elastin and Carotid-Artery-Diseases

Early vascular aging is a process characterized by a reduction in arterial elastin with an increase in collagen that has been related to cardiovascular risk factor and can determine an increased arterial stiffness and central blood pressure. It can be measured by several non invasive methods and in different arterial segment. The present paper will focus on functional (local stiffness parameter) and structural (intima media thickness) carotid arteries alterations typically evaluated by ultrasound methods. Methodological, research and clinical issue has been reviewed.

Topics: Adult; Age Factors; Arterial Pressure; Carotid Arteries; Carotid Artery Diseases; Carotid Intima-Media Thickness; Collagen; Elastin; Humans; Middle Aged; Predictive Value of Tests; Vascular Remodeling; Vascular Stiffness; Young Adult

Sickle cell anemia (SCA) causes chronic inflammation and multiorgan damage. Less understood are the arterial complications, most evident by increased strokes among children. Proteolytic mechanisms, biomechanical consequences, and pharmaceutical inhibitory strategies were studied in a mouse model to provide a platform for mechanistic and intervention studies of large artery damage due to sickle cell disease. Approach and Results: Townes humanized transgenic mouse model of SCA was used to test the hypothesis that elastic lamina and structural damage in carotid arteries increased with age and was accelerated in mice homozygous for SCA (sickle cell anemia homozygous genotype [SS]) due to inflammatory signaling pathways activating proteolytic enzymes. Elastic lamina fragmentation observed by 1 month in SS mice compared with heterozygous littermate controls (sickle cell trait heterozygous genotype [AS]). Positive immunostaining for cathepsin K, a powerful collagenase and elastase, confirmed accelerated proteolytic activity in SS carotids. Larger cross-sectional areas were quantified by magnetic resonance angiography and increased arterial compliance in SS carotids were also measured. Inhibiting JNK (c-jun N-terminal kinase) signaling with SP600125 significantly reduced cathepsin K expression, elastin fragmentation, and carotid artery perimeters in SS mice. By 5 months of age, continued medial thinning and collagen degradation was mitigated by treatment of SS mice with JNK inhibitor.. Arterial remodeling due to SCA is mediated by JNK signaling, cathepsin proteolytic upregulation, and degradation of elastin and collagen. Demonstration in Townes mice establishes their utility for mechanistic studies of arterial vasculopathy, related complications, and therapeutic interventions for large artery damage due to SCA.

Topics: Anemia, Sickle Cell; Animals; Anthracenes; Carotid Arteries; Carotid Artery Diseases; Cathepsin K; Collagen; Disease Models, Animal; Elastin; Hemoglobins; Homozygote; Humans; JNK Mitogen-Activated Protein Kinases; Mice, Transgenic; Mutation; Protein Kinase Inhibitors; Proteolysis; Signal Transduction; Time Factors; Vascular Remodeling

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.. 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.. 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).. 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.

Topics: Adolescent; Adult; Age Factors; Carotid Artery Diseases; Carotid Artery, Common; Elastin; Female; Humans; Male; Middle Aged; Radiography; Risk Factors; Vascular Remodeling; Young Adult

Atherosclerotic plaques with a low content of connective tissue proteins are believed to have an increased risk of rupture and to give rise to clinical events. The aim of the present study was to investigate if the content of elastin, collagen and of the matrix metalloproteinase (MMP) -1, -3, -9 and -12 in plaques removed at surgery can be associated with the occurrence of ipsilateral symptoms.. The atherosclerotic plaques of 221 patients undergoing carotid endarterectomy were analyzed and their composition was related to the incidence of preoperative, intraoperative and postoperative neurological events.. Elastin, collagen and MMP-12 contents were lower in males and diabetic patients. Elastin (P .010), MMP-3 (P .008) and MMP-9 (P < .0001) were lower, while MMP-1 (P .004) and MMP-9 (P .002) were higher in plaques of patients with preoperative symptoms, even after correction for the time between the occurrence of symptoms and surgery. Elastin and MMP-12 decreased (r = -0.17, P .009 and r = -.288, P <.0001 respectively) while MMP-1 (r = 0.17, P .012) and MMP-9 (r = .21 P <.0001) increased with age. After a mean follow-up time of 39.6 ± 16.6 months, 7.7% of patients had suffered one or multiple ipsilateral neurological events. Patients with plaque elastin levels lower than the median (52 mg/g) had increased post-operative incidence of ipsilateral stroke (P for trend 0.009 using Log Rank Chi-square test). This finding was confirmed when controlling for age, gender, hypertension, diabetes, smoking, pre-operative symptoms and statin usage in a Cox Proportional Hazard model (hazard ratio 7.38, 95% C.I. 1.50-36.31).. These observations support the concept that elastin may be important for plaque stability, and suggest that a low plaque content of elastin is associated with a higher risk for ipsilateral stroke.

Topics: Aged; Carotid Artery Diseases; Collagen; Comorbidity; Elastin; Endarterectomy, Carotid; Female; Humans; Incidence; Male; Matrix Metalloproteinases; Plaque, Atherosclerotic; Risk Factors; Stroke

Angiotensin II (AII) receptor 1 (ATR1) and angiotensin converting enzyme 1 (ACE1) blockers have been shown to reduce acute cardiovascular events in patients, improve plaque stability and modify matrix metalloproteinase (MMP) expression. However, the role of the ACE1/AII/ATR1 axis in interstitial collagenase regulation has not been fully explored. In this study, we investigated the effect of ATR1 and ACE1 blockade on the expression and activity of MMP-1, -8 and -13 in human carotid atheroma.. Atheroma samples (n = 24) were obtained from patients undergoing carotid endarterectomy. The effects of ATR1 (irbesartan), ACE1 (quinapril), ACE2 (DX600) and MMP (GM6001) blockade on the expression of AII, the interstitial collagenases and soluble elastin fragments were investigated in explant culture supernatants. Paired atheroma samples were incubated with intervention or media control for 4 days. Protein levels (AII, MMP-1, -8, -13 and soluble elastin) were determined by ELISA.. ATR1, but not ACE1, blockade significantly reduced MMP-1 and -8 concentrations in atheroma supernatants. ACE2 blockade significantly increased MMP-1 and -8 concentrations in atheroma supernatants. AII concentration in atheroma supernatants significantly increased after ATR1, ACE1 and ACE2 blockade. Release of soluble elastin fragments increased after ATR1 and ACE1 blockade, but was not changed by an MMP inhibitor.. Our findings suggest that ATR1 blockade alters AII, MMP-1, MMP-8 expression and a marker of elastin degradation in human atheroma, but that the elastin degradation response is not MMP driven. This data contributes to the recognised ability of ATR1 blockade to modify plaque stability.

Topics: Angiotensin II; Angiotensin II Type 1 Receptor Blockers; Angiotensin-Converting Enzyme Inhibitors; Biphenyl Compounds; Carotid Artery Diseases; Collagenases; Dipeptides; Elastin; Endarterectomy, Carotid; Extracellular Matrix; Humans; Irbesartan; Matrix Metalloproteinase 1; Matrix Metalloproteinase 13; Matrix Metalloproteinase 8; Matrix Metalloproteinase Inhibitors; Organ Culture Techniques; Peptides; Peptidyl-Dipeptidase A; Plaque, Atherosclerotic; Quinapril; Receptor, Angiotensin, Type 1; Signal Transduction; Tetrahydroisoquinolines; Tetrazoles

Elastase incubation was performed in the LCCA in 13 New Zealand white rabbits. Three weeks after incubation, DSA demonstrated that 10 (10/13, 77%) bifurcation-type aneurysms at the origin of the LCCA were present; mean aneurysm neck, width, and height values were 3.7 ± 1.1, 3.8 ± 0.9, and 8.7 ± 2.3 mm, respectively. The LCCA can be used to create bifurcation aneurysms in rabbits.

Topics: Aneurysm; Angiography, Digital Subtraction; Animals; Aorta, Thoracic; Aortic Aneurysm, Thoracic; Carotid Artery Diseases; Carotid Artery, Common; Disease Models, Animal; Elastin; Ligation; Rabbits

Elastin fragmentation is a common characteristic of vascular diseases, such as abdominal aortic aneurysms, peripheral arterial disease, and aortic dissection. Examining growth and remodeling in the presence of dysfunctional elastic fibers provides insight into the adaptive or maladaptive changes that tissues undergo in compensating for structural deficiencies. This study used the maturation of fibulin-5 knockout (KO) and wild-type mice to study the effects of fragmented elastic fibers on the growth and remodeling of carotid arteries. The microstructural content and organization and the biaxial mechanical behavior of common carotid arteries were measured, and parameter estimation performed from KO and WT mice aged 3, 4, 8, and 13 wk. Gross measurements and biaxial tests revealed significant differences in pressure-diameter behavior, in vivo axial stretch, opening angle, compliance, and wall stresses during maturation of wild-type arteries, but little change in these values in KO mice. Multiphoton microscopy used to image collagen fibers across the vessel wall in pressurized and stretched arteries suggests that there is little variation in fiber angles between different ages. Parameter estimation revealed significant differences in material parameters between genotypes and age groups. This study suggests that neonatal formation and cross-linking of functional elastic fibers, followed by increases in artery size due to growth with little remodeling of the elastic fibers, endow arteries with large distensibility and contribute to the evolution of mechanical behavior of arteries during maturation. Dysfunction in neonatal formation of elastic fibers abrogates many of the changes in mechanical response that take place during the maturation.

Topics: Animals; Animals, Newborn; Biomechanical Phenomena; Carotid Arteries; Carotid Artery Diseases; Collagen; Elastic Tissue; Elastin; Extracellular Matrix; Extracellular Matrix Proteins; Male; Mice; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; Models, Cardiovascular; Recombinant Proteins; Stress, Mechanical; Vascular Stiffness

Matrix metalloproteinase-9 is considered to play a pivotal role in aneurismal formation. We showed that gingival fibroblasts (GF) in vitro reduced matrix metalloproteinase-9 activity via increased secretion of tissue inhibitor of metalloproteinase 1. We aimed to evaluate in vivo the efficacy of GF transplantation to reduce aneurism development in a rabbit model.. Seventy rabbit carotid aneurisms were induced by elastase infusion. Four weeks later, GF, dermal fibroblast, or culture medium (DMEM) were infused into established aneurisms. Viable GF were abundantly detected in the transplanted arteries 3 months after seeding. GF engraftment resulted in a significant reduction of carotid aneurisms (decrease of 23.3% [P<0.001] and 17.6% [P=0.01] of vessel diameter in GF-treated arteries, 1 and 3 months after cell therapy, respectively), whereas vessel diameter of control DMEM and dermal fibroblast-treated arteries increased. GF inhibited matrix metalloproteinase-9 activity by tissue inhibitor of metalloproteinase 1 overexpression and matrix metalloproteinase-9/tissue inhibitor of metalloproteinase 1 complex formation, induced elastin repair, and increased elastin density in the media compared with DMEM-treated arteries (38.2 versus 18.0%; P=0.02). Elastin network GF-induced repair was inhibited by tissue inhibitor of metalloproteinase 1 blocking peptide.. Our results demonstrate that GF transplantation results in significant aneurism reduction and elastin repair. This strategy may be attractive because GF are accessible and remain viable within the grafted tissue.

Topics: Aneurysm; Animals; Carotid Artery Diseases; Cell Survival; Cells, Cultured; Elastin; Fibroblasts; Gingiva; Humans; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Rabbits; Tissue Inhibitor of Metalloproteinase-1

Quantifying the time course of load-induced changes in arterial wall geometry, microstructure, and properties is fundamental to developing mathematical models of growth and remodeling. Arteries adapt to altered pressure and flow by modifying wall thickness, inner diameter, and axial length via marked cell and matrix turnover. To estimate particular biomaterial implications of such adaptations, we used a 4-fiber family constitutive relation to quantify passive biaxial mechanical behaviors of mouse carotid arteries 0 (control), 7-10, 10-14, or 35-56 days after an aortic arch banding surgery that increased pulse pressure and pulsatile flow in the right carotid artery. In vivo circumferential and axial stretches at mean arterial pressure were, for example, 11% and 26% lower, respectively, in hypertensive carotids 35-56 days after banding than in normotensive controls; this finding is consistent with observations that hypertension decreases distensibility. Interestingly, the strain energy W stored in the carotids at individual in vivo conditions was also less in hypertensive compared with normotensive carotids. For example, at 35-56 days after banding, W was 24%, 39%, and 47% of normal values at diastolic, mean, and systolic pressures, respectively. The energy stored during the cardiac cycle, W(sys)-W(dias), also tended to be less, but this reduction did not reach significance. When computed at normal in vivo values of biaxial stretch, however, W was well above normal for the hypertensive carotids. This net increase resulted from an overall increase in the collagen-related anisotropic contribution to W despite a decrease in the elastin-related isotropic contribution. The latter was consistent with observed decreases in the mass fraction of elastin.

Topics: Animals; Biomechanical Phenomena; Carotid Artery Diseases; Carotid Artery, Common; Collagen; Disease Models, Animal; Elastin; Hypertension; Male; Mice; Mice, Inbred C57BL; Pulsatile Flow; Stress, Mechanical

Treatment of cerebral aneurysms by endovascular deployment of liquid embolic agents has been proposed as an alternative strategy to conventional coiling, and new materials are being developed for embolization. In this study, the authors used a single-injection, biocompatible, biodegradable and pH-responsive acrylated chitosan (aCHN) with conjugated vascular endothelial growth factor (rhVEGF) in a rat aneurysm model.. The efficacy of the aCHN formulation with rhVEGF was tested using a common carotid artery occlusion model in rats, and the extent of embolization was evaluated using quantitative, qualitative, and histopathological techniques after 14 days of implantation.. The mean occlusion was significantly greater for the rhVEGF/aCHN-treated group (96.8 +/- 3.0%) than for the group receiving aCHN (74.7 +/- 5.6%) (p < 0.01). Through qualitative evaluation, intimal and medial proliferation were significantly greater with rhVEGF/aCHN than with aCHN and controls (p < 0.001). Degradation of the aCHN filler was monitored in concert with the production of extracellular matrix components. Macrophages migrated in and proliferated inside the occluded carotid artery lumens were identified by histological and immunostainings. Results showed resorption of chitosan with concurrent development of collagen and elastin into the vessel lumen, suggesting clot maturation into fibrosis.. Chitosan with a bioactive agent such as rhVEGF showed excellent results in occluding aneurysms in a rat model.

Topics: Animals; Biocompatible Materials; Brain; Carotid Artery Diseases; Carotid Artery, Common; Chitosan; Collagen; Disease Models, Animal; Elastin; Embolization, Therapeutic; Extracellular Matrix; Fibrosis; Hydrogen-Ion Concentration; Intracranial Aneurysm; Macrophages; Mitogens; Rats; Rats, Sprague-Dawley; Vascular Endothelial Growth Factor A

To determine the role of multinucleated giant cells (MGCs) in cardiovascular diseases.. MGCs are a hallmark of giant cell arteritis. They are also described in atherosclerotic plaques from aortic aneurysms and carotid and coronary arteries. Herein, we demonstrate that the cholate-containing Paigen diet yields many MGCs in atherosclerotic plaques of apolipoprotein E-/- mice. These mice revealed a 4-fold increase in MGC numbers when compared with mice on a Western or Paigen diet without cholate. Most of the MGCs stained intensively for cathepsin K and were located at fibrous caps and close to damaged elastic laminae, with associated medial smooth muscle cell depletion. During in vitro experiments, MGCs demonstrated a 6-fold increase in elastolytic activity when compared with macrophages and facilitated transmigration of smooth muscle cells through a collagen-elastin matrix. An elastin-derived hexapeptide (Val-Gly-Val-Ala-Pro-Gly [VGVAPG]) significantly increased the rate of macrophage fusion, providing a possible mechanism of in vivo MGC formation. Comparable to the mouse model, human specimens from carotid arteries and aortic aneurysms contained cathepsin K-positive MGCs.. Apolipoprotein E-/- mice fed a Paigen diet provide a model to analyze the tissue-destructive role of MGCs in vascular diseases.

Topics: Animals; Antigens, Differentiation; Aortic Aneurysm; Apolipoproteins E; Atherosclerosis; Carotid Artery Diseases; Cathepsin K; Cell Fusion; Cell Movement; Cells, Cultured; Cholates; Collagen; Dietary Fats; Disease Models, Animal; Elastin; Endotoxins; Giant Cells, Foreign-Body; Humans; Immunohistochemistry; Interleukin-4; Macrophages, Peritoneal; Mice; Mice, Knockout; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Oligopeptides; Time Factors; Toll-Like Receptor 4

To validate the hypothesis that the toxic heavy metal lead (Pb) may be linked to cardiovascular diseases via the initiation of atherosclerosis, in vivo and in vitro studies were conducted.. During the human study part of this project, serum Pb levels of healthy young women were correlated to carotid intima-media thickness. Multivariate logistic regression analyses showed that increased serum Pb levels were significantly associated with an increased intima-media thickness (P=0.01; odds ratio per SD unit, 1.6 [95% CI, 1.1 to 2.4]). In vitro, Pb induced an increase in interleukin 8 production and secretion by vascular endothelial cells. Nuclear factor erythroid 2-related factor-2 is the crucial transcription factor involved in Pb-induced upregulation of interleukin 8. Endothelial cell-secreted interleukin 8 triggered intimal invasion of smooth muscle cells and enhanced intimal thickening in an arterial organ culture model. This phenomenon was further enhanced by Pb-increased elastin synthesis of smooth muscle cells.. Our data support the hypothesis that Pb is a novel, independent, and significant risk factor for intimal hyperplasia.

Topics: Adolescent; Carotid Artery Diseases; Cell Movement; Cell Proliferation; Cells, Cultured; Dose-Response Relationship, Drug; Elastin; Endothelial Cells; Female; Heat-Shock Proteins; Humans; Hyperplasia; Interleukin-8; Lead; Logistic Models; Mammary Arteries; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; NF-E2-Related Factor 2; Odds Ratio; Organ Culture Techniques; Radial Artery; Risk Assessment; Risk Factors; Severity of Illness Index; Time Factors; Tunica Intima; Ultrasonography; Up-Regulation; Young Adult

We tested the hypothesis that carotid artery stiffening with ageing is associated with transforming growth factor-β1 (TGF-β1)-related increases in adventitial collagen and reductions in medial elastin, which would be reversed by voluntary aerobic exercise. Ex vivo carotid artery incremental stiffness was greater in old (29–32 months, n = 11) vs. young (4–7 months, n = 8) cage control B6D2F1 mice (8.84 ± 1.80 vs. 4.54 ± 1.18 AU, P < 0.05), and was associated with selective increases in collagen I and III and TGF-β1 protein expression in the adventitia (P < 0.05), related to an increase in smooth muscle α-actin (SMαA) (myofibroblast phenotype) (P < 0.05). In cultured adventitial fibroblasts, TGF-β1 induced increases in superoxide and collagen I protein (P < 0.05), which were inhibited by Tempol, a superoxide dismutase. Medial elastin was reduced with ageing, accompanied by decreases in the pro-synthetic elastin enzyme, lysyl oxidase, and increases in the elastin-degrading enzyme, matrix metalloproteinase 2. Fibronectin was unchanged with ageing, but there was a small increase in calcification (P < 0.05). Increased incremental stiffness in old mice was completely reversed (3.98 ± 0.34 AU, n = 5) by 10–14 weeks of modest voluntary wheel running (1.13 ± 0.29 km day−1), whereas greater voluntary wheel running (10.62 ± 0.49 km day−1) had no effect on young mice. The amelioration of carotid artery stiffness by wheel running in old mice was associated with reductions in collagen I and III and TGF-β1, partial reversal of the myofibroblast phenotype (reduced SMαA) and reduced calcification (all P < 0.05 vs. old controls), whereas elastin and its modulating enzymes were unaffected. Adventitial TGF-β1-related oxidative stress may play a key role in collagen deposition and large elastic artery stiffening with ageing and the efficacious effects of voluntary aerobic exercise.

Topics: Aging; Analysis of Variance; Animals; Carotid Artery Diseases; Cells, Cultured; Collagen; Connective Tissue; Cyclic N-Oxides; Elastin; Fibroblasts; Fibronectins; Male; Matrix Metalloproteinase 2; Mice; Myocytes, Smooth Muscle; Physical Conditioning, Animal; Rats; Rats, Sprague-Dawley; Spin Labels; Superoxides; Transforming Growth Factor beta1

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.. 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).. 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).. 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.

Topics: Calcinosis; Carotid Artery Diseases; Carotid Artery, Common; Collagen; Elastin; Endarterectomy, Carotid; Feasibility Studies; Fiber Optic Technology; Fibrosis; Foam Cells; Humans; Lasers; Lipids; Necrosis; Predictive Value of Tests; Reproducibility of Results; Rupture; Sensitivity and Specificity; Spectrometry, Fluorescence; Time Factors

Clinical studies have demonstrated that 50% of individuals with chronic renal disease (CRD) die of cardiovascular causes, including advanced calcific arterial and valvular disease; however, the mechanisms of accelerated calcification in CRD remain obscure, and no therapies can prevent disease progression. We recently demonstrated in vivo that inflammation triggers cardiovascular calcification. In vitro evidence also indicates that elastin degradation products may promote osteogenesis. Here, we used genetically modified mice and molecular imaging to test the hypothesis in vivo that cathepsin S (catS), a potent elastolytic proteinase, accelerates calcification in atherosclerotic mice with CRD induced by 5/6 nephrectomy.. Apolipoprotein-deficient (apoE(-/-))/catS(+/+) (n=24) and apoE(-/-)/catS(-/-) (n=24) mice were assigned to CRD and control groups. CRD mice had significantly higher serum phosphate, creatinine, and cystatin C levels than those without CRD. To visualize catS activity and osteogenesis in vivo, we coadministered catS-activatable and calcification-targeted molecular imaging agents 10 weeks after nephrectomy. Imaging coregistered increased catS and osteogenic activities in the CRD apoE(-/-)/catS(+/+) cohort, whereas CRD apoE(-/-)/catS(-/-) mice exhibited less calcification. Quantitative histology demonstrated greater catS-associated elastin fragmentation and calcification in CRD apoE(-/-)/catS(+/+) than CRD apoE(-/-)/catS(-/-) aortas and aortic valves. Notably, catS deletion did not cause compensatory increases in RNA levels of other elastolytic cathepsins or matrix metalloproteinases. Elastin peptide and recombinant catS significantly increased calcification in smooth muscle cells in vitro, a process further amplified in phosphate-enriched culture medium.. The present study provides direct in vivo evidence that catS-induced elastolysis accelerates arterial and aortic valve calcification in CRD, providing new insight into the pathophysiology of cardiovascular calcification.

Topics: Animals; Aorta; Aortic Valve; Aortic Valve Stenosis; Apolipoproteins E; Calcinosis; Carotid Artery Diseases; Cathepsins; Cells, Cultured; Creatinine; Cystatin C; Elastin; Humans; Kidney Failure, Chronic; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Muscle, Smooth, Vascular; Nephrectomy; Osteogenesis; Phosphates; Tunica Intima; Tunica Media

Arteries experience marked variations in blood pressure and flow during the cardiac cycle that can intensify during exercise, in disease, or with aging. Diverse observations increasingly suggest the importance of such pulsatility in arterial homeostasis and adaptations. We used a transverse aortic arch banding model to quantify chronic effects of increased pulsatile pressure and flow on wall morphology, composition, and biaxial mechanical properties in paired mouse arteries: the highly pulsatile right common carotid artery proximal to the band (RCCA-B) and the nearly normal left common carotid artery distal to the band (LCCA-B). Increased pulsatile mechanical stimuli in RCCA-B increased wall thickness compared with LCCA-B, which correlated more strongly with pulse (r* = 0.632; P < 0.01) than mean (r* = 0.020; P = 0.47) or systolic (r* = 0.466; P < 0.05) pressure. Similarly, inner diameter at mean pressure increased in RCCA-B and correlated slightly more strongly with a normalized index of blood velocity pulsatility (r* = 0.915; P < <0.001) than mean flow (r* = 0.834; P < 0.001). Increased wall thickness and luminal diameter in RCCA-B resulted from significant increases in cell number per cross-sectional area (P < 0.001) and collagen-to-elastin ratio (P < 0.05) as well as a moderate (1.7-fold) increase in glycosaminoglycan content, which appears to have contributed to the significant decrease (P < 0.001) in the in-vivo axial stretch in RCCA-B compared with LCCA-B. Changes in RCCA-B also associated with a signficant increase in monocyte chemoattractant protein-1 (P < 0.05) whereas LCCA-B did not. Pulsatile pressure and flow are thus important stimuli in the observed three-dimensional arterial adaptations, and there is a need for increased attention to the roles of both axial wall stress and adventitial remodeling.

Topics: Animals; Aorta, Thoracic; Blood Pressure; Carotid Artery Diseases; Carotid Artery, Common; Chemokine CCL2; Collagen; Disease Models, Animal; Elastin; Heart Rate; Hypertension; Mice; Mice, Inbred C57BL; Pulsatile Flow; Stress, Mechanical

Cystatin C is a major inhibitor of the elastin- and collagen-degrading cysteine proteases and may therefore have an important role in preserving atherosclerotic plaque stability. In this study we analyzed the associations between human carotid plaque cystatin C expression and the plaque content of collagen and elastin.. Thirty-one plaques were removed by endarterectomy and homogenized. Cystatin C levels were analyzed by densitometry of Western blots and elastin and collagen levels were determined colorimetrically.. The plaque content of cystatin C correlated with total elastin (r = 0.58, p = 0.001) and collagen (r = 0.50, p = 0.004), as well as with cross-linked forms of elastin (r = 0.42, p = 0.022) and collagen (r = 0.52, p = 0.003). Immunohistochemical analysis demonstrated that cystatin C colocalized with elastin and collagen. No correlation was seen between cystatin C and the amount of degraded elastin or collagen in plaques.. The positive correlation between cystatin C levels and collagen and elastin levels in plaques supports the notion that cystatin C plays an important role in maintaining atherosclerotic plaque stability.

Topics: Aged; Blotting, Western; Carotid Arteries; Carotid Artery Diseases; Collagen; Colorimetry; Cystatin C; Cystatins; Elastin; Endarterectomy, Carotid; Female; Humans; Immunohistochemistry; Male; Middle Aged

Cathepsin K (CatK), a potent elastinolytic and collagenolytic cysteine protease, likely participates in the evolution and destabilization of atherosclerotic plaques. To assess better the biology of CatK activity in vivo, we developed a novel near-infrared fluorescence (NIRF) probe for imaging of CatK and evaluated it in mouse and human atherosclerosis.. The NIRF imaging agent consists of the CatK peptide substrate GHPGGPQGKC-NH2 linked to an activatable fluorogenic polymer. In vitro, CatK produced a 2- to 14-fold activation of the agent over other cysteine and matrix metalloproteinases (P<0.0001), as well as a >8-fold activation over a control imaging agent (P<0.001). Optical imaging of atheroma revealed >100% NIRF signal increases in apolipoprotein E-/- mice in vivo (n=13; P<0.05, CatK imaging agent versus control agent) and in human carotid endarterectomy specimens ex vivo (n=14; P<0.05). Fluorescence microscopy of plaque sections demonstrated that enzymatically active CatK (positive NIRF signal) localized primarily in the vicinity of CatK-positive macrophages. Augmented NIRF signal (reflecting CatK activity) colocalized with disrupted elastin fibers within the media underlying plaques.. Use of this novel protease-activatable NIRF agent for optical imaging in vivo demonstrated preferential localization of enzymatically active CatK to macrophages, consistent with their known greater elastinolytic capabilities compared with smooth muscle cells. Augmented CatK proteolysis in atheromata further links CatK to vascular remodeling and plaque vulnerability.

Topics: Animals; Apolipoproteins E; Carotid Arteries; Carotid Artery Diseases; Cathepsin K; Cathepsins; Elastin; Enzyme Activation; Fluorescein-5-isothiocyanate; Fluorescent Dyes; Humans; Immunohistochemistry; Macrophages; Mice; Mice, Mutant Strains; Microscopy, Fluorescence; Peptide Hydrolases; Spectroscopy, Near-Infrared

Arterial tortuosity is a frequent manifestation of vascular disease and collateral vessel growth, but its causes are poorly understood. This study was designed to assess the relationship between the development of tortuosity and the mechanical forces that are imposed on arterial tissue.. Axial strain in rabbit carotid arteries was reduced from 62+/-2% to 33+/-2% by implanting an interposition graft, prepared from the contralateral carotid, at the downstream end of the artery. Axial strain remained unchanged for 12 weeks; however, all vessels became tortuous because of tissue growth and remodeling. After 7 days, there was a marked elevation in proliferation rates of endothelial and smooth muscle cells; however, increased apoptosis was also detected, and no net accumulation of DNA was observed. Significant accumulations of elastin (24%) and total collagen (26%) occurred by 5 weeks. Gelatin zymography detected upregulation and activation of matrix metalloproteinase-2 (MMP-2), and confocal microscopy revealed enlargement of fenestrae in the internal elastic lamina. MMP inhibition by treatment with doxycycline prevented enlargement of fenestrae and development of tortuosity, and it enabled normalization of axial strain by 5 weeks.. These findings indicate that substantial axial strain is necessary to sustain the morphological stability of arteries, and that a reduction in strain results in arterial tortuosity attributable to aberrant MMP activity.

Topics: Animals; Apoptosis; Carotid Arteries; Carotid Artery Diseases; Cell Division; Collagen; Elastin; Endothelium, Vascular; Matrix Metalloproteinase 2; Muscle, Smooth, Vascular; Rabbits; Stress, Mechanical; Tensile Strength; Transplantation, Autologous; Weight-Bearing

Many processes involved in the pathogenesis of atherosclerosis result in modifications of the extracellular matrix. These changes not only determine the mechanical stability of atherosclerotic lesions but can directly or indirectly influence further development of the lesions. The purpose of the present study was to compare the matrix composition of human carotid plaques from symptomatic patients with those obtained from patients without symptoms. Furthermore, matrix changes related to age were studied.. Thirty atherosclerotic carotid plaques were removed by endarterectomy from 27 patients and divided into 2 groups on the basis of the presence of ipsilateral symptoms. The plaques were homogenized, and the total levels of the major components of the extracellular matrix were determined.. Plaques associated with symptoms were characterized by increased levels of elastin (1.58+/-0.46 versus 1.24+/-0.40 mg/g wet wt; P=0.03) and decreased levels of hydroxyapatite (45.1+/-46.3 versus 131.4+/-111.7 mg/g wet wt; P=0.02) compared with asymptomatic plaques. The increase in elastin in plaques from symptomatic patients was due to elevated levels of an intermediate-size fraction, as determined by liquid chromatography. Collagen and sulfated glycosaminoglycans were present in equal amounts in both groups. Elastin content in carotid plaques decreased with age.. Carotid plaques from symptomatic patients have lower levels of hydroxyapatite than those from asymptomatic patients. The present study also raises the possibility that non-cross-linked forms of elastin, increased in plaques associated with symptoms, could be a marker of plaque vulnerability and/or directly induce harmful cellular activities or increase lipoprotein retention in the vascular wall.

Topics: Age Factors; Aged; Aging; Biomarkers; Carotid Artery Diseases; Cerebrovascular Disorders; Collagen; Durapatite; Elastin; Endarterectomy, Carotid; Extracellular Matrix; Female; Glycosaminoglycans; Humans; Male; Risk Factors

To establish the relationship between elastin degradation and aneurysm growth in New Zealand white rabbit model aneurysms, and to explore the potential for pharmacologic inhibition of elastinolysis and aneurysm growth with use of the matrix metalloproteinase (MMP) inhibitor doxycycline.. Elastase-induced, saccular aneurysms created in the right common carotid artery in 30 animals randomly divided into controls (n = 16) and doxycycline treated (n = 14) were studied. Aneurysm growth was determined by angiography and aneurysm specimens were collected at 7 and 14 days for histologic and immunohistochemical analysis.. Aneurysms were characterized by marked elastin degradation and thickening of the arterial wall media in the absence of inflammatory cell markers. There was no evidence for expression of MMPs in the aneurysm wall at any time point. Aneurysm formation and growth were not prevented by the systemic administration of doxycycline. Mean aneurysm width increased from 3.1 +/- 0.7 mm at 3 days to 3.7 +/- 0.8 mm at 7 days and 4.2 +/- 0.8 mm at 14 days (P =.012 and P =.017, respectively). There was no statistically significant difference in aneurysm size and elastin content at any time point between doxycycline treated and control animals.. Elastase-induced rabbit aneurysm formation is accompanied by total elastin destruction that was not inhibited by the administration of doxycycline. Aneurysms in this model may be caused by the initial infusion of elastase, rather than by ongoing degradation from endogenous proteases released by inflammatory cells.

Topics: Animals; Carotid Artery Diseases; Disease Models, Animal; Doxycycline; Elastin; Intracranial Aneurysm; Matrix Metalloproteinase Inhibitors; Pancreatic Elastase; Rabbits; Random Allocation

Calcification has been examined in 250 samples of atherosclerotic lesions (types II to VI) of human carotid arteries using von Kossa and haematoxylin staining. Early calcification described as 'stippling' was first noted in stage III specimens, with intermediate and solid calcifications becoming increasingly prominent within advanced plaques, especially stages Vb and VI. Although the relative frequencies of stippling, intermediate and large calcified deposits varied between plaques of the same stage, the prevalent sites of calcification were recognized as the deeper regions of the intima and the atheroma. Immunolocalization and histochemical techniques were used to identify the associations of mast cells (MCs), macrophages, smooth muscle cells (SMCs), and elastin with the different stages of calcification. Early, dispersed stippling was commonly associated with local accumulations of macrophages (HAM56 and CD68-positive), MCs and extracellular MC tryptase, the presence of immunoreactive elastin, but the relative absence of SMCs. Intermediate stages of calcification described as 'morula' deposits were also associated with local increases in the numbers of macrophages and MCs. Larger calcified deposits, even within the same plaque specimen, showed no regular pattern of cellular or elastin associations. However, in the vast majority of specimens, macrophages represented the predominant cell type associated with different phases of calcification. By contrast, the calcification less frequently observed in the media beneath advanced plaques was commonly associated with SMCs and elastin; only rarely were macrophages or MCs present. These studies are the first to demonstrate that macrophages, MCs, and extracellular tryptase frequently occupy micro-environmental loci showing the first stages of calcification within the atherosclerotic plaque; similar associations with more advanced mineral deposits are discussed in relation to plaque rupture.

Topics: Aged; Aged, 80 and over; Arteriosclerosis; Calcinosis; Carotid Artery Diseases; Elastin; Female; Humans; Macrophages; Male; Mast Cells; Middle Aged; Muscle, Smooth, Vascular

To test the hypothesis that atherosclerosis may be initiated by hypoperfusion or thrombotic occlusion of the adventitial vasa vasonum.. In a new model of atherogenesis, an early atherosclerotic lesion may be initiated by removal of the adventitia from the carotid artery of the New Zealand White rabbit, wherein lie the vasa vasorum.. Animal laboratory, University Department of Surgery and Medicine.. Immunocytochemistry was undertaken to demonstrate the presence of smooth muscle cells and macrophages within the intimal lesions. Smooth muscle cells were labelled with a monoclonal antibody designated HHF35 and macrophages were labelled with a rabbit specific, macrophage specific antibody, RAM11. CHIEF RESULTS: In rabbits fed a normal diet, at day 14, the intimal lesion was composed exclusively of smooth muscle cells. By day 28, such lesions had regressed. In rabbits fed a high cholesterol diet, at day 14, the intimal lesion was composed of a mixture of macrophages and smooth muscle cells. By day 42, the pattern of cellular distribution was such that macrophages (present as foam cells) were predominant. In the presence of persistent hypercholesterolaemia these lesions did not regress.. This new model can produce two different cellular responses that may mimic the intimal lesions seen with re-stenosis after angioplasty or in hypercholesterolaemic man and as such, might be useful in separating out these two different pathophysiologies.

Topics: Animals; Arteriosclerosis; Carotid Arteries; Carotid Artery Diseases; Collagen; Elastic Tissue; Elastin; Endothelium, Vascular; Foam Cells; Hypercholesterolemia; Immunohistochemistry; Ischemia; Macrophages; Male; Microscopy, Electron; Muscle, Smooth, Vascular; Rabbits; Thrombosis; Tunica Intima

High resolution (125-microns lateral, 55-microns axial) images of 16 muscular (femoral) and 15 elastic (common carotid) human arteries were made in vitro with use of a prototype 45-MHz intravascular imaging system. Four distinct regions of scattering, excluding plaque, were identified in the ultrasound images corresponding histologically to the adventitia, media, thickened intima and elastic laminae, both internal and external. Arterial samples imaged under pressure and in a collapsed state underwent dimensional changes but exhibited similar levels of backscatter amplitude. All the elastic arteries displayed a prominent echogenic media, whereas all the muscular arteries displayed an echolucent media. Scattering from the internal elastic lamina in muscular arteries provided an excellent landmark for defining the location and extent of intimal thickening or plaque. In elastic arteries the internal elastic lamina could not be distinguished from the echogenic media; consequently, the boundary between the media and intimal layer was indistinct. Differences in the relative concentration and organization of collagen and elastin were found to provide a consistent explanation for the differences in scattering that were observed between individual layers within an artery as well as between muscular and elastic arteries.

Topics: Arteriosclerosis; Carotid Arteries; Carotid Artery Diseases; Collagen; Elastin; Femoral Artery; Humans; Pressure; Ultrasonography

Despite significant progress in elucidating the pathogenesis of aneurysmal disease, the precise etiology of arterial wall degradation remains unclear. Numerous etiologies have been implicated, including stress-strain factors, structural wall abnormalities, and enzyme imbalance. We have previously shown that collagenase and elastase are increased in aortic aneurysm tissue. Herein we report and characterize a newly described serum compound that is able to hydrolyze an artificial elastase substrate but is not elastase. This substance is elevated in patients with atherosclerotic disease and, following aneurysmectomy, its concentration increases by threefold. Examination of the substance reveals that it is bound to lipid and consists of four subunits of molecular weights: 310,000, 62,000, 40,000, and 10,000 daltons. It has characteristics of thiol, carboxyl, and metalloenzymes and is most active at a pH of 7.0 to 8.0. A relationship between this serum compound and aneurysm tissue enzymatic activity is noted. We postulate that this serum compound may be produced by mononuclear cells and released into the serum. Furthermore, monocytes may enter the arterial wall intima and release this substance, resulting in proteolytic arterial wall degradation and subsequent aneurysm formation.

Topics: Aortic Aneurysm; Aortic Diseases; Arterial Occlusive Diseases; Blood Proteins; Carotid Artery Diseases; Elastin; Female; Humans; Hydrogen-Ion Concentration; Leukocytes; Male; Middle Aged; Molecular Weight; Monocytes; Neutrophils; Pancreatic Elastase; Peptide Hydrolases; Proteoglycans

Topics: Adult; Aged; Autoantibodies; Carotid Artery Diseases; Cerebral Infarction; Cerebrovascular Disorders; Complement System Proteins; Elastin; Giant Cell Arteritis; Humans; Immune Complex Diseases; Immunoglobulins; Male; Middle Aged; Temporal Arteries