Effects of subcutaneously administered dermatan sulfate (MF 701) on the coagulation and fibrinolytic parameters of healthy volunteers

Eight healthy volunteers were given single subcutaneous doses of dermatan sulfate (DS, 100, 200 and 400 mg), heparin (5,000 IU) and placebo in random order. Wash-out between treatments was > 10 days. Serial blood samples were taken before and up to 24 hours after treatment to measure coagulation and fibrinolytic parameters. Thrombin generation was significantly inhibited by DS and heparin as compared to placebo. The effect of DS was dose-dependent. Peak inhibition after 200 mg DS was comparable to that of 5,000 IU heparin, but lasted longer. A small, bordeline significant prolongation of APTT was observed after 400 mg DS and heparin. The changes in PAI and fibrinolytic activities were those of the circadian variation. No changes were seen in the other parameters tested. In conclusion, single s.c. doses of DS (200, or 400 mg) inhibit thrombin generation equally or more than 5,000 IU heparin and for a longer time. The effect of both treatments on fibrinolysis is negligible.     

Activated platelets increase fibrinolysis of mesenchymal progenitor cells

Bone regeneration is initiated by the formation of a blood clot. Activated platelets within this fibrin‐rich matrix release signaling molecules that can attract mesenchymal progenitor cells. To gain insight into the cellular mechanism by which activated platelets can support the immigration of mesenchymal progenitors, we have tested the hypothesis that platelet‐released signaling molecules increase the capacity of bone marrow stromal cells (BMSC) to activate plasminogen. We report herein that platelet‐released supernatants (PRS) elevate total urokinase‐type plasminogen activator (uPA) and total plasminogen activator inhibitor‐1 (PAI‐1) levels in BMSC, as assessed by immunoassay. Quantitative polymerase chain reaction showed an upregulation of uPA, uPA receptor, and PAI‐1. Zymography and kinetic analysis based on casein hydrolysis revealed enhanced activity of cell‐associated uPA upon exposure of BMSC to PRS. Inhibiting c‐Jun N‐terminal kinase (JNK) and phosphatidylinositol 3‐kinase (PI3K) signaling reduced uPA production and decreased plasminogen activation. Corresponding Western blot analysis showed increased phosphorylation of JNK and AKT in BMSC treated with PRS. These results suggest that activated platelets can enhance the plasminogen activation capacity of mesenchymal progenitors through the stimulation of uPA production, requiring JNK and PI3K/AKT signaling. By this mechanism platelets may contribute to the organization of the blood clot during bone regeneration. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 27: 972–980, 2009     

Fibrinolysis and fibrinogenolysis in liver disease

Patients with liver disease frequently have hemostatic abnormalities which include accelerated fibrinolysis. In order to assess the fibrinolytic state in liver disease, plasma levels of fibrinogenolysis products (FgDP), fibrinolysis products (FbDP), and fibrinogenolysis plus fibrinolysis products (TDP) were measured with newly developed enzyme-linked immunosorbent assays based on monoclonal antibodies in 36 patients with liver disease (six patients with acute hepatitis, seven with chronic hepatitis, ten with liver cirrhosis, 11 with hepatocellular carcinoma, and two with intrahepatic cholestasis). As compared with healthy subjects, mean plasma levels of FbDP (1,083 +/- SD 1,254 vs. 236 +/- 100 ng/ml, P = 0.005) and TDP (1,773 +/- 1,814 vs. 669 +/- 212 ng/ml, P = 0.001) were significantly elevated in patients with liver disease, whereas FgDP was normal (389 +/- 202 vs. 396 +/- 132 ng/ml, P = 0.87). Plasma FbDP correlated very well with TDP (r = 0.986, P less than 0.00001) in liver disease. In addition, FbDP and TDP but not FgDP correlated with plasma concentrations of thrombin-antithrombin III complex. When plotted by the disease categories, the magnitude of elevations of FbDP and TDP was the most prominent in acute hepatitis followed by hepatocellular carcinoma. These findings indicate that activation of fibrinolysis occurs following thrombin generation, but increased primary fibrinogenolysis is rare in liver disease.     

Low-frequency ultrasound induces nonenzymatic thrombolysis in vitro.

OBJECTIVE: To evaluate whether ultrasound, applied over a distance of several centimeters and in the absence of thrombolytic agents, may have a thrombolytic effect on blood clots. METHODS: Low-frequency (20 kHz) continuous wave ultrasound at different intensity levels (0.15-1.2 W/cm2) and exposure times (5, 10, and 20 minutes) was assessed for its potential to induce thrombolysis of fresh human blood clots. The ultrasound effect was also studied in combination with recombinant tissue-type plasminogen activator-mediated thrombolysis. Experiments were carried out in a flow model in degassed sodium phosphate buffer at 37 degrees C at a distance of 3 cm from the ultrasonic probe to the blood clots. Regardless of ultrasound exposure times, blood clots in all experimental groups and the control group were left in the flow system for 20 minutes. RESULTS: The use of ultrasound alone showed a significant thrombolytic effect compared with the control group, with a statistically significant effect at 0.15 W/cm2 and exposure of 10 minutes (P = .02). There was a clear correlation between the extent of weight loss and the chosen intensity level and exposure time. Complete disruption in 8 of 10 blood clots occurred at 1.2 W/cm2 within 10 min. Addition of ultrasound to recombinant tissue-type plasminogen activator-mediated thrombolysis significantly enhanced thrombolysis compared with application of recombinant tissue-type plasminogen activator or ultrasound alone (P = .0001), with the results pointing toward a purely additive, nonsynergistic effect of the 2 treatment modalities. Lysis was more effective in fresh thrombi. CONCLUSIONS: The use of low-frequency ultrasound alone, without addition of a thrombolytic drug, has the potential to induce thrombolysis over a distance. Combination of ultrasound with recombinant tissue-type plasminogen activator is superior to either treatment alone. Ultrasound is a promising tool for developing an alternative or additional treatment modality for acute cerebral vessel occlusion.     

静脉血栓的抗磷脂蛋白抗体与抗凝血、纤维蛋白溶解关系

为了探讨静脉血栓的病因病理及其与抗凝血、纤维蛋白溶解的关系 ,对 4 7例静脉血栓 (VT)患者用酶联免疫法检测抗心肌磷脂抗体 (ACA) ,用凝血法测定狼疮抗凝物 (LA)和抗活化蛋白C抗性 (APCR) ,用多聚酶链反应内切酶法鉴定因子VLeiden ,用发色底物法测定抗凝血酶Ⅲ (ATⅢ )、蛋白C(PC)、纤溶酶原 (Plg)、组织纤溶酶原激活物 (tPA)、组织纤溶酶原激活物抑制物 (tPAI)等抗凝血、纤维蛋白溶解活性。结果表明 :VT患者中 3 4 %有ACA和 (或 )LA阳性 ,其中以ACAIgG和LA为主 ;9.5 %的Plg缺乏 ,8.3 %的tPAI升高 (明显高于对照 ,P <0 0 0 5 ) ;ATⅢ、PC、tPA缺乏者依次为 4 .5 %、4 .5 %、2 .8% (与对照无差异性 ,P >0 .0 5 ) ;ATⅢ、PC、Plg联合缺乏者 1例 ;APCR未发现相应的factorVLeiden ;抗磷脂蛋白抗体 (APA)阳性和阴性组之间的各抗凝血和纤维蛋白溶解活性没有明显差异性 ;4例APCR阳性 ,3例ACA和 (或 )LA阳性 ,这 3例血浆和正常血浆混合后 2例APCR并没有完全得到纠正。结论 :抗磷脂蛋白抗体和纤维蛋白溶解异常是VT较多见的相关病理因素 ;LA和 (或 )ACA干扰蛋白C抗凝血途径 ,使之形成获得性APCR ,而此APCR可能是体内导致易栓的病因之一。     

Development of a new test for the global fibrinolytic capacity in whole blood.

BACKGROUND: The development of global tests for the fibrinolytic capacity in blood is hampered by the low base-line fibrinolytic activity in blood, by the involvement of both plasmatic components and blood cells in the fibrinolytic system and by the loss of fibrinolytic activity as a result of the action of plasminogen activator inhibitor-1 (PAI-1). OBJECTIVE: To develop a new test for the global fibrinolytic capacity (GFC) of whole blood samples. METHODS AND RESULTS: Collection of blood in thrombin increased the subsequent generation of fibrin degradation products. This was ascribed to rapid clot formation and concomitant reduction of in vitro neutralization of tissue-type plasminogen activator (tPA) by PAI-1. On the basis of this observation, the following test was designed: blood samples were collected in thrombin with and without aprotinin and clots were incubated for 3 h at 37 degrees C. The GFC was assessed from the difference between the fibrin degradation products in the two sera. The assay was applied to blood samples from patients and healthy subjects. Other hemostasis parameters were determined in plasma samples taken simultaneously. The GFC varied considerably (normal range 0.13-13.6 microg mL(-1)); physical exercise strongly increased the GFC. Statistically significant correlations were found with tPA activity, PAI-1 activity and fibrinogen level. A mixture of antibodies against tPA and urokinase-type plasminogen activator (uPA) completely inhibited the GFC. An inhibitor of activated thrombin-activatable fibrinolysis inhibitor (TAFI) accelerated fibrinolysis 8-fold. CONCLUSION: The new test represents a global assessment of the main fibrinolytic factors in plasma and potentially those associated with blood cells.     

Definition of major bleeding in clinical investigations of antihemostatic medicinal products in non-surgical patients

Summary. A variety of definitions of major bleeding have been used in published clinical studies, and this diversity adds to the difficulty in comparing data between trials and in performing meta-analyses. In the first step towards unified definitions of bleeding complications, the definition of major bleeding in non-surgical patients was discussed at the Control of Anticoagulation Subcommittee of the International Society on Thrombosis and Haemostasis. Arising from that discussion, a definition was developed that should be applicable to studies with all agents that interfere with hemostasis, including anticoagulants, platelet function inhibitors and fibrinolytic drugs. The definition and the text that follows have been reviewed and approved by the cochairs of the subcommittee and the revised version is published here. The intention is to also seek approval of this definition from the regulatory authorities.     

肿瘤患者凝血及纤溶状态的改变

目的 探讨恶性肿瘤患者凝血、抗凝及纤溶指标的变化及其临床意义。方法 用Culter ACL-200全自动血凝仪对35例正常对照,25例子宫肌瘤,98例恶性肿瘤(胃癌20例,结直肠癌21例,食道癌18例,肺癌19例,宫颈癌20例)的凝血酶原时间(PT)、活化部分凝血活酶时间(APTT)、纤维蛋白原(Fbg)、凝血酶凝固时间(TT)、抗凝血酶Ⅲ(AT-Ⅲ)、纤溶酶原(PLG)及α2-纤溶抑制物(α2-PI)进行检测。结果 恶性肿瘤组PT、APTT、TT水平较正常对照组增高,但无显著性差异;Fbg含量增高,AT-Ⅲ活性降低,PLC及2-PI活性增高,有显著性差异(p<0.05或p<0.01);恶性肿瘤有转移组与无转移组比较Fbg舍量增加、TT缩短、AT-Ⅲ、PLG及α2-PI活性均有显著性改变(p<0.05或p<0.01)。子宫肌瘤的上述指标与正常对照组之间无统计学差异。结论 恶性肿瘤凝血功能增强,抗凝及纤溶活性降低,机体处于高凝状态,有血栓形成倾向;恶性肿瘤有转移组纤溶活性增高则有助于恶性肿瘤转移。     

血栓弹力图应用价值的再探讨──在纤溶功能测定中的意义

本文采用在血样中加入一定量尿激酶，然后测定血栓弹力图（ＴＥＧ），并与常规方法进行了比较。本组共测定正常人３４例。各种临床疾病计５０例。结果表明本文介绍方法较常现方法更能敏感反应血块纤溶状况。同时还提出血块开始溶解时间（Ｔ０），最大溶解幅度（Ｐｍａｘ）和溶解速度（Ｖ）三种新的测定参数。并对其意义进行了分析。