Induction of monocyte tissue factor procoagulant activity during coronary artery bypass surgery is reduced with heparin-coated extracorporeal circuit

The possible activation of monocytes to express tissue factor procoagulant activity (TF-PCA) during CPB (cardiopulmonary bypass) was investigated. 22 patients undergoing myocardial revascularization were randomly assigned to two groups. In group C, heparin-coated circuits (Duraflo II) and reduced systemic heparinization (ACT > 250s) were used. In group NC, non-coated circuits and standard heparin administration (ACT > 480s) were used. Adherent monocytes retrieved from the oxygenators immediately after bypass arrest showed a 2-3-fold increase in TF-PCA when compared to circulating cells pre-CPB (P < 0.01). When cell PCA was expressed as percent change from pre-CPB (baseline) values, circulating monocytes in group NC at CPB-arrest showed a 2-fold increase in PCA compared to group C (P < 0.05). Moreover, the percent increase in PCA of oxygenator-retrieved monocytes was 7-fold in group NC and 2-fold in group C (P < 0.008 and P < 0.004, respectively). Thus, heparin-coating of the extracorporeal circuit reduced induction of adherent cell TF-PCA by 70% (P < 0.05). Thus, monocyte TF-PCA may cause activation of the extrinsic coagulation pathway during CPB surgery. It is apparent that heparin-coating enhanced biocompatibility of extracorporeal circuits. Reduced systemic heparinization in group C proved to be safe. However, further reduction of heparin administration may not be advisable, since monocytes were still activated in the coated oxygenator.     

Control of contact activation on end-point immobilized heparin: the role of antithrombin and the specific antithrombin-binding sequence

The uptake and activation of FXII from blood plasma was studied in small-diameter polyethylene tubing, surface-modified by end-point immobilization of heparin. Two preparations of heparin were used to modify the contact-activating properties of the plastic tubing: unfractionated, functionally active heparin and low-affinity heparin, lacking the specific antithrombin-binding sequence and virtually devoid of anticoagulant activity. The uptakes of FXII on the two heparin surfaces were similar. No activated FXII could be demonstrated on the unfractionated heparin surface, whereas on the low-affinity heparin surface nearly all FXII underwent spontaneous activation. The suppression of FXII activation on the unfractionated heparin surface was investigated by using plasma depleted of antithrombin, complement C1 esterase inhibitor, or both. The removal of antithrombin resulted in extensive activation of FXII, whereas the depletion of C1 esterase inhibitor had only a minor effect. Experiments with recalcified plasma showed rapid clot formation during exposure to the low-affinity heparin surface. After depletion of antithrombin, but not complement C1 esterase inhibitor, the recalcified plasma clotted in contact with the unfractionated heparin surface as well. We conclude that antithrombin and the antithrombin-binding sequence in the surface-immobilized heparin are essential for the prevention of surface activation of FXII and triggering of the intrinsic coagulation system.     

Human monocyte binding to fibronectin enhances IFN-gamma-induced early signaling events

Leukocyte integrins are fundamentally important in modulating adhesion to extracellular matrix components and to other cells. This integrin-mediated adhesion controls leukocyte arrest and extravasation during the onset of inflammatory responses. Moreover, integrin-ligand interactions trigger signaling pathways that may influence leukocyte phenotype and function at sites of inflammation. In the current studies, we evaluated the combinatorial effects of monocyte adhesion and IFN-gamma on intracellular signaling pathways. IFN-gamma triggers a well-defined signal transduction pathway, which although not directly stimulated by monocyte adherence to fibronectin or arginine-glycine-aspartate (RGD)-coated substrata, was enhanced significantly in these matrix-adherent cells. Compared with monocytes in suspension or adherent on plastic surfaces, monocytes adherent to fibronectin or RGD exhibited a greater than threefold increase in steady state levels of IFN-gamma-induced mRNA for the high affinity Fc gammaRI receptor. By electrophoretic mobility shift assays, this increase in mRNA was associated with a 5- to 10-fold increase in the STAT1-containing DNA-binding complex that binds to Fc gammaRI promoter elements. Furthermore, the tyrosine phosphorylation of STAT1 and the tyrosine kinases JAK1 and JAK2 was enhanced significantly in RGD-adherent monocytes compared with control cells. These results suggest a novel mechanism by which integrin-mediated cell adhesion can modulate the magnitude of cytokine-induced signal transduction pathways, thereby amplifying cellular events leading to monocyte activation and inflammation.     

Citrate anticoagulation does not correct cuprophane bioincompatibility as evaluated by the expression of leukocyte surface molecules

BACKGROUND: Citrate, used for the anticoagulation of the extracorporeal dialysis circuit, reduces ionized calcium by chelation and has been claimed to attenuate dialyser membrane bioincompatibility. Dialysis with complement-activating cuprophane membranes is associated with leukopenia which has been related to an increase in adhesion molecule expression on the surface of circulating leukocytes. METHODS: The effect of citrate anticoagulation on the expression of CD11b, CD11c and CD45 on the surface of granulocytes and CD14 on monocytes during haemodialysis with cuprophane membranes, was evaluated by flow cytometric analysis. A comparison of standard heparin vs citrate was performed in 14 chronic haemodialysis patients. During citrate anticoagulation a calcium-free dialysate was used and citrate was infused to obtain a concentration of 4.3 mmol/l blood. The unchallenged 'baseline state' expression of the surface molecules and the increase after ex vivo stimulation with phorbol 12-myristate 13-acetate (delta-PMA) or formyl-methionyl-leucyl-phenylalanine (delta-fMLP) was studied. RESULTS: With heparin, as well as with citrate, a sharp fall in granulocyte and monocyte count was observed after 15 min of dialysis, followed by a recovery at the end of the session. The expression of CD11b, CD11c and CD45 on granulocytes increased markedly during cuprophane dialysis with a peak at 15 min; there were no differences in response between heparin and citrate anticoagulation. Delta-PMA and delta-fMLP for CD45, CD11c and CD14 showed a decrease during cuprophane dialysis vs t0; again there were no differences between heparin and citrate. CONCLUSION: We conclude that the use of citrate was not associated with reduced leukocyte activation as measured by the expression of surface molecules during cuprophane dialysis and that no effect on dialysis leukocytopenia could be registered.     

Efficacy of local inhibition of procoagulant activity associated with small-diameter prosthetic vascular grafts

PURPOSE: Graft procoagulant activity is determined by thrombin (IIa) and activated factor X (Xa) that binds to thrombus. Thrombus-associated factor Xa and thrombin are resistant to antithrombin III-dependent therapy (heparin). To avoid complications and costs associated with systemic administration, we evaluated whether locally applied antithrombotic agents inhibit prosthetic graft procoagulant activity under no-flow and low-flow conditions. METHODS: Four-millimeter-diameter collagen-coated grafts were preclotted in recalcified human plasma, washed, immersed in antithrombotic agents (either 100 nm hirudin, 20 microns D-Phe-L-Pro-L-Arg chloromethylketone, 5 microns tick anticoagulant peptide or 5 or 10 micrograms/ml tissue factor pathway inhibitor) or saline solution, and extensively rewashed. Grafts were exposed to recalcified plasma either in multiwell plates or underwent perfusion at 1 ml/min flow rate. Fibrinopeptide A, which reflects fibrin elaboration, was measured as a marker of thrombin activity. RESULTS: Inhibitors reduced fibrinopeptide generation at 8 minutes by 55% (tissue factor pathway inhibitor), 57% (hirudin), or 63% (tick anticoagulant peptide and D-Phe-L-Pro-L-Argchloromethylketone) compared with the control agents (p < 0.05). Under low-flow conditions tissue factor pathway inhibitor and hirudin reduced fibrinopeptide generation at 13 minutes by 61% and 49%, respectively, when compared with control agents (p < 0.05). CONCLUSIONS: Graft-associated inhibitors targeted at factors IIa, Xa, or tissue factor/VIIa/Xa complex effectively reduce procoagulant activity on prosthetic grafts. The success of local application of antithrombotic agents in attenuating early fibrin formation suggests that this strategy could favorably influence acute graft patency, and we speculate these agents may improve long-term graft patency as well.     

Inhibition of human monocyte adhesion to endothelial cells by the coumarin derivative, cloricromene

1. The ability of the coumarin derivative cloricromene (8-monochloro-3-beta-diethylaminoethyl-4-methyl-7-ethoxy- carbonylmethoxycoumarin) to inhibit monocyte adhesion to human cultured umbilical vein endothelial cells (HUVEC) was investigated. 2. Cloricromene (10-200 microM) inhibited, in a concentration-dependent manner, the adhesion of both resting and activated monocytes to HUVEC. Significant inhibition was reached with drug concentrations ranging between 15 to 30 microM. 3. The inhibitory activity was, at least in large part, directed to monocytes since no inhibition was observed after selective preincubation of HUVEC with cloricromene and the drug maintained its effect also on monocyte adhesion to paraformaldehyde-treated HUVEC. 4. Inhibition was maximal after 1 min of exposure of monocytes to cloricromene and persisted even in the absence of the drug. 5. Both basal and chemoattractant-mediated monocyte adhesion was inhibited by cloricromene as it was by TS1/18, a monoclonal antibody (mAb) directed to beta 2 integrins; however, cytofluorimetric analysis showed that cloricromene was unable to modulate the expression of beta 2 integrins on the monocyte surface. 6. When monocyte adhesion was mediated by a large set of adhesive receptors, as obtained after treatment of HUVEC with either interleukin 1 beta (IL-1; 50 ng ml-1) or tumour necrosis factor-alpha (TNF; 100 u ml-1), the inhibitory effect of cloricromene was considerably reduced. 7. The results of this study show that cloricromene may regulate monocyte adhesion to HUVEC, an event relevant in vivo in the pathogenesis of inflammatory and atherosclerotic processes.     

Antioxidant properties of the decarboxylated dimer of aminoethylcysteine ketimine: assessment of its ability to scavenge peroxynitrite

In the present study we used an in-vitro technique to examine initiation and propagation of blood coagulation at the surface of tantalum coronary stents exposed to flowing platelet-rich and platelet-free plasma. The time course of factor IXa production at the surface of the stent was not influenced by platelets. In spite of a significant factor IXa production, no thrombin activity was detected when the tantalum stent was exposed to platelet-free plasma; only when the stent was exposed to platelet-rich plasma was extensive thrombin production observed. These findings indicate that tantalum triggers blood coagulation, but that (adherent) platelets are essential for thrombin generation. Heparin-coated tantalum stents exposed to flowing platelet-rich plasma showed that factor IXa generation was slightly reduced compared with the bare stent. However, the heparin coating drastically delayed the onset of thrombin generation and largely reduced the steady-state production of thrombin. We found a clear relationship between the antithrombin binding capacity and the antithrombogenic potential of the heparin-coated stents. The mode of action of immobilized heparin is thought to abrogate thrombin generation by inhibiting thrombin-dependent positive feedback reactions at the surface of the coronary stent.     

Depletion of intravascular pools of tissue factor pathway inhibitor (TFPI) during repeated or continuous intravenous infusion of heparin in man

Tissue factor pathway inhibitor (TFPI) is a potent inhibitor of the extrinsic coagulation system. TFPI is increased several-fold in postheparin plasma and thereby thought to contribute significantly to the antithrombotic action of heparin. The present study was conducted to investigate how repeated (n = 8) and continuous (n = 6) administration of heparin affect plasma TFPI and the inhibition of tissue factor (TF)-induced coagulation ex vivo in humans. Free TFPI antigen (TFPI Ag) increased from 19.2 +/- 4.0 ng/ml to 204.7 +/- 31.7 ng/ml after intravenous injection of 5000 IU of unfractionated heparin. Five repeated injections of 5000 IU of heparin at 4 h intervals caused a progressive decrease (-45 +/- 8%, p < 0.0001 for time effect) in heparin-releasable TFPI and a progressive shortening of the clotting time as determined in a dilute prothrombin time assay (dPT) (-8.7 +/- 6.1 s, p < 0.0001). The basal concentration of TFPI Ag in plasma collected immediately before each heparin injection was decreased by 29 +/- 15% (p < 0.0001), whereas the dPT was decreased by 6.9 +/- 3.5 s (p < 0.0001). During a 24 h continuous infusion of heparin TFPI Ag decreased from 161.5 +/- 26.0 ng/ml to 35.6 +/- 4.7 ng/ml (-77.3 +/- 5.1%) (p < 0.0001). The contribution of TFPI to the inhibition of TF-induced coagulation during heparin infusion was estimated to decrease from 60 +/- 15% to 20 +/- 10% (p < 0.0001). The present data indicate partial depletion of intravascular pools of TFPI by repeated and continuous heparin administration and thereby attenuation of its contribution to the antithrombotic action of heparin.     

Interaction of heparin with canine coagulation proteins: in vivo and in vitro studies

The effects of heparin on the coagulation profile and on specific factor activity in canine plasma have been examined both in vivo and in vitro. The results show that the prolongation of the partial thromboplastin time of plasma produced by heparin is, at least in part, the result of the interaction of heparin with the intrinsic Factors VIII, IX and XI and the inhibition of their procoagulant activity by heparin. A significant correlation was found between the partial thromboplastin time assay and the circulating heparin activity following intravenous administration of heparin to dogs. The results confirm the suitability of the partial thromboplastin time assay for monitoring heparin therapy in the dog.     

Lipoproteins and the haemostatic system in atherothrombotic disorders

The early belief that the haemostatic system has no active role in the formation of the atheromatous plaque is no longer tenable. Rather, the association between hypercholesterolaemia and atherosclerosis appears to arise in part because of various effects of high concentrations of LDL and VLDL particles on the cellular and humoral components of the system, thereby promoting plaque growth and thrombosis. These may be summarized as follows: 1. High concentrations of native LDL have been reported to promote the adhesion of monocytes to the endothelial cell, suggesting that the latter undergoes a form of activation upon such exposure. Oxidized LDL is more potent in this respect, and persistent exposure of endothelium to such particles can eventually lead to cell injury. 2. Activated endothelial cells acquire characteristics on their luminal surface conducive to thrombin generation and fibrin production. Thrombin has several actions on the endothelial cell, monocyte, smooth muscle cell and platelet which in the presence of hypercholesterolaemia will promote the formation of atheroma. 3. Oxidatively modified LDL can activate circulating monocytes, when they also acquire procoagulant properties which favour thrombin production. 4. Platelets show an increased tendency to aggregate when exposed to hypercholesterolaemic plasma. This effect may arise in part because the platelet of the hypercholesterolaemic patient expresses an increased number of fibrinogen binding sites on its surface following activation by agonists such as ADP. These hyperaggregable platelets adhere to activated endothelial cells which express von Willebrand factor on their surface, and to subendothelial proteins exposed in the gaps that open between injured endothelial cells. Platelets exposed to raised LDL levels also show a reduced sensitivity to prostacyclin, an antiaggregatory agent. Oxidatively modified LDL has been reported to stimulate aggregation of platelets in the absence of other agonists such as ADP or thrombin (spontaneous aggregation). 5. Platelet aggregation and fibrin deposition at sites of endothelial injury will create microthrombi which become incorporated into the lesion by organization, thereby increasing the fibrous and cellular content of the atheromatous plaque. 6. Lipolysis of triglyceride-rich lipoproteins at the endothelial cell surface leads to transient activation of the coagulation mechanism with activation of factor VII. Activated factor VII is a potent procoagulant when it forms a complex with tissue factor in the atheromatous lesion. Persistent hypertriglyceridaemia is accompanied by raised concentrations of factor X, factor IX, factor VII and prothrombin. 7. Hypertriglyceridaemia is associated with an increased plasma concentration of PAI-1 and a reduction in plasma fibrinolytic activity.(ABSTRACT TRUNCATED AT 400 WORDS)     

Natural anticoagulants, aging, and thromboembolism

The normal aging process alters blood coagulation system in humans; this may be of great concern in the view of the known association of vascular disease with advancing age. The plasma concentration of several coagulation factors, namely fibrinogen, factor VII, factor VIII, factor IX, high molecular-weight kininogen, and prekallikrein, increase in healthy humans, paralleling the physiological aging process. Plasma parameters of clotting activation in vivo, such as prothrombin fragment 1 + 2, fibrinopeptide A, thrombin-antithrombin III complex, and D-dimer, are positively correlated with age. Nevertheless, among centenarians, biochemical signs of marked hypercoagulability are associated with a healthy state. Natural anticoagulants, including antithrombin III, heparin cofactor II, protein C, protein S, and tissue factor pathway inhibitor, can modulate the reactions of blood coagulation system. The occurrence of menopause is accompanied by a significant increase in antithrombin III plasma level; the mean antithrombin III levels in older women exceed levels in male contemporaries. In healthy elderly subjects heparin cofactor II plasma concentrations are lower than in young subjects, independently of gender. Protein C levels raise with age in both sexes, as well as free protein S levels. In women, statistically significant increases in the plasma concentration of the tissue factor pathway inhibitor have been observed, whereas no significant age-related change has been found in men. The fact that many subjects with congenital defects of natural anticoagulants do not undergo thromboembolic events in young age suggests that in healthy individuals a raise in natural anticoagulants can balance the age-related increase of procoagulant factors.     

Somitogenesis: segmenting a vertebrate

OBJECTIVE: To evaluate the effect of gliclazide administration to NIDDM patients on 1) monocyte adhesion to cultured endothelial cells, 2) plasma cytokine and lipid peroxide levels, and 3) monocyte cytokine production. RESEARCH DESIGN AND METHODS: Poorly controlled glyburide-treated diabetic patients (n = 8) and healthy control subjects (n = 8) were recruited. At the beginning of the study, glyburide was replaced by an equivalent hypoglycemic dose of gliclazide. Serum and monocytes were isolated from blood obtained from control and diabetic subjects before and after 3 months of treatment with gliclazide. RESULTS: Plasma lipid peroxide levels and monocyte adhesion to endothelial cells are enhanced in NIDDM patients, and gliclazide administration totally reverses these abnormalities. Before gliclazide treatment, serum levels of cytokines did not differ in the control and the diabetic groups, with the exception of an enhancement of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL)-6 in NIDDM subjects. Basal and lipopolysaccharide (LPS)-stimulated monocyte production of interleukin-1 beta, IL-6, and IL-8 did not differ between the two groups. Furthermore, basal monocyte production of TNF-alpha was similar in the control and the diabetic groups, whereas a marked increase in the LPS-stimulated monocyte production of TNF-alpha was observed in the NIDDM group. Gliclazide treatment lowered LPS-stimulated TNF-alpha production by diabetic monocytes to levels similar to those observed in control subjects. CONCLUSIONS: Gliclazide administration to NIDDM patients inhibits the increased adhesiveness of diabetic monocytes to endothelial cells and reduces the production of TNF-alpha by these cells. These results suggest that treatment of NIDDM subjects with gliclazide may be beneficial in the prevention of atherosclerosis associated with NIDDM.     

Apoptotic vascular endothelial cells become procoagulant

Whereas unperturbed endothelial cells provide potent anticoagulant properties, exposure to inflammatory and atherogenic stimuli can rapidly lead to a procoagulant behavior. Because recent studies provide evidence that apoptosis of vascular cells may occur under conditions such as atherosclerosis and inflammation, we investigated whether apoptotic endothelial cells may contribute to the development of a prothrombotic state. In this report, it is shown that both adherent and detached apoptotic human umbilical vein endothelial cells (HUVECs) become procoagulant. Apoptosis was induced by staurosporine, a nonspecific protein kinase inhibitor, or by culture in suspension with serum deprivation. Both methods resulted in similar findings. As assessed by flow cytometric determination of annexin V binding, HUVECs undergoing cell death exhibited typically a more rapid exposure of membrane phosphatidylserine (PS) than DNA fragmentation. Depending on the stage of apoptosis, this redistribution of phospholipids was found to induce an increase of the activity of the intrinsic tenase complex by 25% to 60%. Although apoptotic cells did not show antigenic or functional tissue factor (TF) activity, when preactivated with lipopolysaccharide, TF procoagulant activity increased by 50% to 70%. At 8 hours after apoptosis induction, antigenic thrombomodulin, heparan sulfates, and TF pathway inhibitor decreased by about 83%, 80%, and 59%, respectively. The functional activity of these components was reduced by about 36%, 52%, and 39%, respectively. Moreover, the presence of apoptotic HUVECs led to a significant increase of thrombin formation in recalcified citrated plasma. In conclusion, apoptotic HUVECs, either adherent or in suspension, become procoagulant by increased expression of PS and the loss of anticoagulant membrane components.     

Covalent immobilization of hirudin improves the haemocompatibility of polylactide-polyglycolide in vitro

A biodegradable polymer, poly(D,L-lactide-co-glycolide) RESOMER RG756, was modified by surface immobilization of recombinant hirudin (r-Hir) with glutaraldehyde as coupling reagent to improve the blood contacting properties of the polymer. The activity of immobilized hirudin on the polymer was estimated by a chromogenic assay to about 2.5 ATU r-Hir cm-2. The improvement of the haemocompatibility of the modified RG756 was evaluated in terms of platelet adhesion/activation, whole blood clotting times and clot formation rate. Fluorescence microscopy revealed that surface modification with r-Hir resulted in decreased platelet adhesion and activation. An ELISA for P-selectin, a marker of platelet activation, was used to confirm this result. Clotting time experiments demonstrated significantly prolonged non-activated partial thromboplastin times, and a decreased clot formation rate of whole blood in contact with r-Hir modified RG756 compared with the plain polymer. Comparison of immobilized r-Hir with bound heparin yielded equivalent improvement of blood-contacting properties of the investigated polymers. These in vitro investigations indicate that the immobilization of r-Hir on RG756 is a useful method to improve the blood contacting properties of polylactides/polyglycolides and other polymers as well.     

Monocyte diapedesis through an in vitro vessel wall construct: inhibition with monoclonal antibodies to thrombospondin

Human peripheral blood monocytes were examined for migration across an endothelial cell monolayer in an in vitro vessel wall construct. Few monocytes invaded in the absence of a chemotactic gradient, despite significant adhesion to the endothelial monolayer. However, the addition of zymosan-activated human plasma to the lower compartment, to create a chemotactic gradient across the vessel wall, resulted in significantly enhanced monocyte migration. Pretreatment of the monocytes with monoclonal antibodies to thrombospondin (TSP) dramatically inhibited monocyte diapedesis into the vessel wall. The same treatment inhibited monocyte adhesion to endothelial cells in two-dimensional monolayer cultures as well as in vessel wall constructs (no chemotactic gradient). Of interest, however, the monoclonal antibodies had no inhibitory effect on monocyte migration into collagen gels devoid of endothelial cells in response to the same chemotactic gradient, suggesting the importance of TSP in monocyte-endothelial cell interactions. Monoclonal antibodies to fibronectin and normal mouse immunoglobulin G did not inhibit migration in this model of a vessel wall. Furthermore, monoclonal antibodies to TSP showed no inhibition of human peripheral blood neutrophil migration. Previous studies have shown that monocytes synthesize TSP and express this moiety on their surface. The present data suggest that monocytes may utilize TSP to interact with endothelial cells lining the vessel wall during diapedesis.     

The role of air plethysmography in the diagnosis of chronic venous insufficiency

The role of protein and cellular components of thrombi in mediating bacterial adhesion on artificial surfaces was investigated in this study. The attachment of Staphylococcus aureus on polyurethane surfaces was observed directly using an automated video microscopy system. Surfaces were preconditioned with components of platelet-fibrin thrombi, including fibrinogen, thrombin, plasma, and isolated platelets. Experiments were performed in a radial flow chamber, and attachment rate constants were compared on the preconditioned surfaces in an effort to understand the complex relationship that exists between bacterial infection and thrombosis on synthetic biomaterials. Preadsorption of fibrinogen to surfaces significantly increased S. aureus adhesion compared to those preadsorbed with albumin alone while the presence of fibrin dramatically increased bacterial attachment compared to plasma preadsorbed surfaces. While the presence of adherent platelets also increased bacterial attachment, fibrin appeared to play a larger role in mediating bacterial adhesion on polyurethane surfaces. Striking results were obtained on the zwitterionic phosphonated polyurethane for a number of pretreatment conditions with regard to decreased bacterial adhesion and fibrinogen deposition.     

Aprotinin reduces the expression of p-selectin on the surface of platelet and leukocyte-platelet conjugates

P-Selectin, an adhesion molecule expressed on the surfaces of activated platelets and the vascular endothelium, mediates platelet binding to monocytes and neutrophils. Monocytes and neutrophils produce superoxide anion by activated platelets through p-selectin. Aprotinin, a serine protease inhibitor, inhibits plasmin to activate platelets during cardiopulmonary bypass (CPB). A total of 25 patients were studied to clarify the effects of aprotinin on p-selectin expression during CPB. Nine patients were not given aprotinin (control group), and 16 were given aprotinin of 2 million U in the priming solution (aprotinin group). The platelet count and soluble p-selectin in the plasma, p-selectin on the surface membranes of platelets, and leukocyte-platelet conjugate levels were measured during and after CPB. The platelet count was maintained well in the aprotinin group. The increases of soluble p-selectin in the plasma, platelet surface p-selectin, and leukocyte-platelet conjugates were less in the aprotinin group than in the control group (p < 0.05). In conclusion, aprotinin in patients undergoing CPB may reduce the early inflammatory reactions induced by p-selectin.     

Enhancement of monocyte procoagulant activity by adhesion on vascular smooth muscle cells and intercellular adhesion molecule-1-transfected Chinese hamster ovary cells

BACKGROUND: Plaque erosion is a frequent finding in sudden death due to coronary thrombosis. The present study sought to investigate whether monocyte adhesion to human aortic vascular smooth muscle cells (VSMCs) induces procoagulant activity (PCA) and whether this could be mediated by intercellular adhesion molecule-1 (ICAM-1). METHODS AND RESULTS: We incubated mononuclear cells (MNCs) with VSMCs and ICAM-1-transfected Chinese hamster ovary (CHO) cells, investigated monocyte tissue factor (TF) mRNA expression by Northern blot analysis and TF protein expression by ELISA, and measured PCA. Incubation of MNCs with VSMCs for 6 hours increased PCA from 0.7+/-0.1 to 166.0+/-37.9 mU/105 cells (P=0.007), which could be inhibited in a dose-dependent manner by the addition of blocking anti-ICAM-1 monoclonal antibodies. Prestimulation of VSMCs with interleukin-1beta enhanced surface ICAM-1 expression significantly but did not induce PCA in VSMCs. Incubation of MNCs with prestimulated VSMCs led to a further increase in PCA to 239.9+/-27.9 mU/10(5) cells (P=0.02 compared with incubation with unstimulated VSMCs). Incubation of MNCs with VSMCs enhanced TF mRNA after 2 hours and significantly increased TF protein content after 6 hours. Incubation of purified monocytes with ICAM-1-transfected CHO cells increased PCA from 1.2+/-0.2 to 81.9+/-3.3 mU/10(5) cells (P<0.001 compared with incubation with untransfected CHO cells) after 6 hours. This effect could be inhibited significantly by the addition of blocking anti-CD18, anti-CD11b, or anti-CD11c monoclonal antibodies. Similar results were obtained for MNCs. CONCLUSIONS: Monocyte adhesion to VSMCs induces TF mRNA and protein expression and monocyte PCA, which is regulated by beta2-integrin-mediated monocyte adhesion to ICAM-1 on VSMCs.     

Heparinization for routine hemodialysis

Routine hemodialysis requires anticoagulation with heparin to prevent clotting in the extracorporeal circuit. Activated whole blood clotting times are used to measure the level of anticoagulation during dialysis. The magnitude of the clotting time obtained for a given level of heparin depends on the test used, and each dialysis unit must establish its own target range. The preferred dosage regimen for heparin is an initial loading dose, followed by a constant infusion. Dosage regimens based on intermittent boluses of heparin are associated with periods of under- and over-anticoagulation and require more staff attention during dialysis. Because patient responses to heparin differ markedly, the doses required to achieve suitable anticoagulation must be determined on an individual basis. The doses of heparin for an individual patient are best determined using a simple pharmacodynamic model, although they can be found empirically through trial and error. In spite of careful anticoagulation, an unsatisfactory outcome may still result. Patient-specific factors may be responsible for some of these adverse outcomes; however, in most cases they result from poor technique, including failure to deliver the prescribed dose of heparin, turbulence and foam formation in the extracorporeal circuit, and excessive access recirculation.