Adhesion of activated platelets to endothelial cells: evidence for a GPIIbIIIa-dependent bridging mechanism and novel roles for endothelial intercellular adhesion molecule 1 (ICAM-1), alphavbeta3 integrin, and GPIbalpha.

Although it has been reported that activated platelets can adhere to intact endothelium, the receptors involved have not been fully characterized. Also, it is not clear whether activated platelets bind primarily to matrix proteins at sites of endothelial cell denudation or directly to endothelial cells. Thus, this study was designed to further clarify the mechanisms of activated platelet adhesion to endothelium. Unstimulated human umbilical vein endothelial cell (HUVEC) monolayers were incubated with washed, stained, and thrombin-activated human platelets. To exclude matrix involvement, HUVEC were harvested mechanically and platelet binding was measured by flow cytometry. Before the adhesion assay, platelets or HUVEC were treated with different receptor antagonists. Whereas blockade of platelet beta1 integrins, GPIbalpha, GPIV, P-selectin, and platelet-endothelial cell adhesion molecule (PECAM)-1 did not reduce platelet adhesion to HUVEC, blockade of platelet GPIIbIIIa by antibodies or Arg-Gly-Asp (RGD) peptides markedly decreased adhesion. Moreover, when platelets were treated with blocking antibodies to GPIIbIIIa-binding adhesive proteins, including fibrinogen and fibronectin, and von Willebrand factor (vWF), platelet binding was also reduced markedly. Addition of fibrinogen, fibronectin, or vWF further increased platelet adhesion, indicating that both endogenous platelet-exposed and exogenous adhesive proteins can participate in the binding process. Evaluation of the HUVEC receptors revealed predominant involvement of intercellular adhesion molecule (ICAM)-1 and alphavbeta3 integrin. Blockade of these two receptors by antibodies decreased platelet binding significantly. Also, there was evidence that a component of platelet adhesion was mediated by endothelial GPIbalpha. Blockade of beta1 integrins, E-selectin, P-selectin, PECAM-1, vascular cell adhesion molecule (VCAM)-1 and different matrix proteins on HUVEC did not affect platelet adhesion. In conclusion, we show that activated platelet binding to HUVEC monolayers is mediated by a GPIIbIIIa-dependent bridging mechanism involving platelet-bound adhesive proteins and the endothelial cell receptors ICAM-1, alphavbeta3 integrin, and, to a lesser extent, GPIbalpha.

The suppression of the coagulation of nonanticoagulated whole blood in vitro by human umbilical endothelial cells cultivated on microcarriers is not dependent on protein C activation.

Human umbilical vein endothelial cells (HUVEC) were cultivated on globular microcarriers in order to improve the endothelial cell surface to blood-volume ratio over the conventional flat bed cultures. HUVEC-beads were tested for their modulation of blood coagulation using a combination of two steps: HUVEC-beads were added into the syringe used for the venipuncture, in order to achieve immediate contact between cells and blood, and no anticoagulant was used during the incubation time of HUVEC-beads with whole blood. The coagulation initiation produced by venipuncture was almost completely suppressed as judged by thrombin measurements over a period of 60 min. The activated partial thromboplastin time showed a prolongation by a factor > 3. Direct measurements of activated protein C (APC) were negative. Moreover, inhibition of APC-generation with a monoclonal anti-human protein C antibody did not affect the anticoagulant properties of endothelial cells. Therefore the anticoagulant properties exerted by HUVEC-beads are not dependent on APC.

Factor V Leiden mutation in postthrombotic and non-postthrombotic venous ulcers.

OBJECTIVE: To determine the prevalence of the factor V Leiden mutation in patients with postthrombotic and non-postthrombotic venous ulcers. DESIGN: Case-control study. SETTING: Department of Dermatology, University Hospital of Zurich, Zurich, Switzerland. PARTICIPANTS: Seventy-three consecutive outpatients and inpatients with venous ulcers and 45 age- and sex-matched control subjects (matched to the 42 patients with postthrombotic syndrome). MAIN OUTCOME MEASURES: Frequency of postthrombotic and non-postthrombotic findings in patients with venous ulcers. Prevalence of the factor V Leiden mutation in these different subgroups. RESULTS: Postthrombotic syndrome was identified as the cause of 42 (58%; 95% confidence interval [CI], 45%-69%) of 73 venous ulcers, and the remainder were caused by primary valvular insufficiency. In postthrombotic ulcers, the prevalence of the factor V Leiden mutation was 38% (95% CI, 24%-54%) (16/42), which corresponds to an odds ratio of 13.2 (95% CI, 2.8-62.3; P<.001). In non-postthrombotic venous ulcers, the prevalence was 16% (95% CI, 5%-34%) (5/31), which corresponds to an odds ratio of 3.2 (95% CI, 1.0-10.0; P =.07). CONCLUSIONS: The factor V Leiden mutation is highly prevalent in patients with postthrombotic venous ulcers. Even patients with non-postthrombotic venous ulcers show a moderately elevated prevalence of the factor V Leiden mutation. Some of the latter might be misclassified because of near-to-perfect revascularization after asymptomatic deep venous thrombosis. However, as long as the therapeutic consequences of the factor V Leiden mutation are not established, systematic screening cannot be recommended in patients with venous ulcers.

In patients with deep-vein thrombosis elevated levels of factor VIII correlate only with von Willebrand factor but not other endothelial cell-derived coagulation and fibrinolysis proteins.

Several studies have shown that patients with venous thrombosis have elevated levels of factor VIII (FVIII) at an increased frequency. Most such patients also have high von Willebrand factor (vWF) levels. Since vWF is synthesized by the vascular endothelium, we hypothesized that elevated FVIII levels would also be associated with an increase of other endothelial cell-derived coagulation proteins suggesting perturbation of the endothelium. In 100 healthy individuals and 129 patients with venous thromboembolism, we have determined antigenic FVIII levels along with several endothelial proteins including vWF, soluble thrombomodulin (sTM), tissue-type plasminogen activator (t-PA), and plasminogen activator inhibitor 1 (PAI-1). Levels of FVIII, vWF, PAI-1 and t-PA were significantly increased in patients compared with the controls (FVIII, vWF, and PAI-1,P < 0.001; t-PA, P < 0.05). Levels of sTM, however, were higher in the controls than in the patients (P < 0.001). Whereas the FVIII levels correlated well with the vWF levels in the patients (correlation, 0.61; P < 0.001) and the controls (correlation, 0.70; P < 0.001), there was neither a relevant correlation between FVIII and sTM, PAI-1, and t-PA, nor between vWF and sTM, PAI-1, and t-PA in the patients and the controls. In conclusion, although levels of PAI-1 and t-PA can be found, on average, at increased levels in patients with thrombosis, FVIII levels correlate only with vWF but not other endothelial cell-derived coagulation and fibrinolysis proteins including sTM, PAI-1, and t-PA.

Molar substitution and C2/C6 ratio of hydroxyethyl starch: influence on blood coagulation.

BACKGROUND: Development of hydroxyethyl starches (HES) with a low impact on blood coagulation but a long intravascular persistence is of clinical interest. A previous in vitro study showed that low substituted high molecular weight HES does not compromise blood coagulation more than medium molecular weight HES. In the present study we assessed the individual effects on blood coagulation of molar substitution and C2/C6 ratio of a high molecular weight HES. METHODS: Blood was obtained from 30 healthy patients undergoing elective surgery and mixed with six high molecular weight (700 kDa) HES solutions differing in their molar substitution (0.42 and 0.51) and C2/C6 ratio (2.7, 7 and 14) to achieve 20, 40 and 60% dilution. Blood coagulation was assessed by Thrombelastograph analysis (TEG) and plasma coagulation tests. Data were compared using a three-way analysis of variance model with repeated measures on the three factors. RESULTS: Higher molar substitution compromised blood coagulation most (for all TEG parameters, P<0.05). The lowest C2/C6 ratio was associated with the lowest effect on blood coagulation; r (P<0.001), angle alpha (P=0.003) and coagulation index (P<0.001). No effect on k and maximum amplitude was observed (P for both >0.50). The higher molar substitution was associated with a lesser increase in PT (P=0.007) and a greater decrease in factor VIII (P=0.010). PTT, functional and antigenic von Willebrand factors were not significantly influenced by molar substitution (P for all >0.20). No significant differences between solutions with the same molar substitution but different C2/C6 ratios were found in plasma coagulation parameters (P for all >0.05). CONCLUSIONS: TEG analysis indicates that high molecular HES with a molar substitution of 0.42 and a C2/C6 ratio of 2.7 has the lowest effect on in vitro human blood coagulation.

Molecular weight of hydroxyethyl starch: is there an effect on blood coagulation and pharmacokinetics?

BACKGROUND: The development of hydroxyethyl starches (HES) with low impact on blood coagulation but higher volume effect compared with the currently used HES solutions is of clinical interest. We hypothesized that high molecular weight, low-substituted HES might possess these properties. METHODS: Thirty pigs were infused with three different HES solutions (20 ml kg(-1)) with the same degree of molar substitution (0.42) but different molecular weights (130, 500 and 900 kDa). Serial blood samples were taken over 24 h and blood coagulation was assessed by Thromboelastograph analysis and analysis of plasma coagulation. In addition, plasma concentration and in vivo molecular weight were determined and pharmacokinetic data were computed based on a two-compartment model. RESULTS: Thromboelastograph analysis and plasma coagulation tests did not reveal a more pronounced alteration of blood coagulation with HES 500 and HES 900 compared with HES 130. In contrast, HES 500 and HES 900 had a greater area under the plasma concentration-time curve [1542 (142) g min litre(-1), P<0.001, 1701 (321) g min litre(-1), P<0.001] than HES 130 [1156 (223) g min litre(-1)] and alpha half life (t(alpha)(1/2)) was longer for HES 500 [53.8 (8.6) min, P<0.01] and HES 900 [57.1 (12.3) min, P<0.01] than for HES 130 [39.9 (10.7) min]. Beta half life (t(beta)(1/2)), however, was similar for all three types of HES [from 332 (100) to 381 (63) min]. CONCLUSIONS: In low-substituted HES, molecular weight is not a key factor in compromising blood coagulation. The longer initial intravascular persistence of high molecular weight low-substituted HES might result in a longer lasting volume effect.

[Toxic shock syndrome in infection due to Streptococcus pyogenes]. / Toxisches Schock-Syndrom bei Streptokokkus-pyogenes-Infektionen.

Acute infections with group A beta-hemolytic streptococcus normally take a favourable course under therapy with penicillin. Only in a few cases has a completely different manifestation been described with multisystem failure similar to toxic shock syndrome induced by Staphylococcus aureus. We report on 4 patients (1990/91) who showed this manifestation in spite of immediate antibiotic therapy. In 3 patients the suspected portal of entry was the skin, in 1 patient it was unknown. Group A streptococci were grown from blood cultures from all 4 patients. Without an underlying immune deficiency all 4 patients (age 22, 24, 38 and 51) went into septic shock with high fever, hepatic and renal impairment, diarrhea, DIC and cerebral confusion. 2 patients died within a few days after developing acute respiratory distress syndrome and cerebral edema. All strains isolated from the patients were penicillin-sensitive, group A streptococci. 3 of them were M-type 1, which are known to be more invasive. The bacteremia by itself is not sufficient to explain all complications and the high mortality rate. It is probable that streptococcal toxins, such as pyrogenic exotoxin A, streptolysin O, or a new unknown one, play a decisive role.

Updates in perioperative coagulation: physiology and management of thromboembolism and haemorrhage.

Understanding of blood coagulation has evolved significantly in recent years. Both new coagulation proteins and inhibitors have been found and new interactions among previously known components of the coagulation system have been discovered. This increased knowledge has led to the development of various new diagnostic coagulation tests and promising antithrombotic and haemostatic drugs. Several such agents are currently being introduced into clinical medicine for both the treatment or prophylaxis of thromboembolic disease and for the treatment of bleeding. This review aims to elucidate these new concepts and to outline some consequences for clinical anaesthesia and perioperative medicine.

Coagulation activation markers do not correlate with the clinical risk of thrombosis in pregnant women.

OBJECTIVE: Because coagulation activation markers have been shown to indicate an increased risk of thrombosis, we tested whether thrombin-antithrombin III complexes and D-dimers correlated with the risk assessment in pregnant women on the basis of clinical data. STUDY DESIGN: We divided a group of 261 pregnant women (305 pregnancies) into low- and high-risk groups according to the personal and family histories of thrombosis and the presence of a hereditary or an acquired thrombophilia. Women with a thrombotic event in the current pregnancy formed a separate group. All pregnancies with or without heparin therapy were closely monitored with thrombin-antithrombin III and D-dimer values for the entire course of the pregnancy. Retrospectively, the data were then correlated with the different groups and subgroups. RESULTS: The course of the mean thrombin-antithrombin III values of all 305 pregnancies was close to or slightly above the upper cutoff line, whereas the D-dimer values were well within the normal range. Independent of heparin, there was no difference in the course of the thrombin-antithrombin III and D-dimer values between the low- and high-risk groups. Only women with ongoing thrombosis during pregnancy had significantly higher thrombin-antithrombin III and D-dimer values with or without heparin therapy. Among those individuals with elevated thrombin-antithrombin III or D-dimer values, there were no specific, recognizable patients who had elevated markers more often than others. CONCLUSIONS: Thrombin-antithrombin III and D-dimer values do not correlate with a risk stratification assessed by clinical criteria. There are many women at low clinical risk who have elevated markers, and there are many women at very high clinical risk who have normal markers. Thus thromboprophylaxis would often be used inadequately if the indication were based on coagulation markers.

Evaluation of an optimal dose of low-molecular-weight heparin for thromboprophylaxis in pregnant women at risk of thrombosis using coagulation activation markers.

There is no consensus on the dose of low-molecular-weight (LMW) heparin for thromboprophylaxis in pregnant women at increased risk of thrombosis. Based upon monitoring with anti-factor Xa activity, the studies showed conflicting results suggesting either fixed dosages throughout the pregnancy or dosages adapted to the gestational age. We tested whether monitoring thromboprophylaxis with D-dimers and thrombin-antithrombin complexes (TAT) would provide additional information on the optimal dose of LMW heparin. Women (165 women and 202 pregnancies) with hereditary or acquired thrombophilia or a history of thrombosis were considered to receive prophylactic LMW heparin therapy. All women received initially 5,000 IU/day of dalteparin s.c. All further dosages were determined solely on the basis of TAT and/or D-dimer values which were determined every 2-3 weeks. As soon as one of these values increased above the normal range, the dose of LMW heparin was adjusted. In 84.6% of all pregnancies, TAT and/or D-dimer values increased above the normal range once or more during the pregnancy. Consequently, the dose of LMW heparin had to be adjusted at least once over the course of the pregnancy. The mean daily dose of LMW heparin increased from 5,000 to 11,200 U between the 6th and 40th week of gestation. Adverse effects included one major bleeding and six local complications, but no thromboembolic event. In conclusion, increasing doses of LMW heparin are needed to keep TAT and D-dimers within the normal range during pregnancy. Hence, to suppress thrombin generation, apparently, the dosage of LMW heparin should be adjusted to the gestational age rather than using fixed doses throughout the pregnancy. However, it remains to be further established whether elevated TAT and D-dimers truly reflect a prothrombotic state during pregnancy.

Cyclosporine-induced detachment of vascular endothelial cells initiates the intrinsic coagulation system in plasma and whole blood.

Cyclosporine A (CsA) is supposed to alter the metabolism of vascular endothelial cells, leading to a prothrombotic state. We examined by which mechanism human umbilical vein endothelial cells (HUVECs) treated with CsA would promote coagulation in human plasma and in whole blood. Treatment of HUVECs with CsA at concentrations clinically used led to dose-dependent cell detachment, with the subsequent exposure of the highly procoagulant connective tissue. As determined by scanning electron microscopy, cell counting of detached and adherent cells, and antigenic measurement of collagen exposure, HUVECs treated with 0.4 micrograms/ml CsA (or more) for 4 days exhibited significant amounts of subendothelial areas. On CsA-treated HUVEC monolayers, the clotting time of recalcified citrated platelet-rich plasma (PRP), but not platelet-poor plasma (PPP), was dose-dependently shortened. Likewise, the onset of thrombin generation was significantly earlier. Except at a high concentration of 8.0 micrograms/ml CsA, there was no procoagulant effect when PPP was used. To investigate CsA-treated HUVECs in whole blood, cells were cultivated on globular microcarriers and were incubated with nonanticoagulated whole blood. When untreated cells were used, generation of factor Xa, thrombin, and kallikrein was completely suppressed for 30 minutes. HUVEC beads treated with 0.4 and 0.8 micrograms/ml CsA, however, led to a dose-dependent generation of all three coagulation factors, with peak values at 2.5 to 5 minutes. Extrinsic activation was excluded, since CsA treatment did not induce tissue factor activity in HUVECs. Furthermore, the thrombomodulin activity of HUVECs w as not altered by CsA. In conclusion, treatment of HUVECs with CsA for 4 days at concentrations clinically used leads to the exposure of subendothelial areas that induce activation of the intrinsic coagulation in recalcified PRP and nonanticoagulated whole blood.

Endothelial cells undergoing apoptosis become proadhesive for nonactivated platelets.

Under normal conditions, platelets do not adhere to endothelium. However, when platelets or endothelial cells are stimulated by thrombin or cytokines, respectively, platelets bind avidly to endothelium. Because there is accumulating evidence that endothelial cells may become apoptotic under certain proinflammatory or prothrombotic conditions, we investigated whether endothelial cells undergoing apoptosis may become proadhesive for nonactivated platelets. Human umbilical vein endothelial cells (HUVEC) were induced to undergo apoptosis by staurosporine, a nonspecific protein kinase inhibitor, or by culture in suspension with serum-deprivation. After treatment of HUVEC or platelets with different receptor antagonists, nonactivated, washed human platelets were allowed to adhere to HUVEC for 20 minutes. To exclude matrix involvement, platelet binding was measured in suspension by using flow cytometry. Independent of the method of apoptosis induction, there was a marked increase in platelet binding to apoptotic HUVEC. Although HUVEC exhibited maximal adhesiveness for platelets after 2 to 4 hours, complete DNA fragmentation of HUVEC occurred only several hours later. Adhesion assays after blockade of different platelet receptors showed only involvement of beta1-integrins. Platelet binding to apoptotic HUVEC was inhibited by more than 70% when platelets were treated with blocking anti-beta1 antibodies. Treatment of apoptotic HUVEC with blocking antibodies to different potential platelet receptors, including known ligands for beta1-integrins, did not affect platelet binding. As assessed by determination of beta-thromboglobulin and platelet factor 4 in the supernatants, platelets bound to apoptotic HUVEC became slightly activated. However, significant expression of platelet P-selectin (CD62P) was not found. These data provide further evidence that endothelial cells undergoing apoptosis may contribute to thrombotic events.

Dose- and shear rate-dependent effects of heparin on thrombogenesis induced by rabbit aorta subendothelium exposed to flowing human blood.

The effect of heparin on thrombogenesis induced by the subendothelium of rabbit aorta was investigated in 24 healthy volunteers after intravenous injection of different doses (0, 1000, 2500, and 5000 IU). By using an ex vivo perfusion chamber system, the interaction between flowing blood and exposed subendothelium was measured at low (50 s-1), intermediate (650 s-1), and high (2600 s-1) wall shear rates. The low shear rate simulated blood flow in venous, the intermediate shear rate in arterial, and the high shear rate in small or stenosed arterial vessels. Deposition of fibrin, platelets, and platelet thrombi on vascular subendothelium (SE) was quantified by morphometrical and immunological techniques. Fibrin deposition prevailed at low shear rates and was only minimal at high shear rates (30 +/- 1% vs. 1 +/- 0.4% coverage of SE with fibrin, means +/- SEM, p less than 0.001). In contrast, the interaction of platelets with SE was more intense at high compared to low shear rates, as indicated by higher platelet adhesion (54 +/- 5% vs. 4 +/- 1% coverage of SE with platelets, p less than 0.001) and platelet thrombus volumes (4.8 +/- 1.3 vs. 0.5 +/- 0.1 microns 3/microns 2, p less than 0.001). Fibrin deposition on SE was inhibited by heparin in a dose-dependent manner and was abolished after high doses. In addition, high doses of heparin reduced the height and volume of platelet thrombi at low and intermediate wall shear rates, but no effect was found at the high shear rate. Our data show that heparin inhibits the formation of both fibrin and platelet thrombi on vascular subendothelium. The lack of effect of heparin on platelet thrombus formation at high shear rates indicates that thrombin modulates the growth rate and/or stability of platelet thrombi at low and intermediate shear rates, whereas additional factors may control platelet thrombus growth and stability at high shear conditions.

Effects of the oral anticoagulant phenprocoumon on blood coagulation and thrombogenesis induced by rabbit aorta subendothelium exposed to flowing human blood: role of dose and shear rate.

We investigated the effect of oral anticoagulation on thrombogenesis induced by the subendothelium of rabbit aorta. Eighteen healthy volunteers underwent a 2-week treatment with the oral coumarin preparation phenprocoumon to a target international normalized ratio (INR) of 5. By using an ex-vivo perfusion chamber system, the interaction between flowing blood and exposed subendothelium was measured at low (50 sec-1) and high (650 sec-1) wall shear rates. The low shear rate simulated blood flow in venous vessels and the high shear rate simulated blood flow in arterial vessels. Deposition of fibrin, platelets, and platelet thrombi on subendothelium was quantified by morphometric and immunologic techniques. Fibrin deposition prevailed at the low shear rate (183 +/- 57 ng/mm2 vs 46 +/- 16 ng/mm2, low vs high shear rate; mean +/- SEM; p less than 0.01). In contrast, the interaction of platelets with subendothelium was more intense at high shear rates when compared with low shear rates, as indicated by higher platelet adhesion (44% +/- 2% vs 9% +/- 1% coverage of subendothelium with platelets, p less than 0.001) and platelet thrombus volumes (3.6 +/- 0.4 microns 3/microns 2 vs 1.3 +/- 0.3 microns 3/microns 2, p less than 0.001). Fibrin deposition on subendothelium was substantially reduced even at a low intensity of anticoagulation (reduction by 60% at INR 2.1 and by 75% at INR 3.7) and was abolished after high coumarin doses (INR 5.2). In contrast, a significant inhibition of platelet thrombus formation could be achieved only by high doses of phenprocoumon (INR 5.2). Our data indicate that relatively low doses of oral anticoagulants (INR between 2 and 3.5) substantially inhibit the fibrin formation on subendothelium prevailing at venous shear conditions, whereas a high intensity of anticoagulation (INR 4 to 5) is necessary to inhibit the formation of platelet-rich thrombi prevailing at arterial shear conditions.

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.

Extensive thrombus formation in the right ventricle due to a rare combination of arrhythmogenic right ventricular cardiomyopathy and heterozygous prothrombin gene mutation G20210 A.

Arrhythmogenic right ventricular cardiomyopathy is a rare heart muscle disease characterized by right and often left ventricular myocardial atrophy and fibrofatty replacement. Heart failure, arrhythmias and sudden death are characteristic complications. We observed a female in whom arrhythmogenic right ventricular cardiomyopathy was diagnosed due to presyncopes and dyspnea on exertion. A left ventricular thrombus was found echocardiographically, which disappeared with oral anticoagulation. Subsequently, however, extensive thrombus formation in the dilated akinetic right ventricle occurred which was resistant to combined treatment with heparin and oral anticoagulation. Thrombophilia screening showed a mutant prothrombin 20210A allele which is an inherited coagulopathy associated with increased plasma levels of prothrombin and increased risks of mainly venous thrombosis. The patient developed endstage biventricular heart failure and underwent heart transplantation within 3 months after thrombus formation in the right ventricle was diagnosed. In the explanted heart, the thrombus in the right ventricle was impressively large and calcified. In patients with unusual thrombus formation in the heart, coagulopathy may be associated and should be excluded.

A novel beta 1 integrin-dependent mechanism of leukocyte adherence to apoptotic cells.

Adherence of leukocytes to cells undergoing apoptosis has been reported to be dependent on a variety of recognition pathways. These include alpha V beta 3 (CD51/CD61, vitronectin receptor), CD36 (thrombospondin receptor), macrophage class A scavenger receptor, phosphatidylserine translocated to the outer leaflet of apoptotic cell membranes, and CD14 (LPS-binding protein). We investigated the mechanism by which leukocytes adhere to apoptotic endothelial cells (EC). Peripheral blood mononuclear leukocytes and U937 monocytic cells adhered to human or bovine aortic EC induced to undergo apoptosis by withdrawal of growth factors, treatment with the promiscuous protein kinase inhibitor staurosporine, with the protein synthesis inhibitor and protein kinase activator anisomycin, or with the combination of cycloheximide and TNF-alpha. Expression of endothelial adherence molecules such as CD62E (E-selectin), CD54 (ICAM-1), and CD106 (VCAM-1) was not induced or increased by these treatments. A mAb to alpha V beta 3, exogenous thrombospondin, or blockade of phosphatidylserine by annexin V did not inhibit leukocyte adherence. Further, leukocyte binding to apoptotic EC was completely blocked by treatment of leukocytes but not EC with mAb to beta 1 integrin. These results define a novel pathway for the recognition of apoptotic cells.

Low- and medium-molecular-weight hydroxyethyl starches: comparison of their effect on blood coagulation.

BACKGROUND: High-molecular-weight hydroxyethyl starch (HES) compromises blood coagulation more than medium-molecular-weight HES. The authors compared medium molecular weight HES (200 kd [HES200]) and low-molecular-weight HES (70 kd [HES70]). METHODS: In a prospective, double-blind, randomized-sequence crossover study, 22 male volunteers received 15 ml/kg HES200 and HES70. Blood samples were taken before and 5 min, 30 min, 1 h, 2 h, 4 h, 8 h, and 24 h after infusion. The following parameters were analyzed at all time points: prothrombin time, activated partial thromboplastin time, fibrinogen, factor VIII, antigenetic and functional von Willebrand factor, platelets, Thrombelastograph analysis parameters (reaction time, coagulation time, maximum amplitude, angle alpha, and clot lysis 30 and 60 min after maximum amplitude), ionized calcium, hematocrit, HES plasma concentration, molecular weight (weight average and number average), molar substitution, and polydispersity (weight average/number average). Repeated-measures analysis of variance (P < 0.05) was used to compare the response of the aforementioned parameters to the infusion of HES70 and HES200. RESULTS: Both HES solutions had a significant impact on all parameters. A slightly greater compromise with HES200 was found in activated partial thromboplastin time (P = 0.010), factor VIII (P = 0.009), antigenetic von Willebrand factor (P = 0.041), functional von Willebrand factor (P = 0.026), maximum amplitude (P = 0.008), and angle alpha (P = 0.003). No difference was established with the other parameters. HES concentration (P < 0.001), weight average (P < 0.001), number average (P < 0.001), and polydispersity (P < 0.001) were higher with HES200. There was no difference with molar substitution (P = 0.091). CONCLUSIONS: Low-molecular-weight hydroxyethyl starch (70 kd) compromises blood coagulation slightly less than HES200, but it is unclear whether this is clinically relevant.