Magnesium and manganese ions accelerate tissue factor-induced coagulation independently of factor IX.

The purpose of the present study was to assess the effect of magnesium and manganese ions on tissue factor (TF)-induced coagulation and the possible role of factor IX therein. When magnesium chloride or manganese chloride were added in low concentrations to normal human plasma, the human (recombinant) TF-induced coagulation time was shortened. At higher concentrations, magnesium and manganese prolonged the TF-induced coagulation time. Maximum shortening of the coagulation time was obtained at a concentration of 0.5 mmol/l Mn or 2 mmol/l Mg in plasma. Shortening of the TF-induced coagulation time by magnesium and manganese was also observed in factor IX-deficient plasma. A comparison was made between TF preparations from human, rabbit, and bovine brain. The accelerating effect of magnesium was greater with human than with rabbit brain TF. Using bovine brain TF, the clotting time was not shortened by magnesium. Activated factor X-induced coagulation of normal plasma was not accelerated by magnesium. From these experiments, it is inferred that activation of factor X by factor VII-TF can be accelerated by magnesium and manganese ions independently of factor IX.     

Plasma antigen levels of the lipoprotein-associated coagulation inhibitor in patient samples

Human plasma contains an inhibitor of tissue factor-initiated coagulation known as the lipoprotein-associated coagulation inhibitor (LACI) or also known as the extrinsic pathway inhibitor (EPI). A competitive fluorescent immunoassay was developed to measure the plasma concentration of LACI in samples from normal individuals and patients with a variety of diseases. The LACI concentration in an adult control population varied from 60% to 160% of the mean with a mean value corresponding to 89 ng/mL or 2.25 nmol/L. Plasma LACI levels were not decreased in patients with severe chronic hepatic failure, warfarin therapy, primary pulmonary hypertension, thrombosis, or the lupus anticoagulant. Plasma LACI antigen was decreased in some, but not all patients with gram-negative bacteremia and evidence for disseminated intravascular coagulation. Plasma LACI levels were elevated in women undergoing the early stages of labor (29%), in patients receiving intravenous tissue-type plasminogen activator (45%), and in patients receiving intravenous heparin (375%). A radioligand blot of the pre- and post-heparin plasma samples shows the increase to be in a 40-Kd form of LACI. Very low levels of plasma LACI antigen were found in patients with homozygous abetalipoproteinemia and hypobetalipoproteinemia, diseases associated with low plasma levels of apolipoprotein B containing lipoproteins. Following the injection of heparin into one patient with homozygous abetalipoproteinemia, the plasma LACI antigen level increased to a level comparable with that in normal individuals after heparin treatment.     

Automated amidolytic method for evaluating the activated partial thromboplastin time compared with a conventional coagulation method

We report studies of the validity and clinical application of a new amidolytic method for evaluating the activated partial thromboplastin time (APTT) compared with a conventional clotting method. The results with the two methods were well correlated for normal plasma and plasma from hemophilia patients. Congenital deficiencies of of the intrinsic coagulation pathway other than hypo- and dysfibrinogenemia detected by the amidolytic method agreed with those detected by the clotting APTT. The results with the two methods for plasma from patients under heparin treatment were statistically different, suggesting a lesser sensitivity of the amidolytic method to heparinization. The use of the amidolytic APTT reagent in combination with factor VIII and IX deficient plasma allowed the measurement of both factors. The results obtained with normal and hemophilic plasma were in agreement with those obtained with the one-stage clotting method in all except two occasions. Even though the amidolytic method demonstrated the presence of the lupus anticoagulant in the majority of tested samples of known lupus subjects, it was less sensitive to the abnormality than the clotting method.     

The lipoprotein-associated coagulation inhibitor that inhibits the factor VII-tissue factor complex also inhibits factor Xa: insight into its possible mechanism of action

Blood coagulation is initiated when plasma factor VII(a) binds to its essential cofactor tissue factor (TF) and proteolytically activates factors X and IX. Progressive inhibition of TF activity occurs upon its addition to plasma. This process is reversible and requires the presence of VII(a), catalytically active Xa, Ca2+, and another component that appears to be associated with the lipoproteins in plasma, a lipoprotein-associated coagulation inhibitor (LACI). A protein, LACI(HG2), possessing the same inhibitory properties as LACI, has recently been isolated from the conditioned media of cultured human liver cells (HepG2). Rabbit antisera raised against a synthetic peptide based on the N-terminal sequence of LACI(HG2) and purified IgG from a rabbit immunized with intact LACI(HG2) inhibit the LACI activity in human serum. In a reaction mixture containing VIIa, Xa, Ca2+, and purified LACI(HG2), the apparent half-life (t1/2) for TF activity was 20 seconds. The presence of heparin accelerated the initial rate of inhibition threefold. Antithrombin III alpha alone had no effect, but antithrombin III alpha with heparin abrogated the TF inhibition. LACI(HG2) also inhibited Xa with an apparent t1/2 of 50 seconds. Heparin enhanced the rate of Xa inhibition 2.5-fold, whereas phospholipids and Ca2+ slowed the reaction 2.5-fold. Xa inhibition was demonstrable with both chromogenic substrate (S-2222) and bioassays, but no complex between Xa and LACI(HG2) could be visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Nondenaturing PAGE, however, showed that LACI(HG2) bound to Xa but not to X or Xa inactivated by diisopropyl fluorophosphate. Thus, LACI(HG2) appears to bind to Xa at or near its active site. Bovine factor Xa lacking its gamma-carboxyglutamic acid-containing domain, BXa(-GD), through treatment with alpha-chymotrypsin, was used to further investigate the Xa requirement for VIIa/TF inhibition by LACI(HG2). LACI(HG2) bound to BXa(-GD) and inhibited its catalytic activity against a small molecular substrate (Spectrozyme Xa), though at a rate approximately sevenfold slower than native BXa. Preincubation of LACI(HG2) with saturating concentrations of BXa(-GD) markedly retarded the subsequent inhibition of BXa. The VII(a)/TF complex was not inhibited by LACI(HG2) in the presence of BXa(-GD), and further, preincubation of LACI(HG2) with BXa(-GD) slowed the inhibition of VIIa/TF after the addition of native Xa. The results are consistent with the hypothesis that inhibition of VII(a)/TF involves the formation of a VIIa-TF-XA-LACI complex that requires the GD of XA.(ABSTRACT TRUNCATED AT 400 WORDS)     

Hypothermia and blood coagulation: dissociation between enzyme activity and clotting factor levels

Previous studies of hypothermia and blood coagulation have focused on alterations in the levels of blood clotting elements using coagulation tests performed under normothermic conditions. However, because of the enzymatic nature of activated clotting factors, hypothermia should also be expected to affect clotting factor activities. Multiple determinations of activated partial thromboplastin times (APTT), prothrombin times (PT), and thrombin times (TT) were performed on commercially available normal human plasma at assay temperatures similar to those encountered clinically (25-37 degrees C). Both the APTT and the PT were significantly prolonged at temperatures below 35 degrees C (P less than 0.05). Clotting time correlated significantly with assay temperature in a negative exponential fashion for all three tests (r = -0.97 for APTT, -0.93 for PT, -0.71 for TT, P less than 0.001 for all regressions). Clotting time prolongation appears proportional to the number of enzymatic steps involved. These data indicate that the coagulopathy observed during hypothermia is, in part, independent of clotting factor levels.     

Effects of α-tocopherol on platelets and the coagulation system

Vitamin E is one of the most widely used antioxidants in cryopreservation and preservation technology. The objective of this study is to examine the effect of vitamin E on platelets and the coagulation system. Vitamin E was added at different concentrations in the range between 0.25 and 5 mM to donor plasma. Using a STA/STA Compact coagulation analyzer the following clotting tests were performed: prothrombin time (PT), activated partial thromboplastin time (APTT), thrombin time (TT). The control clotting times PT (13.80 - 0.4 s), APTT (27.4 - 2.4 s) and TT (17.6 - 0.4 s) remained unchanged in the presence of vitamin E. The effect of vitamin E on platelets was assessed by platelet-induced clot retraction (PICR) and aggregation by thrombin. PICR was unaffected by vitamin E. Platelet aggregation, however, was profoundly inhibited by vitamin E. We found that inhibition of platelet aggregation by vitamin E was concentration dependent: increasing with increasing vitamin E concentration. This inhibitory effect, however, was widely reversible upon dilution of vitamin E with autologus platelet-poor plasma.     

联合输注新鲜冰冻血浆和冷沉淀凝血因子的止血效果观察

目的:探讨联合输注新鲜冰冻血浆(FFP)和冷沉淀凝血因子用于肝肿瘤术后凝血功能异常患者的止血效果。方法:将120例肝肿瘤术后凝血功能异常患者随机分为3组:FFP单独输注组、冷沉淀凝血因子单独输注组及FFP和冷沉淀凝血因子联合输注组,每组各40例。所有患者住院期间均行肝肿瘤切除术,术后进行常规治疗,联合或单独输注FFP和冷沉淀凝血因子,3组分别于输注前1h及输注后24h静脉采血,检测输注前后凝血酶原时间(PT)、部分凝血活酶时间(APTT)、凝血酶时间(TT)和纤维蛋白原(FIB)。结果:3组输注后PT、APTT、TT、FIB和输注前1h比较均差异有统计学意义(均P0.01);与2个单独输注组比较,联合输注组PT、APTT、TT数值均差异有统计学意义(均P0.05),FIB数值与FFP单独输注组比较也有明显提高(P0.05)。结论:联合或单独给肝肿瘤术后凝血功能异常患者进行输注均能改善患者的凝血功能,联合输注的止血效果要好于单独输注一种成分。     

A step change in oral anticoagulation: lack of coagulation monitoring with ximelagatran

The clinical development of ximelagatran for the treatment and prevention of various arterial and venous thromboembolic disorders has used fixed-dose regimens without coagulation monitoring in all indications. Although monitoring is not required, effects on the various coagulation assays that are available are seen with its active form melagatran, and there are situations where an assessment of anticoagulant effect may help to inform clinical decisions. However, the sensitivity of different coagulation assays varies considerably. The thrombin clotting time (TT) and ecarin clotting time (ECT) are highly sensitive to plasma melagatran concentrations (IC 50 approximately 0.01 micromol/L and approximately 0.15 micromol/L, respectively), with an approximate linear relationship between plasma melagatran concentration and prolongation of clotting time. In comparison, the activated partial thromboplastin time (APTT) (IC 50 approximately 0.3 to 0.8 micromol/L) and prothrombin time (PT) (IC 50 approximately 0.9 to 2.9 micromol/L) are relatively insensitive, and the concentration-response relationship shows a flattening with increasing plasma melagatran concentration. Commercially available APTT and PT reagents varied considerably in their sensitivity to melagatran. Comparing the various coagulation assays, the APTT, ECT, and TT are suitable choices when an indicator of the anticoagulant effect of ximelagatran is required, although the absence of international standards requires calibration of each test in individual laboratories and the ECT is not widely available.     

Coagulation protein function: enhancement of the anticoagulant effect of acetaldehyde by sulfated glycosaminoglycans

In view of the increased anticoagulant effect of acetaldehyde-treated heparin, other glycosaminoglycans (GAGs) such as chondroitin sulfates A and C, dermatan sulfate (chondroitin sulfate B), heparan sulfate, and hyaluronic acid were tested for anticoagulant activity before and after exposure to acetaldehyde. Clotting times of human plasma Ci-Trol coagulation control, level I (Baxter Healthcare Corp.), were tested in the presence of 1.8, 3.0, 3.6, or 4.5 g heparin (0.32, 0.54, 0.64, 0.81 units heparin). Additionally, 9, 27, or 90 g of chondroitin sulfates A, B, or C was utilized in lieu of heparin. The effects of 2 g heparin (0.36 units), chondroitin sulfates A, B, and C, (20 g each), 2 g heparan sulfate, and 2 g hyaluronic acid, respectively, in the presence of 44.7 mM acetaldehyde on the clotting time of plasma were studied. It was observed that chondroitin sulfate B (dermatan sulfate) prolonged the clotting time of plasma, although to a lesser extent than heparin. Chondroitin sulfates A and C, heparan sulfate, and hyaluronic acid did not prolong clotting time. However, pretreatment of all the sulfated GAGs with acetaldehyde gave products that enhanced the anticoagulant effect of acetaldehyde, notwithstanding the lack of anticoagulant effect of the GAGs. In contrast, hyaluronic acid exhibited no effect upon clotting time nor did its acetaldehyde-treated product. Furthermore, ethanol exhibited no effect upon the clotting times of the GAG–plasma mixtures. These results suggest that sulfated GAGs may be modified by acetaldehyde, a component of plasma in chronic alcoholics, and that the resultant products may contribute to the prolonged clotting times.     

比较国产比伐卢定与肝素对大鼠颈总动脉血栓模型凝血系统的影响

目的比较国产比伐卢定（bivalirudin,B）与肝素（heparin,H）对大鼠颈总动脉血栓模型凝血系统的影响。方法以50％FeCl3诱导大鼠左颈总动脉血栓形成建立模型。42只SD大鼠分为假手术组（S组,n=6）、血栓模型组（M组,n=12）、国产比伐卢定联合尿激酶（urokinase,UK）组（B＆UK组,n=12）、肝素联合UK组（H＆UK组,n=12）,观察4组给药前与用药后2h活化部分凝血活酶时间（activated partial thromboplastin time,APTT）、凝血酶原时间（prothrombin time,PT）及血栓质量的变化。结果与S组比较,M组、B＆UK组及H＆UK组给药前APTT、PT均缩短,但后3组间比较差异无统计学意义（P〉0．05）;与S组比较,给药后M组APTT、PT明显缩短：与S组及M组比较,给药后B＆UK组及H＆UK组APTT、PT均明显延长（P〈0．01）,而H＆UK组APTT比B＆UK组显著延长,相反,B＆UK组盯比H＆UK组显著延长。S组血管内无血栓形成．B＆UK组及H＆UK组血栓质量均显著低于M组[（2．14±0．40）mg比（3．28±0．58）mg,（2．41±0．58）mg比（3．28±0．58）mg,P均〈0．01]。结论国产比伐卢定与肝素均能在大鼠UK溶栓过程中发挥抗凝及抑制血栓形成作用,且国产比伐卢定比肝素抑制外源性凝血系统的作用更明显,能更有效减少出血风险。     

Coagulation Protein Function VI (Augmentation of Anticoagulant Function by Acetaldehyde-Treated Heparin)

Acetaldehyde (AcH) at preincubationconcentrations of 447, 89.4, and 17.9 mM potentiates theeffects of heparin on the clotting time of plasma. Whilecontrol plasma clotted in the range of 12.6 ± 0.1 to 13.8 ± 0.1 sec, and heparin-treatedplasma clotted in a range from 131.5 ± 2.5 to168.2 ± 1.2 sec, heparin that was preincubated atroom temperature for 30 min with 89.4 or 447 mM AcH didnot clot plasma in 300 sec. Heparin exposed to 17.9 mMAcH clotted plasma in 193 ± 1.1 sec. Ethanol ata 404 mM concentration also prolonged the clotting timeof heparin-treated plasma >300 sec, while 202 mM ethanol prolonged the clotting time ofheparin-treated plasma from 149.0 ± 2.0 sec to219.5 ± 1.7 sec. It is suggested that AcH altersthe tertiary structure of heparin by adduct formation,possibly by formation of cyclic acetals with iduronicand glucuronic acids, thereby more readily affectingbinding of the glycosaminoglycan to antithrombin IIIand/or thrombin, prolonging clotting time. Ethanol, which does not react covalently with heparin,might affect its conformation as a consequence of anorganic solvent effect. Protamine sulfate prolonged theclotting time of plasma from 13.6 ± 0.1 sec to 17.9 ± 0.2 sec. Protaminesulfate-treated heparin clotted plasma in 21.0 ±0.4 sec relative to heparin-treated plasma (160.4± 1.7 sec). In subsequent experiments,AcH-treated protamine sulfate extended the clotting time of protamine sulfate from17.9 ± 0 sec to 33.7 ± 0.6 sec. Prioraddition of protamine sulfate to AcH- heparin mixturesor heparin to protamine sulfate-AcH mixtures beforeaddition to plasma resulted in clotting times of 22.0± 0.4 sec and 24.1 ± 0.5 sec,respectively, relative to control clotting times of162.3 ± 2.6 sec for plasma-heparin mixtures.These results confirm both the reduction in coagulation time ofheparin-treated plasma by protamine sulfate and theprolongation of clotting time of plasma by protaminesulfate. Furthermore, and importantly, they indicatethat acetaldehyde-treated protamine sulfate is a more effectiveanticoagulant than protamine sulfate. It is suggestedthat reversible adduct formation between acetaldehyde,heparin, and protamine sulfate may occur as a meansexplaining the essentially identical coagulation time ofthese mixtures when added to plasma regardless of theorder of premixing. Ethanol (404 mM) did not influenceprotamine sulfate effects. Lastly, the potentiation of the anticoagulant function of heparin byacetaldehyde suggests that a structural modification ofthe glycosaminoglycan may occur in alcoholics.     

Relative neutralization of the biological actions of sulfaminoheparosans (K5 derivatives) and heparins by protamine sulfate.

Biosynthetic, semisynthetic, and synthetic analogues of heparins are currently developed as substitute antithrombotic agents for heparin. Sulfaminoheparosan (SAH) represents a bacterial polysaccharide (K5)-derived antithrombotic polymer from which pharmacologically active products with varying molecular weights (5-25 kDa) can be derived. These agents have been shown to exhibit pharmacologic effects comparable to heparins. The objective of this investigation is to determine the relative neutralization profile of various SAH derivatives, also called as bioheparins, by protamine sulfate. Four SAH fractions with varying molecular weights (20, 9, 7, and 6 kDa), a low molecular weight heparin (LMWH), tinzaparin, and unfractionated heparin (UFH) were supplemented to normal human pool plasma over a concentration range of 6.2 to 100 microg/mL. A fixed amount of protamine sulfate at 25 microg/mL (final concentration) was supplemented to determine the neutralization profile by performing tests such as prothrombin time (PT), activated partial thromboplastin time (APTT), Heptest, prothrombin-induced clotting time (PiCT), and amidolytic anti-Xa and anti-IIa assays. Protamine sulfate produced varying degrees of neutralization of all bioheparin fractions in the clotting assays. In the amidolytic anti-IIa assay relatively stronger inhibition was noted for all agents than inhibition of FXa. On a cumulative basis the neutralization profile of SAHs was comparable with heparins. These results suggest that the anticoagulant activities of SAH derivatives can be antagonized by protamine sulfate. These studies warrant further in vivo investigation to validate the relative neutralization profile of sulfaminoheparosans.     

Inhibition of chymotrypsin by heparin cofactor II.

Human heparin cofactor II is a plasma protein that is known to inhibit thrombin. The rate of thrombin inhibition by heparin cofactor II is accelerated (greater than or equal to 1000-fold) in the presence of the glycosaminoglycans, heparin and dermatan sulfate. We have found that chymotrypsin A alpha is also inhibited by heparin cofactor II with a second-order rate constant value of 1.8 X 10(6) M-1 X min-1 at pH 8.0 and 25 degrees C. However, there was no measurable effect of heparin or dermatan sulfate on the rate of chymotrypsin inhibition. Arginine-modified heparin cofactor II showed a comparable percentage loss of both antichymotrypsin and antithrombin activities. Heparin cofactor II and chymotrypsin formed a stable complex with a Mr value near 90,000 when analyzed by NaDodSO4/polyacrylamide gel electrophoresis; this suggests a 1:1 reaction stoichiometry. The chymotrypsin cleavage site in heparin cofactor II was the same as that for thrombin, and primary structure analysis of the inhibitor showed a P'1-P'8 sequence of Ser-Thr-Gln-Val-Arg-Phe-Thr-Val ... . The results indicate that, in contrast to alpha 1-antichymotrypsin, which does not inhibit trypsin-like enzymes, including thrombin, heparin cofactor II can effectively inhibit both thrombin and chymotrypsin.     

Respective role of antithrombin III and heparin cofactor II in the in vitro anticoagulant effect of heparin and of various sulphated polysaccharides

The in vitro anticoagulant effects of standard heparin (SH) and of seven other sulphated polysaccharides (SPS) were investigated by measuring activated partial thromboplastin time (APTT) prolongation of normal plasma and of plasmas selectively depleted of antithrombin III (AT III), of heparin cofactor II (HC II) and of both heparin cofactors. This allowed the determination of the relative contribution of each of the two heparin cofactors to the SPS anticoagulant effect. The SPS varied in their relative activities as catalysts of thrombin inhibition by purified AT III or HC II. The anticoagulant activities of heparin and dermatan sulphate were primarily attributable to their ability to enhance thrombin inhibition by AT III and HC II respectively. Heparin had an additional minor anticoagulant activity which was independent of both AT III and HC II. Pentosan polysulphate, high molecular weight dextran sulphate, heparin with low affinity for AT III and a sulphated heparin derivative had weaker anticoagulant activities in normal plasma than standard heparin. The anticoagulant activities of these last four SPS in plasma depleted of both AT III and HC II were similar to their respective activities in normal plasma. This suggests that these SPS act by directly preventing thrombin generation rather than by enhancing thrombin inhibition.     

肝素治疗在三硝基苯磺酸诱导的结肠炎模型中的作用

目的 探讨三硝基苯磺酸(TNBS)诱导的大鼠实验性结肠炎模型中凝血异常与炎症的关系.方法 将40只SD大鼠分为4组:正常对照组、结肠炎组、肝素治疗组和柳氮磺吡啶(SASP)治疗组.检测各组的PT、APTT、抗凝血酶(AT)活性,及组织大体评分和病理评分、血中TNFα水平,并对结果进行统计学分析.结果 TNBS诱导的结肠炎模型中PT及APTT均比正常对照组缩短[(14.83±0.45)s比(16.68±1.08)s及(12.49±1.30)s比(29.06±1.60)8,P值均<0.05],AT活性较正常对照组下降[(111.33±8.50)%比(122.13±3.52)%,P<0.05].肝素治疗组的PT及APTT.均较结肠炎组延长[(17.83±0.78)s比(14.83±0.45)s及(53.34.±9.49)8比(12.49±1.30)s,P值均<0.05],AT活性较结肠炎组为高[(131.67±6.92)%比(111.33±8.50)%,P<0.05].SASP治疗组与结肠炎组比较,PT和APTT差别无统计学意义(P>0.05),AT活性较结肠炎组高[(122.33±5.82)%比(111.33±8.50)%,P<0.05].肝素治疗组大体评分低于结肠炎组(2.50±0.55比4.75±1.16,P<0.05),组织病理评分明显低于结肠炎组(3.83±0.41比7.75±1.04,P<0.05),TNFα水平比结肠炎组减低[(84.75±18.03)ag/L比(149.93±23.52)ng/L,P<0.05].结论 TNBS诱导的实验性结肠炎模型中存在凝血异常;肝素治疗TNBS诱导的实验性结肠炎模型有效.这提示凝血系统异常在实验性结肠炎发生发展中可能起到一定作用.     

Utilization of a continuous flow reactor to study the lipoprotein-associated coagulation inhibitor (LACI) that inhibits tissue factor

A microperfusion system containing a glass capillary, the inner surface of which is coated with a phospholipid bilayer containing tissue factor, was used to explore the requirement for factors VIIa and Xa in the complex formed with the lipoprotein-associated coagulation inhibitor (LACI). Various combinations of factors VIIa, Xa, and LACI were perfused together or sequentially at a wall shear rate of 300 sec-1; a final perfusion of factors X and VIIa was performed to evaluate the residual tissue factor activity. Factor Xa concentration at the outlet of the tube was determined using a chromogenic substrate. In the presence of factors VIIa, Xa, and LACI, complete inhibition of tissue factor was observed on both phosphatidylcholine (neutral surfaces) and on phosphatidylserine/phosphatidylcholine (acidic) surfaces; omission of factors Xa or LACI resulted in no inhibition. The absence of factor VIIa in the initial perfusion steps resulted in no inhibition on neutral surfaces whereas about 90% inhibition was observed on acidic surfaces. Initial perfusion with factor Xa, but not LACI, followed by the remaining protein components, resulted in an inhibitory complex. Thus, it appears that a tissue factor:factor Xa:LACI complex can form in the absence of factor VIIa on acidic surfaces; moreover, our data imply a tissue factor binding site for factor Xa, but not for LACI.     

Inhibition of coagulation and thrombin-induced platelet activities by a synthetic dodecapeptide modeled on the carboxy-terminus of hirudin

A synthetic, tyrosine-sulfated, dodecapeptide (BG8865) modeled on residues 53-64 of hirudin was found to elevate the activated partial thromboplastin time (APTT), prothrombin time (PT), and thrombin time (TT) of human plasma in a dose-dependent manner. The most sensitive assay was the TT, which was prolonged 2 and 3 times control values at 2.2 and 4.1 micrograms/mL hirudin peptide, respectively. The sulfated dodecapeptide exhibited no dependency on antithrombin III as monitored by the APTT in the presence of sheep anti-human antithrombin III antibodies, and its activity was not neutralized by platelet releasates or platelet factor 4. In studies of thrombin-induced platelet activation, the hirudin peptide was found to block aggregation, serotonin release and thromboxane A2 generation. At thrombin concentrations of 0.25 U/mL, the IC50 (concentration resulting in 50% inhibition) for inhibition of platelet aggregation was 0.72 micrograms/mL peptide. Inhibition of TXA2 generation and serotonin release correlated closely with inhibition of aggregation. Using platelets from patients with clinically documented heparin-induced thrombocytopenia anticoagulant doses of heparin were found to induce platelet aggregation and thromboxane A2 generation. In sharp contrast, anticoagulant-equivalent doses of hirudin peptide had no effect on patient platelets, as evidenced by a lack of platelet aggregation and thromboxane A2 generation. These data provide compelling in vitro evidence that the hirudin peptide has several potential advantages over heparin, namely effective inhibition of thrombin-induced platelet activities, co-factor independence, insensitivity to endogenous heparin-neutralizing factors, and an apparent lack of direct or immune-mediated platelet stimulating properties.     

Vitamin K deficiency amplifies anticoagulation response to ximelagatran: possible implications for direct thrombin inhibitors and their clinical safety

Dietary vitamin K is known to influence the anticoagulation response to warfarin. It is possible that dietary vitamin K availability also influences the pharmacological activity of other oral anticoagulants, which target the vitamin-K dependent clotting proteins in the coagulation cascade. This study examined whether vitamin K insufficiency affected anticoagulation response to the direct thrombin inhibitor, ximelagatran. Anticoagulation response to ximelagatran and warfarin in rats on a normal diet was compared to those on a vitamin K deficient diet. Ximelagatran and warfarin increased prothrombin time (PT) by 1.4- and 1.3-fold, activated partial thromboplastin time (APTT) by 1.8- and 1.4-fold and ecarin clotting time (ECT) by 6.8- and 1.2-fold, respectively, in rats on normal diet. Vitamin K deficient diet alone caused modest increases in PT, APTT and ECT. The anticoagulant activity of both ximelagatran and warfarin was significantly greater in rats on vitamin K deficient diet (6.1- and 12.3-fold for PT, 2.6- and 5.1-fold for APTT and 2.9- and 1.6-fold for ECT, respectively) compared to those on normal diet. Factor II activity was reduced by both ximelagatran (58%) and warfarin (44%) in rats on normal diet. However, factor II activity was virtually abolished (<0.1%) by both drugs in rats on vitamin K deficient diet. The results suggest that oral anticoagulant drugs, whose primary site of action is not within the vitamin K cycle, may also exhibit variability in clinical response due to dietary variation as the established coumarin drugs such as warfarin.     

国产注射用比伐卢定对冠状动脉介入治疗患者凝血功能的影响

目的探讨国产注射用比伐卢定对经皮冠状动脉介入(PCI)治疗术患者凝血功能的影响。方法随机选择50例择期行PCI的患者,术中用肝素(对照组,25例)或国产注射用比伐卢定(比伐卢定组,25例)抗凝。分别于PCI术前、用药后5 min、术后即刻、停药后30 min、停药后2 h测活化凝血时间(ACT)。用药前、用药结束后6、24、72 h,静脉采血,检测活化凝血酶时间(APTT)、凝血酶原时间(PT)、凝血酶时间(TT)和纤维蛋白原(FIB)。结果用药后5 min及术后即刻比伐卢定组ACT显著高于对照组(均为P<0.001);PCI术前和停药后30 min两组患者ACT差异无统计学意义(P=0.362、P=0.732),停药后2 h比伐卢定组ACT显著低于对照组[(208.27±34.84)s比(241.48±41.34)s,P=0.01]。术后凝血功能4项与对照组比较差异无统计学意义(均为P>0.05),两组之间均无心血管临床事件发生(P=1.00),大出血事件两组之间差异也无统计学意义(P=1.00)。结论与常规肝素抗凝相比,国产注射用比伐卢定作为抗凝剂应用于PCI术中,起效更快,效果更强,而且半衰期更短,提示其有效性和安全性比肝素钠更佳。     

高同型半胱氨酸血症与老年冠心病

目的 :探讨高同型半胱氨酸 (HCY)血症与冠心病 (CHD)的关系及其可能的机制。方法 :选择 112例CHD患者 (CHD组 ) ,34例健康体检者 (对照组 ) ,分别测定血浆 HCY、活化的部分凝血活酶时间 (APTT)、血浆凝血酶原时间 (PT)、纤维蛋白原 (FIB)。结果 :CHD组血浆 HCY较对照组显著升高。高 HCY的 CHD组中 APTT、PT较正常 HCY的 CHD组及对照组组显著减低 ,而 FIB则显著升高。结论 :高 HCY血症与 CHD密切相关 ,并通过影响血凝状态而参与 CH D的发生及发展