Clinical and therapeutical implications of EPC biology in atherosclerosis

Bone marrow-derived circulating endothelial progenitor cells have been successfully used to enhance angiogenesis after tissue ischemia. The role of endothelial progenitor cells in endothelial cell homeostasis and their putative role in atherogenesis have been recently investigated. Cardiovascular risk factors negatively influence endothelial progenitor cell number and function while vasculoprotection e.g. by statins, estrogens and physical activity may be partly mediated by progenitor cells. Endogenous mobilization or injection of ex-vivo generated endothelial progenitor cells is associated with an enhanced reendothelialization, an improvement of endothelial function and reduced atherosclerotic burden. In contrast, endothelial progenitor cells may promote plaque angiogenesis in animal models and may negatively influence plaque development and stability. However, in humans with coronary atherosclerotic disease, endothelial progenitor cells are a novel risk predictor for cardiovascular mortality and morbidity. In this review we focus on the role of circulating endothelial progenitor cells in endothelial cell repair mechanisms at the vascular wall and their potentially protective and therapeutic role in atherosclerotic disease.     

Monocytic microparticles promote atherogenesis by modulating inflammatory cells in mice

Microparticles (MP) are generated during a vast number of biological processes such as inflammation, cell activation and apoptosis. Increasing evidence points towards an important role of MP as intercellular messengers of biological information. During atherogenesis, monocytes infiltrate the vascular wall and foster inflammation, accompanied by the release of monocytic MP (mono-MP). To date, only little is known about the biological function of mono-MP in the vascular wall. Here, we investigated the role of mono-MP during atherogenesis. Mono-MP were generated by starvation of THP-1 monocytes and isolated by ultracentrifugation. To investigate whether mono-MP influence atherogenesis, ApoE^−/− mice were fed a high-fat, cholesterol-rich diet for 8 weeks and simultaneously treated with mono-MP or vehicle twice a week. Mice treated with mono-MP showed significantly increased monocyte and T-cell infiltration into the vessel wall, as assessed by Moma-2 and CD3 staining, and enhanced plaque formation, as assessed by oil-red-O staining. However, atherosclerotic plaque composition was not influenced by mono-MP application. In vitro, incubation of mono-MP with murine macrophages and endothelial cells resulted in the uptake of calcein-labelled mono-MP. Mono-MP uptake initiated the generation of intracellular reactive oxygen species. Murine macrophages pre-treated with mono-MP showed significantly enhanced expression of CCR2, migration to MCP-1 and increased release of pro-inflammatory interleukin-6. Co-incubation of mono-MP with endothelial cells resulted in significantly increased expression of ICAM-1, as assessed by RT-PCR and ELISA. Mono-MP act as paracrine messengers that intensify inflammation during atherogenesis by stimulating vascular-bound and inflammatory cells in their vicinity.     

Scavenger receptor regulation and atherosclerosis

Atherosclerosis and its complications, such as coronary heart disease, heart infarction and stroke, are the leading causes of death in the developed world. High blood pressure, diabetes, smoking and a diet high in cholesterol and lipids clearly increase the likelihood of premature atherosclerosis, albeit other factors, such as the individual genetic makeup, may play an additional role. During atherosclerosis, uncontrolled cholesterol and lipid accumulation in macrophages and smooth muscle cells leads to foam cell formation and to the progression of the atherosclerotic plaque. This review will focus on foam cell formation within the atherosclerotic lesion, the involvement of the scavenger receptor genes in this process, and the possibility to interfere with scavenger receptor function to reduce the progression of atherosclerosis. To date, the regulatory mechanisms for the expression of scavenger receptor genes and their role in atherosclerosis are not well characterized. Knowledge on this subject could lead to a better understanding of the process, prevention and therapy of this disease.     

Endothelial caveolin-1 plays a major role in the development of atherosclerosis

Clinical studies have established the important impact of atherosclerotic disease in Western societies. This disease is characterized by the accumulation of lipids and the migration of various cell types in the sub-endothelial space of blood vessels. As demonstrated by many studies, endothelial cells play an essential role in the development of this disease. The endothelium acts as a gatekeeper of blood vessel integrity and cardiovascular health status. For instance, the transfer of lipids via the transport of lipoproteins in the arterial intima is believed to be mediated by endothelial cells through a process termed transcytosis. In addition, lipoproteins that accumulate in the sub-endothelial space may also be modified, in a process that can direct the activation of endothelial cells. These steps are essential for the initiation of an atherosclerotic plaque and may be mediated, at least in part, by caveolae and their associated protein caveolin-1. In the present study, we evaluate the role of caveolin-1/caveolae in the regulation of these two steps in endothelial cells. Our data clearly demonstrate that caveolin-1 is involved in the regulation of lipoprotein transcytosis across endothelial cells and in the regulation of vascular inflammation.     

Elevated cell-specific microparticles are a biological marker for cerebral dysfunctions in human severe malaria

Cerebral malaria (CM) and severe anemia (SA) are the most severe complications of Plasmodium falciparum infections. Although increased release of endothelial microparticles (MP) correlates with malaria severity, the full extent of vascular cell vesiculation remains unknown. Here, we characterize the pattern of cell-specific MP in patients with severe malaria. We tested the hypothesis that systemic vascular activation contributes to CM by examining origins and levels of plasma MP in relation to clinical syndromes, disease severity and outcome. Patients recruited in Douala, Cameroon, were assigned to clinical groups following WHO criteria. MP quantitation and phenotyping were carried out using cell-specific markers by flow cytometry using antibodies recognizing cell-specific surface markers. Platelet, erythrocytic, endothelial and leukocytic MP levels were elevated in patients with cerebral dysfunctions and returned to normal by discharge. In CM patients, platelet MP were the most abundant and their levels significantly correlated with coma depth and thrombocytopenia. This study shows for the first time a widespread enhancement of vesiculation in the vascular compartment appears to be a feature of CM but not of SA. Our data underpin the role of MP as a biomarker of neurological involvement in severe malaria. Therefore, intervention to block MP production in severe malaria may provide a new therapeutic pathway.     

Production,Fate and Pathogenicity of Plasma Microparticles in Murine Cerebral Malaria

In patients with cerebral malaria (CM), higher levels of cell-specific microparticles (MP) correlate with the presence of neurological symptoms. MP are submicron plasma membrane-derived vesicles that express antigens of their cell of origin and phosphatidylserine (PS) on their surface, facilitating their role in coagulation, inflammation and cell adhesion. In this study, the in vivo production, fate and pathogenicity of cell-specific MP during Plasmodium berghei infection of mice were evaluated. Using annexin V, a PS ligand, and flow cytometry, analysis of platelet-free plasma from infected mice with cerebral involvement showed a peak of MP levels at the time of the neurological onset. Phenotypic analyses showed that MP from infected mice were predominantly of platelet, endothelial and erythrocytic origins. To determine the in vivo fate of MP, we adoptively transferred fluorescently labelled MP from mice with CM into healthy or infected recipient mice. MP were quickly cleared following intravenous injection, but microscopic examination revealed arrested MP lining the endothelium of brain vessels of infected, but not healthy, recipient mice. To determine the pathogenicity of MP, we transferred MP from activated endothelial cells into healthy recipient mice and this induced CM-like brain and lung pathology. This study supports a pathogenic role for MP in the aggravation of the neurological lesion and suggests a causal relationship between MP and the development of CM.     

Tobacco mosaic virus movement protein enhances the spread of RNA silencing

Eukaryotic cells restrain the activity of foreign genetic elements, including viruses, through RNA silencing. Although viruses encode suppressors of silencing to support their propagation, viruses may also exploit silencing to regulate host gene expression or to control the level of their accumulation and thus to reduce damage to the host. RNA silencing in plants propagates from cell to cell and systemically via a sequence-specific signal. Since the signal spreads between cells through plasmodesmata like the viruses themselves, virus-encoded plasmodesmata-manipulating movement proteins (MP) may have a central role in compatible virus:host interactions by suppressing or enhancing the spread of the signal. Here, we have addressed the propagation of GFP silencing in the presence and absence of MP and MP mutants. We show that the protein enhances the spread of silencing. Small RNA analysis indicates that MP does not enhance the silencing pathway but rather enhances the transport of the signal through plasmodesmata. The ability to enhance the spread of silencing is maintained by certain MP mutants that can move between cells but which have defects in subcellular localization and do not support the spread of viral RNA. Using MP expressing and non-expressing virus mutants with a disabled silencing suppressing function, we provide evidence indicating that viral MP contributes to anti-viral silencing during infection. Our results suggest a role of MP in controlling virus propagation in the infected host by supporting the spread of silencing signal. This activity of MP involves only a subset of its properties implicated in the spread of viral RNA.     

Matrix metalloproteinases at key junctions in the pathomechanism of stroke

Matrix metalloproteinases play a crucial role in the remodelling of the extracellular matrix through direct degradation of its structural proteins and control of extracellular signalling. The most common cause of ischemic brain damage is an atherothrombotic lesion in the supplying arteries. The progress of the atherosclerotic plaque development and the related thrombotic complications are mediated in part by matrix metalloproteinases. In addition to their role in the underlying disease, various members of this protease family are upregulated in the acute phase of ischemic brain damage as well as in the post-ischemic brain recovery following stroke. This review summarizes the current understanding of the matrix metalloproteinase-related molecular events at three stages of the ischemic cerebrovascular disease (in the atherosclerotic plaque, in the neurovascular unit of the brain and in the regenerating brain tissue).     

Endothelial dysfunction in acute and chronic coronary syndromes: evidence for a pathogenetic role of oxidative stress

The past two decades have highlighted the pivotal role of the endothelium in preserving vascular homeostasis. Among others, nitric oxide (NO) is currently believed to be the main component responsible for endothelium dependent vasorelaxation and therefore for endothelial function integrity. Reduced NO bioavailability causes the so-called "endothelial dysfunction," which seems to be the common molecular disorder comprising stable atherosclerotic narrowing lesions or acute plaque rupture causing unstable angina or myocardial infarction. Compelling evidence is accumulating, stressing the role of oxidative stress in causing reduced NO bioavailability and subsequently endothelial dysfunction (ED). More recently, the role of endothelial cell (EC) apoptosis as a possible final stage of ED and plaque activation has been suggested. In vitro and in vivo evidence suggests a role of oxidative stress also as a putative mechanism finally leading to plaque denudation and activation through increased EC apoptosis. Thus, oxidative stress, irrespective of atherosclerotic disease stages, seems to represent a key phenomenon in vascular disease progression and possible prevention.     

三磷酸腺苷结合盒转运体和细胞胆固醇外排

胆固醇是细胞质膜的重要组成成分。然而,过多的胆固醇累积可导致细胞中毒。异常的胆固醇胞内迁移与蓄积是造成许多心血管疾病如动脉粥样硬化的分子基础。细胞内胆固醇稳态由胆固醇的吸收、合成及外排等一系列过程调控。在哺乳动物细胞中,调节胆固醇合成、吸收和外排是维持体内胆固醇平衡的必要生理过程。本综述着重概述了三磷酸腺苷结合盒转运体(ABC)家族,如ABCA1、ABCG1、ABCG5和ABCG8的细胞功能及生理作用,以及这些转运体在调控胆固醇胞外转运中的分子机制。     

Cardiovascular co-morbidity in patients with rheumatic diseases

During recent years atherosclerosis, the major cause of cardiovascular disease (CVD), has been recognised as a chronic inflammatory condition in which rupture of atherosclerotic lesions appears to play a major role. The risk of CVD is raised in many rheumatic diseases. This risk is high in systemic lupus erythematosus - as much as a 50-times increase among middle-aged women has been reported. Studies on CVD and atherosclerosis in rheumatic disease could thus provide interesting information about CVD and atherosclerosis in addition to being an important clinical problem. A combination of traditional and nontraditional risk factors accounts for the increased risk of CVD and atherosclerosis in rheumatic disease. One interesting possibility is that atherosclerotic lesions in rheumatic disease are more prone to rupture than normal atherosclerotic lesions. It is also likely that increased risk of thrombosis may play an important role, not least in systemic lupus erythematosus. Further, it is not clear whether an increased risk of CVD is a general feature of rheumatic disease, or whether this only occurs among subgroups of patients. It should be emphasised that there is an apparent lack of treatment studies where CVD in rheumatic disease is the end point. Control of disease activity and of traditional risk factors, however, appears to be well founded in relation to CVD in rheumatic disease. Further studies are needed to determine the exact role of lipid-lowering drugs as statins. Hopefully novel therapies can be developed that target the causes of the inflammation in atherosclerotic lesions both in rheumatic patients and in the general population.     

Neovascularization in human atherosclerosis

In the absence of disease, microvessels provide vessel wall nutrients to the tunica media, while the intima is fed by oxygen diffusion from the lumen. As disease evolves and the tunica intima thickens, oxygen diffusion is impaired, and microvessels become the major source for nutrients to the vessel wall. Microvessels serve as a port of entry for inflammatory cells, from the systemic circulation to the nascent atherosclerotic lesion. As disease progress, microvessels also play a role in intraplaque hemorrhage, lipid core expansion, and plaque rupture. In addition, microvessels are also involved in stent restenosis, and plaque regression. Therefore, microvessels are a pivotal component of atherosclerosis, and proper patient risk-stratification in the near future may include the detection of increased neovascularization in atherosclerotic lesions. This review divided in two parts summarizes the current understanding of atherosclerosis neovascularization, starting with the normal anatomy and physiology and progressing to more advanced stages of the disease. We will review the structure and function of vasa vasorum in health and disease, the mechanisms responsible for the angiogenic process, the role of the immune system, including inflammation and Toll-like receptors, and the pathology of microvessels in early atherosclerotic plaques. Furthermore, the review addresses the advanced stages of atherosclerosis, summarizing the progressive role for microvessels during disease progression, red blood cell extravasation, lipid core expansion, plaque rupture, healing, repair, restenosis, and disease regression, offering the clinician a state-of-the-art, "bench to bedside" approach to neovascularization in human atherosclerosis.     

Defensins and cathelicidins: neutrophil peptides with roles in inflammation, hyperlipidemia and atherosclerosis

Atherosclerosis is a disease that begins in fetal life and represents a leading cause of morbidity and mortality associated with significant socioeconomic consequences. A central concept with regard to its pathogenesis is that of endothelial cell dysfunction, which is associated with the release of a large number of mediators secreted by leukocytes that are present in large numbers at the sites of atheroma formation. Neutrophil peptides defensins and cathelicidins are essential elements of the innate immunity and have been present in high concentrations in atherosclerotic plaques in humans. Recently, their role as potential mediators of vascular disease was investigated. Defensins are involved in the lipoprotein metabolism in the vessel wall, favoring LDL and lipoprotein (a) accumulation and modification in the endothelium and the extracellular matrix. They also interfere with the vascular smooth muscle cell function, exhibit prothrombotic activity, and play an inhibitory role in various phases of angiogenesis. Cathelicidins were recently found to enhance endothelial proliferation in cultures, induce functionally significant angiogenesis in animal models, and regulate endothelial cell apoptosis. Further study of these peptides could provide useful insight in the relationship between inflammation and atherosclerosis and is anticipated to have therapeutic and prognostic ramifications.     

Abnormal in vitro proliferation of splenic mononuclear phagocytes from autoimmune motheaten mice

Motheaten mice develop combined immunodeficiency and fatal autoimmune disease that follow autosomal recessive inheritance. In splenocyte cultures of motheaten mice, supplemented with 5% normal serum proliferating cells (MP) were present exhibiting morphologic characteristics of mononuclear phagocytes at light and electron microscopic levels. The macrophage nature of these cells was confirmed by the lack of Thy-1 antigen and immunoglobulins; the expression of Mac-1 antigen, FcR for IgG, and Ia antigens on their cell surfaces; their ability to phagocytize EA and adhere to plastic; the presence of nonspecific esterase and lysomal enzymes in their cytoplasm; and the pattern of peroxidase localization similar to monocyte-derived macrophages. MP from motheaten mice exponentially grew in culture in the absence of exogenous growth factors with a doubling time of approximately 76 hr. Although these cells were present in splenocyte cultures of normal controls, their number did not increase during the culture period under the same conditions. The addition of dextran sulfate further enhanced the proliferation of MP from motheaten mice, and induced exponential growth of these cells from normal controls, reaching only the level of unstimulated cells from motheaten mice. Radioautographic analysis demonstrated that MP substantially contributed to the elevated spontaneous and dextran sulfate-induced DNA synthesis in splenocyte cultures. Therefore, the in vitro abnormality of MP may be indicative of in vivo aberrancies of macrophages from motheaten mice and lends credence for investigating the role of macrophages in immunodeficiency and autoimmunity that develop very early in motheaten mice.     

CD4+CD25+ 调节性T 细胞与冠状动脉粥样硬化病变的研究进展

CD4+CD25+调节性T细胞是一个具有独特免疫调节功能的T细胞亚群,人体主要通过CD4+CD25+调节性T细胞以免疫负向调节的方式来抑制自身反应性T细胞的作用,减少免疫性疾病的发生,从而维持机体内环境的稳定,维持免疫耐受。CD4+CD25+Treg已被证实其与肿瘤、感染、自身免疫病、移植免疫等多种疾病的发生、发展及转归均相关。随着社会的进步和人民生活水平的提高冠状动脉粥样硬化性病变作为一种慢性病变,其发病率越来越高,已经成为严重危害人类健康的常见病,近年来越来越多的证据表明炎症及免疫反应机制在冠状动脉粥样硬化性心脏病的发生、发展及预后过程中具有重要的作用。而CD4+CD25+调节性T细胞在冠状动脉粥样硬化性病变中所起的作用也受到越来越多的关注。本文就CD4+CD25+调节性T细胞与冠状动脉粥样硬化病变之间的关联做一综述。     

Differential Expression of Cerebellar Metabotropic Glutamate Receptors mGLUR2/3 and mGLUR4a after the Administration of a Convulsant Drug and the Adenosine Analogue Cyclopentyladenosine

Metabotropic glutamate receptors (mGluR) play a role in synaptic transmission, neuronal modulation and plasticity but their action in epileptic activity is still controversial. On the other hand adenosine acts as a neuromodulator with endogenous anticonvulsive properties. Since cerebellum from epileptic patients has shown neuronal damage, sometimes associated with Purkinje cells loss, we have explored the effect of repetitive seizures on two types of mGluR in the cerebellum. Seizures were induced by the convulsant drug 3-mercaptopropionic acid (MP) and the effect of the adenosine analogue cyclopentyladenosine (CPA) alone or before MP administration (CPA+MP) were also evaluated. The expression of the receptors subtypes 2/3 (mGluR2/3) and 4a (mGluR4a) was assessed by immunocitochemistry. Granular cell layer was labeled with mGluR2/3 antibody and increased immunoreactivity was observed after MP (60%), CPA (53%) and CPA + MP (85%) treatments. Control cerebellum slices showed mGluR4a reactivity around Purkinje cells, while MP, CPA and CPA+MP treatment decreased this immunostaining. Repetitive administration of MP and CPA induces an increased cerebellar mGluR2/3 and a decreased mGluR4a immunostaining, suggesting a distinct participation of both receptors that may be related to the type of cell involved. A protective action and /or an apoptotic effect may not be discarded. CPA repetitive administration although increase seizure latency, cannot prevent seizure activity.     

Myosin phosphatase target subunit: Many roles in cell function

Phosphorylation of myosin II is important in many aspects of cell function and involves a myosin kinase, e.g. myosin light chain kinase, and a myosin phosphatase (MP). MP is regulated by the myosin phosphatase target subunit (MYPT1). The domain structure, properties, and genetic analyses of MYPT1 and its isoforms are outlined. MYPT1 binds the catalytic subunit of type 1 phosphatase, delta isoform, and also acts as an interactive platform for many other proteins. A key reaction for MP is with phosphorylated myosin II and the first process shown to be regulated by MP was contractile activity of smooth muscle. In cell division and cell migration myosin II phosphorylation also plays a critical role and these are discussed. However, based on the wide range of partners for MYPT1 it is likely that MP is implicated with substrates other than myosin II. Open questions are whether the diverse functions of MP reflect different cellular locations and/or specific roles for the MYPT1 isoforms.     

Distinct proteome features of plasma microparticles

Plasma microparticles (MPs) are spherical cell membrane fragments derived from either apoptotic or activated cells. Characterized by a rich phospholipid moiety and many protein constituents, MPs normally circulate in the blood and contribute to numerous physiological processes. In disease states, MPs derived from the injured organ likely contain valuable markers for determining the site, type, and extent of disease pathology. However, the basic protein characteristics of plasma MPs have yet to be described. In this study, MPs from a pooled plasma sample derived from 16 healthy donors, all of group A blood type, were prepared by ultracentrifugation. Flow cytometry confirmed that a majority of these MPs are smaller than 1 microm. Factor Xa generation assay revealed the presence of tissue factor activity in these MPs, confirming MPs' role in initiating blood coagulation. The MP proteome was analyzed by two-dimensional (2-D) gel electrophoresis performed in triplicate, and compared with a 2-D gel of pooled whole plasma and blood platelets. Overall, plasma MPs displayed distinct protein features and a greater number of protein spots (1021-1055) than that detected in whole plasma (331-370). Protein spots expressed in high abundance in the MP proteome were then excised and submitted for protein identity determination. This process provided protein identification for 169 protein spots and reported their relative protein quantities within the MP proteome. These 169 protein spots represented 83 different proteins and their respective isoforms. Thirty of these proteins have never before been reported in previous proteome analyses of human plasma. These results provide unprecedented information on the MP proteome and create a basis for future studies to understand MP biology and pathophysiology.     

Atherosclerosis: role of chemokines and macrophages

Atherosclerosis is a pathological process that takes place in the major arteries and is the underlying cause of heart attacks, stroke and peripheral artery disease. The earliest detectable lesions, called fatty streaks, contain macrophage foam cells that are derived from recruited monocytes. More-advanced atherosclerotic lesions, called fibro-fatty plaques, are the result of continued monocyte recruitment and smooth muscle cell migration and proliferation. Variable numbers of CD4+ T cells are found in atherosclerotic lesions, and cytokines secreted by T helper 1 (Th1)- or Th2-type cells can have a profound influence on macrophage gene expression within atherosclerotic plaques. This review briefly addresses the key features of macrophage biology and discusses the factors that influence the growth and development of atherosclerotic lesions (atherogenesis). It then considers the potential role of chemokines in mediating monocyte recruitment and macrophage differentiation within atherosclerotic lesions.