Glutathione oxidation in calcium- and p38 MAPK-dependent membrane blebbing of endothelial cells.

Under conditions where apoptosis is prevented, peroxides disrupt the endothelial monolayer by inducing cytoskeletal rearrangements, cell retraction and formation of arrays of membrane blebs. In human umbilical vein endothelial cells (HUVEC), the H(2)O(2)-induced membrane blebbing was found to be a transient process executed by two parallel signaling mechanisms: (i) mobilization of cytosolic [Ca(2+)](i) through a pathway requiring oxidation of reduced glutathione (GSH), and (ii) activation of p38 mitogen-activated protein kinases (MAPK) independently of GSH oxidation and Ca(2+) mobilization. In the HUVEC, membrane blebbing was thus blocked by inhibition of GSH oxidation, Ca(2+) mobilization or p38 MAPK activation.

Function of glutathione peroxidase in endothelial cell vitality.

The two human umbilical vein endothelial cell-derived lines, ECRF24 and ECV304, differ in responsiveness to oxidative stress. In confluent monolayers of ECRF24, but not in ECV304, peroxides induce stress responses such as plasma membrane blebbing and nuclear condensation. The peroxide effect on ECRF24 was preceded by oxidation of reduced glutathione (GSH) and of NAD(P)H, and by oxidation of the redox-sensitive probe, chloromethyl 2',7'-dichlorofluorescin (DCFH).

Entry of human cytomegalovirus into retinal pigment epithelial and endothelial cells by endocytosis.

Purpose: Human retinal pigment epithelial (RPE) cells and endothelial cells (HUVECs) are targets of human cytomegalovirus (HCMV) infection in vivo with significantly protracted replication in vitro compared with that in fibroblasts. This study analyzes the kinetics and mechanisms of HCMV entry into both cell types.

Methods: RPE cells were obtained from donor eyes.

Peroxide-induced membrane blebbing in endothelial cells associated with glutathione oxidation but not apoptosis.

Cells under oxidative stress induced by peroxides undergo functional and morphological changes, which often resemble those observed during apoptosis. Peroxides, however, also cause the oxidation of intracellular reduced glutathione (GSH). We investigated the relation between these peroxide-induced effects by using human umbilical vein endothelial cells (HUVEC) and two HUVEC-derived cell lines, ECRF24 and ECV304.

Nuclear import as a barrier to infection of human umbilical vein endothelial cells by human cytomegalovirus strain AD169.

Human embryonal fibroblasts (HEF) are fully permissive for infection by human cytomegalovirus (HCMV) strain AD169, whereas human umbilical vein endothelial cells (HUVEC) seem to form an almost complete barrier to infection with this virus. To investigate this difference in permissiveness, HCMV infection of both cell types was studied using in situ hybridisation (ISH) as well as immunocytochemistry to detect viral DNA and viral proteins. At 2 h post-infection (p.

In vitro tumor angiogenesis assays: plasminogen lysine binding site 1 inhibits in vitro tumor-induced angiogenesis.

It is generally accepted that tumors are angiogenesis-dependent. For research and clinical purposes it would be very attractive to have a simple in vitro model that allows a rapid screening of the angiogenic potential of tumors and to study the effect of angiogenic inhibitors. In vitro angiogenesis models were developed, based on endothelial sprouting/tube formation on a collagen gel, using both tumor cell lines and tumor biopsies.

Multiple organ involvement during experimental cytomegalovirus infection is associated with disseminated vascular pathology.

Since much of the pathogenesis of cytomegalovirus (CMV) disease is still unknown and vascular involvement may be of importance a rat model was used to study the nature and course of CMV-induced vascular pathology. In this model, local CMV infection was established by subcutaneous inoculation of rat-specific CMV (RCMV) in the sole of the foot. Sings of endothelial activation, including leucocyte adhesion, preceded detectable RCMV infection of these cells.

Cellular arrangement of human breast cancer cell lines determines hemostatic parameters.

Two in vitro models are compared to investigate whether cellular configuration or composition of the matrix in which the cells are cultured influences growth and/or prognostic parameters. T47D, MCF-7 and Hs578T breast cancer cell lines were cultured on two different matrices (agarose and collagen). Growth curves, biological markers (Ki-67, p53 and bcl-2) and the expression of hemostatic parameters were studied.

Effect of membrane-permeable sulfhydryl reagents and depletion of glutathione on calcium mobilisation in human platelets.

Exposure to peroxides is known to increase the sensitivity of platelets towards activation by agonists. Similar platelet-activating effects are induced by sulfhydryl reagents that evoke Ca2+-induced Ca2+ release (CICR) by stimulating the Ca2+-releasing property of the inositol-1,4,5-trisphosphate receptor. We questioned whether these compounds may act by mobilising intracellular calcium in platelets by altering the intracellular glutathione redox state.

Activation of protein kinase C enhances the infection of endothelial cells by human cytomegalovirus.

The infection of cultured endothelial cells with human cytomegalovirus (HCMV) is generally limited to less than 10% of the cells in contrast to HCMV infection of fibroblasts, where essentially all cells can be infected. It is known that HCMV infection influences a number of signal transduction pathways of infected cells. We therefore questioned whether, conversely, the infectivity of human umbilical vein endothelial cells could be influenced by the deliberate activation of these pathways.

Intracellular thiol redox status affects rat cytomegalovirus infection of vascular cells.

There is increasing evidence for cytomegalovirus (CMV) induced vascular pathology during acute infection in the immunocompromised host. Inflammation is involved in such processes, which is frequently associated with increased levels of oxidative mediators and reduced anti-oxidant protection. A relation between viral infection and oxidative stress has been recognized for human immunodeficiency virus and herpes simplex virus-1 infections, but little is known in this respect for CMV infections.

A dual role for endothelial cells in cytomegalovirus infection? A study of cytomegalovirus infection in a series of rat endothelial cell lines.

Several clinical findings point to the involvement of microvascular endothelial cells in cytomegalovirus-related pathology. In this study the interactions of cytomegalovirus (CMV) with microvascular endothelial cells was investigated in an in vitro rat model. A series of rat endothelial cell lines, considered representative for the heterogeneity of heart microvascular endothelium in vivo, were infected with rat CMV (RCMV).

Tumour angiogenesis: pathophysiology and clinical significance.

In the 1970s it was first proposed that tumours depended on the establishment of a microcirculation in order to grow beyond a few millimetres. Thereafter, the search to prove this hypothesis increased strongly and by the end of the 1980s, evidence was given that tumours were angiogenesis-dependent and metastatic cells were only shed after the tumour had established its microcirculation. The process of neovascularization is regulated by numerous growth factors, vascular endothelial cells, and matrix proteins released from host stromal cells such as macrophages and mast cells.

Polyunsaturated fatty acids and function of platelets and endothelial cells.

Fish oil diets, rich in n-3 polyunsaturated fatty acids, are considered to have an antithrombotic effect. Both platelets and endothelial cells play a crucial role in the regulation of thrombosis and haemostasis. There is substantial evidence that, in these cells, fish oil-derived polyunsaturated fatty acids can replace the n-6 polyunsaturated fatty acid, arachidonic acid, in the membrane phospholipids and can modify those cellular reactions in which the latter fatty acid participates.

Cytomegalovirus infection and vessel wall pathology.

This review focuses on information regarding the cytomegalovirus (CMV) in relation to vessel wall pathology including clinical symptoms, pathogenesis and latency. Evidence obtained for involvement of CMV in vascular pathology will be summarized. CMV infection of vascular cells induces cell activation, which leads to expression of adhesion proteins, MHC molecules, cytokine receptors and the production of cytokines and growth factors.

Differential effects of endothelial cell fatty acid modification on the sensitivity of their membrane phospholipids to peroxidation.

In order to study the relationship between the fatty acid (FA) composition of human umbilical vein endothelial cells (HUVEC) and their susceptibility to oxidative stress, we modified their FA composition by long-term culturing in media supplemented with a saturated, monounsaturated, or polyunsaturated FA. Sensitivity of the cellular phospholipids to peroxidation was monitored by measuring conjugated diene formation and decrease of polyunsaturated FAs induced by CuSO4 and H2O2 in liposomes prepared from the respective phospholipid extracts. The extent of phospholipid peroxidation was found to increase with increasing content of polyunsaturated FAs.

Membrane related effects in endothelial cells induced by human cytomegalovirus.

Previously, we have reported on the increase in procoagulant activity of human umbilical vein endothelial cells (HUVEC) after infection with human cytomegalovirus (HCMV). When using microvascular endothelial cells from foreskin (MVEC), we also observe a significant increase in membrane perturbation and a concomittant increase in procoagulant activity. This effect is both observed with a laboratory HCMV strain (AD169) with low pathogenicity for endothelium and a HUVEC adapted strain (VHL-E) that readily infects endothelial cells.

Tumor spheroid-induced vesicle formation on endothelial cells is associated with procoagulant properties.

Fibrin deposits in tumor beds are an intriguing phenomenon. It has been suggested that fibrin plays a role as a provisional matrix in which the tumor grows and induces development of a vascular network. On the other hand fibrin possibly protects the tumor nodule from host defense mechanisms.

Continuous monitoring of lipid peroxidation by measuring conjugated diene formation in an aqueous liposome suspension.

A method is described for the direct and continuous monitoring of lipid peroxidation in an aqueous suspension of sonicated liposomes. By means of ultraviolet difference spectroscopy using tandem cuvettes, the formation of conjugated dienes during liposome peroxidation can be followed. Using this technique, the effect of the fatty acid composition of liposomes on lipid peroxidation can be studied.

Long-term fatty acid modification of endothelial cells: implications for arachidonic acid distribution in phospholipid classes.

Human umbilical vein endothelial cells were cultured in various fatty acid-modified media until equilibrium conditions were reached (7-8 days). The effects on the fatty acid composition of phospholipid classes and on the metabolism of arachidonic acid (20:4(n-6)) were studied. The results showed that in every phospholipid class large changes in fatty acid composition, including 20:4(n-6) content, were induced by long-term modification with unsaturated as well as saturated fatty acids.

The procoagulant response of cytomegalovirus infected endothelial cells.

The report describes the effect of an in vitro infection of human umbilical vein endothelial cells with human Cytomegalovirus (CMV). The parameters studied are cellular procoagulant activity, secretion of plasminogen activator inhibitor (PAI-1) and urokinase-type plasminogen activator (u-PA), activation and internalization of factor X and Merocyanine 540 staining. The infection does not result in an increase in PAI-1 and u-PA secretion, but it brings about a procoagulant response, which is relatively rapid compared to the tissue factor mediated response induced by inflammatory mediators.

The effect of viral infection on eicosanoid formation and procoagulant activity of rat peritoneal macrophages. Role of the dietary fat type.

The effect of dietary manipulation on eicosanoid formation in rat peritoneal macrophages was studied in relation to some of their effector functions: cellular procoagulant activity, production of reactive oxygen species (measured as chemiluminescence), and phagocytosis of antibody-coated erythrocytes. Rats were fed adequate diets for eight weeks containing mackerel oil (MO), sunflowerseed oil (SO) or hydrogenated coconut oil (HCO). The release of eicosanoids from resident macrophages stimulated by opsonized zymosan was significantly lower for the MO group as compared to the other dietary groups.

Membrane fatty acid composition and endothelial cell functional properties.

In order to study the influence of endothelial cell fatty acid composition on various membrane related parameters, several in vitro methods were developed for manipulating the fatty acid content of human endothelial cell membranes. Changes in membrane fatty acid profile were induced by using fatty acid modified lipoproteins or free fatty acids. The largest changes in endothelial fatty acid composition were obtained by culturing the cells in media supplemented with specific free fatty acids.

The effect of virus infection on the adherence of leukocytes or platelets to endothelial cells.

It has been reported that atherosclerotic lesions contain genomic material belonging to members of the herpes family. This suggests that latent viral infection may be one of the atherogenic triggers. In this study we show that early infection of endothelial cell monolayers with Herpes Simplex virus type 1 (HSV-1) or Cytomegalovirus (CMV) results in an increased monocyte (MC) and polymorphonuclear leukocyte (PMN) adherence, but not in an increased platelet adhesion.