteries.

Characterization of lipid parameters in diabetic and non-diabetic atherosclerotic patients.

Background & ObjectiveThe relationship between lipid profile perturbation and diabetes associated complications has long been an area of interest. Dyslipidemia is a potent predictor of cardiovascular morbidity and mortality in diabetic patients. The aim of present study was to investigate relationship between aging and lipid profiles in diabetic and non-diabetic atherosclerotic patients.MethodsFive hundred and seventy six individuals (45–75 year age) participated in this study. Among these, 192 were having history of diabetes mellitus and atherosclerosis. Individuals are categorized on the base of health (normal, non-diabetic atherosclerosis, diabetic atherosclerosis) and age (45–55 years, 56–65 years, and 66–75 years). All the participants were subjected to the procedures like a detailed history, biochemical analysis for fasting blood sugar, hemoglobin A1c, total cholesterol (TC), triglycerides (TG), low-density lipoprotein-(LDL), very low-density lipoprotein (VLDL) and high-density lipoprotein (HDL). All these parameters were compared between diabetic and non-diabetic atherosclerotic patients of all three age groups. TC/HDL and LDL/HDL were also calculated.ResultsDiabetic atherosclerotic individuals (both males and females) had high level of TC, TG, LDL, VLDL and low level of HDL in comparison to non-diabetic atherosclerotic and normal control individuals. Among all three age groups, lipoprotein abnormality was observed to be more frequent in females than males. There was a significant increase in TC/HDL and LDL/HDL ratio in diabetic atherosclerotic subjects compared to age and sex matched non-diabetic atherosclerotic and normal control.ConclusionsDegree of dyslipidemia increases with increase in age in both genders. Female are more prone to diabetic dyslipidemia and hence have more risk of developing atherosclerosis with increasing age.

Arterial stiffness, pulse pressure, and the kidney.

Classical studies indicate that the contribution of kidneys to hypertension is almost exclusively related to the association between mean arterial pressure (MAP) and vascular resistance. Recent reports including estimates of glomerular filtration rate (GFR) have shown that pulse pressure (PP) and pulse wave velocity, 2 major indices of arterial stiffness, now emerge as significant predictors of cardiovascular risk and age-associated decline in GFR. Such findings are mainly observed in patients with hypertension and renal failure and in atherosclerotic subjects undergoing coronary angiography. In such patients, amplification of PP between ascending and terminal aorta at the renal site is constantly increased over 10mm Hg (P < 0.001), whereas MAP level remains continuously unmodified. This PP amplification is significantly associated with presence of proteinuria. Furthermore, increases in plasma creatinine and aortic stiffness are independently and positively correlated (P < 0.001) both in cross-sectional and longitudinal studies. All these relationships associating PP, arterial stiffness, and renal function are mainly observed in patients 60 years of age or older. Furthermore, in renal transplant patients and their donors, subjects have been recruited for evaluations of arterial stiffness and posttransplant decline in GFR. Determinants of GFR decline were evaluated 1 and 9 years after transplantation. The first year GFR decline was related to smoking and acute rejection, whereas the later was significantly and exclusively associated with donor age and aortic stiffness. Thus, in hypertensive humans, the observed association between PP and GFR suggests that the 2 parameters are substantially mediated by arterial stiffness, not exclusively by vascular resistance.

Bioinformatics approach to evaluate differential gene expression of M1/M2 macrophage phenotypes and antioxidant genes in atherosclerosis.

Atherosclerosis is a pro-inflammatory process intrinsically related to systemic redox impairments. Macrophages play a major role on disease development. The specific involvement of classically activated, M1 (pro-inflammatory), or the alternatively activated, M2 (anti-inflammatory), on plaque formation and disease progression are still not established. Thus, based on meta-data analysis of public micro-array datasets, we compared differential gene expression levels of the human antioxidant genes (HAG) and M1/M2 genes between early and advanced human atherosclerotic plaques, and among peripheric macrophages (with or without foam cells induction by oxidized low density lipoprotein, oxLDL) from healthy and atherosclerotic subjects. Two independent datasets, GSE28829 and GSE9874, were selected from gene expression omnibus (http://www.ncbi.nlm.nih.gov/geo/) repository. Functional interactions were obtained with STRING (http://string-db.org/) and Medusa (http://coot.embl.de/medusa/). Statistical analysis was performed with ViaComplex(®) (http://lief.if.ufrgs.br/pub/biosoftwares/viacomplex/) and gene score enrichment analysis (http://www.broadinstitute.org/gsea/index.jsp). Bootstrap analysis demonstrated that the activity (expression) of HAG and M1 gene sets were significantly increased in advance compared to early atherosclerotic plaque. Increased expressions of HAG, M1, and M2 gene sets were found in peripheric macrophages from atherosclerotic subjects compared to peripheric macrophages from healthy subjects, while only M1 gene set was increased in foam cells from atherosclerotic subjects compared to foam cells from healthy subjects. However, M1 gene set was decreased in foam cells from healthy subjects compared to peripheric macrophages from healthy subjects, while no differences were found in foam cells from atherosclerotic subjects compared to peripheric macrophages from atherosclerotic subjects. Our data suggest that, different to cancer, in atherosclerosis there is no M1 or M2 polarization of macrophages. Actually, M1 and M2 phenotype are equally induced, what is an important aspect to better understand the disease progression, and can help to develop new therapeutic approaches.

Secreted proteins from carotid endarterectomy: an untargeted approach to disclose molecular clues of plaque progression

BackgroundAtherosclerosis is the main cause of morbidity and mortality in Western countries and carotid plaque rupture is associated to acute events and responsible of 15-20% of all ischemic strokes. Several proteomics approaches have been up to now used to elucidate the molecular mechanisms involved in plaque formation as well as to identify markers of pathology severity for early diagnosis or target of therapy. The aim of this study was to characterize the plaque secretome. The advantage of this approach is that secretome mimics the in vivo condition and implies a reduced complexity compared to the whole tissue proteomics allowing the detection of under-represented potential biomarkers.MethodsSecretomes from carotid endarterectomy specimens of 14 patients were analyzed by a liquid chromatography approach coupled with label free mass spectrometry. Differential expression of proteins released from plaques and from their downstream distal side segments were evaluated in each specimen. Results were validated by Western blot analysis and ELISA assays. Histology and immunohistochemistry were performed to characterize plaques and to localise the molecular factors highlighted by proteomics.ResultsA total of 463 proteins were identified and 31 proteins resulted differentially secreted from plaques and corresponding downstream segments. A clear-cut distinction in the distribution of cellular- and extracellular-derived proteins, evidently related to the higher cellularity of distal side segments, was observed along the longitudinal axis of carotid endarterectomy samples. The expressions of thrombospondin-1, vitamin D binding protein, and vinculin, as examples of extracellular and intracellular proteins, were immunohistologically compared between adjacent segments and validated by antibody assays. ELISA assays of plasma samples from 34 patients and 10 healthy volunteers confirmed a significantly higher concentration of thrombospondin-1 and vitamin D binding protein in atherosclerotic subjects.ConclusionsTaking advantage of the optimized workflow, a detailed protein profile related to carotid plaque secretome has been produced which may assist and improve biomarker discovery of molecular factors in blood. Distinctive signatures of proteins secreted by adjacent segments of carotid plaques were evidenced and they may help discriminating markers of plaque complication from those of plaque growth.