Angiopoietin-1 reduces VEGF-stimulated leukocyte adhesion to endothelial cells by reducing ICAM-1, VCAM-1, and E-selectin expression

Vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang1) are potent vasculogenic and angiogenic factors that hold promise as a means to produce therapeutic vascularization and angiogenesis. However, VEGF also acts as a proinflammatory cytokine by inducing adhesion molecules that bind leukocytes to endothelial cells, an initial and essential step toward inflammation. In the present study, we used human umbilical vascular endothelial cells (HUVECs) to examine the effect of Ang1 on VEGF-induced expression of three adhesion molecules: intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin. Interestingly, Ang1 suppressed VEGF-induced expression of these adhesion molecules. Furthermore, Ang1 reduced VEGF-induced leukocyte adhesion to HUVECs. These results demonstrate that Ang1 counteracts VEGF-induced inflammation by reducing VEGF-induced endothelial adhesiveness.     

Endothelial tetraspanin microdomains regulate leukocyte firm adhesion during extravasation

Tetraspanins associate with several transmembrane proteins forming microdomains involved in intercellular adhesion and migration. Here, we show that endothelial tetraspanins relocalize to the contact site with transmigrating leukocytes and associate laterally with both intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Alteration of endothelial tetraspanin microdomains by CD9-large extracellular loop (LEL)-glutathione S-transferase (GST) peptides or CD9/CD151 siRNA oligonucleotides interfered with ICAM-1 and VCAM-1 function, preventing lymphocyte transendothelial migration and increasing lymphocyte detachment under shear flow. Heterotypic intercellular adhesion mediated by VCAM-1 or ICAM-1 was augmented when expressed exogenously in the appropriate tetraspanin environment. Therefore, tetraspanin microdomains have a crucial role in the proper adhesive function of ICAM-1 and VCAM-1 during leukocyte adhesion and transendothelial migration.     

Fluid shear stress modulates surface expression of adhesion molecules by endothelial cells

We investigated the effect of hemodynamic shear forces on the expression of adhesive molecules, E-selectin, and intercellular adhesion molecule-1 (ICAM-1) on human umbilical vein endothelial cells (HUVEC) exposed to laminar (8 dynes/cm2) or turbulent shear stress (8.6 dynes/cm2 average), or to a static condition. Laminar flow induced a significant time-dependent increase in the surface expression of ICAM- 1, as documented by flow cytometry studies. Endothelial cell surface expression of ICAM-1 in supernatants of HUVEC exposed to laminar flow was not modified, excluding the possibility that HUVEC exposed to laminar flow synthetize factors that upregulate ICAM-1. The effect of laminar flow was specific for ICAM-1, while E-selectin expression was not modulated by the flow condition. Turbulent flow did not affect surface expression of either E-selectin or ICAM-1. To evaluate the functional significance of the laminar-flow-induced increase in ICAM-1 expression, we studied the dynamic interaction of total leukocyte suspension with HUVEC exposed to laminar flow (8 dynes/cm2 for 6 hours) in a parallel-plate flow chamber or to static condition. Leukocyte adhesion to HUVEC pre-exposed to flow was significantly enhanced, compared with HUVEC maintained in static condition (233 +/- 67 v 43 +/- 16 leukocytes/mm2, respectively), and comparable with that of interleukin-1 beta treated HUVEC. Mouse monoclonal antibody anti-ICAM-1 completely blocked flow-induced upregulation of leukocyte adhesion. Interleukin-1 beta, which upregulated E-selectin expression, caused leukocyte rolling on HUVEC that was significantly lower on flow- conditioned HUVEC and almost absent on untreated static endothelial cells. Thus, laminar flow directly and selectively upregulates ICAM-1 expression on the surface of endothelial cells and promotes leukocyte adhesion. These data are relevant to the current understanding of basic mechanisms that govern local inflammatory reactions and tissue injury.     

Human mast cell progenitors use alpha4-integrin, VCAM-1, and PSGL-1 E-selectin for adhesive interactions with human vascular endothelium under flow conditions

Mast cells (MCs) are central to asthma and other allergic diseases, and for responses to infection and tissue injuries. MCs arise from committed progenitors (PrMCs) that migrate from the circulation to tissues by incompletely characterized mechanisms, and differentiate in situ in perivascular connective tissues of multiple organs. PrMCs derived in vitro from human cord blood were examined for adhesion molecule expression and their ability to adhere to human umbilical vein endothelial cells (HUVECs) under conditions that mimic physiologic shear flow. The PrMCs expressed alpha(4)beta(1), low levels of beta7, and the beta2-integrins alphaLbeta2 and alphaMbeta2. The PrMCs also expressed PSGL-1, but not L-selectin. At low (0.5 dynes/cm(2)-1.0 dynes/cm(2)) shear stress, PrMCs attached and rolled on recombinant E-selectin and P-selectin and VCAM-1. An anti-PSGL-1 monoclonal antibody (mAb) blocked essentially all adhesion to P-selectin but reduced adhesion to E-selectin by only 40%, suggesting PrMCs express other ligands for E-selectin. PrMCs adhered strongly to tumor necrosis factor-alpha (TNF-alpha)-activated HUVECs, whereas adhesion to interleukin 4 (IL-4)-activated HUVECs was lower. PrMC adhesion to IL-4-activated HUVECs was totally alpha4-integrin- and VCAM-1-dependent. Adhesion to TNF-alpha-activated HUVECs was blocked by 50% by mAbs against alpha4-integrin, vascular cell adhesion molecule-1 (VCAM-1), E-selectin, or PSGL-1, whereas combinations of mAbs to alpha4-integrin plus PSGL-1, or VCAM-1 plus E-selectin, blocked adhesion by greater than 70%. Thus, PrMCs derived in vitro predominantly use alpha4-integrin, VCAM-1, PSGL-1, and other ligands that bind E-selectin for adhesion to cytokine-activated HUVEC monolayers. These observations may explain the abundance of MCs at sites of mucosal inflammation, where VCAM-1 and E-selectin are important inducible receptors.     

Cytokines in Sickle Cell Disease

Sickle red cells express adhesion molecules including integrin ₄β₁, CD36, band 3 protein, sulfated glycolipid, Lutheran protein, phosphatidylserine and integrin-associated protein. The proadhesive sickle cells may bind to endothelial cell P-selectin, E-selectin, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), CD36 and integrins leading to its activation. Monocytes also activate endothelium by releasing proinflammatory cytokines like tumor necrosis factor alpha (TNF-) and interleukin 1β (IL-1β). Sickle monocytes also express increased surface CD11b and cytoplasmic cytokines TNF and IL-1β indicating activated state. Polymorphonuclear leukocytes (PMNs) are also activated with reduced L-selectin expression, enhanced CD64 expression and elevated levels of sL-selectin, sCD16 and elastase resulting in increased adhesiveness to the endothelium. Platelets are also activated and secrete thrombospondin (TSP) and cytokine IL-1. They also form platelet- monocytes aggregates causing endothelial cell P-selectin expression. Endothelial cell activation by these multiple mechanisms leads to a loss of vascular integrity, expression of leukocyte adhesion molecules, change in the surface phenotype from antithrombotic to prothrombotic, excessive cytokine production and upregulation of HLA molecules. Furthermore, contraction of these activated endothelial cells leads to exposure of extracellular matrix proteins, such as TSP, laminin, and fibronectin and their participation in adhesive interactions with bridging molecules from the plasma such as von Willebrand factor (vWf) released from endothelial cells, ultimately culminating in vasoocclusion and local tissue ischemia, the pathognomonic basis of vasoocclusive crisis.     

Tumor necrosis factor alpha induces endothelial galactosyl transferase activity and verocytotoxin receptors. Role of specific tumor necrosis factor receptors and protein kinase C

Infections with verocytotoxin (VT) producing Escherichia coli have been strongly implicated in the epidemic form of hemolytic uremic syndrome (HUS). Endothelial damage plays a central role in the pathogenesis of HUS. In vitro studies have shown that VT can damage endothelial cells after interaction with its cellular receptor globotriaosylceramide (GbOse3cer). Cytokines, such as tumor necrosis factor alpha (TNF alpha) and interleukin-1 (IL-1) can potentiate the toxic effect of VT by inducing a protein-synthesis dependent increase in VT receptors on endothelial cells. In this study, the mechanisms underlying the increase in endothelial VT receptors induced by TNF alpha were studied in more detail. To investigate which proteins were involved in this induction, endothelial cells were incubated with and without TNF alpha in the presence of 14C-galactose or 14C-glucose. Thin-layer chromatography (TLC) analysis of the glycolipid extracts of these cells demonstrated a markedly enhanced incorporation of 14C-galactose in GbOse3cer and other galactose-containing glycolipids, suggesting that TNF alpha enhanced galactosyl-transferase activity. To examine the role of the two recently cloned TNF-receptors (TNFR-p75 and TNFR-p55) in the TNF alpha-induced increase in GbOse3cer in human endothelial cells, cells were incubated with TNF alpha, the TNFR-p55 selective R32W-S86T- TNF alpha-mutant, or the TNFR-p75 selective D143N-A145R-TNF alpha- mutant. The effect of TNF alpha activation, determined by binding- experiments with 125I-VT-1, could be largely, but not completely mimicked by R32W-S86T-TNF alpha. Although incubation of cells with D143N-A145R-TNF alpha did not show an increase in VT-1 binding, the monoclonal antibody utr-1, which prevents binding to TNFR-p75, decreased the TNF alpha-induced VT-1 binding. Activation of protein kinase C (PKC) by phorbol ester increases the expression of VT-1 receptors; this effect was prevented by the PKC inhibitor Ro31-8220 and by homologous desensitization by pretreatment with phorbol ester. In contrast, the presence of the protein kinase inhibitor Ro31-8220 or desensitization of PKC activity reduced the TNF alpha-induced increase in VT-1 receptors maximally by 50% and 24%, respectively. Comparable reductions in overall protein synthesis and the synthesis of E-selectin and plasminogen activator inhibitor-1 (PAI-1) were observed. This suggests an effect on general protein synthesis rather than a specific effect of PKC in the signal transduction pathway, by which TNF alpha induces VT-1 receptors. Our results indicate that TNF alpha can increase the VT-1 receptors on endothelial cells by inducing galactosyl- transferase activity, that this action of TNF alpha mainly occurs via the TNFR-p55; and that PKC activation increases expression of VT-1 receptors by a separate mechanism that acts additively to the TNF alpha- induced increase in VT-1 receptors.     

Patterns of vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 expression in rabbit and mouse atherosclerotic lesions and at sites predisposed to lesion formation.

The recruitment of mononuclear leukocytes and formation of intimal macrophage-rich lesions at specific sites of the arterial tree are key events in atherogenesis. Inducible endothelial cell adhesion molecules may participate in this process. In aortas of normal chow-fed wild-type mice and rabbits, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1), but not E-selectin, were expressed by endothelial cells in regions predisposed to atherosclerotic lesion formation. En face confocal microscopy of the mouse ascending aorta and proximal arch demonstrated that VCAM-1 expression was increased on the endothelial cell surface in lesion-prone areas. ICAM-1 expression extended into areas protected from lesion formation. Hypercholesterolemia induced atherosclerotic lesion formation in rabbits, LDL receptor and apolipoprotein E knockout mice, and Northern blot analysis demonstrated increased steady-state mRNA levels of VCAM-1 and ICAM-1, but not of E-selectin. Immunohistochemical staining revealed that VCAM-1 and ICAM-1 were expressed predominantly by endothelium in early lesions and by intimal cells in more advanced lesions. In early and advanced lesions, staining was most intense in endothelial cells at and adjacent to lesion borders. ICAM-1 staining extended into the uninvolved aorta. These expression patterns were highly reproducible in both species. The only difference was that VCAM-1 expression in endothelium over the central portions of lesions was found frequently in rabbits and rarely in mice. The expression of VCAM-1 by arterial endothelium in normal animals may represent a pathogenic mechanism or a phenotypic marker of predisposition to atherogenesis.     

The effects of dipyridamole stress test on plasma levels of soluble adhesion molecules intracellular adhesion molecule-1, vascular cell adhesion molecule-1, E-selectin and L-selectin in patients with ischemic heart disease and patients with syndrome X.

BACKGROUND: Adhesion of activated leukocytes to the endothelium as a result of myocardial ischemia/reperfusion has been shown to be involved in the development of tissue injury. Leukocyte adhesion to the endothelium occurs via adhesion molecules expressed on the surface of both cell types. Upon cell activation these proteins may be released into the circulation and measured in a soluble form. AIM: To verify whether the dipyridamole stress test, performed in patients with ischemic heart disease (IHD) and in patients with syndrome X, modifies plasma levels of the soluble adhesion molecules vascular cell adhesion molecule-1 (VCAM-1), intracellular adhesion molecule-1 (ICAM-1), E-selectin and L-selectin. METHODS: Plasma levels of the soluble endothelial adhesion molecules ICAM-1, VCAM-1 and E-selectin, as well as of the soluble leukocyte adhesion molecule L-selectin, were measured in venous blood samples taken before and 7 min after administration of dipyridamole in patients with IHD, patients with syndrome X and healthy individuals. Myocardial perfusion was evaluated using single photon emission tomography. The plasma levels of soluble VCAM-1, ICAM-1, E-selectin and L-selectin were all measured using enzyme-linked immunosorbent assays. RESULTS: After infusion of dipyridamole, plasma levels of ICAM-1 increased significantly in patients with IHD, whereas they remained unchanged in patients with syndrome X and in the control group. In patients with IHD, the initial plasma levels of VCAM-1, E-selectin and L-selectin, before administration of dipyridamole, were higher than those observed in patients with syndrome X and than those in the control group. Plasma levels of soluble VCAM-1, E-selectin and L-selectin decreased significantly in patients with IHD following the dipyridamole stress test, whereas they remained unchanged in patients with syndrome X, and in the control group. CONCLUSION: In patients with IHD, administration of dipyridamole induces myocardial ischemia resulting in modification of plasma levels of the soluble adhesion molecules.     

Effect of Tribulus Terrestris L. on Expression of ICAM-1, VCAM-1, E-Selectin and Proteome Profile of Human Endothelial Cells In-Vitro

Background: Atherosclerosis is a chronic inflammation that interferes with blood arteries functions due to the accumulation of low density lipids and cholesterol. Objective: To investigate the effect of aqueous extract and saponin fraction of Tribulus terrestris L. (TT) on the proteome and expression of intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin in the human umbilical vein endothelial cells (HUVEC) and human bone marrow endothelial cell (HBMEC) lines. Methods: Two cell lines were cultured and induced with lipopolysaccharide (LPS). The primed cells were then treated with aqueous extract and saponin fraction of TT. The protein profile of the endothelial cells was assessed under normal and LPS-induced conditions using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and 2D gel electrophoresis (2-DE). The levels of VCAM-1, ICAM-1, and E-selectin were estimated by use of western blotting. Results: LPS-induced HUVECs and HBMECs were shown to significantly increase the expression of ICAM-1, VCAM-1, and E-selectin in comparison to control groups. Our findings revealed that TT extract resulted in significantly more reduced levels of proteom (80 spots) as well as all the three mentioned proteins compared with the effect of saponin fraction alone. Conclusion: TT extract and its saponin fraction exerted anti-inflammatory effects on HUVEC and HBMEC lines and reduced the expression of ICAM-1, VCAM-1, and E-selectin. However, the anti-inflammatory effect of aqueous extract was greater than that of saponin fraction. Therefore, TT could be considered as a potential candidate for the treatment or prevention of atherosclerosis.     

Regulation of adhesion molecule expression by human synovial microvascular endothelial cells in vitro

Objective. To examine the in vitro expression of E-selectin, P-selectin, intercellular adhesion molecule 1 (ICAM-1), ICAM-2, vascular cell adhesion molecule 1 (VCAM-1), and platelet–endothelial cell adhesion molecule 1 (PECAM-1) by synovial microvascular endothelial cells (SMEC) in comparison with microvascular neonatal foreskin endothelial cells (FSE) and macrovascular human umbilical vein endothelial cells (HUVE). Methods. Cultured endothelial cells were treated for 4 hours with medium alone or tumor necrosis factor α (TNF α). The expression of endothelial adhesion molecules was evaluated by flow cytometry, cell enzyme-linked immunosorbent assay, and Northern blot analysis. Results. SMEC continuously expressed E-selectin under basal culture conditions, whereas FSE and HUVE did not. TNF α treatment of rheumatoid arthritis (RA) SMEC resulted in sustained peak expression of E-selectin for up to 24 hours, which subsequently declined but remained elevated even at 72 hours. In contrast, peak E-selectin expression in FSE and HUVE occurred between 4 hours and 16 hours after TNF α treatment and then declined to near basal levels by 24–48 hours. SMEC expressed significantly higher levels of ICAM-1 compared with HUVE under basal culture conditions. There was no difference between SMEC, FSE, and HUVE in the expression of P-selectin, VCAM-1, ICAM-2, or PECAM-1. Northern blot analysis demonstrated that the levels of E-selectin expression by TNF α-stimulated endothelial cells correlated with their respective messenger RNA levels. Conclusion. Regulation of E-selectin and ICAM-1 expression in RA synovial endothelium is different from that in neonatal foreskin and human umbilical vein endothelium. The augmented expression of adhesion molecules in RA synovial endothelium may facilitate the recruitment of leukocytes to this site.     

氧化低密度脂蛋白通过血凝素样氧化低密度脂蛋白受体1诱导血管内皮细胞粘附分子的表达

目的探讨血凝素样氧化低密度脂蛋白受体1（LOX-1）在氧化低密度脂蛋白（ox—LDL）诱导血管内皮细胞粘附分子表达中的作用。方法用不同浓度ox-LDL培养人脐静脉内皮细胞（HUVECs）,用RrealtimeRT—PCR测定LOX-1 mRNA的表达;RT—PCR测定细胞间粘附分子1（ICAM-1）、血管细胞粘附分子1（VCAM-1）以及E选择素的mRNA表达;用Westernblot测定LOX-1、ICAM-1、VCAM-1以及E选择素蛋白的表达,观察ox-LDL对血管内皮细胞粘附分子表达的影响。HUVECs预先用聚肌苷酸[poly（I）]和爱兰苔胶处理,再用ox—LDL培养,再分别测定上述粘附分子的表达水平,比较加入LOX-1阻断剂前后粘附分子表达的变化。结果ox-LDL各剂量组皆可上调LOX-1、ICAM-1、E选择素的mRNA和蛋白表达（P〈0．01）,在10～50μm/ml剂量范围内呈现明显的剂量一效应关系（P〈0．01）,但ox—LDL对VCAM-1的表达没影响。HUVECs预先与250μg／ml的poly（Ⅰ）或爱兰苔胶作用2h,然后加入50μg/ml的ox—LDL作用24h。poly（Ⅰ）和爱兰苔胶都能抑制ox—LDL诱导的LOX-1、ICAM-1和E选择素的mRNA和蛋白表达水平,与未阻断组相比,差异皆有统计学意义（P〈0．01）。结论ox—LDL可以上调血管内皮细胞粘附分子的表达,LOX-1受体阻断剂可以部分阻断ox—LDL的上调作用,ox-LDL诱导血管内皮细胞粘附分子的表过是通过LOX-1介导的。     

Toll样受体4/核因子κB和氧化低密度脂蛋白受体LOX-1对单核内皮细胞黏附的影响

目的近年研究表明介导先天免疫反应的受体Toll样受体4（TLR4）参与动脉粥样硬化的发生发展。TLR4激动后通过诱导核因子κB（NF—κB）激活,调控炎症因子的表达。研究观察TLR4／NF—κB激活对人脐静脉内皮细胞（HUVECs）氧化低密度脂蛋白受体（lectin—like oxidized lowdensity lipoprotein receptor-1,LOX-1）、细胞间黏附分子1（intercellular adhesion molecule-1,ICAM-1）、E-选择素（E—selectin）表达的调节,以及它们在介导单核内皮细胞黏附中的作用。方法应用脂多糖（LPS,1mg／L）刺激HUVECs 24h。采用RT—PCR方法检测TLR4、LOX-1、ICAM-1、E—selecfin mRNA表达水平;采用蛋白质印迹技术检测TLR4、LOX-1蛋白表达水平及核蛋白NF—κB p65表达的变化;采用直接计数法观察HUVECs与单核细胞的黏附率。结果LPS上调TLR4表达,激活NF—κB,上调HUVECs LOX-1、ICAM-1、E—selectin表达,增加单核细胞与内皮细胞的黏附率;抗LOX-1、ICAM-1、E—selectin抗体均部分抑制LPS介导的单核与内皮细胞黏附率的增加;NF—κB抑制剂一咖啡酸苯乙酯（CAPE）抑制了LPS介导的上述效应。结论TLR4／NF—κB激活上调LOX-1、ICAM-1、E—selectin表达,它们均在LPS介导的单核内皮细胞黏附过程中起部分作用,NF—κB在LPS介导的上述效应中起关键调节作用,干预NF—κB激活可能成为防治动脉粥样硬化的靶目标。     

Modulation by high glucose of adhesion molecule expression in cultured endothelial cells

Summary We evaluated the influence of high ambient glucose on cellular expression of adhesion molecules, known to mediate endothelial interaction of leucocytes and monocytes. Paired cultures of individual isolates of human umbilical vein endothelial cells (HUVECs) were studied by fluorescence activated cell sorter analysis after exposure to 30 vs 5 mmol/l glucose. Incubation of HUVECs for 24 h in 30 mmol/l glucose increased ICAM-1 (intercellular adhesion molecule-1; 116.4±16.9% of control, p 0.05), but not PECAM (platelet endothelial cell adhesion molecule) expression, compared to cultures kept in 5 mmol/l glucose. Long-term exposure (13±1 days) of HUVECs to 30 mmol/l glucose increased expression of ICAM-1 to 122.5±32.2% (p<0.002) and reduced that of PECAM to 86.9±21.3% vs the respective control culture in 5 mmol/l glucose (p<0.02). Stimulation of confluent HUVECs, kept in 30 vs 5 mmol/l glucose for 13±1 days, with 20 U/ ml interleukin-1 for 24 h (ICAM-1) and 4 h (endothelial leukocyte adhesion molecule 1) resulted in reduced ICAM-1 (84.8±27.0%, p<0.05) and endothelial leukocyte adhesion molecule-1 (87.6±22.4%, p<0.05) expression vs control cells, while that of PECAM (t: 24 h) and vascular cell adhesion molecule-1 (t: 16 h) remained unchanged. In conclusion, it appears that differences in expression of adhesion molecules on HUVECs in response to high glucose reflects endothelial glucose toxicity, which may also induce endothelial dysfunction in diabetes.Abbreviations HUVECs Human umbilical vein endothelial cells - ICAM-1 intercellular adhesion molecule-1 - PECAM platelet endothelial cell adhesion molecule - ELAM-1 endothelial leukocyte adhesion molecule-1 - VCAM-1 vascular cell adhesion molecule-1 - IL-1 interleukin-1 - FACS fluorescence activated cell sorter - FCS fetal calf serum - PBS phosphate buffered saline     

Cytokines in sickle cell disease

Sickle red cells express adhesion molecules including integrin alpha4beta1, CD36, band 3 protein, sulfated glycolipid, Lutheran protein, phosphatidylserine and integrin-associated protein. The proadhesive sickle cells may bind to endothelial cell P-selectin, E-selectin, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), CD36 and integrins leading to its activation. Monocytes also activate endothelium by releasing proinflammatory cytokines like tumor necrosis factor alpha (TNF-alpha) and interleukin 1beta (IL-1beta). Sickle monocytes also express increased surface CD11b and cytoplasmic cytokines TNFalpha and IL-1beta indicating activated state. Polymorphonuclear leukocytes (PMNs) are also activated with reduced L-selectin expression, enhanced CD64 expression and elevated levels of sL-selectin, sCD16 and elastase resulting in increased adhesiveness to the endothelium. Platelets are also activated and secrete thrombospondin (TSP) and cytokine IL-1. They also form platelet- monocytes aggregates causing endothelial cell P-selectin expression. Endothelial cell activation by these multiple mechanisms leads to a loss of vascular integrity, expression of leukocyte adhesion molecules, change in the surface phenotype from antithrombotic to prothrombotic, excessive cytokine production and upregulation of HLA molecules. Furthermore, contraction of these activated endothelial cells leads to exposure of extracellular matrix proteins, such as TSP, laminin, and fibronectin and their participation in adhesive interactions with bridging molecules from the plasma such as von Willebrand factor (vWf) released from endothelial cells, ultimately culminating in vasoocclusion and local tissue ischemia, the pathognomonic basis of vasoocclusive crisis.     

胰高血糖素样肽-1抗动脉粥样硬化的效果和机制研究

目的 探索胰高血糖素样肽（GLP）-1对氧化低密度脂蛋白（ox-LDL）诱导的人脐静脉内皮细胞（HUVECs）损伤的作用及机制。方法 将体外培养HUVECs分为对照组（无处理）和GLP-1预处理组（分别给予0、2.5、5和10 nmol/L GLP-1预处理24 h后,再以100 μg/ml ox-LDL氧化24 h）。应用MTT法检测细胞生存率,评估GLP-1在ox-LDL诱导HUVECs损伤中的作用;应用免疫荧光技术评估GLP-1在ox-LDL诱导单核细胞对HUVECs的黏附的影响;应用ELISA法检测HUVECs在接受ox-LDL和GLP-1处理后E-选择素,细胞间黏附分子（ICAM）-1、血管细胞黏附分子（VCAM）-1的分泌,并应用RT-PCR法检测E-选择素、ICAM-1、VCAM-1基因表达。结果 MTT试验显示（2.5、5和10 μmol/L）的GLP-1均可降低ox-LDL对HUVECs的杀伤作用,保护作用与浓度呈正相关（P<0.05）;免疫荧光检测结果显示,GLP-1可以降低单核细胞对ox-LDL损伤HUVECs的黏附作用,且和浓度呈正相关（P<0.05）。GLP-1处理后的ICAM-1、VCAM-1及E-选择素的表达和分泌均有显著下降（均有P<0.05）。结论 GLP-1可能通过对抗HUVECs氧化损伤,下调ICAM-1、VCAM-1和E-选择素的表达与分泌,减少单核细胞趋化和黏附,发挥抗动脉粥样硬化作用。     

Acute myeloid leukaemia blast cells bind to human endothelium in vitro utilizing E-selectin and vascular cell adhesion molecule-1 (VCAM-1)

Summary The adhesion of acute myeloid leukaemia (AML) blast cells to human umbilical vein endothelial cells (HUVECs) was investigated in vitro . Adhesion of blast cells from 10 cases of AML to unstimulated and interleukin-1β (IL-1) stimulated HUVECs was similar to or greater than that of control neutrophils. The extent to which endothelial E-selectin and vascular cell adhesion molecule-1 (VCAM-1) were involved in this adhesive process was investigated using blocking monoclonal antibodies to these proteins. In the majority of cases studied (7/8), anti-E-selectin significantly inhibited adhesion to IL-1 stimulated endothelium (26–65% inhibition) and in 5/8 cases so did anti-VCAM-1 (maximum of 31% inhibition). All cases were found to express the sialylated Lewis x antigen and very late activation antigen-4, ligands for E-selectin and VCAM-1 respectively. Our results indicate that leukaemic blast cells adhere to human endothelium and that there are E-selectin and, to a lesser extent. VCAM-1-dependent components to this process. Such adhesive interactions are likely to confer on AML blast cells the ability to migrate across the vascular wall and so to establish extravascular disease.