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Background: Considering the longevity of the worldwide population, the cardiovascular diseases deserve particular attention, especially the carotid artery disease in the ≥80-year-old population. The stiffness of the common carotid artery, for example, has been showed in numerous clinical studies as a marker of increased risk of stroke, dementia, and depression. Besides, with the emergence of new surgical techniques such as the transcarotid artery revascularization that uses the common carotid artery as a workstation, the biomechanical and histological features of this vessel, more than ever, must be detailed., Methods: Left common carotid artery fragments from 9 cadaver donors (≥80 years old) were evaluated. Biomechanical (failure stress, tension, and strain) and histological (percentage of collagen and elastic fibers) features of these samples were analyzed with special focus on gender differences., Results: Statistically significant differences in biomechanical and histological features between the genders were observed. The percentage of collagen fiber in intima (P = 0.008) and media (P = 0.041) layers was significantly lower in men than in women. A higher elasticity (failure strain) of the specimens in male gender was also observed (P = 0.025). No significant difference was observed in the layers thickness between the genders regardless which part of the arterial wall was considered., Conclusions: These biomechanical and histological findings could be the responsible for the higher left common carotid artery stiffness observed among ≥80-year-old women when compared with men in numerous clinical studies in literature., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The polyherbal blend Zyflamend™ has been shown to have anti-inflammatory properties and attenuate inflammatory-modulated pathologies. Fish oils have also been shown to have cardioprotective properties. However, the beneficial effects of their combination have not been investigated. Intimal hyperplasia (IH), a pathological remodeling response of a vessel to injury, is heavily regulated by an immune-mediated reaction. The objective of this study was to determine if dietary supplementation with Zyflamend and/or Wholemega could affect inflammatory-dependent vascular remodeling mechanisms when provided at human equivalent doses. Based on their anti-inflammatory properties and protective benefits demonstrated in previous pre-clinical studies, we hypothesized administration of these supplements would prevent IH in an animal model of vascular injury. The diets of aged male rats were supplemented with human equivalent doses of Zyflamend (Zyf) and/or Wholemega (WMega) or placebo (Plac) for 1wk prior to balloon angioplasty (BA)-induced injury of the left carotid artery. At 28d post-injury morphometric analysis of carotid tissue revealed IH was decreased in Zyf + WMega animals compared to placebo, while Zyf or WMega independently had no significant effect. Serum cytokine screening indicated injury-induced interleukin family isoforms, interferon-γ, and macrophage inflammatory proteins were downregulated by Zyf + WMega. Immunohistochemical staining for monocyte/macrophage phenotypic markers revealed that while overall monocyte/macrophage vessel infiltration was not affected, Zyf + WMega limited the alternative differentiation of M2 macrophages and reduced the presence of myofibroblasts in the injured vessel wall. In summary, dietary supplementation with Zyf + WMega attenuated the acute inflammatory response following vascular injury and inhibited IH development in vivo., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Objectives: The aim of this study was to investigate whether and what carotid plaque characteristics predict systemic cardiovascular outcomes in patients with clinically established atherosclerotic disease., Background: Advancements in atherosclerosis imaging have allowed assessment of various plaque characteristics, some of which are more directly linked to the pathogenesis of acute cardiovascular events compared to plaque burden., Methods: As part of the event-driven clinical trial AIM-HIGH (Atherothrombosis Intervention in Metabolic Syndrome with Low HDL/High Triglycerides: Impact on Global Health Outcomes), subjects with clinically established atherosclerotic disease underwent multicontrast carotid magnetic resonance imaging (MRI) to detect plaque tissue composition and high-risk features. Prospective associations between MRI measurements and the AIM-HIGH primary endpoint (fatal and nonfatal myocardial infarction, ischemic stroke, hospitalization for acute coronary syndrome, and symptom-driven revascularization) were analyzed using Cox proportional hazards survival models., Results: Of the 232 subjects recruited, 214 (92.2%) with diagnostic image quality constituted the study population (82% male, mean age 61 ± 9 years, 94% statin use). During median follow-up of 35.1 months, 18 subjects (8.4%) reached the AIM-HIGH endpoint. High lipid content (hazard ratio [HR] per 1 SD increase in percent lipid core volume: 1.57; p = 0.002) and thin/ruptured fibrous cap (HR: 4.31; p = 0.003) in carotid plaques were strongly associated with the AIM-HIGH endpoint. Intraplaque hemorrhage had a low prevalence (8%) and was marginally associated with the AIM-HIGH endpoint (HR: 3.00; p = 0.053). High calcification content (HR per 1 SD increase in percent calcification volume: 0.66; p = 0.20), plaque burden metrics, and clinical risk factors were not significantly associated with the AIM-HIGH endpoint. The associations between carotid plaque characteristics and the AIM-HIGH endpoint changed little after adjusting for clinical risk factors, plaque burden, or AIM-HIGH randomized treatment assignment., Conclusions: Among patients with clinically established atherosclerotic disease, carotid plaque lipid content and fibrous cap status were strongly associated with systemic cardiovascular outcomes. Markers of carotid plaque vulnerability may serve as novel surrogate markers for systemic atherothrombotic risk., (Copyright © 2017 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.)

Background: Carotid artery geometry has been suggested as a risk factor for atherosclerotic carotid artery disease (ACD). Although normal aging and development of disease can both lead to geometric changes in the artery, whether geometric changes in a given artery actually predispose to disease or are just a consequence of remodeling during aging is unclear. We investigated carotid artery geometric changes with aging to identify geometric features associated with the presence of ACD., Methods: Carotid artery geometry was quantified by measuring carotid artery diameter, tortuosity, and bifurcation angle using three-dimensional reconstructions of thin-section computed tomography angiography scans in 15 healthy individuals (average age, 43 ± 18 years; range, 15-64 years). The same geometric features were measured in 17 patients (68 ± 10 years old) with unilateral ACD. Geometric features associated with presence of ACD were determined by using the nondiseased contralateral carotid artery as an intrinsic control. Elastin-stained carotid arteries were analyzed to assess age-related structural changes in 12 deceased individuals., Results: Increases were noted in bulb diameter (0.64 mm), bifurcation angle (10°), and tortuosity of the common carotid (CCA; 0.03) and internal carotid arteries (ICA; 0.04) for every decade of life. Density and continuity of circumferential and longitudinal elastin in the CCA and ICA decreased with age. Compared with normal carotid arteries, those with ACD demonstrated larger bulb diameters (P = .001) but smaller bifurcation angles (P = .001). CCA tortuosity (P = .038) increased in ACD arteries compared with normal carotid arteries, but ICA tortuosity was decreased (P = .026)., Conclusions: With increasing age, bulb diameter, tortuosity, and bifurcation angle increases in carotid arteries. These geometric changes may be related to degradation and fragmentation of intramural elastin. Arteries with atherosclerotic occlusive disease demonstrate decreased ICA tortuosity and smaller bifurcation angles compared with nondiseased carotid arteries., (Copyright © 2015 Society for Vascular Surgery. Published by Elsevier Inc. All rights reserved.)

Highly diluted solutions of Gentian Violet and Evans Blue were used to visualize the elastin network in viable porcine right common carotid artery (RCCA) preparations. The two simple, alternative methods of staining were applied to proximal, intermediate, and distal sections of RCCA under various experimental conditions. These included the state of the vessel wall soon after excision, under relaxed smooth muscle condition after preconditioning, and during vasoconstriction. Micrographs of arterial rings, sectors, and axial strips show that the RCCA is an artery of the elastic type at the proximal end and of the muscular type at the distal end. While in sections of freshly dissected or KCl-constricted arteries the elastic lamellae show the well-known waviness, those in sections from arteries with relaxed smooth muscle and after preconditioning appear nearly straight. It is hoped that the inexpensive staining tools will contribute to solve conflicting interpretations existing on elastin structures in the arterial wall.

Objective: To establish rabbit model of restenosis after carotid endarterectomy surgery, and to study tissue inflammatory cytokines (TNF-α, IL-6) involved in restenosis., Methods: A total of 32 rabbits were randomly divided into two groups: model group and control group. The right common carotid artery in rabbits was damaged by carotid endar terectomy in model group. The tissues were harvested at different time points respectively, the pathological changes of the vascular wall after operation were observed at different time points. The changes of expression of tissue vascular wall inflammatory cytokines (TNF-α, IL-6) at different time points after the surgery was observed by RT-PCR, and the changes of serum inflammatory cytokines (TNF-α, IL -6) were detected by ELISA., Results: The new intima appeared after 7 days of the injury and reached the peak on 28 d which is uneven and significantly thicker than the control group (P<0.01). The tissue inflammatory cytokines (TNF-α, IL-6) were significantly increased after the rabbit common carotid artery injury, which was significant difference compared with normal control group (P<0.05)., Conclusions: The tissue inflammatory factors significantly increase after the rabbit carotid artery injury, which suggests the mutual concurrent effects of inflammatory cytokines can result in the proliferation of vascular restenosis., (Copyright © 2014 Hainan Medical College. Published by Elsevier B.V. All rights reserved.)

Background: Restenosis of the carotid artery is common following carotid endarterectomy, but analysis of lesion composition has mostly been based on histological study of explanted restenotic lesions. This study investigated the ability of 3T cardiovascular magnetic resonance (CMR) to determine the components of recurrent carotid artery disease and examined whether these differed from primary atherosclerotic plaque., Methods: 50 patients underwent 3T CMR of both carotid arteries using a standard multicontrast protocol: time-of-flight (TOF), T1-weighted (T1W), T2-weighted (T2W), and PD-weighted (PDW) Turbo-Spin-Echo (TSE) sequences. 25 patients had previously undergone carotid endarterectomy (mean time since surgery 1580 days, range 45-6560 days), and 25 with primary asymptomatic atherosclerotic plaques served as controls. Two experienced reviewers analysed the multicontrast CMR images according to the presence or absence of major plaque features and assigned an overall classification type., Results: In patients with recurrent carotid disease following endarterectomy, the mean degree of restenosis was 51% (range 30-90%). Three distinct types of restenosis were identified: 5 patients (20%) showed CMR characteristics of fibro-atheromatous tissue, 11 patients (44%) had plaque features consistent with possible myointimal (fibromuscular) hyperplasia, and 6 patients (24%) had recurrent plaque suggestive of further lipid accumulation. Three patients (12%) showed evidence of post-surgical dissection of the carotid intima. Compared to primary atherosclerotic plaques, restenotic plaques were more likely to contain fibro-atheromatous tissue (p = 0.05) and smooth muscle (p < 0.01), and less likely to contain lipid (p < 0.01). Composition did not differ significantly between patients with early and late restenosis., Conclusions: As defined by CMR, restenotic lesions of the carotid artery fall into three distinct types and differ in composition from primary atherosclerotic plaques. If validated by subsequent histological studies, these findings could suggest a role for CMR in detecting high-risk (i.e. lipid-rich) restenotic lesions.

Chronic inflammation that occurs with aging is one of the risk factors for cardiovascular disease. Regular exercise may prevent cardiovascular morbidity by decreasing chronic systematic inflammation. Additionally, excess inflammation can be reduced by the anti-inflammatory protein pentraxin 3 (PTX3). Thus, both habitual exercise and PTX3 have an anti-inflammatory effect. However, it is unclear whether regular exercise leads to increased plasma PTX3 concentration. In the present study, we investigated the effects of regular aerobic exercise on plasma PTX3 concentration in middle-aged and elderly women. Twenty-two postmenopausal women (60 ± 6 years) were randomly divided evenly into 2 groups (i.e., exercise intervention and control). Subjects in the exercise group completed 2 months of regular aerobic exercise training (walking and cycling, 30-45 min, 3-5 days·week⁻¹). Before and after the intervention, we evaluated plasma PTX3 concentration, peak oxygen uptake, blood chemistry, and arterial distensibility (carotid arterial compliance and β-stiffness) in all participants. There were no significant differences in baseline parameters between the 2 groups. Plasma PTX3 concentration was significantly increased in the exercise group after the intervention (p < 0.05). High-density lipoprotein cholesterol, peak oxygen uptake, and arterial compliance were also significantly increased (p < 0.05), while β-stiffness was markedly decreased (p < 0.01) after the intervention. On the other hand, there was no change in the parameters tested in the control group. This study demonstrates that regular aerobic exercise increases plasma PTX3 concentration with improvement of high-density lipoprotein cholesterol, peak oxygen uptake, and arterial distensibility in postmenopausal women.

Diabetes is associated with increased risk of cardiovascular disease. Advanced glycation end-products (AGEs) are considered to be a major pathogenic factor for diabetic vascular complications. The levels of AGEs are increased in diabetic patients. We have studied the presence of the major AGE methylglyoxal (MGO)-derived hydroimidazolone in human aorta and carotid arteries, using immunohistochemistry (IHC), western blotting and mass spectrometry. By IHC, MGO-derived modifications were detected mainly associated with cells in intimal thickenings and cells in microvessels in adventitia. In type V lesions MGO-derived AGE was also present, extracellular in the necrotic core and in cells at the border of the core. The highest degree of modification was probably associated with cell nuclei. By western blotting and mass spectrometry fibrin(ogen), the cytoskeleton-associated protein moesin and the nuclear proteins lamin A and C were identified as putative main targets for MGO-derived modification. LC-MS/MS studies of fibrin(ogen) modified in vitro with low concentrations of MGO identified the sites that were most prone to modification. These results indicate that AGE modifications occur preferentially on specific proteins. The modification of these proteins may play a role in vascular dysfunction and development of atherosclerosis in diabetes.

The selection of a mathematical descriptor for the passive arterial mechanical behavior has been long debated in the literature and customarily constrained by lack of pertinent data on the underlying microstructure. Our objective was to analyze the response of carotid artery subjected to inflation/extension with phenomenological and microstructure-based candidate strain-energy functions (SEFs), according to species (rabbit vs. pig) and region (proximal vs. distal). Histological variations among segments were examined, aiming to explicitly relate them with the differential material response. The Fung-type model could not capture the biphasic response alone. Combining a neo-Hookean with a two-fiber family term alleviated this restraint, but force data were poorly captured, while consideration of low-stress anisotropy via a quadratic term allowed improved simulation of both pressure and force data. The best fitting was achieved with the quadratic and Fung-type or four-fiber family SEF. The latter simulated more closely than the two-fiber family the high-stress response, being structurally justified for all artery types, whereas the quadratic term was justified for transitional and muscular arteries exhibiting notable elastin anisotropy. Diagonally arranged fibers were associated with pericellular medial collagen, and circumferentially and longitudinally arranged fibers with medial and adventitial collagen bundles, evidenced by the significant correlations of SEF parameters with quantitative histology.

The mechanisms by which dietary sodium modulates cardiovascular risk are not fully understood. This study investigated whether sodium intake is related to carotid structure and hemodynamics and to plasma matrix metalloproteinase (MMP) activity in hypertensive adults. One hundred thirty-four participants were cross-sectionally evaluated by clinical history, anthropometry, carotid ultrasound, and analysis of hemodynamic, inflammatory, and metabolic variables. Daily sodium intake (DSI) was estimated by 24-h recall, discretionary sodium, and a FFQ. In 42 patients, plasma MMP-2 and MMP-9 activities were also analyzed. The mean DSI was 5.52 ± 0.29 g/d. Univariate analysis showed that DSI correlated with common carotid artery systolic and diastolic diameter (r = 0.36 and 0.34; both P < 0.001), peak and mean circumferential tension (r = 0.44 and 0.39; both P < 0.001), Young's Elastic Modulus (r = 0.40; P < 0.001), intima-media thickness (r = 0.19; P < 0.05), and internal carotid artery resistive index (r = 0.20; P < 0.05). Multivariate analyses revealed that only artery diameter, circumferential wall tension, and Young's Elastic Modulus were independently associated with DSI. Conversely, plasma MMP-9 activity was associated with DSI (r = 0.53; P < 0.001) as well as with common carotid systolic diameter (r = 0.33; P < 0.05) and Young's Elastic Modulus (r = 0.38; P < 0.01). In conclusion, sodium intake is associated with carotid alterations in hypertensive adults independently of systemic hemodynamic variables. The present findings also suggest that increased MMP-9 activity might play a role in sodium-induced vascular remodeling.

Limited information is available that examines the interrelationships between glucose tolerance, serum albumin, subclinical inflammation, and carotid atherosclerosis (CA) in patients with spinal cord injury (SCI). We prospectively recruited 110 male patients with traumatic SCI, 57 with SCI at thoracic level 6 or above (SCI-T6) and 53 with SCI at T7 or below (SCI-T7), and 62 age-matched able-bodied controls from the National Taiwan University Hospital. The associations among glucose levels after oral glucose tolerance tests (OGTT), serum albumin, high-sensitivity C-reactive protein (hs-CRP), and CA in terms of the extracranial carotid artery (ECCA) plaque score and common carotid artery (CCA) intima-media thickness (IMT) were examined. Results showed significantly higher post-challenge glucose levels and carotid plaque scores and lower serum albumin in the SCI-T6 patients. In addition, serum albumin was negatively associated with CA and post-challenge glucose levels. The higher post-challenge glucose levels at 120min (Glu120) were associated with higher serum hs-CRP levels and lower serum albumin levels. In addition, lower serum albumin levels were associated with a thicker CCA IMT and a higher prevalence of ECCA plaque. Mixed models revealed that body mass index, age, LDL-cholesterol, Glu120, homeostasis model assessment for insulin resistance (HOMA-IR), lower serum albumin and smoking habits were positively associated with CCA IMT. Age, HOMA-IR, LDL-cholesterol, and lower serum albumin were identified as the important factors for the presence of carotid plaque by multiple linear regression analyses. In conclusion, post-challenge hyperglycemia and serum albumin levels are important indicators of CV health in men with SCI.

Objective: Plaque with dense inflammatory cells, including macrophages, thin fibrous cap and superficial necrotic/lipid core is thought to be prone-to-rupture. We report a time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) technique for detection of such markers of plaque vulnerability in human plaques., Methods: The autofluorescence of carotid plaques (65 endarterectomy patients) induced by a pulsed laser (337 nm, 0.7 ns) was measured from 831 distinct areas. The emission was resolved spectrally (360-550 nm range) and temporally (0.3 ns resolution) using a prototype fiber-optic TR-LIFS apparatus. Lesions were evaluated microscopically and quantified as to the % of different components (fibrous cap, necrotic core, inflammatory cells, foam cells, mature and degraded collagen, elastic fibers, calcification, and smooth muscle cell of the vessel wall)., Results: We determined that the spectral intensities and time-dependent parameters at discrete emission wavelengths (1) allow for discrimination (sensitivity >81%, specificity >94%) of various compositional and pathological features associated with plaque vulnerability including infiltration of macrophages into intima and necrotic/lipid core under a thin fibrous cap, and (2) show a linear correlation with plaque biochemical content: elastin (P<0.008), collagen (P<0.02), inflammatory cells (P<0.003), necrosis (P<0.004)., Conclusion: Our results demonstrate the feasibility of TR-LIFS as a method for the identification of markers of plaque vulnerability. Current findings enable future development of TR-LIFS-based clinical devices for rapid investigation of atherosclerotic plaques and detection of those at high-risk.

Background and Purpose: Echogenicity of carotid plaque well reflects the risk of ischemic stroke and may be predictive of the histologic content of the plaque. However, objective evaluation of plaque echogenicity has been hampered by a lack of established quantitative measures. This study examined the relation between echogenicity assessed by integrated backscatter (IBS) analysis and (1) symptomatic history and (2) histologic features of carotid plaques., Methods: We used acoustic densitometry to quantify by IBS analysis the echogenicity of 31 carotid plaques of 26 patients undergoing carotid endarterectomy or stenting. IBS was subsequently compared with histologic findings of the respective tissue in 10 patients who underwent endarterectomy. The IBS value was calibrated with 2 reference structures (vessel lumen and adventitia) as the IBS index., Results: The IBS index of symptomatic plaques was lower than that of asymptomatic plaques (23.1 +/- 12.5 vs. 36.5 +/- 18.2, p < 0.05). The IBS index in fatty/necrotic atheromatous sites (n = 20, 16.6 +/- 10.7) was lower than that in fibrous (n = 26, 42.4 +/- 13.6, p < 0.01) or calcified (n = 11, 87.7 +/- 17.4, p < 0.01) sites and the same as that in intraplaque hemorrhagic sites (n = 50, 23.6 +/- 16.9)., Conclusions: Carotid plaque echogenicity, as quantitatively assessed by IBS analysis, correlates well with the presence or absence of prior symptoms and histologic contents of the plaques. IBS analysis may aid in the assessment of carotid plaque-related risk of stroke., (2008 S. Karger AG, Basel.)

Increased common carotid artery (CCA) intima-media thickness (IMT) measured by B-mode ultrasound is an early marker of the atherosclerotic process. Arterial calcification is not clearly understood. Using the particle-induced X-ray emission (PIXE) method, we have looked for the location in the artery wall where calcium accumulated in the early phase of atherosclerosis. Twelve segments of CCAs of deceased stroke patients were investigated. In-vivo, carotid duplex ultrasound was performed with bilateral CCA IMT measurement at plaque-free sections. During autopsy, segments of carotid arteries were collected and filled under pressure with a stained histologic embedding material. The frozen arteries were cut into 60-microm-thick slices. Calcium distribution maps from the segments of arteries were determined by PIXE method. IMT measured by ultrasound and calcium distribution maps measured by PIXE were compared. In our cross-sectional study, using the PIXE analysis and ultrasound images, we could demonstrate early calcium accumulation in the media layer. Our results have also shown a significant relationship between calcium content of distributional maps measured by PIXE analysis and corresponding IMT on B-mode ultrasound images of human CCAs.

Background: Rupture of vulnerable atheromatous plaque in the carotid and coronary arteries often leads to stroke and heart attack respectively. The role of calcium deposition and its contribution to plaque stability is controversial. This study uses both an idealized and a patient-specific model to evaluate the effect of a calcium deposit on the stress distribution within an atheromatous plaque., Methods: Using a finite-element method, structural analysis was performed on an idealized plaque model and the location of a calcium deposit within it was varied. In addition to the idealized model, in vivo high-resolution MR imaging was performed on 3 patients with carotid atheroma and stress distributions were generated. The individual plaques were chosen as they had calcium at varying locations with respect to the lumen and the fibrous cap., Results: The predicted maximum stress was increased by 47.5% when the calcium deposit was located in the thin fibrous cap in the model when compared with that in a model without a deposit. The result of adding a calcium deposit either to the lipid core or remote from the lumen resulted in almost no increase in maximal stress., Conclusion: Calcification at the thin fibrous cap may result in high stress concentrations, ultimately increasing the risk of plaque rupture. Assessing the location of calcification may, in the future, aid in the risk stratification of patients with carotid stenosis., ((c) 2007 S. Karger AG, Basel)

To explore the biological effects of shear stress on intact artery and the change of growth factor during stress-induced vascular remodeling, we established an artery organ-cultured system under stress in vitro, and the common carotid arteries of pigs were cultured under shear stress of 20, 5 and 0 dyn/cm2. PDGF-A synthesis of vascular smooth muscle cells (VSMCs) cultured for 1, 4 and 7 days were studied by immunohistochemical and computer image processing methods, and PDGF-B secretion of endothelial cells (ECs) cultured within 12 h were studied by ELISA. Results showed that PDGF-B increased obviously under shear stress of 5 dyn/cm2, and reached the highest point at about 3 h; PDGF-A synthesis also obviously increased under low shear stress in 7 days. Increasing of PDGF synthesis promotes phenotype switch and proliferation of VSMC. It may have important influence on artery remodeling under low shear stress.

Objective: We investigated the potential role of ADAMTS-1 (a disintegrin and metalloprotease with thrombospondin motif type I) in atherogenesis., Methods and Results: ADAMTS-1 is expressed at the highest levels in the aorta when compared with other human tissues examined. Immunolocalization studies in human aorta and coronary artery indicate that ADAMTS-1 expression is mainly seen at low levels in the medial layer, but upregulated in the intima when plaque is present. We found that ADAMTS-1 mRNA levels are significantly higher in proliferating/migrating cultured primary aortic vascular smooth muscle cells (VSMCs) compared with resting/confluent cells. Using the mouse carotid artery flow cessation model, we show that there are differences in vessel remodeling in ADAMTS-1 transgenic/apoE-deficient mice compared with apoE deficiency alone, particularly a significant increase in intimal hyperplasia. We show that ADAMTS-1 can cleave the large versican containing proteoglycan population purified from cultured human aortic VSMCs. Finally, using versican peptide substrates, we show data suggesting that ADAMTS-1 cleaves versican at multiple sites., Conclusions: We hypothesize that ADAMTS-1 may promote atherogenesis by cleaving extracellular matrix proteins such as versican and promoting VSMC migration.

Monocyte chemoattractant protein-1 (also known as CC chemokine ligand 2 [CCL2]) and its receptor CC chemokine receptor 2 (CCR2) play a central role in the inflammatory response and neointimal formation after vascular injury. In the context of hyperlipidemia, this appears to involve neointimal monocyte infiltration. Hence, we investigated the function of the CCL2/CCR2 axis in early monocyte recruitment to injured arteries. Wire-induced injury of the carotid artery in apoE-/- mice caused a rapid increase of JE/CCL2 protein in the vessel wall peaking at 24 hours after injury, whereas serum JE/CCL2 was increased solely at 6 hours and blood cell-associated levels were unaltered, as demonstrated by enzyme-linked immunosorbent assay. Immunohistochemistry revealed intense staining for JE/CCL2 in smooth muscle cells (SMCs) and in association with platelets adherent to the denuded vessel wall 24 hours after injury. In vitro, exogenous or SMC-derived JE/CCL2 binds to the platelet surface and triggers monocyte arrest on adherent platelets but not on SMCs in flow assays. Accordingly, monocyte arrest in ex vivo perfused apoE-/- carotid arteries isolated 24 hours after injury was profoundly inhibited by pretreatment with a JE/CCL2 antibody. In CCR2-/-/apoE-/- mice, neointimal plaque area was reduced by 47% compared with CCR2+/+/apoE-/- mice. Moreover, CCR2 deletion markedly decreased neointimal macrophage content while expanding SMC content. Vascular JE/CCL2 expressed by SMCs and immobilized by adherent platelets after endothelial denudation is crucial for mediating early monocyte recruitment to injured arteries in hyperlipidemic mice. This mechanism may explain reduced neointimal macrophage infiltration and lesion formation in CCR2-deficient apoE-/- mice.

To test the hypothesis that concentration polarization of atherogenic lipids may occur in the arterial system and play an important role in the localization of atherogenesis, we measured in vitro the luminal surface concentration of bovine serum albumin (as a tracer macromolecule) in the canine carotid artery by directly taking liquid samples from the luminal surface of the artery. The experimental results show that the luminal surface albumin concentration, c(w), was higher than the bulk concentration, c(0) as predicted by our theory. The relative luminal surface albumin concentration, c(w)/c(0), decreased very sharply at low wall shear rate, G, but gradually approached the value of 1.0 asymptotically as G was increased. The experiment shows that water flux rate across the vessel wall, v(w), has a profound impact on concentration polarization. For instance, at G = 0 and 185 s(-1), when v(w) = 8.9 +/- 1.7 x 10(-6) cm/s, c(w) was 65% and 15% higher than c(0), respectively, meanwhile when v(w) = 4.8 +/- 0.6 x 10(-6)cm/s, c(w) was only 42% and 5% higher than c(0), respectively. The experiment also revealed that concentration polarization occurred in a thin layer close to the luminal surface of the artery. The thickness of this layer was water flux rate-dependent. The higher the water flux rate, the thicker was the layer. The present study therefore confirms that concentration polarization can indeed occur in the arterial system and our theoretical analysis is accurate in predicting this mass transfer phenomenon., (Copyright 2002 Elsevier Science Ltd.)

Unlabelled: OBJECTIVES; Living human carotid atherosclerotic plaques were examined in vitro by near-infrared (NIR) spectroscopy to determine the spectral features of plaque vulnerability., Background: Plaque disruption, a major cause of heart attacks and strokes, cannot generally be predicted, but is thought to depend mainly on plaque composition. Near-infrared spectroscopy has been used to detect components in tissues noninvasively., Methods: Using an NIR spectrometer fitted with a fiberoptic probe, living human carotid atherosclerotic plaques (from 25 patients) were examined ex vivo for plaque vulnerability. The plaques were cut into smaller sections according to their gross pathologic features, and NIR measurements were done at 20 degrees C, usually within 10 min., Results: According to the American Heart Association's recommended classification scheme, the lesions were classified into three groups: the first group comprised of vulnerable type V/VI lesions; the second group, stable type I/II lesions; and the third (intermediate) group, mainly type III/IV lesions. Cluster analysis of the specimens' NIR spectra identified three major composition groups in each of the three NIR spectral regions: 2,200 to 2,330 nm, 1,620 to 1,820 nm and 1,130 to 1,260 nm. Calculation of the lipid/protein ratios in each group at two NIR regions (2,200 to 2,330 nm) revealed ratios of 1.49 +/- 1.20, 2.12 +/- 1.00 and 3.37 +/- 0.88 for type I/II, type III/IV and advanced type V/VI lesions, respectively (p < 0.03). At 1,620 to 1,820 nm, the respective ratios for these histologic groups were 0.57 +/- 0.21, 1.54 +/- 0.46 and 2.40 +/- 0.44 (p < 0.00003)., Conclusions: The good ex vivo discrimination of histologically vulnerable and stable plaques in this study suggests that NIR spectroscopy has the potential to identify vulnerable atherosclerotic plaques in vivo.

Unlabelled: objectives; The aim of this study was to determine whether in patients with congestive heart failure (CHF) a distensibility (Dist) reduction: 1) similarly occurs in different arteries; 2) is related to CHF severity; and 3) is reversible with treatment. background: Several studies suggest that CHF is accompanied by a reduced arterial Dist., Methods: We measured diameter in radial artery, carotid artery (CA) and abdominal aorta (AO) by echotracking. Distensibility was obtained by relating it to blood pressure. Data were collected in 30 patients with CHF (New York Heart Association functional class I to III) under standard treatment with diuretic, digitalis and angiotensin-converting enzyme (ACE) inhibitor in whom CHF severity was assessed by maximum oxygen consumption (VO(2)max) percentage and in 30 age- and gender-matched controls. Patients with CHF were then randomized to maintain standard treatment (n = 10), double the ACE inhibitor dose (n = 10) or add an angiotensin II antagonist (n = 10) and restudied after two months., Results: Distensibility was markedly reduced in the CHF group in all three vessels (p < 0.01), CA and AO Dist being related to CHF severity (p < 0.05). After two months, Dist did not change in the group maintained under standard treatment, but it increased significantly (p < 0.05) and similarly when the ACE inhibitor dose was doubled or an angiotensin II antagonist was added., Conclusions: Congestive heart failure is characterized by a reduction of Dist of large-elastic and middle-sized muscular arteries. The reduction of large-elastic artery Dist is related to the CHF severity. These alterations can be reversed by drugs, effectively interfering with the renin-angiotensin system either at the ACE or at the angiotensin receptor level.

Background: C-reactive protein (CRP) has been linked to cardiovascular disease and atherosclerosis. Large-scale epidemiological studies have shown a correlation of CRP level with risk of stroke, myocardial infarction and peripheral arterial disease. Nevertheless, the question whether serum CRP itself is an independent indicator of the atherosclerotic process remains unanswered., Methods: In a community-based sample free of advanced atherosclerotic disease (n = 1018; mean age +/- SD, 54.1 +/- 12.0 years; 49.7% women) we examined the relationship between carotid intimal medial thickness (IMT), conventional vascular risk factors (that is, smoking, obesity, elevated blood pressure, diabetes mellitus, hypercholesterolaemia) and serum CRP., Results: We found an association between increasing IMT values with increasing CRP values for all sites within the carotid system (for example, common carotid artery [CCA-] IMT, beta = 0.174, P < 0.001). The relationship was weakened after accounting for the above-mentioned conventional risk factors (linear regression), particularly body mass index, but remained significant (for example, mean CCA-IMT beta = 0.02, P = 0.042). Including fibrinogen in the regression made the relationship no longer significant (mean CCA-IMT beta = 0.01, P = 0.277)., Conclusion: It is unlikely that CRP per se is a major independent cause of early arteriosclerosis. Elevations of CRP, or less specifically chronic inflammation, may mediate the effect of certain conventional risk factors on promoting atherogenesis, especially obesity.

Dissolution of the fibrin blood clot is regulated in large part by plasminogen activator inhibitor-1 (PAI-1). Elevated levels of plasma PAI-1 may be an important risk factor for atherosclerotic vascular disease and are associated with premature myocardial infarction. The role of the endogenous plasminogen activation system in limiting thrombus formation following atherosclerotic plaque disruption is unknown. This study found that genetic deficiency for PAI-1, the primary physiologic regulator of tissue-type plasminogen activator (tPA), prolonged the time to occlusive thrombosis following photochemical injury to carotid atherosclerotic plaque in apolipoprotein E-deficient (apoE(-/-)) mice. However, anatomic analysis revealed a striking difference in the extent of atherosclerosis at the carotid artery bifurcation between apoE(-/-) mice and mice doubly deficient for apoE and PAI-1 (PAI-1(-/-)/apoE(-/-)). Consistent with a previous report, PAI-1(+/+)/apoE(-/-)and PAI-1(-/-)/apoE(-/-) mice developed similar atherosclerosis in the aortic arch. The marked protection from atherosclerosis progression at the carotid bifurcation conferred by PAI-1 deficiency suggests a critical role for PAI-1 in the pathogenesis of atherosclerosis at sites of turbulent flow, potentially through the inhibition of fibrin clearance. Consistent with this hypothesis, intense fibrinogen/fibrin staining was observed in atherosclerotic lesions at the carotid bifurcation compared to the aortic arch. These observations identify significant differences in the pathogenesis of atherosclerosis at varying sites in the vascular tree and suggest a previously unappreciated role for the plasminogen activation system in atherosclerosis progression at sites of turbulent flow. (Blood. 2000;96:4212-4215)

Purpose: This study investigated the distribution of collagen III and IV in experimental microvascular anastomoses and correlated the findings with the quality of the healing process., Materials and Methods: A previously excised and reanastomosed section of the left common carotid artery of 21 female Wistar rats were explanted 4 weeks after surgery. Collagens III and IV were identified in perfusion-fixed specimens by immunohistochemistry., Results: The identification and distribution of collagens reflected the quality of each suturing procedure. A myointimal hyperplasia was present, extending into the vascular lumen, and was closely related to the ends of the vessel segments. The extent of myointimal hyperplasia was dependent on the length and the vitality of the vessel segments. The media was the weakest part of the tissue layers with regard to regenerative potential in the axial direction. The marked cuff of the adventitia around the anastomosis was rich in collagen III-positive vasa vasorum. Collagen IV was distributed in net-like patterns in the media. The distribution pattern in the media was not observed in anastomoses with incomplete approximation and necrotic segment ends. The collagen IV-positive subendothelial basal membrane was incomplete in the aneurysms because of partial necrosis., Conclusions: The experimental findings support the need for atraumatic suturing in microvascular surgery. Immunohistochemical detection of collagens can be a valuable tool for studying the basic processes of wound healing and the integration of microvascular flaps into their recipient sites.

Because of renewed interest in the progression, stabilization, and regression of atherosclerotic plaques, it has become important to develop methods for characterizing structural features of plaques in situ and noninvasively. We present a nondestructive method for ex vivo quantification of 2 solid-phase components of plaques: crystalline cholesterol and calcium phosphate salts. Magic angle spinning (MAS) nuclear magnetic resonance (NMR) spectra of human carotid endarterectomy plaques revealed (13)C resonances of crystalline cholesterol monohydrate and a (31)P resonance of calcium phosphate hydroxyapatite (CPH). The spectra were obtained under conditions in which there was little or no interference from other chemical components and were suitable for quantification in situ of the crystalline cholesterol and CPH. Carotid atherosclerotic plaques showed a wide variation in their crystalline cholesterol content. The calculated molar ratio of liquid-crystalline cholesterol to phospholipid ranged from 1.1 to 1.7, demonstrating different capabilities of the phospholipids to reduce crystallization of cholesterol. The spectral properties of the phosphate groups in CPH in carotid plaques were identical to those of CPH in bone. (31)P MAS NMR is a simple, rapid method for quantification of calcium phosphate salts in tissue without extraction and time-consuming chemical analysis. Crystalline phases in intact atherosclerotic plaques (ex vivo) can be quantified accurately by solid-state (13)C and (31)P MAS NMR spectroscopy.

We have previously demonstrated that high-flow (HF) conditions inhibit experimental intimal hyperplasia. We hypothesized that such flow conditions may alter transforming growth factor-beta1 (TGF-beta1) after mural injury. The right common carotid artery (CCA) was balloon-injured in 54 New Zealand White male rabbits. Flow was thereafter preserved (normal flow [NF]), reduced by partial outflow occlusion (low flow [LF]), or increased by ligation of the left CCA (HF). Four sham-operated animals served as uninjured controls. Mean blood flow and pressure in the right CCA were measured before and after flow modulation and before euthanasia (3, 7, and 14 days). TGF-beta1 mRNA and protein levels in the right CCA were determined by Northern and ELISA analyses at each time point. At 7 and 14 days, intimal hyperplasia was quantified, and the transmural localization of TGF-beta1 was determined by immunohistochemical analysis. Mean flow was reduced from 22+/-1 to 10+/-3 mL/min in the LF group and increased to 34+/-2 mL/min in the HF group (P<0.001). Blood pressure was not different among the flow groups for all time points. Wall shear stress was markedly decreased in the LF group to 14+/-4 dyne/cm(2) and increased in the HF group to 63+/-6 dyne/cm(2) at 7 days compared with values in uninjured controls (39+/-2 dyne/cm(2), P<0.001) and the NF group (44+/-7 dyne/cm(2), P<0.001). At 14 days, wall shear stress was similar among the flow groups. The intima-to-media ratio was 5- and 2-fold greater in the LF group than in the HF and NF groups at 14 days. mRNA levels for TGF-beta1 and its active ligand were increased in the HF group by at least 2- and 3-fold, respectively, at 3 and 7 days compared with levels in uninjured controls and the LF group (P<0.05) but were not different among the flow groups at 14 days. TGF-beta1 preferentially localized in the abluminal vascular smooth muscle cells of the HF arterial segments. Flow- and shear-mediated release of TGF-beta1 may therefore play a role in abrogating the proliferative and migratory response of vascular smooth muscle cells in the early stages after mural injury.

The atheroprotective effects of estrogen during the process of atherogenesis is well documented, whereas limited information is available about the effect of estrogen on pre-existing atherosclerotic lesions. After bilateral ovariectomy, 24 New Zealand White rabbits were randomized into three groups of eight animals each and subsequently fed a 0.5% cholesterol diet. In group I, the vessels were excised at day 84, whereas in group II, the cholesterol diet was continued for a total of 168 days. In group III, the animals were first fed with a cholesterol diet for 84 days; in the second phase of the experiment, the cholesterol diet was continued for a further 84 days with a combined estrogen treatment (1 mg estradiol valerate per kg body weight per week intramuscularly). At the end of the experiment, the proximal aortic arch, right carotid artery, thoracical aorta and abdominal aorta of each animal were excised and prepared for histological and immunohistological examination. By day 168, morphometrical analysis displayed a significantly lower plaque development under estrogen therapy in the carotid artery (0.08+/-0.18 mm(2) vs. 0.60+/-0.39 mm(2)), the thoracic aorta (0.56+/-0.94 mm(2) vs. 3.63+/-2.06 mm(2)), and in the abdominal aorta (0.55+/-0.70 mm(2) vs. 1.71+/-1.05 mm(2)) in comparison with the corresponding 168 day control group. However, estrogen treatment has failed to reduce further atherosclerotic plaque development in the aortic arch (9.42+/-1.79 mm(2) vs. 11. 64+/-3.29 mm(2)). Immunohistological detection of the 'anti-human factor VIII related antigen', i.e. the 'von Willebrand factor' (vWF), showed a significantly lower number of luminal cells positive for vWF in the aortic arch in the 84-day cholesterol group, compared with the corresponding controls of normocholesterolemic rabbits (65. 9+/-12.4% vs. 83.1+/-6.2%; P<0.05). Estradiol was able to inhibit the further progression of atherosclerosis when moderate vessel wall alterations were present, whereas pre-existing severe atherosclerosis was associated with a failure of the anti-atherosclerotic estrogen action. As suggested by the in situ detection of vWF as a morphological marker for endothelial cells, an intact endothelial layer might play an important role in mediating the beneficial effect of estrogen in the process of atherosclerosis.

In the chicken, glomus cells are widely distributed in the carotid body and in the wall of the common carotid artery and around its branches. The cells located in the wall of the common carotid artery express intense immunoreactivity for neuropeptide Y (NPY). They contain abundant dense-cored vesicles ranging from 70 to 220 nm in diameter. In this study, we examined ultrastructural localization of NPY in the glomus cells by using the postembedding immunogold method. Gold particles representing immunoreactivity for NPY were specifically localized on the dense-cored vesicles of the glomus cells. In addition, the localization of NPY mRNA in the glomus cells was examined by in situ hybridization with digoxigenin-labeled chicken NPY cRNA probe. A strong hybridization signal for NPY mRNA was detected in the glomus cells located in the wall of the common carotid artery. Few glomus cells of the carotid body, however, displayed labeling for NPY mRNA. Northern blot analysis with the chicken NPY exon 4 probe demonstrated that a single band for NPY mRNA was present in the poly (A) + RNA isolated from the common carotid artery where the glomus cells were distributed. Furthermore, the expression of NPY mRNA in the common carotid artery was confirmed by the reverse transcription-polymerase chain reaction. These results indicate that the chicken glomus cells are able to produce NPY but that the level of its translation varies according to the location of the cells.

Background: Growth factors such as platelet-derived growth factor (PDGF) have been postulated to be important mediators of neointimal formation in balloon-injured artery. Binding of growth factors to their receptors activates intrinsic receptor tyrosine kinase, resulting in tyrosine phosphorylation of receptors themselves and cellular substrate proteins. We investigated in vivo activities of growth factors by determining the extent of tyrosine phosphorylation of growth factor receptors and substrate proteins in injured artery., Methods and Results: Rat balloon-injured carotid artery was analyzed for phosphotyrosine content of PDGF alpha- and beta-receptors, epidermal growth factor (EGF) receptors, and insulin receptor substrate-1 (IRS-1) by immunoprecipitation and anti-phosphotyrosine Western blot. The development of intimal thickening after deendothelializing balloon catheterization of rat carotid artery was accompanied by transient twofold to threefold increases in the extent of tyrosyl phosphorylation of PDGF alpha- and beta-receptors but not EGF receptor or IRS-1. The AT1 angiotensin II (Ang II) receptor antagonist TCV-116 markedly inhibited both tyrosyl phosphorylation of PDGF alpha- and beta-receptors and intimal thickening. The AT1 antagonist reduced mRNA levels of both PDGF-A and -B chains in injured arteries., Conclusions: The present study provides direct evidence for increased PDGF activities in injured artery in situ and the involvement of Ang II in stimulated activation of PDGF receptors. These results are consistent with the pathogenetic role for PDGF in intimal thickening.

The mechanical properties and structure of a human common carotid artery were studied in order to develop criteria for designing and manufacturing compliant textile vascular grafts. The arterial wall comprised a composite of elastin and collagen fibers with the collagen fibers crimped. This structure led to a unique pressure-circumferential stretch ratio curve, the slope of which increased with an increase in strain. The increase in slope was particularly rapid at a stretch ratio above 1.4 or pressure above 120 mmHg. Based on the knowledge gained, a criteria for the design of biomechanically compliant arterial grafts was developed. An elastomeric prestretched polyurethane monofilament yarn with a low modulus of elasticity and a bulked polyester multifilament yarn with a high modulus of elasticity were combined and used as threads in the manufacture of grafts. Tubular structures of diameters in the range 4-6 mm were made by weaving. Transverse compliance and morphological and permeability properties of these grafts were determined and compared with those of a currently available woven commercial grafts and human carotid arteries. Results indicated that the compliance values of the hybrid grafts were comparable with those of the human carotid artery. Preliminary in vivo studies in dogs showed promising results: a thin, stable neointima developed within 6 months of implantation on the flow surface.

An immunocytochemical and cytochemical study has been made on the ultrastructural localisation of type III (endothelial) nitric oxide synthase, endothelin-1 and the binding sites of lectin from Bandeirea simplicifolia to the endothelium surface-associated glycoproteins in the rat left common carotid artery at 1 and 28 d after Fogarty embolectomy balloon catheter-induced injury. Controls were carotid arteries from sham operated rats. In the controls, the immunoreactivity to nitric oxide synthase-III and endothelin-1 was localised in different proportions in vascular endothelial cells (36.9% +/- 4.3 and 7.6% +/- 2.7, respectively); immunoreactivity was confined to the cytoplasm and the membranes of intracellular organelles and structures. In contrast, staining with lectin was localised on the luminal surface of all endothelial cells. 1 d after injury, platelets were adherent to the endothelium-denuded intima. Some of the platelets displayed, immunoreactivity to nitric oxide synthase-III and endothelin-1 and were stained with lectin. 28 d after injury, a neointimal thickening of substantial size was present. Subpopulations of the regrown endothelial cells covering the luminal surface of the neointima showed positive immunoreactivity to nitric oxide synthase-III and endothelin-1 but there was a significant decrease in the proportion of nitric oxide synthase-III-containing endothelial cells (17.2% +/- 1.9; P < 0.001) and a significant increase in the proportion of endothelin-1-containing endothelial cells (36.9% +/- 4.7; P < 0.001) compared with the controls. Staining with lectin was associated with the cell membrane of all endothelial cells and in addition with cells located 'deeper' in the neointima which showed lectin-positive plasmalemma, Golgi complex and multivesicular bodies/lysosomes. In conclusion, regenerated endothelial cells of the neointima showed reduced population (2-fold) of nitric oxide synthase-III-and increased population (5-fold) endothelin-1-positive cells. The subendothelial location of some lectin-stained cells after balloon catheter injury indicates the heterogeneity of the neointima and suggests that some of these cells are involved in early angiogenesis. 24 h and 28 d after injury some platelets showed positive immunoreactivity for nitric oxide synthase-III and endothelin-1.

The vascular responses to 5-hydroxytrypamine (5-HT), 5-carboxamidotryptamine (5-CT, a selective 5-HT1-like receptor agonist), alpha-methyl-5-HT (alpha-M-5-HT, a relatively selective 5-HT2 receptor agonist), noradrenaline (NA), and KCl were examined in isolated, cannulated, and perfused canine common carotid arterial preparations. They caused strong vasoconstrictions. The rank order of vasoconstrictive potency was 5-HT > alpha-M-5-HT > or = NA > 5-CT >> KCl. The 5-HT-induced vasoconstriction was significantly depressed by methysergide (a 5-HT1 and 5-HT2 receptor antagonist), ketanserin (a selective 5-HT2 receptor antagonist), and spiperone (a selective 5-HT2 receptor antagonist). The 5-CT- and alpha-M-5-HT-induced vasoconstrictions were also significantly inhibited by methysergide, spiperone, and ketanserin. The NA-induced vasoconstriction was readily inhibited by bunazosin (an alpha-adrenoceptor antagonist) and ketanserin but not significantly inhibited by spiperone and methysergide. KCl has a weak potency for producing a vasoconstriction of the canine common carotid artery. A relatively large dose of diltiazem (a calcium channel-blocker) did not modify 5-HT-induced vasoconstrictions.(ABSTRACT TRUNCATED AT 250 WORDS)

Photodynamic therapy, the light activation of photosensitizers into cytotoxic mediators, has been a successful treatment for experimental intimal hyperplasia (IH). To understand the basis of the photosensitizer chloroaluminum sulfonated phthalocyanine (CASPc)-mediated photoinhibition of intimal hyperplasia in the rat common carotid artery model, we studied photosensitizer partitioning in hyperplastic as compared to normal arterial tissue. Serum clearance of CASPc is exponential with, a half-life of 300 minutes. Laser-induced fluorescence and spectrofluorimetric analyses of artery tissue demonstrated an approximately 60% lower uptake and retention of CASPc by normal arterial tissue as compared to arteries with IH; the differences become more pronounced at 24 h. Fluorescent microscopy of arterial tissue demonstrated increased uptake of the CASPc by the artery with IH. However, by 24 h it is primarily the IH tissue that has retained the CASPc, with clearance of the dye from the media of normal or hyperplastic arteries. These data demonstrate that IH, like neoplastic tissue, has an increased accumulation of CASPc compared to normal artery. The preferential partitioning into hyperplastic tissue has implications for therapeutic targeting of this cellular population with photodynamic therapy.