The effect of humic acid on the adhesibility of neutrophils

Humic acid (HA), a fluorescent allomelanin, has been implicated as an etiological agent of Blackfoot disease (BFD), a peripheral vascular disease prevailing in the southwest of Taiwan. Clinical and pathological studies reveal that it is similar to atherosclerosis. In this report, the effect of HA on human neutrophils is studied because prolonged and enhanced activation of neutrophils adhered on endothelium may damage the endothelium and initiate the process of thrombosis and vasculitis. METHODS: Neutrophils, treated with various concentrations of HA, were added to culture plates, cultured human umbilical vein endothelial cells (HUVECs), or human umbilical vein endothelium tissue culture for 15 or 30 min. The adhesion of neutrophils was measured qualitatively and quantitatively. The mechanism of neutrophil activation was studied with free radical production and various kinase measurements and their activities' assays. RESULTS: HA was shown to enhance, in a dose-dependent manner, the adhesion of neutrophils on the culture plates, cultured human umbilical vein endothelial cells, and human umbilical vein endothelium tissue culture. The adhesion-enhancing ability of HA is elicited through activation of ERK, P38 mitogen-activated kinase (P38MAPK), and phosphoinositide 3 kinase (PI3K) in neutrophils. HA also induces the NF-kappaB activation in neutrophils. CONCLUSION: HA treatment markedly enhanced adhesion and superoxide radical production of neutrophils, the characteristics of activated neutrophils; and all these stimulation effects were blocked by several kinase inhibitors, reflecting the involvement of the ERK, P38MAPK, and PI3K on the activation of neutrophils. The induction of NF-kappaB implied that the consequence of neutrophil activation by HA were similar to other stimulants. The prolonged neutrophil activation will further damage endothelium cell and cause thrombosis, vaculitis, as well as arteriosclerosis. This may partially explain why HA consumption will cause BFD.     

Reduction of the anticoagulant activity of glycosaminoglycans on the surface of the vascular endothelium by endotoxin and neutrophils: Evaluation by an amidolytic assay

The processes that underlie the coagulopathy observed in severe infection are not fully understood, but seem to be due to an imbalance in the antithrombotic, and prothrombotic properties of the vascular endothelium. Sulphated glycosaminoglycans (GAGs) present on the vessel wall represent an important component of the non-thrombogenic nature of the endothelium. We have modified an amidolytic assay to study the functional ability of GAGs on human umbilical vein endothelial cells (HUVECS), and investigate the effect of E. coli endotoxin and neutrophils on HUVEC surface anticoagulant activity (SAA). Neither endotoxin alone, nor separated neutrophils at lower concentrations (less than 10⁶ neutrophils per ml), had major effects on endothelial SAA. When activated neutrophils were incubated with HUVECS pre-stimulated with endotoxin, a significant decrease in SAA was seen using either plasma (mean percentage of control 67.8% ± sem 7.8; p< 0.02) or purified ATIII (mean percentage of control 69% ± sem 4.6; p< 0.001). We suggest that alterations in endothelial surface GAGs may occur during sepsis and inflammation, and that this may have important consequences for vascular function. This system will allow the further study of the role of GAGs in the intravascular thrombosis of severe sepsis, and other inflammatory diseases.     

NO reduces PMN adhesion to human vascular endothelial cells due to downregulation of ICAM-1 mRNA and surface expression

Reperfusion damage is largely due to the adherence of polymorphonuclear leukocytes to the endothelium initiated by adhesion molecule upregulation. The reduced endothelial nitric oxide release during ischemia may be involved in the upregulation of intercellular adhesion molecule 1. In this study, we tested if nitric oxide donors suppress polymorphonuclear leukocyte adherence to activated endothelial cells by inhibition of the intercellular adhesion molecule 1 surface expression. Confluent human umbilical vein endothelial cells were stimulated with tumor necrosis factor alpha (300 U/mL) after preincubation with increasing concentrations of the nitric oxide donors CAS 1609 (0.005-5 mM/L) and 3-(4-morpholinyl)-sydnonimine (0.01-1 mM/L). Intercellular adhesion molecule 1 surface expression was measured in a cell surface enzyme-linked immunosorbent assay, intercellular adhesion molecule 1 mRNA by Northern analysis. Human saphenous vein endothelial cells were transfected with the inducible nitric oxide synthase gene and stimulated with tumor necrosis factor alpha (300 U/mL). Fluorescein green-labeled polymorphonuclear leukocytes adhering to activated human umbilical vein endothelial cells/human saphenous vein endothelial cells were quantified by epifluorescent microscopy. The intercellular adhesion molecule 1 surface expression of activated human umbilical vein endothelial cells/human saphenous vein endothelial cells was significantly diminished to 40 to 60% of the maximum after treatment with CAS 1609, 3-(4-morpholinyl)-sydnonimine, or transfection with the inducible nitric oxide synthase gene. Intercellular adhesion molecule 1 mRNA was diminished by CAS 1609 and 3-(4-morpholinyl)-sydnonimine in the same manner. The functional relevance of our data was shown by reduction of polymorphonuclear leukocyte adherence to activated human umbilical vein endothelial cells/human saphenous vein endothelial cells following treatment with CAS 1609 and 3-(4-morpholinyl)-sydnonimine or transfection with inducible nitric oxide synthase. Tumor necrosis factor-induced polymorphonuclear leukocyte adherence was abolished by blocking antibody against intercellular adhesion molecule 1. Thus, exogenous or endogenous substitution of nitric oxide diminishes the expression of endothelial intercellular adhesion molecule 1 and its mRNA following tumor necrosis factor alpha stimulation. This results in a reduced polymorphonuclear leukocyte adherence to activated endothelium.     

Malignant cell attachment to endothelium of ex vivo perfused human umbilical vein. Modulation by platelets, plasma and fibronectin

The success of blood-born metastatic spread depends upon a key event: the tumor cell arrest and attachment to the host organ vasculature. In the present study, we have investigated interactions between several normal and cancer cell lines and vascular endothelium in a model of ex vivo perfusion of human umbilical vein. In this system, hydrodynamic parameters are monitored and endothelial cells are kept in their original environment known to modulate their phenotype. Metastatic tumor cell adhesion to the perfused endothelium was found to be significantly higher than that of normal cells tested. Platelets and soluble plasma factors including fibronectin promoted tumor cell arrest and adhesion to endothelium. Altogether our results indicate that the ex vivo perfusion of human umbilical vein allows the study of the interactions between malignant tumor cells, circulating plasma or blood cells and the endothelium during blood-born metastatic spread.     

Induction of tissue factor synthesis in human umbilical vein endothelial cells involves protein kinase C.

Incubation of human umbilical vein endothelial cells with one of the following compounds: endotoxin, recombinant interleukin-1 beta, recombinant tumor necrosis factor alpha, allogenic lymphocyte subpopulations or phorbol ester resulted in significant induction of tissue factor synthesis. Diacylglycerol had the same effect and also enhanced synergistically the induction caused by endotoxin and interleukin-1 beta. Two different inhibitors of protein kinase C, H7 and sphingosine, inhibited tissue factor synthesis at concentrations which did not depress protein synthesis in general, suggesting that protein kinase C is involved in the processes leading to tissue factor synthesis. Cells down-regulated for the tissue factor response to TPA responded essentially normally to endotoxin and interleukin-1 with regard to tissue factor synthesis.     

Pentoxifylline prevents tumor necrosis factor-induced suppression of endothelial cell surface thrombomodulin

Thrombomodulin (TM) expression has been reported to be down-regulated by cytokines (endotoxin, interleukin-1, and tumor necrosis factor). We report, in the present study, up-regulation of surface TM antigen of human umbilical vein endothelial cells (HUVECs) by pentoxifylline (PTX) which is one of the agents that can increase intracellular cyclic AMP in HUVECs at therapeutic concentrations. Surface TM antigen was measured by an enzyme immunoassay. PTX increased surface TM antigen and intracellular cAMP in HUVECs in a dose dependent manner. Upregulation of TM by PTX was due to de novo synthesis of TM protein resulting from increased TM mRNA levels. PTX counterbalanced the TNF-induced suppression of TM expression. These results suggest that protein kinase A may be involved in cellular regulatory mechanism for TM expression and PTX may protect partially against TNF-induced endothelial cell injury and restore anticoagulant state of endothelium.     

Heparan sulfate proteoglycans modulate monocyte migration across cerebral endothelium

Heparan sulfate proteoglycans (HSPGs) are known to participate in a wide range of biological events, including cellular trafficking. In this study we report that in situ cerebral blood vessels highly express HSPGs. Of the syndecan family, syndecan-2 is highly expressed on virtually all brain vessels and syndecan-1 and -3 are only present on larger blood vessels. These endothelial HSPGs have a functional role in monocyte diapedesis across brain endothelium, as assessed in our in vitro adhesion and migration assays. Our data indicate that heparin prevents monocyte adhesion to brain endothelium by interacting solely with the monocyte. Transendothelial migration of monocytes can be prevented by preincubation of brain endothelium with heparin by enzymatic removal of heparan sulphate side chains or by inhibition of cellular sulfation. Blocking of G-protein-dependent signaling in the monocytes prevented monocyte adhesion and migration to similar extent, suggesting that G-dependent signaling may be involved in HSPG-mediated monocyte adhesion and transendothelial migration. Our data demonstrate that brain endothelial HSPGs have a modulatory role in the transendothelial migration of monocytes in a direct and indirect fashion and may therefore contribute to the formation of neuroinflammatory lesions.     

Platelet-endothelial cell interaction in pulmonary micro-circulation: the role of PARS

Accumulation of platelets might contribute to acute lung injury during systemic inflammation. The aim of the study was to elucidate the role of the poly (ADP-ribose) synthetase, a nucleotide-polymerizising enzyme, in mediation of platelet-endothelial cell interaction through regulation of adhesion molecules within the pulmonary microcirculation during endotoxemia. We used in vivo fluorescence microscopy to quantify kinetics of fluorescently labeled erythrocytes and platelets in rabbit pulmonary arterioles and venules. Six hours after onset of endotoxin infusion we observed a massive interaction of platelets with the microvascular endothelial cells, whereas under control conditions, no platelet sequestration was measured. An up-regulation of P- and E-selectin was detected in lung tissue following endotoxin infusion by immunohistochemistry and Western blot analysis. Blockade of endothelial P-selectin with fucoidin resulted in a reduction of the endotoxin-induced platelet-endothelial cell interaction. Inhibition of poly (ADP-ribose) synthetase by 3-aminobenzamide inhibited the endotoxin-induced expression of endothelial P- and E-selectin and the subsequent recruitment of platelets. In summary, we provide first in vivo evidence that platelets accumulate in pulmonary microcirculation following endotoxemia. Poly (ADP-ribose) synthetase seems to mediate this platelet-endothelial cell interaction via P- and E-selectin expressed on the surface of microvascular endothelium.     

Inhibition of thrombin-induced signaling by resveratrol and quercetin: effects on adenosine nucleotide metabolism in endothelial cells and platelet-neutrophil interactions

BACKGROUND: Thrombin downregulates endothelial ectonucleotidase activity resulting in high levels of adenosine diphosphate (ADP) and adenosine triphosphate (ATP) which lead to platelet, leukocyte and endothelial activation. Depending on adenosine nucleotide levels, resting platelets inhibit and thrombin-activated platelets increase respiratory burst of neutrophils. Whether the red wine polyphenols quercetin and resveratrol affect thrombin-dependent adenosine nucleotide, metabolism and thrombin-induced signaling is unknown. MATERIALS AND METHODS: ATP and ADP secretion by platelets, the impact on subsequent oxidative burst activity in neutrophils and CD39/ATPdase function in endothelial cells (ECs)was studied. Cell migration was measured in modified Boyden chambers; adenosine metabolites were quantified by high-performance liquid chromatography (HPLC). Signal transduction was studied by Western blotting. RESULTS: Quercetin and resveratrol inhibited thrombin-induced ADP and ATP secretion from platelets in a concentration-dependent manner. Augmented respiratory burst of neutrophils in response to thrombin-activated platelets was also inhibited by the two polyphenols as was neutrophil migration toward thrombin-induced supernatants of platelets. Quercetin and resveratrol restored the decreased CD39/ATPdase activity in human umbilical vein endothelial cells, in response to thrombin as demonstrated by adenosine monophosphate (AMP) and adenosine increases in endothelial culture supernatants. Both polyphenols inhibited thrombin-induced MAPK, JNK and focal adhesion kinase activities in endothelial cells. CONCLUSION: Quercetin and resveratrol interfere with the proinflammatory signaling of thrombin resulting in the inhibition of adenosine nucleotide secretion from activated platelets and decreased neutrophil function. Moreover, the polyphenols protect endothelial adenosine nucleotide metabolism when downregulated by thrombin. These observations may explain cardioprotective effects of grape products.     

On the control of the plasma contact activation system on human endothelium: comparisons with heparin surface

The endothelial lining in segments of the human saphenous vein harvested before and shortly after systemic heparinization of patients was investigated with regard to the ability to inhibit activated FXII. Comparisons were made with a surface modified by endpoint attached heparin, which, like the endothelium, is negatively charged and exposes the specific antithrombin binding sequence and also binds FXII and antithrombin. The heparin surface preincubated with plasma or blood and endothelium in vivo having been exposed to non-heparinzed blood and expressed similar amounts of FXII but no FXIIa. On both surfaces activation of bound FXII with ellagic acid resulted in consumption of the proenzyme and in inhibition of formed FXIIa. On both heparin surface and the endothelium consumption of antithrombin occurred concomitantly, demonstrating the importance of this inhibitor in the control of contact activation system. On the endothelium, in vivo or ex vivo, exposed to heparinized blood, most FXII was present in the active form and only incubation with purified antithrombin could restore the FXIIa inhibitory capacity. It is suggested that antithrombin bound by the specific antithrombin binding sequence of endothelial heparan sulphate, like antithrombin bound to the specific antithrombin binding sequence on heparin surface, inhibits alpha-FXIIa. This makes the endothelium consonant with the plasma contact activation system. It is suggested that systemic heparinzation of patients may both activate endothelial bound FXII and interfere in the inhibition of formed FXIIa by depriving the endothelium of required amounts of antithrombin.     

Reduced adhesion of PBMNCs to endothelium in methylprednisolone-treated MS patients: preliminary results

Methylprednisolone (MP) is a synthetic steroid commonly used in the treatment of multiple sclerosis (MS) relapses. It has a wide spectrum of activities on immune cells: it might also act by preventing mononuclear cell/endothelium adhesion. We studied adhesion phenomena between cultured human umbilical vein endothelial cells (HUVECs) and PBMNCs (CD45⁺, CD14⁺) from 6 MS patients treated in vivo with MP. We also studied fluctuations in CD11a and CD18 levels on lymphocytes and monocytes, as well as changes in serum sICAM-1 and sVCAM-1 concentrations. After MP treatment, PBMNCs adhesion to endothelium decreased at 3 h, while it went back to baseline levels at 24 h. A tendency to increase in both CD11a and CD18 on the surface of lymphocytes was detected, while an increase in serum sVCAM-1 was seen at 3 h.     

An enzyme-linked immunosorbent assay (ELISA) used to study the cellular secretion of endothelial plasminogen activator inhibitor (PAI-1)

A two-site sandwich ELISA was developed to measure PAI-1 antigen and utilised a polyclonal antiserum produced against PAI-1 purified from human endothelial cell secretory products. The assay was calibrated against a preparation of pure PAI-1 whose protein concentration had been determined by amino acid analysis and the detection limit was 30 pg PAI-1 ml-1 sample. PAI-1 was detected in primate sera but not in a wide range of non-primate sera and no cross-reactivity with alpha 2-antiplasmin or antithrombin III was observed. The ELISA was used to study cellular secretion of PAI-1 which was confirmed as a major secretory protein in human umbilical vein endothelial cells (HUVEC). PAI-1 antigen accumulated in the medium in a linear fashion with time and accounted for approximately equal to 10% of total secreted protein. Specific activity of intracellular PAI-1 was typically 20-fold greater than that of PAI-1 in 24 h conditioned medium and a t1/2 for inactivation of secreted PAI-1 of 0.53 h was calculated. Purified endotoxin stimulated the secretion of PAI-1 antigen and raised the intracellular levels in HUVEC cultures showing that the anti-fibrinolytic actions of endotoxin are effected by increasing the rate of synthesis and secretion of PAI-1.     

Methylprednisolone acts on peripheral blood mononuclear cells and endothelium in inhibiting migration phenomena in patients with multiple sclerosis

BACKGROUND: Intravenous methylprednisolone hemisuccinate is administered to patients with multiple sclerosis (MS) during exacerbations to improve the rate of recovery. Corticosteroids could be beneficial in MS exacerbations also by decreasing transmigration of peripheral blood mononuclear cells (PBMNCs) through the blood-brain barrier. OBJECTIVES: To evaluate how in vivo intravenous methylprednisolone treatment in patients with MS could influence transmigration of PBMNCs in an in vitro model; to perform transmigration experiments through a methylprednisolone-treated endothelium with PBMNCs from untreated healthy control subjects to evaluate putative selective effects of corticosteroids on endothelium; concomitantly, to quantify the concentration of matrix metalloproteinases 2 and 9 in supernatants of PBMNCs and in serum samples from methylprednisolone-treated patients with MS; to evaluate monokine induced by interferon-gamma release in the supernatants of human umbilical vein endothelial cells treated with interferon-gamma alone or interferon-gamma and methylprednisolone; and to perform gene expression studies of matrix metalloproteinases 2 and 9 in human umbilical vein endothelial cells and PBMNCs from methylprednisolone-treated patients with MS. PATIENTS: Eight patients with MS in exacerbation were studied before and 3 and 24 hours after intravenous methylprednisolone treatment, 1 g. RESULTS: The absolute number of transmigrated PBMNCs from methylprednisolone-treated patients with MS significantly (P<.01) decreased at 3 hours and increased again at 24 hours, reaching values higher than those before treatment onset. Methylprednisolone was also able to significantly (P<.03) reduce the number of PBMNCs from healthy controls migrating through interferon-gamma-stimulated or unstimulated endothelium. In vitro methylprednisolone treatment decreased monokine induced by interferon-gamma production in human umbilical vein endothelial cells. CONCLUSIONS: Methylprednisolone may be able to decrease transmigration of PBMNCs through the blood-brain barrier, exerting its inhibitory effects on PBMNCs and endothelium. A "rebound" of transmigration at 24 hours suggests that a single infusion is not optimal for achieving a persistent reduction in transmigration.     

Immunocompetence of human microvascular brain endothelial cells: cytokine regulation of IL-1β, MCP-1, IL-10, sICAM-1 and sVCAM-1

Endothelia from the brains of four patients undergoing neurosurgery, including one multiple sclerosis (MS) patient, were studied in vitro to determine cytokine and chemokine production; the release of soluble adhesion molecules was also investigated. The same procedure was repeated on human umbilical vein endothelial cells (HUVECs) in order to detect possible district-specific differences. After isolation, the endothelium was cultured and stimulated with γ-interferon (IFN), tumour necrosis factor alpha (TNF-α) and LPS. The results showed that brain endothelium, in our experimental conditions, does not produce interleukin (IL)-10 and produces lower amounts of IL-1β and soluble intercellular adhesion molecule-1 (sICAM-1) than HUVECs do; no differences were detected in soluble vascular cell adhesion molecule-1 (sVCAM-1) production. MCP-1 mRNA was detected both without and after stimulation with TNF-α and γ-IFN in HUVECs and MS human brain endothelial cells (HBECs), while in non-MS-HBECs it was found only after γ-IFN stimulation. Received: 24 June 1997 Received in revised form: 20 February 1998 Accepted: 5 March 1988     

Endothelial cell apoptosis induced by bacteria-activated platelets requires caspase-8 and -9 and generation of reactive oxygen species

A common feature of severe sepsis is vascular inflammation and damage to the endothelium. Because platelets can be directly activated by bacteria and endotoxin, these cells may play an important role in determining the outcome of sepsis. For example, inhibiting platelet interactions with the endothelium has been shown to attenuate endothelial cell damage and improve survival during sepsis. Although not entirely understood, the interactions between bacteria-activated platelets and the endothelium may play a key role in the vascular pathology of bacterial sepsis. Haemophilus somnus is a bacterial pathogen that causes diffuse vascular inflammation and endothelial damage. In some cases H. somnus infection results in an acute and fatal form of vasculitis in the cerebral microvasculature known as thrombotic meningoencephalitis (TME). In this study, we have characterized the mechanisms involved in endothelial cell apoptosis induced by activated platelets. We observed that direct contact between H. somnus-activated platelets and endothelial cells induced significant levels of apoptosis; however, Fas receptor activation on bovine endothelial cells was not able to induce apoptosis unless protein synthesis was disrupted. Endothelial cell apoptosis by H. somnus-activated platelets required activation of both caspase-8 and caspase-9, as inhibitors of either caspase inhibited apoptosis. Furthermore, activated platelets induced endothelial cell production of reactive oxygen species (ROS) and disrupting ROS activity in endothelial cells significantly inhibited apoptosis. These findings suggest that bacterial activation of platelets may contribute to endothelial cell dysfunction observed during sepsis, specifically by inducing endothelial cell apoptosis.     

组织工程化类金刚石膜复合材料与人血管内皮细胞的相容性研究

目的：验证纳米相类金刚石薄膜复合材料与人血管内皮细胞相容性,为组织工程化机械瓣膜材料的构建提供依据。 方法：利用脉冲激光沉积法在人工心脏机械瓣膜上沉积纳米相类金刚石薄膜,将人脐静脉血管内皮细胞与纳米相类金刚石薄膜复合材料体外复合培养。倒置显微镜及扫描电镜观察细胞在材料表面的生长、附着情况;MTT 法检测细胞在材料上增殖情况;同时分别测定人脐静脉血管内皮细胞在类金刚石薄膜材料和空白对照组中一氧化氮及前列环素分泌水平,以评价其活性。 结果：人脐静脉血管内皮细胞能在纳米相类金刚石薄膜复合材料上良好地黏附、增殖、生长。人脐静脉血管内皮细胞分泌一氧化氮和前列环素水平在类金刚石薄膜材料和空白对照组没有显著性差异(P > 0.05),说明纳米相类金刚石薄膜材料对人脐静脉血管内皮细胞的活性没有影响。 结论：纳米相类金刚石薄膜复合材料具有良好的细胞相容性,有可能作为组织工程化机械瓣膜材料。     

脑梗死患者中性粒细胞与血管内皮细胞黏附性的研究

目的　分析脑梗死后外周血中性粒细胞 (PMN)与血管内皮细胞 (EC)黏附性的变化规律 ,探讨抗细胞间黏附分子 1单克隆抗体 (ICAM 1mAb)对其影响。方法　用细胞计数法检测 30例脑梗死患者梗死后 1周以内、2 1天时PMN与经 (或不经 )ICAM 1mAb处理的人类脐静脉内皮细胞细胞株ECV 30 4的黏附率。结果(1)脑梗死患者PMN与ECV 30 4的黏附率在 1周以内明显增高 ,到第 2 1天时显著下降 ;(2 )ICAM 1mAb可以明显降低 1周以内脑梗死患者PMN与ECV 30 4的黏附 (P <0 .0 1) ,而 2 1天时脑梗死患者和健康对照组PMN与ECV 30 4黏附则影响较小 (P <0 .0 5 )。结论　 (1)急性期脑梗死患者PMN与EC黏附增强 ,到恢复期时基本正常 ;(2 )ICAM 1参与介导脑梗死急性期PMN与EC的黏附 ,ICAM 1mAb可部分阻断PMN与EC黏附     

Effect of platelet activating factor on leukocyte-endothelial cell interactions

The proinflammatory effects of platelet activating factor (PAF) that impact upon tissue inflammation were studied in vitro using the adherence of human neutrophils to endothelium and the increase in macromolecule permeability of endothelial monolayers. PAF produced both a time- and dose-dependent increase in neutrophil-endothelial cell adhesion. The adhesion promoting properties observed were primarily due to an effect of PAF on endothelium and not on neutrophils and was independent of endothelial cell cyclooxygenase products. The PAF receptor antagonist kadsurenone inhibited the adhesion response suggesting endothelial surface PAF receptors are involved in these responses. Whereas PAF alone did not alter endothelial cell barrier properties, leukocyte activation (neutrophil and platelets) with PAF produced significant increases in 125I-albumin clearance across endothelial monolayers. These studies suggest that PAF has potent proinflammatory effects and that it can play a significant role in the endothelial response to injury.     

Platelet and shear rate promote tumor cell adhesion to human endothelial extracellular matrix--absence of a role for platelet cyclooxygenase

Blood platelets are thought to be involved in certain aspects of malignant dissemination. To study the role of platelets in tumor cell adherence to vascular endothelium we performed studies under static and flow conditions, measuring tumor cell adhesion in the absence or presence of platelets. We used highly metastatic human adenocarcinoma cells of the lung, cultured human umbilical vein endothelial cells (ECs) and extracellular matrices (ECM) prepared from confluent EC monolayers. Our results indicated that under static conditions platelets do not significantly increase tumor cell adhesion to either intact ECs or to exposed ECM. Conversely, the studies performed under flow conditions using the flat chamber perfusion system indicated that the presence of 2 x 10(5) pl/microliters in the perfusate significantly increased the number of tumor cells adhered to ECM, and that this effect was shear rate dependent. The maximal values of tumor cell adhesion were obtained, in presence of platelets, at a shear rate of 1,300 sec-1. Furthermore, our results with ASA-treated platelets suggest that the role of platelets in enhancing tumor cell adhesion to ECM is independent of the activation of the platelet cyclooxygenase pathway.