Increased expression of inhibitor of apoptosis proteins in atherosclerotic plaques of symptomatic patients with carotid stenosis

Vascular remodeling and atheromatous lesion formation are determined in part by the balance between apoptosis and survival of vascular smooth muscle cells (VSMCs). In the chronic stages, apoptosis of VSMCs in the atherosclerotic plaques contributes to the weakening and potential rupture of the plaque causing pathologies such as acute coronary syndrome. The higher incidence of apoptosis in the plaques of symptomatic than in asymptomatic patients has been demonstrated, but the expression of survival proteins, including the inhibitor of apoptosis proteins (IAPs), has not been thoroughly examined. The aim of this study was to investigate the immunohistochemical expression of cellular inhibitor of apoptosis protein-2 (cIAP2), x-linked inhibitor of apoptosis protein (XIAP), and survivin in normal carotid arteries, and carotid endarterectomy specimens of symptomatic and asymptomatic patients with carotid stenosis. The results demonstrated stronger immunopositivity to smooth muscle myosin heavy chain antigen (SM-MHC) (sm2), proliferating cell nuclear antigen (PCNA), and p50 subunit of NF-kappabeta in the asymptomatic plaques than in symptomatic plaques. Furthermore, there was higher expression of cIAP2, XIAP, and survivin in the symptomatic than in the asymptomatic plaques and this paralleled caspase-3 expression. The increased expression of IAPs in symptomatic plaques could be due to endogenous defense mechanism to protect against the pro-apoptotic effect of the inflammatory stimuli that are released in the plaques. This could be involved in the stabilization of symptomatic atheromatous plaques and may prove a potential therapeutic target.     

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.     

Vascular reactivity and atheromatous plaques in post-menopausal women on tibolone treatment. Open prospective study with Doppler ultrasonography in internal carotid artery

Background: Arteriosclerosis is the main cause of ischaemic ictus. The middle cerebral and anterior cerebral arteries, which irrigate over 70% of the entire cerebral tissue, spring from the internal carotid. Additionally, it is in the extracraneal vessels that embolism, thrombosis and stenosis originate more frequently. Aim: To evaluate the variations in blood flow (pulsatility index: PI) and to assess the evolution of atherogenic lesions (thickening of the vascular intima and the presence of atheromatous plaques) in the internal carotid artery after tibolone therapy. Subjects and methods: A total of 116 healthy menopausal women were included in this open, prospective and comparative study. Of them, 101 subjects completed the 48 weeks follow-up. Subjects were allocated in two groups: group T (n = 55) received 2.5 mg/day of tibolone daily and group C (n = 61) was a free-treatment control group. To evaluate both resistance to blood flow and the existence and evolution of atheromatous plaques in the internal carotid, an ultrasonograph with a pulsed Doppler was used. The PI was used as the parameter of vascular tone. To study atherosclerotic lesions in the internal carotid artery, we used morphological criteria. Measurements were done before entering in the study, and at 12, 24, 36 and 48 weeks of treatment. Results: After tibolone treatment the PI in the internal carotid artery was observed markedly diminished. Moreover, tibolone reduces both the thickness and length of atheromatous plaque and the degree of vascular stenosis. Conclusion: tibolone administration reduced the carotid atheromatous plaque in thickness and length and improved cerebral perfusion.     

稳定性,不稳定性心绞痛及急性心肌梗死病人冠状动脉病变的？…

比较稳定型心绞痛、不稳定型心绞痛及急性心肌梗死病人冠状动病变的组织学差异,以阐明其发生的病理学机制。方法选天临床诊明确的ＳＡ、ＵＡ及ＡＭＩ病人死后的尸检收脏标本,对其冠状动脉取材并做组织学及免疫组织化学观察。     

Expression of collagen, interstitial collagenase, and tissue inhibitor of metalloproteinases-1 in restenosis after carotid endarterectomy.

Extracellular matrix is the principal component of the fibrous caps of atherosclerotic plaques and intimal hyperplastic lesions of reconstructed arteries. Interstitial collagen form an important part of the matrix, and the balance between collagen synthesis and degradation by interstitial collagenase (matrix metalloproteinase-1, MMP-1) may determine whether plaques rupture or vessels develop stenosis. We examined type I procollagen gene expression in human atherosclerotic and restenotic carotid arteries using in situ messenger RNA (mRNA) hybridization and the expression of MMP-1 and its endogenous inhibitor (tissue inhibitor of metalloproteinases-1, TIMP-1) by immunohistochemistry. Compared with normal arteries, atherosclerotic plaques bed increased expression of immunoreactive MMP-1 and TIMP-1 with modest increase of type 1 procollagen mRNA. Early restenotic lesions (< 1.5 years) contained abundant type I procollagen mRNA but little immunoreactive MMP-1 and TIMP-1. Late restenotic lesions (> 4 years) resembled atheroma and exhibited increased immunoreactive MMP-1 and TIMP-1 as well as abundant type I procollagen mRNA. Compared with atherosclerotic plaques, type I procollagen is increased and MMP-1 is decreased in early restenotic lesions. MMP-1 and TIMP-1 expressions are upregulated in lesions with a clear atheroma. These findings suggest that the balance between proteolysis and matrix synthesis may influence both the stability of atheromatous plaques and the development of restenotic lesions.     

Coexpression of cyclooxygenase-2 and matrix metalloproteinases in human aortic atherosclerotic lesions

Inflammation appears to have a major role in the development of atherosclerosis. Cyclooxygenase-2 (COX-2) is involved in the inflammatory response via the generation of prostanoids that, in turn, are involved in the production of matrix metalloproteinases (MMPs). This study aimed to investigate atherosclerosis in human aortas for in situ tissue distribution of COX-2, MMPs including MMP-9 and membrane type 1 MMP (MT1-MMP), and tissue inhibitor of metalloproteinase-2 (TIMP-2). Immunohistochemical studies were performed on atherosclerotic lesions of aortas from patients with aortic aneurysms (n = 4) and dissections (n = 3) by using antibodies to COX-2, MMP-9, MT1-MMP, and TIMP-2. Control tissues were obtained from traumatically dissected aortas (n = 2). All specimens from diseased aortas had atherosclerotic lesions ranging from fatty streak to atheromatous plaques. In control, there was no expression of COX-2, MMP-9, and MT1-MMP in all aortic layers. Immunoreactivity for COX-2 was predominantly noted in macrophages and smooth muscle cells (SMCs) of the intima including atherosclerotic plaque itself and the medial layer of the plaque base, as well as in SMCs and endothelial lining of the vasa vasorum in the adventitia. Immunoreactivity for MMP-9 and MT1-MMP was found in the same distribution as that of COX-2. Additionally, the expression of TIMP-2 increased in relation to MMP-9 expression. This study demonstrates that COX-2 is coexpressed with MMP-9 and MT1-MMP, not only by macrophages and SMCs in atherosclerotic lesions, but also in endothelial lining of the vasa vasorum of human aortas. Thus, vascular inflammatory reactions may influence extracellular matrix remodeling by coactivation of MMPs in the development of atherosclerosis and, in turn, the progression of disease.     

脑梗死患者间歇使用低分子肝素对颈动脉斑块和血清炎性标志物的影响

目的:脑梗死患者间断使用低分子肝素,观察其对颈动脉斑块和血清炎性标志物的影响.方法:选择脑梗死并具有颈动脉斑块的患者78例作为对象,随机分为常规组(37例)和低分子肝素组(41例),分别于治疗前后测定颈动脉斑块情况及血清高敏C反应蛋白(hs-CRP)、基质金属蛋白酶-9(MMP-9)水平.结果:①治疗前比较,两组间颈动...     

Cytoskeletal features of normal and atheromatous human arterial smooth muscle cells

The distribution of actin, vimentin, desmin, and tropomyosin was studied in the media of the human aorta and femoral and coronary arteries, as well as in atheromatous plaques from the same arteries, by means of immunofluorescence, densitometric analysis of sodium dodecylsulfate-polyacrylamide gel electrophoresis, and bidimensional gel electrophoresis. The proportions of desmin-containing cells varied in the media of different arteries; 4 per cent of the cells in the aorta, 11 per cent in the coronary artery, and 37 per cent in the femoral artery contained desmin. In fibrous atheromatous plaques, independently of the artery, desmin-containing cells were almost absent, but they reappeared in complicated lesions. The content of vimentin per smooth muscle cell increased in fibrous atheromatous plaques, whereas the content of actin and tropomyosin was less than in normal media. Moreover, the alpha-actin predominance observed in the media was transformed to beta-actin predominance in the atheromatous plaques. These cytoskeletal changes provide new, possibly useful, biochemical markers for the characterization of smooth muscle cells during early and advanced phases of atheroma formation.     

基质金属蛋白酶-9及其组织抑制物-1与颈动脉粥样斑块稳定性相关研究

目的: 探讨基质金属蛋白酶-9（MMP-9）、基质金属蛋白酶抑制剂-1（TIMP-1）、MMP-9/TIMP-1与脑梗死患者颈动脉粥样斑块稳定性的相关性。方法: 80例动脉粥样硬化性脑梗死患者按TCD微栓子检测结果分为微栓子阴性组70例和微栓子阳性组10例，20例正常人作为对照组，分别测定血浆MMP-9、TIMP-1水平。结果: 脑梗死患者血浆MMP-9、TIMP-1水平明显高于正常对照组(P<0.01)，相关性分析发现MMP-9水平与TIMP-1水平呈正相关（r=0.76，P<0.01）。MMP-9/TIMP-1比值仅在微栓子阳性组明显增高。结论: 血浆MMP-9参与了脑梗死的病理生理过程，血浆MMP-9和MMP-9/TIMP-1与颈动脉粥样斑块的不稳定性呈正相关。     

C-reactive protein immunohistochemical localization in normal and atherosclerotic human aortas

We examined immunohistochemically 104 formaldehyde-fixed, paraffin-embedded, human autopsy aortic specimens for C-reactive protein (CRP) presence and localization. In addition, we correlated immunoreactivity with a spectrum of the following histologic categories: normal aorta, fatty streak, atheromatous plaque, and fibrous plaque. Using appropriate controls, we confirmed CRP immunoreactivity in 3.3% of normal specimens, 75% of fatty streaks, 90.2% of atheromatous plaques, and 64.6% of fibrous plaques. Immunoreactivity in fatty streaks was located around collections of foam cells. Immunoreactivity in atheromatous plaques was in a bandlike distribution corresponding to the pale-staining insudative zone frequently seen in such lesions. The correlation and localization of CRP immunoreactivity in atherosclerotic lesions presented here suggests a functional role for CRP in the pathogenesis of atherosclerosis. Our results encourage efforts to determine more precisely the physiologic contributions of CRP to the development and exacerbation of atherosclerosis.     

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。     

Smooth muscle homeostasis in human atherosclerotic plaques through interleukin 15 signalling

Interleukin (IL)-15 is a cytokine that has a broad tissue distribution and is important in maintaining homeostasis of cells and stability of tissues. When II-15 is also expressed by vascular smooth muscle cells (SMC), which are the dominant type of cells in most atherosclerotic plaques, it could be important in maintaining plaque tissue integrity and hence resistance of plaques towards development of clinically relevant complications such as plaque rupture and thrombosis. In this study, IL-15 and IL-15Rα in vitro expression by coronary artery SMC was investigated using RT -PCR and FACS analysis. Immunohistochemistry was used to study in situ expression of IL-15 and IL-15R by SMC of human carotid artery atherosclerotic plaques. Multiplex ligand-dependent probe amplification (MLPA) was used to investigate the mRNA expression of 40 pro- and anti inflammatory genes after stimulating coronary SMC with IL-15. We found that atherosclerotic SMC express both IL-15 and its receptor IL-15R, and TNF-γ and TNF-α enhance IL-15R expression in cultured SMC. MLPA studies on SMC revealed enhanced expression of PDGF beta mRNA after IL15 stimulation. In conclusion, our data suggest that IL-15 may contribute to atherosclerotic plaque integrity by stimulation of smooth muscle cells, probably in a PDGF dependent fashion.     

MicroRNA-21 is a unique signature associated with coronary plaque instability in humans by regulating matrix metalloproteinase-9 via reversion-inducing cysteine-rich protein with Kazal motifs

BackgroundCoronary atherosclerotic unstable plaque is one of the leading causes of cardiovascular death. Macrophage-derived matrix metalloproteinase (MMP) 9 is considered for degrading extracellular matrix and collagen, thereby thinning the fibrous cap in plaques. miR-21 is implicated to play an important role in the progression of atherosclerosis. Nevertheless, miR-21 as the biomarker for coronary atherosclerotic unstable plaque remains unknown. We aimed to investigate the prediction role of miR-21 for unstable plaque by pathway study of miR-21 on MMPs and its inhibitor RECK in macrophages.MethodsExpression of miR-21 in macrophages and serum miR-21 as well as MMP-9 was measured in patients with coronary non-calcified plaque, calcified plaque and controls. In vitro experiment was done in human macrophages by over-expressing miR-21 or down-regulating RECK. The regulation of RECK and MMP-9 by miR-21 was evaluated by western blotting and siRNA strategy.ResultsPatients with non-calcified coronary artery lesions had significantly higher miR-21 in macrophages and lower miR-21 serum levels compared to the control and calcified plaque patients. At the same time, the serum levels of MMP-9 were significantly elevated in non-calcified patients. Experiments in vitro indicated that over-expressing miR-21 could induce the expression and secretion of pro-MMP-9 and active-MMP-9 in human macrophages via targeting gene RECK, and knocking down RECK expression by specific siRNA can resemble that of miR-21 over-expression.ConclusionsmiR-21 might be a biomarker for plaque instability by suppressing target gene RECK to promote the expression and secretion of MMP-9 in macrophages.     

Coronary artery remodelling and the assessment of stenosis by pathologists

When atherosclerotic plaques develop, the cross- sectional area of the artery at that point often increases to accommodate the plaque without any reduction in lumen size. In consequence the angiogram does not detect a high proportion of atherosclerotic plaques. The increase in size of the artery (compensatory dilatation − arterial remodelling) varies widely in degree between different plaques even in the same artery. Dilatation of a degree to prevent any loss of lumen size is regarded as adequate compensatory dilatation. In contrast, other plaques are associated with no or minimal increase in the vessel cross-sectional area and a reduction in lumen size is present (inadequate compensation). High-grade stenosis is in particular associated with a total failure of remodelling. Such plaques may have had a rapid growth phase, out-pacing the ability of the medial smooth muscle cells to undergo a rearrangement.
The phenomenon of remodelling has important consequences for pathologists who use the traditional method of comparing the lumen size relative to the cross-sectional area of the vessel at the site of a plaque to measure stenosis. The area of the vessel at this point may be anything up to 60% above its size before the plaque developed. An error is introduced which on average overestimates diameter stenosis by 30% when compared to an angiographic equivalent method in which the lumen size at the lesion is compared to the lumen size at an adjacent segment of artery without a plaque.     

256层CT对颈动脉粥样硬化斑块成分及性质的分析

目的　探讨256层螺旋CT 血管造影（CT angiography,CTA）评价颈动脉粥样硬化斑块的价值。 方法　 对234例拟诊颈动脉硬化狭窄的患者行头颈部CTA检查, 根据颈动脉有无斑块分为病变组和正常对照组,通过CT值分析斑块成分和性质。 结果　病变组266支动脉,其中脂肪斑块组51支、纤维斑块组26支、钙化斑块组67支、混合斑块组122支;正常组163支。脂肪斑块组脑梗死出现率明显大于纤维斑块组和钙化斑块组（P＜0.01）;脂肪合并钙化斑块组脑梗死出现率明显大于纤维合并钙化斑块组（P＜0.01）;含脂肪斑块组溃疡出现率大于含纤维斑块组（P＜0.05）。 结论　256层螺旋CT能够分析颈动脉斑块的成分、性质,初步评价颈动脉斑块的稳定性。     

ADAM-10 could mediate cleavage of N-cadherin promoting apoptosis in human atherosclerotic lesions leading to vulnerable plaque: A morphological and immunohistochemical study

Atherosclerosis remains a major cause of mortality. Whereas the histopathological progression of atherosclerotic lesions is well documented, much less is known about the development of unstable or vulnerable plaque, which can rupture leading to thrombus, luminal occlusion and infarct. Apoptosis in the fibrous cap, which is rich in vascular smooth muscle cells (VSMCs) and macrophages, and its subsequent weakening or erosion seems to be an important regulator of plaque stability. The aim of our study was to improve our knowledge on the biological mechanisms that cause plaque instability in order to develop new therapies to maintain atherosclerotic plaque stability and avoid its rupture. In our study, we collected surgical specimens from atherosclerotic plaques in the right or left internal carotid artery of 62 patients with evident clinical symptoms. Histopathology and histochemistry were performed on wax-embedded sections. Immunohistochemical localization of caspase-3, N-cadherin and ADAM-10 was undertaken in order to highlight links between apoptosis, as expressed by caspase-3 immunostaining, and possible roles of N-cadherin, a cell-cell junction protein in VSMCs and macrophages that provides a pro-survival signal reducing apoptosis, and ADAM-10, a “disintegrin and metalloproteases” that is able to cleave N-cadherin in glioblastomas. Our results showed that when apoptosis, expressed by caspase-3 immunostaining, increased in the fibrous cap, rich in VSMCs and macrophages, the expression of N-cadherin decreased. The decreased N-cadherin expression, in turn, was linked to increased ADAM-10 expression. This study shows that apoptotic events are probably involved in the vulnerability of atherosclerotic plaque.     

Genetic loss of Gas6 induces plaque stability in experimental atherosclerosis

The growth arrest-specific gene 6 (Gas6) plays a role in pro-atherogenic processes such as endothelial and leukocyte activation, smooth muscle cell migration and thrombosis, but its role in atherosclerosis remains uninvestigated. Here, we report that Gas6 is expressed in all stages of human and mouse atherosclerosis, in plaque endothelial cells, smooth muscle cells and macrophages. Gas6 expression is most abundant in lesions containing high amounts of macrophages, ie thin fibrous cap atheroma and ruptured plaque. Genetic loss of Gas6 does not affect the number and size of initial and advanced plaques in ApoE(-/-) mice, but alters its plaque composition. Compared to Gas6(+/+): ApoE(-/-) mice, initial and advanced plaques of Gas6(-/-): ApoE(-/-) mice contained more smooth muscle cells and more collagen and developed smaller lipid cores, while the expression of TGFbeta was increased. In addition, fewer macrophages were found in advanced plaques of Gas6(-/-): ApoE(-/-) mice. Hence, loss of Gas6 promotes the formation of more stable atherosclerotic lesions by increasing plaque fibrosis and by attenuating plaque inflammation. These findings identify a role for Gas6 in plaque composition and stability.