Atherosclerosis in Macaca mulatta: Histopathological,morphometric, and histochemical studies in aorta and coronary arteries of spontaneous and induced atherosclerosis

A comparative study of morphology, size, and histochemistry of the intimal lesions in aorta and coronary arteries of spontaneously occurring and cholesterol-induced atherosclerosis in rhesus monkeys has been carried out. A group of 30 normal monkeys was also investigated. Spontaneous atherosclerosis was noted in 10 of 55 adult monkeys autopsied serially; fatty streaks or atheroma in the aorta was noted in seven, fibrous plaque was noted in two, and diffuse intimal thickening was observed on one animal only. The coronary arteries showed fibrous intimal thickening without lipid in 8 of these 10 monkeys. There was fair to heavy deposition of acid mucopolysaccharides in the thickened intima along with proliferation of myointimal cells and collagen fibers. In the seven monkeys which were fed an atherogenic diet for 6 months, the aorta showed fatty streaking and atheroma in all animals. The coronary arteries also showed a variable degree of atherosclerosis but the lipid in the thickened intima was not marked. The atherosclerotic plaque height was not significantly greater than that in the spontaneous disease. These differences between spontaneously occurring and cholesterol-induced atherosclerosis in monkeys tend to indicate that the basic mechanism of lesion formation in the two states may be different.     

Role of smooth-muscle cells in the formation of the atherosclerotic plaque

The role of smooth muscle cells in the formation of atherosclerotic plaques was studied by the fluorescent antibody method on autopsy material with the aid of antiserum against smooth-muscle actomyosin. The principal forms of atherosclerotic lesion (lipid stain, fibrous plaque, atheromatous plaque) were investigated in the aorta, the brain vessels, and the coronary arteries. Smooth-muscle cells were found in the intima along with atherosclerotic foci, in the lipid stain and fibrous tissue of the plaque, but not in the atheromatous masses. Proliferation and migration of smooth-muscle cells are regarded as an essential factor in the morphogenesis of atherosclerosis.Presented by Academician of the Academy of Medical Sciences of the USSR A. P. Avtsyn.Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 79, No. 6, pp. 110–113, June, 1975.     

Pathology of myocardial infarction,coronary artery disease,plaque disruption,and the vulnerable atherosclerotic plaque

Despite scientific advances, cardiovascular disease remains the leading cause of death in developed countries. The pathologic process responsible for the majority of this mortality is atherosclerosis. Human atherosclerosis is characterized by the transition of arteries through distinct pathologic stages. Initially, there is vascular wall activation, characterized by the formation of a smooth muscle cell rich intimal hyperplasia/thickening. The thickened intima promotes the lipid and macrophage accumulation characteristic of atherosclerosis. In some patients, the atherosclerotic plaque becomes disrupted stimulating formation of luminal thrombus and acute clinical events. Understanding the pathology of such vulnerable plaques has been a challenging and controversial area of investigation. Recent prospective longitudinal imaging studies of human coronary arteries have confirmed earlier pathologic observations reporting pathologic features that predispose to acute events in some patients include plaques with a thin fibrous cap overlying a large lipid-rich necrotic core as well as plaques with severe stenosis.     

Spontaneous plaque rupture and secondary thrombosis in apolipoprotein E-deficient and LDL receptor-deficient mice

Apolipoprotein E-deficient (apoE(-/-)) and LDL receptor-deficient (LDLR(-/-)) mice develop extensive atherosclerosis, but the occurrence of spontaneous plaque rupture and secondary thrombosis in these models has not been established. The goal of this study was to provide histological evidence of acute complications of atherosclerotic lesions in these mice and to assess their prevalence. Complications of atherosclerosis were initially studied in aortas of control mice which died during previous intervention studies. Coronary arteries and the aortic origin were then systematically assessed in serial sections through the heart of apoE(-/-) and LDLR(-/-) mice. Aortic plaque rupture and/or thrombi were seen in 3 of 82 untreated mice from past intervention studies. Screening of heart sections of 33 older apoE(-/-) mice (age 9-20 months) showed extensive atherosclerosis in one or more coronary arteries of 18 animals. In three coronary arteries, the presence of blood-filled channels within advanced atherosclerotic lesions suggested previous plaque disruption/thrombotic events followed by recanalization. In the aortic origin of the same mice, four deep plaque ruptures (or erosions reaching necrotic core areas) and a large thrombus originating from the core of a disrupted atherosclerotic lesion were observed. Although plaque ruptures/deep erosions were far less frequent than in human populations, these observations demonstrate that spontaneous plaque rupture and secondary thrombosis do occur in apoE(-/-) and LDLR(-/-) mice. These mice may therefore be suitable for studying factors contributing to thrombotic complications of atherosclerosis. However, the frequent absence of a clearly defined single fibrous cap in murine coronary lesions limits their usefulness as a model of fibrous cap rupture.     

Immuno-morphologic detection of specific APO-B antigenic determinants in normal intima and atherosclerotic plaques in the human aorta

Various apo-B antigenic determinants exposures in the normal aortic intima and atherosclerotic plaques were examined by immunofluorescence using 5 clones from monoclonal antibodies (BAb) to apo-B. As a rule, 4 clones were found to react with apo-B in the normal aortic intima, whereas the clone 12G10 failed to respond to it. All the 5 determinants of apo-B were exposed in the fibrous tissue of most atherosclerotic plaques. However, there were atherosclerotic plaques, in whose fibrous tissue only did 1 to 3 clones positively react with apo-B. The similar expression of apo-B antigenic determinants was observed in the atheronecrotic zone of fibrous plaques. Apo-B did not respond to apo-B MAb at all in some atherosclerotic plaques present in the zone of atheronecrosis. Whether it is possible to modify low-density lipoproteins just in the arterial wall, especially in atherosclerotic plaques and whether this process is significant in the pathogenesis of atherosclerosis are discussed in the paper.     

Clonal mapping of the human aorta. Relationship of monoclonal characteristics, lesion thickness, and age in normal intima and atherosclerotic lesions.

The surfaces of 8 aortas from women heterozygous for G-6-PD isoenzymes were mapped for an examination of the relationships of monoclonality, lesion type, lesion thickness, and age of the patient. The percent B isoenzyme value of samples of normal intima (n = 315), fatty steak (n = 68), or fibrous plaque (n = 64) was used to define monoclonality, expressed as the [Z] score, the number of standard deviations from the mean percent B isoenzyme of samples of underlying media. Intimal thickness increased significantly with type of lesion, such that intima less than fatty streak less than fibrous plaque, and with the age of the patient. The percentage of monoclonal portions also increased with lesion type, such that 1% of samples of normal intima were monoclonal, compared with 4.4% of fatty streaks and 12.5% of fibrous plaques (P less than 0.005). Monoclonality increased with intimal thickness when normal intima, fatty streaks, and fibrous plaques were combined (P = 0.0001). When examined separately, normal intima showed a direct correlation between monoclonality and intimal thickness. In contrast, the monoclonality of fatty streaks was inversely associated with thickness (P = 0.016) and the monoclonality of fibrous plaques not related to thickness. When entered into a multiple regression model, lesion type and age, but not lesion thickness, significantly predicted monoclonality. The lack of association of intimal thickness with monoclonality suggests that it is the type of lesion that determines monoclonality and not merely its thickness. This implies that mechanisms other than clonal selection are responsible for the monoclonal characteristics of human atherosclerotic lesions.     

Absence of atherosclerosis evolution in the coronary arterial segment covered by myocardial tissue in cholesterol-fed rabbits

The evolution of atherosclerotic lesions is suppressed in the intima of the human coronary artery, beneath myocardial bridges. To elucidate the mechanism of the protective effect, we investigated morphological changes using the rabbit coronary artery as a model. Rabbits fed a 1%-cholesterol diet were killed at intervals up to 20 weeks. Two short segments of the left coronary arteries running in the epicardial adipose tissue (EpiLAD) and subsequently running in the myocardium (MyoLAD) were compared morphologically. The intima of the EpiLAD had flat endothelial cells with a polygonal shape, and demonstrated raised atherosclerotic lesions with increase in serum cholesterol level. In contrast, the intima of the MyoLAD was free of atherosclerotic lesions throughout the study, and the endothelial cells were spindle-shaped and engorged. While ferritin particles reached only the surroundings of the internal elastic lamina in the MyoLAD, they permeated into the media of the EpiLAD. We suggest that myocardial bridges suppress coronary atherosclerosis by an alteration of endothelial permeability, which may be due to changes in haemodynamic force tending towards a higher shear stress. The data provide an insight into the relationship between haemodynamics and the development of coronary atherosclerosis.     

Proteoglycans in human atherosclerotic lesions- a pilot qualitative and quantitative study by ruthenium red

A pilot study was carried out to test whether normal human aorta and aortic atherosclerotic lesions obtained at post-mortem examination were suitable for the demonstration of proteoglycans (PGs) by ruthenium red (RR) staining, and whether by this method qualitative and quantitative differences of PGs might be detectable in various types of lesions and between lesions and the normal aortic intima. The results indicate that the PGs of the above tissues obtained at post mortem were clearly visualized by electron microscopy using the RR-indicator and were thus suitable for quantitative and qualitative evaluation. Of the RR-positive granules only those 20 nm and larger were assessed. RR-granules measuring 20-50 nm were present in normal intima and media, in increased concentration in the innermost part of the mixed fatty-gelatinous lesion, and in a decreased concentration in the fibrous cap of an atherosclerotic plaque. The same RR-granules were observed in a fatty streak and in addition 50-100 nm granules were present in this type of lesion; the overall RR-granule-concentration was reduced here. Granules of both sizes were interconnected by filamentous network. The differential presence of the large (and hitherto unreported) as well as the smaller granules in the various lesions is briefly discussed in the light of the present-day knowledge of tissue-PGs, and the importance of an examination in situ of the compound PGs-complexes in their native form, is emphasized.     

Preliminary and early stages of atherosclerosis in childhood

According to the unified theory of atherosclerosis, endothelial cell injury and lipid infiltration play an important role in atherogenesis. Newborn babies may suffer endothelial cell damage, as may be detected by electron microscopy. Connective tissue elements are occasionally abundant already in newborns. Chondroitin sulfate A and C increase with age. The children may exhibit continuous accumulation of cholesterol esters in the intima of coronary arteries. Cholesteryl ester fatty acid composition, along with age, tends to approach that of serum low-density lipoproteins. Fatty streaks appear in coronary arteries in puberty, and fibrous plaques are recordable beyond the age of 20 years. The topography of myo-intimal thickenings, fatty streaks, and fibrous plaques is similar to complicated atherosclerotic lesions. Even newborn babies have obstructive myo-intimal thickenings in their coronary arteries. One fifth of all infants under one week of age suffer 20% stenosis, with percentile manifestation of stenosis in the arterial cross-section being established as ratio of intimal area to luminal area of a dilated coronary artery multiplied by 100. Occasionally, the intima is very thick, in our series initiating up to 57% of all narrowing. There are probably noxious factors which temporarily damage the endothelial cells and initiate a rapid, partially reversible thickening reaction. Some of this response of the intima to exogenous stimuli might be genetically determined. A thickened intima is susceptible to lipid deposition and atherosclerosis.     

Localization of fibronectin in atherosclerotic lesions by immunohistochemistry and immunoelectron microscopy]

This paper reports the results of a study on the distribution of fibronectin (FN), its form, character and source in atherosclerotic lesions, using immunohistochemistry (PAP method) and immunoelectron microscopic technique. The results showed that large amounts of FN were localized in fatty streaks, gelatinous lesions and early atherosclerotic plaques. The intima smooth muscle cells in atherosclerotic lesions synthesized more FN, and it is likely that FN represents a new marker of smooth muscle cell modulated from "contractile" to "synthetic" state. With the maturation of atherosclerotic plaque, FN did not fill the whole plaque but was concentrated only in the fibrous cap surface and basocentral part of the atheroma. We also proved that procollagen III peptide (PIIIP) distribution in atherosclerotic plaque was similar to that of FN.     

Determining the temperature distribution of swine aorta with simulated atheromatous plaque under pulsed laser irradiation: an experimental attempt to detect the vulnerability of atherosclerosis.

We developed a method to determine the temperature distribution of swine aortas with simulated atheromatous plaques in order to measure the temperature of atherosclerotic lesions. The inflammation associated with temperature elevation is considered to be one of the aggravating mechanisms of atherosclerosis resulting in fissuring or rupture of atheromatous plaques. The temperature distribution of plaques covered by fibrous caps cannot be measured by conventional thermistors. Indocyanine green (ICG) solution was injected into the subintima of swine aorta to simulate the light absorption coefficient of human atheromatous plaques. The temperature distribution was calculated from measured temperature changes of the aortic intima under pulsed laser irradiation. The aorta was heated from the adventitial side with a halogen lamp to simulate the temperature elevation derived from inflammation. The temperature distribution of the aorta was obtained by solving the heat transfer equation using the surface layer thickness (corresponding to the fibrous cap thickness). The surface layer thickness can be calculated using the following working formula: D(microm)=1363-398DeltaTs+35DeltaTs(2), where AT, denotes intimal surface temperature change under pulsed laser irradiation. The calculated temperature of the ICG layer (corresponding to the atheromatous core) correlated well with the measured temperature (r=0. 97, p<0.0001).     

Detection of fibronectin and collagen types I, III, IV and V in semithin sections of the human aorta

Localization of fibronectin and types I, III, IV and V collagen was investigated in semithin sections of fibrous atherosclerotic plaques and apparently normal intima of human aorta. The effect of different techniques of fixation and processing of the sections on immunostaining under peroxidase-antiperoxidase techniques have been examined. The tissue fixation in paraformaldehyde solution, removing the resin with sodium ethoxide and enzymatic pronase digestion of the sections resulted in successful specific staining of the antigens. It was found that some cells in fibrous plaques formed a cap of multiple layers of dense connective tissue containing fibronectin and type III, IV and V collagen in the absence of collagen type I.     

Comparative morphological study of the structure of the normal and arteriosclerotic internal mammary, coronary, and renal artery wall]

Morphological comparative study of the normal anatomy of the internal mammary artery, coronary artery and renal artery, and their atherosclerotic alterations. We report in this paper the comparative results of a morphological and morphometrical study of the normal anatomy of the Left Coronary Artery (interventricular descending branch) (LCA), Internal Mammary Artery (IMA) and Renal Artery (RA) and their atherosclerotic alterations in 27 unselected people of both sexes aging from 19 to 76 years (average 59 +/- 14.3). Sections from three different segments of each vessel were examined by measuring the thickness of the intima and media calculated at the maximum intimal thickening. The lumen was also measured at the level of its highest pathological reduction; the same measurements were carried out on sections free from atherosclerotic damage. Then the intima to media ratio was assumed as major indicator of atherosclerosis involvement according to the literature. All these parameters were statistically analysed. With these procedures we found that LCA displayed more severely atherosclerotic changes than IMA. The degree of damage in RA fell in between when compared to the other arteries. The intima to media ratio average was 0.061 mm. (+/- 0.084) in I.M.A.; 0.882 mm. (+/- 0.753) in L.C.A. and 0.272 (+/- 0.315) in RA with statistically significant differences between IMA and LCA (p less than 0.001) and between IMA and RA (p less than 0.001). In the lesion free sections intima to media ratios were 0.178 (+/- 0.90) in LCA, 0.053 (+/- 0.28) in IMA and 0.082 (+/- 0.127) in RA. The difference was statistically significative between LCA and IMA, but not between RA and IMA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Determining the temperature distribution of swine aorta with simulated atheromatous plaque under pulsed laser irradiation: an experimental attempt to detect the vulnerability of atherosclerosis

We developed a method to determine the temperature distribution of swine aortas with simulated atheromatous plaques in order to measure the temperature of atherosclerotic lesions. The inflammation associated with temperature elevation is considered to be one of the aggravating mechanisms of atherosclerosis resulting in fissuring or rupture of atheromatous plaques. The temperature distribution of plaques covered by fibrous caps cannot be measured by conventional thermistors. Indocyanine green (ICG) solution was injected into the subintima of swine aorta to simulate the light absorption coefficient of human atheromatous plaques. The temperature distribution was calculated from measured temperature changes of the aortic intima under pulsed laser irradiation. The aorta was heated from the adventitial side with a halogen lamp to simulate the temperature elevation derived from inflammation. The temperature distribution of the aorta was obtained by solving the heat transfer equation using the surface layer thickness (corresponding to the fibrous cap thickness). The surface layer thickness can be calculated using the following working formula: D( µm)=1363-398 &#150 T +35 &#150 T 2, where s s &#150 T s denotes intimal surface temperature change under pulsed laser irradiation. The calculated temperature of the ICG layer (corresponding to the atheromatous core) correlated well with the measured temperature (r=0.97, p<0.0001).     

Characterization of fractalkine (CX3CL1) and CX3CR1 in human coronary arteries with native atherosclerosis, diabetes mellitus, and transplant vascular disease

Background: Fractalkine is a novel chemokine that mediates both firm adhesion of leukocytes to the endothelium via CX3CR1 and leukocyte transmigration out of the bloodstream. Fractalkine has recently been shown to play a role in the pathogenesis of acute organ rejection. Since its expression is regulated by inflammatory agents such as LPS, IL-1, and TNF-, fractalkine involvement in atherosclerosis and transplant vascular disease (TVD) is of particular interest. In this study, we characterized the presence of fractalkine and its receptor CX3CR1 in human coronary arteries from normal, atherosclerotic, diabetic, and TVD settings. Method: Polyclonal rabbit antibodies were used to immunostain human fractalkine and CX3CR1 to localize their presence in transverse sections of the proximal left anterior descending and/or right coronary arteries. Slides were scored in a blinded fashion for intensity of staining (0 to 4+) and for localization in vessel walls. Results: Normal coronary arteries showed no fractalkine staining. In atherosclerotic coronary arteries, staining was localized to the intima, media, and adventitia. Within the media, fractalkine expression was seen in macrophages, foam cells, and smooth muscle cells (SMCs). Diabetic vessels showed similar staining patterns to atherosclerotic coronaries, with much stronger staining in the deep intima. Transplanted coronaries showed staining in the endothelium, intima, and adventitia in early disease, and intimal, medial, and adventitial staining in late disease. CX3CR1 staining was seen in the coronary arteries of all cases, with specific localization to regions with fractalkine staining. Conclusion: The distinctive staining patterns in native atherosclerosis, diabetes mellitus with atherosclerosis, and TVD indicate that the expression of fractalkine and CX3CR1 may be important in the pathogenesis of these diseases.     

Survival and cardiovascular pathology of heterozygous Watanabe heritable hyperlipidaemic rabbits treated with pravastatin and probucol on a low-cholesterol (0.03%)-enriched diet

 This study was aimed at determining the effects of a combined pravastatin and probucol regimen on survival and vascular pathology of heterozygous Wa- tanabe heritable hyperlipidaemic (WHHL) rabbits fed a low-cholesterol (0.03%)-enriched diet. Pravastatin monotherapy preceded the combined treatment. In animals receiving pravastatin and the enriched diet (verum group; n = 6), mean total serum cholesterol levels were consistently lowered at a dosage of 5 mg/kg pravastatin and with the combined treatment. Survival was increased (median 45 vs 25 months), while coronary atherosclerosis was less obstructive and altered to a more fibrous type than in controls (n = 8). The extent of aortic lesions, as determined by the relative plaque volume, was not related to survival in either group. However, aortic plaque types in verum group animals revealed less severe stages with a different composition and architecture, with a lower relative content of macrophage-derived foam cells and necrosis and a higher relative content of extracellular matrix. There was also a thicker fibrous cap than in control animals of similar age. Our data reveal a beneficial effect on survival of heterozygous WHHL rabbits when lipid-lowering and antioxidative treatment are combined. This appears to be due both to reduced coronary atherosclerosis and to a different, more stable type of atherosclerotic disease in this animal model. Received: 4 July 1997 / Accepted: 10 December 1997     

Coronary artery dynamics in vivo

There is considerable evidence that the localization and evolution of vascular disease are mediated, at least in part, by mechanical factors. The mechanical environment of the coronary arteries, which are tethered to the beating heart, is influenced by cardiac motion; the motion of the vessels must be described quantitatively to characterize fully the mechanical forces acting on and in the vessel wall. Several techniques that have been used to characterize coronary artery dynamics from biplane cineangiograms are described and illustrated. There is considerable variability in dynamic geometric parameters from site to site along a vessel, between the right and left anterior descending arteries, and among individuals, consistent with the hypotheses that variations in stresses mediated by geometry and dynamics affect the localization of atherosclerosis and individual risk of coronary heart disease. The few frankly atherosclerotic vessels that have been examined exhibit high torsions in the neighborhood of lesions, an observation which may have etiologic or diagnostic implications. © 2002 Biomedical Engineering Society. PAC2002: 8759Dj, 8757Gg, 8719Uv, 8719Hh     

Interstitial collagens in fibrous atherosclerotic lesions of human aorta

The present study was undertaken to clarify the existing controversy on the collagenous content and composition of human fibrous atherosclerotic versus normal aortic tissues. Several analytic procedures (slab gel electrophoresis; cyanogen bromide peptide mapping; high performance liquid chromatography; ion exchange chromatography) revealed that the amount of the interstitial collagens, i.e. types I and III, was similar in fibrous atherosclerotic lesions and control tissues (70% and 30% respectively). Moreover, when fibrous lesions were analyzed as serial fractions there was a uniform distribution of type I and type III throughout the lesion. Small increases in type III were observed only beneath the lesion where it interfaced with the normal media. The results suggest that contrary to some previous studies no major shifts in the ratio of the interstitial collagens are evident in atherosclerotic lesions as compared to normal intima-media preparations.     

股动脉粥样硬化的病理学特点探讨

目的通过与冠状动脉、颈总动脉粥样硬化比较,探讨股动脉粥样硬化的病理学特点。方法收集解放军总医院老年尸体解剖病例15例,将每例之两侧股动脉、两侧颈总动脉、左冠状动脉前降支进行连续取材,常规病理检查,选取部分节段行平滑肌肌动蛋白、CD68、bax免疫组织化学SP法染色。结果股动脉粥样硬化与冠状动脉粥样硬化在病变类型、斑块中平滑肌细胞、巨噬细胞的分布方面基本相同。但股动脉粥样硬化的范围较小、狭窄程度较轻,其斑块中的平滑肌细胞相对多,巨噬细胞相对少;bax在巨噬细胞的表达多,在平滑肌细胞的表达少。股动脉粥样硬化与颈动脉粥样硬化的病理学特点相似。结论股动脉粥样硬化与冠状动脉粥样硬化病理特点大体相同,但在某些量化指标上存在差异。     

Fenestral Pore Size in the Internal Elastic Lamina Affects Transmural Flow Distribution in the Artery Wall

Interstitial flow through the subendothelial intima and media of an artery wall was simulated numerically to investigate the water flow distribution through fenestral pores which affects the wall shear stress on smooth muscle cells right beneath the internal elastic lamina (IEL). A two-dimensional analysis using the Brinkman model of porous media flow was performed. It was observed that the hydraulic permeability of the intimal layer should be much greater than that of the media in order to predict a reasonable magnitude for the pressure drop across the subendothelial intima and IEL (about 23 mostly at a 70 mm Hg luminal pressure). When K_i was set equal to the value in the media, this pressure drop was unrealistically high. Furthermore, the higher value of K_i produced a nearly uniform distribution of water flow through a simple array of fenestral pores all having the same diameters (1.2 m), whereas when K_i was set at the value in the media, the flow distribution through fenestral pores was highly nonuniform and nonphysiologic. A deformable intima model predicted a nonuniform flow distribution at high pressure (180 mm Hg). Damage to the IEL was simulated by introducing a large fenestral pore (up to 17.8 m) into the array. A dramatic increase in flow through the large pore was observed implying an altered fluid mechanical environment on the smooth muscle cells near the large pore which has implications for intimal hyperplasia and atherosclerosis. The model also predicted that the fluid shear stress on the bottom surface of an endothelial cell is on the order of 10 dyne/cm² a level which can affect cell function. © 2001 Biomedical Engineering Society. PAC01: 8719Tt, 8380Lz, 8716Uv, 8719Ff