Vascular Endothelial Cells In Culture Research Articles

Disaccharide Composition of Glycosaminoglycan Chains in Growing Vascular Endothelial Cells in Culture after Exposure to Lead

Heparan sulfate (HS) chains bind and activate fibroblast growth factor-2 (FGF-2) depending on their microstructure. The glycosaminoglycan chains present particularly as a large HS proteoglycan perlecan in vascular endothelial cells. A heavy metal lead induces a lower response to FGF-2 by inhibition of perlecan synthesis and inhibits the proliferation of the cells. The present study was undertaken to address the question whether lead influences the formation of HS chains in vascular endothelial cells. The data indicate that lead decreases the amount of disaccharide units including hexuronic acid-N-acetylglucosamine, hexuronic acid-N-sulfated glucosamine and 2-O-sulfated hexuronic acid-N-sulfated glucosamine in HS chains accumulated in the cell layer and the conditioned medium. However, no percentage of any disaccharide unit was affected by the metal. The percentage of disaccharide units in chondroitin/dermatan sulfate was also unaffected by lead. The present data support the hypothesis that lead-induced lower response of vascular endothelial cells to endogenous FGF-2 mainly results from a decrease in perlecan molecules of extracellular matrix as a result of selective inhibition of perlecan core protein synthesis rather than a change in the HS structure.

Neutrophil elastase in human atherosclerotic plaques: production by macrophages.

Catabolism of the extracellular matrix (ECM) contributes to vascular remodeling in health and disease. Although metalloenzymes and cysteinyl proteinases have garnered much attention in this regard, the role of serine-dependent proteinases in vascular ECM degradation during atherogenesis remains unknown. We recently discovered the presence of the metalloproteinase MMP-8, traditionally associated only with neutrophils, in atheroma-related cells. Human neutrophil elastase (NE) plays a critical role in lung disease, but the paucity of neutrophils in the atheromatous plaque has led to neglect of its potential role in vascular biology. NE can digest elastin, fibrillar and nonfibrillar collagens, and other ECM components in addition to its ability to modify lipoproteins and modulate cytokine and MMP activity. Fibrous and atheromatous plaques but not normal arteries contained NE. In particular, NE abounded in the macrophage-rich shoulders of atheromatous plaques with histological features of vulnerability. Neutrophil elastase and macrophages colocalized in such vulnerable plaques (n=7). In situ hybridization revealed NE mRNA in macrophage-rich areas, indicating local production of this enzyme. Freshly isolated blood monocytes, monocyte-derived macrophages, and vascular endothelial cells in culture produced active NE and contained NE mRNA. Monocytes produced NE constitutively, with little regulation by cytokines IL-1beta, TNF-alpha, or IFN-gamma but released it when stimulated by CD40 ligand, a cytokine found in atheroma. These findings point to a novel role for the serine protease, neutrophil elastase, in matrix breakdown by macrophages, a critical process in adaptive remodeling of vessels and in the pathogenesis of arterial diseases.

Bacterial lipopolysaccharide induces retinoic acid-inducible gene-I in vascular endothelial cells

We found that retinoic acid-inducible gene-I (RIG-I) is induced in cultures of vascular endothelial cells stimulated with bacterial lipopolysaccharide (LPS). RIG-I encodes a protein belonging to the DExH box family that has diverse roles in the regulation of gene expression and cellular functions. Overexpression of RIG-I resulted in upregulation of cyclooxygenase-2 (COX-2) expression. RIG-I may have important roles in vascular pathology through its activity in regulating the COX-2 gene.

The effect of contrast material on transitional cell carcinoma viability.

In this study, increasing contrast concentration and/or exposure time decreased the growth and adherence of transitional cell carcinoma cells. Similarly, several authors have shown that the toxicity of contrast material to vascular endothelial cells in culture is related to concentration and time of exposure; this toxicity is minimized if non-ionic, low osmolar contrast is utilized. Identical trends of increasing toxicity with increasing osmolality have been documented with kidney cell lines, vascular smooth muscle cells, myocardial cells, neural cells, human fibroblasts, rat mesenchymal cells, and rat hepatocytes. The toxicity of contrast material in malignant cells (human cervical carcinoma cell lines and prostate cancer cell lines) has also been documented. Factors other than direct cellular toxicity may also affect the ability of malignant transitional cells transported with contrast material to implant on other sites in the collecting system. Zhan et al have evaluated neutrophil adhesion to contrast-exposed vascular endothelial cells. These authors observed increasing cellular adhesion with decreasing contrast dose. This was not related to osmolality as there was no similar endothelial adhesion activation with equal osmolality solutions of non-contrast solutions. The authors felt that contrast material, therefore, caused some form of modulation of cell adhesion molecules. This theory is supported by research by Owens et al that shows increases in the adhesive properties of endothelial cells to platelets after contrast exposure, and Rasmussen et al who found decreased granulocyte adherence with contrast exposure. In our study, transitional cell carcinoma cultures were placed on a collagen substrate to encourage adherence. There was an increasing number of non-adherent cells with increasing contrast concentration and exposure time, which suggests an effect of the contrast on cellular adhesion as well (Table 1). Potier et al found that rat mesangial cells were more sensitive to the cytotoxic effects of contrast agents than the less differentiated human fibroblasts were. This might suggest that the more poorly-differentiated transitional cell carcinoma cells would not be as sensitive to contrast material as low-grade tumors. However, we did not observe any differences in the responses of high- versus low-grade transitional cell tumors to exposure to contrast material. Intravenous contrast administration can lead to multiple adverse reactions, such as cardiotoxicy, nephrotoxicy, and hemodynamic toxicity. This makes non-ionic, low-osmolality agents preferable to ionic, high-osmolality agents. However, direct introduction of material into the urinary collecting system for the evaluation of transitional cell carcinoma may be less prone to seeding of viable malignant cells if ionic, high-osmolar contrast agents are utilized. Additional considerations include potential masking of very small filling defects in the upper urinary tract through opacification with extremely concentrated contrast material. Also, cytologic abnormalities are worsened by exposure to ionic, high-osmolar contrast agents, thus, any specimens for cytologic analysis should be collected prior to utilization of any contrast agents.

Effects of tibolone on human breast cancer cells and human vascular coronary cells.

Tibolone is a synthetic progestin with estrogenic and progestogenic properties, used to alleviate menopausal syndromes and for osteoporosis prophylaxis in postmenopausal women. However, only little data are available on tibolone and breast cancer risk and on the effects of tibolone on the cardiovascular system. Therefore, in the present in vitro study, we investigated the effect of tibolone on the growth of the human breast cancer cell line, MCF-7, and on the direct effects of tibolone on the vasculature, i.e. human female coronary endothelial and smooth muscle cells. In the breast cancer cell experiments, tibolone was examined alone and in the presence of 0.1 nM estradiol in the concentration range from 0.001 microM to 1 microM. Tibolone lead to significant cell growth in the concentration range of 0.01 micro M to 1 microM and was not able to inhibit estradiol-induced proliferation at the concentrations of 0.01 microM and 0.1 microM. In the vascular endothelial cell culture experiments, tibolone was tested at the concentrations 0.1 microM, 1 microM and 10 microM. Tibolone reduced the synthesis of endothelin as well as the concentrations of E-selectin, PAI-1 and pro-MMP-1. The magnitude of the effects on these markers varied and was in the range of 11%-42%. Concerning smooth muscle cells, tibolone elicited no changes in the proliferation compared to control values. These data suggest that tibolone does have tumour cell-growth promoting effects in vitro. However, tibolone can positively influence the synthesis of markers in cell cultures of human female coronary artery, which modulate vascular tone and which play a decisive role in the various stages of atherosclerosis. Drawing a clinical consequence from our experiments would result in not recommending the use of tibolone in postmenopausal women at high risk for breast cancer development until long-term controlled clinical studies have been performed on the effect of tibolone administration and breast cancer risk. Experimental studies, such as the present one, are useful to explore mechanisms, but clearly cannot replace clinical studies.

Inhibition by acharan sulphate of angiogenesis in experimental inflammation models.

1. The effects of acharan sulphate, a glycosaminoglycan isolated from the giant African snail Achatina fulica, on angiogenesis in the granulation tissue were analysed using an air pouch-type carrageenin-induced inflammation model in rats and a cotton thread-induced inflammation model in mice. 2. In the carrageenin-induced inflammation model in rats, intra-pouch injections of acharan sulphate (5 and 50 micro g) inhibited the pouch fluid accumulation and the granulation tissue formation as well as the angiogenesis in the granulation tissue at day 6 in a dose-dependent manner. 3. The inhibitory effects of acharan sulphate at 50 micro g on the pouch fluid accumulation and the leucocyte infiltration into the pouch fluid was not so effective as that of the cyclo-oxygenase inhibitor indomethacin at 100 micro g, but the inhibitory effects of acharan sulphate at 50 micro g on the granulation tissue formation and angiogenesis in the granulation tissue were almost the same as those of indomethacin at 100 micro g. 4. Acharan sulphate did not affect levels of vascular endothelial growth factor (VEGF) in the granulation tissue and in the pouch fluid at day 6, but indomethacin significantly lowered them. 5. In the cotton thread-induced inflammation model in mice, injections of acharan sulphate (10 micro g) at the site of the cotton thread implantation inhibited the granulation tissue formation and angiogenesis as indomethacin (20 micro g) did. Acharan sulphate (10 micro g) did not affect levels of VEGF in the cotton thread-induced granulation tissue at day 5, but indomethacin (20 micro g) significantly lowered them. 6. In culture of human vascular endothelial cells, acharan sulphate at 10 and 100 micro g ml(-1) inhibited VEGF-induced capillary tube formation. 7. These findings suggest that the inhibitory effect of acharan sulphate on angiogenesis in carrageenin- and cotton thread-induced granulation tissues is not due to the inhibition of VEGF protein induction, but is due to the inhibition of VEGF-induced vascular tube formation.

Tilapia mast cell lysates enhance neutrophil adhesion to cultured vascular endothelial cells

The possible role of fish mast cells in regulating neutrophil adhesion to vascular endothelial cells was studied using primary cultures of tilapia vascular endothelial cells. The endothelial cell monolayer, which was cultured in 96 well plates, was stimulated for appropriate periods with tilapia mast cell (tMC)-lysates or with Leibovitz-15 (L-15) medium, as a control, and peripheral neutrophils were added into each well after removal of the lysates. After 30min incubation, cells in the wells were fixed with formalin and non-adherent neutrophils were removed. The cells were stained with Giemsa and neutrophil adhesion was observed microscopically. Although some neutrophils attached to the endothelial cells without stimulation, neutrophil adhesion was enhanced after the incubation of the endothelial cells with tMC-lysates. Neutrophil adhesion was maximal 6h after the lysate stimulation, with a six-fold increase compared to the control. Neutrophil adhesion also increased when the endothelial cells were stimulated with neutrophil lysates, lipopolysaccharide and zymosan-treated tilapia sera. These results indicate that fish vascular endothelial cells express some neutrophil adhesion molecule(s) after stimulation with various substances.

Isoflurane Pretreatment Inhibits Cytokine-induced Cell Death in Cultured Rat Smooth Muscle Cells and Human Endothelial Cells

Anesthetics are protective during ischemic-reperfusion injury and associated inflammation; therefore, the authors hypothesized that anesthetic pretreatment may provide protection in culture from cytokine-induced cell death. Rat vascular smooth muscle (VSM) cell and human umbilical vascular endothelial cell (HUVEC) cultures were used to determine whether pretreatment with 30 min of isoflurane decreases cell death from tumor necrosis factor alpha (TNF-alpha), interleukin 1 (IL-1 beta), and interferon (IFN-gamma) alone or in combination. Cell survival and viability were determined by trypan blue staining and cell proliferation assay, as well as by DNA fragmentation assays. The roles of protein kinase C (PKC) and adenosine triphosphate-sensitive potassium (K(ATP)) channels in mediating isoflurane (and halothane) protection were evaluated with the antagonists staurosporine or glibenclamide in cytokine- and also hydrogen peroxide (H(2)O(2))-induced cell death. Pretreatment with 1.5% isoflurane immediately prior to cytokine exposure increased cell survival and viability from cytokines by 10-60% for 24, 48, 72, and 96 h in VSMs and up to 72 h in HUVECs. DNA fragmentation (TUNEL) was also attenuated by isoflurane. Isoflurane was equally effective in VSMs at 0.75, 1.5, and 2.5%, whereas in HUVECs, 1.5 and 2.5% were more effective than 0.75%. In VSMs, isoflurane administered 1 h prior to or simultaneously with cytokines was also effective, whereas isoflurane 2 h prior to cytokines was less effective, and either 4 h prior to or 30 min after cytokines was not effective. In both cytokine- and H(2)O(2)-induced cell death, isoflurane and halothane pretreatment were equally protective, and staurosporine and glibenclamide attenuated the protective effect. Thirty minutes of isoflurane attenuates cytokine-induced cell death and increases cell viability in VSMs for 96 h and in HUVECs for 72 h. Isoflurane must be administered less than 2 h prior to or simultaneously with the cytokines to be protective. These initial inhibitor studies suggest involvement of PKC and K(ATP) channels in isoflurane and halothane protection against both cytokine- and H(2)O(2)-induced cell death of VSMs and HUVECs.

Human Vascular Endothelial Cells Express Pattern Recognition Receptors for Fungal Glucans Which Stimulates Nuclear Factor κB Activation and Interleukin 8 Production

Fungal cell wall glucans nonspecifically stimulate various aspects of innate immunity via interaction with membrane receptors on macrophages, neutrophils, and natural killer cells. We investigated the binding of water-soluble glucans in primary cultures of normal human coronary or dermal vascular endothelial cells (VECs). Membranes from VECs exhibited saturable binding. Competition studies demonstrated the presence of at least two glucan binding sites on VECs. Glucan phosphate competed for all binding sites with a KDof 3.7 μm for coronary VECs and 11 μm for dermal VECs, respectively. Laminarin, a low molecular weight glucan, competed for 47 to 51 per cent of binding ( KD= 2.8–2.9 μm), indicating the presence of at least two binding sites. Glucan (1 μg/mL) stimulated VEC nuclear factor κB nuclear binding activity and Interleukin 8 expression—but not that of vascular endothelial growth factor—in a time-dependent manner. This is the first report of pattern recognition receptors for glucan on human VECs. It also provides the first evidence that glucans can directly modulate the functional activity of VECs by stimulating cytokine gene. These results provide new insights into the mechanisms by which the host recognizes and responds to fungal cell wall products and suggests that the response to glucans may not be confined to leukocytes.

Differential roles of ICAM-1 and E-selectin in polymorphonuclear leukocyte-induced angiogenesis

Ets-1, which stimulates metalloproteinase gene transcription, has a key role in angiogenesis. We first examined whether activated polymorphonuclear leukocytes (PMNs) enhanced angiogenesis through the induction of Ets-1. Addition of activated PMNs to endothelial cells stimulated both in vitro angiogenesis in collagen gel and Ets-1 expression. Both angiogenesis and Ets-1 expression induced by PMNs were reduced by ets-1 antisense oligonucleotide, suggesting that Ets-1 is an important factor in PMN-induced angiogenesis. Although intercellular adhesion molecule (ICAM)-1 and E-selectin are involved in PMN-induced angiogenesis, the mechanisms underlying their roles in angiogenesis have yet to be elucidated. PMN-induced Ets-1 expression was reduced by a monoclonal antibody against ICAM-1 but not E-selectin despite the inhibition of PMN-induced angiogenesis by both antibodies. Moreover, the stimulation of angiogenesis by H(2)O(2) without PMNs was inhibited by a monoclonal antibody to E-selectin but not ICAM-1. These findings suggested that ICAM-1 in endothelial cells may act as a signaling receptor to induce Ets-1 expression, whereas E-selectin seems to function in the formation of tubelike structures in vascular endothelial cell cultures.

Modulation of Insulin-like Growth Factor-I Production of Cultured Retinal Vascular Endothelial Cells by Oxygen, Glucose and Growth Hormone

Purpose: To evaluate the interaction of oxygen, glucose, and growth hormone (GH) on insulin-like growth factor-I (IGF-I) production of cultured bovine retinal vascular endothelial cells. Methods: Confluent cultures of bovine retinal vascular endothelial cells were incubated under 3% or 20% oxygen in Dulbecco's modified Eagle's medium (containing 1 g/L or 4.5 g/L glucose), with or without the addition of 10 ng/mL GH. After incubation times of 0, 24, 48, and 72 hours, IGF-I was measured in the supernatant, and cells were counted. Results: Highest levels of IGF-I were reached after 72 hours with 4.5 g/L glucose and 10 ng/mL GH under 3% O 2. All IGF-I levels found in 3% O 2 samples were significantly higher than those found in 20% O 2 samples. No statistical significance was found regarding glucose concentration or GH supplement. Conclusions: Oxygen turned out to be the sole modulating factor for IGF-I production of cultured retinal vascular endothelial cells.

Shear-Induced Changes in Endothelin-1 Secretion of Microvascular Endothelial Cells

Human glomerular microvascular endothelial cell (HGMEC) culture monolayers were maintained in static culture as controls or subjected to steady laminar shear stress of 0.5, 1.0, or 1.5 N/m2. Over 25 h of shear, the cumulative secretion of ET-1 was 705.4 pg/cm2 in the control, 820.7 pg/cm2 at 0.5 N/m2, 1063.2 pg/cm2 at 1.0 N/m2, and 644.7 pg/cm2 at 1.5 N/m2. The average ET-1 secretion rate for the HGMEC monolayers exposed to 0.5, 1.0, or 1.5 N/m2 of shear stress was 32.83 ± 2.01 pg/cm2·h, 42.53 ± 3.74 pg/cm2·h, and 25.79 ± 1.29 pg/cm2·h, respectively. The average ET-1 secretion rate of the static controls was 28.22 ± 3.11 pg/cm2·h. The results showed that low shear stress (0.5 N/m2) elevated and high shear stress (1.5 N/m2) suppressed secretion of ET-1, while an intermediate level of shear stress (1.0 N/m2) led to the maximum secretion of ET-1, and furthermore, ET-1 secretion varied with the duration of shear in a nonlinear fashion, and the logistic equations may be used to describe relationship between the duration of shear and the ET-1 secretion. The major secretion period of ET-1 occurred between 5.3 and 22.3 h, with the peak secretion rate occurring at approximately 10.7–15.2 h. Our findings showed also that the major secretion period and peak secretion rate of HGMECs varied with the level of shear stress. Thus, the response of cultured human microvascular endothelial cells to shear stress differed from that of large-vessel endothelial cell cultures in terms of ET-1 secretion. In addition to the level of shear stress, the duration of shear is an important determinant in ET-1 secretion. Consequently, the heterogeneity of vascular endothelial cells and the duration of shear should both be considered in future research on the secretion of vascular endothelial cell cultures.

Induction of Plasminogen Activator Inhibitor Type 1 Synthesis by Cadmium in Human Vascular Endothelial Cells in Culture.

Cadmium is a unique heavy metal that stimulates the secretion of plasminogen activator inhibitor type 1 (PAI-1) from vascular endothelial cells. However, it has been incompletely understood whether cadmium stimulation of PAI-1 secretion actually results in a reduction of endothelial fibrinolytic activity and whether the stimulation results from an induction of endothelial PAI-1 synthesis. To address these questions, human umbilical vein endothelial cells were cultured with cadmium chloride in the presence or absence of actinomycin D, H-7 or HA1004. The activity of tissue type and urokinase type plasminogen activators (t-PA and u-PA, respectively) in the conditioned medium was analyzed by fibrin zymography and mRNAs coding t-PA, u-PA and PAI-1 were determined by quantitative reverse transcription-polymerase chain reaction. Results of the experiments indicate that cadmium reduces the activity of both t-PA and u-PA in vascular endothelial cells through induction of PAI-1 synthesis which is mediated by protein kinase C activation. Since the PAI-1 induction by cadmium was observed in neither human vascular smooth muscle cells nor human fibroblasts, it was suggested that vascular endothelial cells are a particular cell type of which PAI-1 synthesis is stimulated by cadmium.

Further evidence for direct vascular actions of statins: effect on endothelial nitric oxide synthase and adhesion molecules.

The effect of the synthetic statin fluvastatin was investigated on the concentrations of endothelial nitric oxide synthase (eNOS) and of soluble adhesion molecules in human vascular endothelial cell cultures. Fluvastatin was able to increase dose-dependently eNOS concentration. In addition, fluvastatin reduced concentrations of E-Selectin and ICAM-1. These results further support the evidence that statins elicit important effects on the vascular endothelium which may contribute to the reduction of morbidity and mortality of cardiovascular diseases.

Mechanical brush-catheter abrasion method for the isolation and culture of human umbilical vein endothelial cells. First in vitro results

Mechanical brush-catheter abrasion method for the isolation and culture of human umbilical vein endothelial cells. First in vitro results. To determine the value of a nylon brush (Cragg thrombolytic catheter system) with regard to the isolation and culture of viable human vascular umbilical vein endothelial cells (HUVEC). Fresh human umbilical cords were collected after a maximum of 6 - 8 h post delivery. They were stored in sterile tubes till further preparation. After predilation the umbilical cord veins, a 6 French brush catheter was inserted, expanded, and removed by hand using rotating movements. The nylon brush was shaken out in cell medium containing serum proteins by a connected motor drive. After repeating these procedures, the detached endothelial cells were centrifugated and resuspended. The cell numbers were assessed and cell viability controlled. The endothelial cell (EC) viability depended on the umbilical cord's age and was the highest (29.8 +/- 11.4 %) within 3 h after delivery. The brush abrasion (3.32 x 10(5) +/- 52016 viable EC/10 cm vein segment) enabled us to culture these cells under different cell concentrations. They grew to confluence after 2 - 3 weeks. The brush-catheter abrasion is an effective method for collecting viable human endothelial cells in vitro.

S-nitrosothiols do not induce oxidative stress, contrary to other nitric oxide donors, in cultures of vascular endothelial or smooth muscle cells

Nitric oxide (NO) has been described to exert various anti-atherogenic actions. However, NO, in some cases, has been shown to stimulate the oxidation of low-density lipoprotein (LDL), which constitute an important triggering event in atherosclerosis. Thus, some NO donors, despite their advantages, might also induce oxidative stress. Therefore, the purpose of this study is to examine the effect of three different NO donors on LDL oxidation, in acellular system as well as in cultures of normal endothelial cells or smooth muscle cells, which constitute the two major cellular components of the arterial wall. Sodium nitroprusside oxidized strongly LDL in medium alone as well as in endothelial or smooth muscle cell cultures. Sydnonimine-1 (SIN-1) oxidized LDL already in the absence of cells and enhanced clearly the LDL oxidation in the cultures. S-nitroso- N-acetylpenicillamine was unable to oxidize LDL in synthetic medium alone as well as in the presence of cells, showing that the amount of superoxide and other reactive oxygen species released by these cells did not suffice, contrary to those liberated by macrophages, to combine to NO providing oxidant activity.

Transforming Growth Factor-β Regulation of Endothelin Expression in Rat Vascular Cell and Organ Cultures

Transforming growth factor (TGF)-beta increases the production of the vasoactive peptide endothelin (ET) in cultures of vascular endothelial cells (EC) and vascular smooth muscle cells (VSMC), but the physiologic or pathologic significance of this regulation has not been determined. The present studies test the hypothesis that when EC and VSMC are in direct contact or close proximity, ET expression is, at least in part, dependent on TGF-beta. The effects of TGF-beta on ET-1 mRNA (Northern analysis and reverse transcription polymerase chain reaction) and peptide (radioimmunoassay) levels were assessed in rat EC and VSMC and vascular organ cultures. TGF-beta2 (1 ng/ml) increased ET-1 mRNA in VSMC and EC plus VSMC cultures and increased ET-1 peptide in EC, VSMC, and EC plus VSMC cultures. TGF-beta2 also increased ET-1 mRNA and peptide in vascular organ cultures. Antibodies that neutralized the activities of TGF-beta1 and TGF-beta2 decreased ET-1 mRNA in EC plus VSMC cultures and in vascular organ cultures. These data indicate that when EC and VSMC are in direct contact or close proximity, TGF-beta increases ET expression and active TGF-beta is present and promotes ET expression. These data suggest that TGF-beta is a determinant of vascular ET expression in vivo, and that TGF-beta regulation of ET expression would affect cardiovascular function in health and disease.

Construction of cell-compatible layer and culture of human umbilical vascular endothelial cells on porous polystyrene membranes

Porous polystyrene (PS) membranes with pore sizes ranged from nanometer scale to several decades' micrometers were fabricated by the thermally induced phase-separation technique, and were characterized by scanning electron microscopy (SEM). Oxidation of PS membranes by concentrated sulfuric acid induced a decrease of the carbon amount on the membrane surfaces accompanying with an increase of the oxygen amount. The hydrophilic surfaces thus constructed proved quite effective to promote endothelial cell adhesion and growth. After culturing for 4 days, a confluent endothelial cells (ECs) layer was observed on nonporous PS membrane treated previously with sulfuric acid for 15 min at 28.5°C. The observation under SEM showed that ECs on the PS membrane surfaces at the fourth day were inversely proportional to the pore size: the smaller the pore size, the larger the amount of ECs on the membrane surfaces. It is supposed that the existence of pores on the membrane surface might generate some disadvantageous resistance to the spreading of cells. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 81: 3523–3529, 2001

Tumor angiogenesis: past, present and the near future.

The concept of treating solid tumors by inhibiting tumor angiogenesis was first articulated almost 30 years ago. For the next 10 years it attracted little scientific interest. This situation changed, relatively slowly, over the succeeding decade with the discovery of the first pro-angiogenic molecules such as basic fibroblast growth factor and vascular endothelial growth factor (VEGF), and the development of methods of successfully growing vascular endothelial cells in culture as well as in vivo assays of angiogenesis. However, the 1990s have witnessed a striking change in both attitude and interest in tumor angiogenesis and anti-angiogenic drug development, to the point where a remarkably diverse group of over 24 such drugs is currently undergoing evaluation in phase I, II or III clinical trials. In this review I will discuss the many reasons for this. These features, together with other recent discoveries have created intense interest in initiating and expanding anti-angiogenic drug discovery programs in both academia and industry, and the testing of such newly developed drugs, either alone, or in various combinations with conventional cytotoxic therapeutics. However, significant problems remain in the clinical application of angiogenesis inhibitors such as the need for surrogate markers to monitor the effects of such drugs when they do not cause tumor regressions, and the design of clinical trials. Also of concern is that the expected need to use anti-angiogenic drugs chronically will lead to delayed toxic side effects in humans, which do not appear in rodents, especially in short-term studies.

Alloantigen profile of graft vascular endothelial cells in rat heart allograft rejection: expression of MHC and non-MHC alloantigens

Summary The alloantigen profile of vascular endothelial cells has been studied in a rat experimental model for MHC-mismatched and MHC-matched (non-MHC mismatched) allogeneic heart transplantation (TX). Hearts were studied for alloantibody deposition in the graft vasculature and the occurrence of vasculopathy. In addition reactivity of allosera with in vitro cultures of graft-type vascular endothelial cells was examined. Alloantibodies were seen deposited in the graft vasculature in MHC-mismatched as well as in MHC-matched TX. In MHC-matched hearts the most dominant vasculopathy consisted of early arteriosclerotic lesions. In vitro studies demonstrated that, in addition to MHC alloantigens, heart vascular endothelial cells express non-MHC alloantigens. These endothelial non-MHC alloantigens are constitutively expressed by heart endothelial cells, and expression is not influenced by cytokine stimulation. We conclude that humoral immune response against these endothelial non-MHC alloantigens may play an important role in development of graft vasculopathy and particularly of arteriosclerotic lesions.