静脉注射肝素钠和达肝素钠对血小板激活影响的对比研究

目的了解肝素钠(UFH)与低分子肝素(LWMH)对血小板激活的影响有无差别。方法59例新入院冠心病患者,随机分为UFH组和达肝素钠组,分别于基线状态下、给药(UFH或达肝素钠5000 IU)后30 min、1 h取血,测定CD62P、GPⅡb/Ⅲa、血管性假血友病因子(vWF)。结果UFH组静脉给药后30 min GPⅡb/Ⅲa、血浆CD62P及vWF的水平均明显高于给药前(P<0.05);给药后1 h GPⅡb/Ⅲa及vWF的水平仍显著高于给药前(P<0.05);血浆CD62P的水平虽仍较给药前增高,但差异无统计学意义(P>0.05)。静脉给予达肝素钠5000 IU30min后GPⅡb/Ⅲa及血浆CD62P的水平均明显高于给药前(P<0.05),但在给药后1 h即与基线值差异无统计学意义(P>0.05);vWF的水平在给予达肝素钠后30 min、1 h均与给药前相比,差异无统计学意义(P>0.05)。结论静脉注射UFH及达肝素钠均可激活血小板;但达肝素钠对血小板活化的影响较小。     

Prevalance of heparin-dependent platelet-activating antibodies in preterm newborns after exposure to unfractionated heparin.

Heparin is frequently used in preterm infants to prolong the patency of intravascular catheters. The aim of this study was to evaluate the prevalence of heparin-dependent platelet-activating antibodies in newborns. A cross-section of all preterm newborn infants expected to require heparin to maintain patency of a central venous access line were enrolled. A blood sample was obtained soon after birth before heparin exposure to exclude the possibility of placental transfer of maternal heparin-dependent platelet-activating antibodies. A second sample was obtained at termination of heparin use (mean duration of heparin exposure was 23 +/- 13 days; range, 6-67). Paired samples, at birth and after heparin use, were available for 42 infants with a mean gestational age of 27.8 +/- 2.2 weeks and birth weight of 1036 +/- 267 g. Thrombocytopenia developed in 57% (24/42) of the infants. None of these infants had clinical suspicion of thrombosis during the study period. The etiology of thrombocytopenia was confirmed sepsis in six, presumed sepsis in three, necrotizing enterocolitis in one, and unclear in 14 infants. Anti-heparin/platelet factor 4 antibodies measured using the standard assays for heparin-induced thrombocytopenia (two commercially available enzyme-linked immunosorbent assay tests and the functional platelet serotonin release assay) were negative on all infants. Although it could be related to the poor ability of these infants to mount an immunologic response, further research is necessary to fully understand this lack of response to heparin and to elucidate further the reasons for thrombocytopenia in very-low-birth-weight infants.     

Greater inhibitory effects of bivalirudin compared with unfractionated heparin plus eptifibitide on thrombin-induced platelet activation

OBJECTIVES: The effects of antithrombotic agents on the activation of platelets by thrombin were determined in blood from patients (n=12) with symptomatic coronary artery disease. METHODS: Blood obtained immediately before cardiac catheterization was incubated in vitro with therapeutic concentrations of unfractionated heparin (1.2 and 2.0 U/ml) alone and in combination with eptifibatide (unfractionated heparin 1.2 U/ml+eptifibatide, 1.7 microg/ml) or bivalirudin (8 and 14 microg/ml). An activated clotting time was determined. Platelet activation was induced with thrombin (0, 5, 15, and 40 U/ml) and assessed with the use of flow cytometry. Fibrin polymerization was prevented by the peptide Gly-Pro-Arg-Pro. The activation of glycoprotein IIb-IIIa and surface expression of P-selectin were identified with antibodies (PAC-1 and anti-CD62). RESULTS: The activated clotting time was 258+/-13 s with 1.2 U/ml unfractionated heparin alone, 396+/-8 s with unfractionated heparin+eptifibatide, and 348+/-9 s with 8 microg/ml bivalirudin. The binding of PAC-1 (reflecting the potential to aggregate) was decreased most effectively by bivalirudin (percentage of platelets binding PAC-1 in response to 15 U/ml thrombin-unfractionated heparin=54+/-7%, unfractionated heparin+eptifibatide=12+/-4%, bivalirudin=1+/-0.3% of platelets, P<0.05 for bivalirudin compared with unfractionated heparin or unfractionated heparin+eptifibatide and unfractionated heparin+eptifibatide compared with unfractionated heparin). Bivalirudin prevented thrombin-induced surface expression of P-selectin more effectively than unfractionated heparin alone or unfractionated heparin+eptifibatide (percentage of platelets expressing P-selectin in response to 15 U/ml thrombin-unfractionated heparin alone=74+/-5%, unfractionated heparin+eptifibatide=53+/-7%, bivalirudin=1+/-0.3%, P<0.05 for bivalirudin compared with unfractionated heparin or unfractionated heparin+eptifibatide). CONCLUSION: Comparison between pharmacologic concentrations shown to be therapeutic demonstrated that bivalirudin inhibited thrombin-induced activation of platelets to a greater extent than unfractionated heparin alone or in combination with eptifibatide.     

低分子肝素与普通肝素在冠脉介入治疗中应用的对比研究

目的 比较低分子肝素(LMWH)与普通肝素(UH)在经皮冠脉介入治疗(PCI)中的抗凝效果及安全性.方法 将适合PCI的冠心病患者分为三组:A组,LMWH组,即PCI手术前、手术中、手术后均应用LMWH;B组,UH组,即PCI手术前、手术中、手术后均应用UH;C组,交叉抗凝组,即PCI手术前用LMWH,手术中应用UH,手术后用LMWH.观察30 d内三组临床效果及出血等不良事件.结果 C组反复心绞痛发作患者较A组、B组增多,差异有统计学意义,而心脏性死亡、中风、急性及亚急性支架内血栓形成或急性再次血运重建等差异无统计学意义.C组少量出血患者较A组、B组增多,差异有统计学意义.严重出血,特别是需要输血的患者三组间差异无统计学意义.导管室使用LMWH的A组患者出现2例导管内血栓形成.结论 LMWH和UH都各有优缺点.LMWH无需监测活化凝血时间,无需静脉持续输注,而UH用药期间需要监测APTT,有时可引起血小板减少症(HIT).应用LMWH的个别患者会出现导管内血栓形成,交叉抗凝临床效果,如反复心绞痛发作较高,在出血不良事件方面也有增加趋势.     

Affinity purification of heparin-dependent antibodies to platelet factor 4 developed in heparin-induced thrombocytopenia: biological characteristics and effects on platelet activation

Antibodies to heparin platelet factor 4 (H-PF4) complexes were purified from the plasma of three patients with heparin-induced thrombocytopenia (HIT) using affinity chromatography. From each plasma, the largest amount of antibodies was eluted with 2 M NaCl at pH 7.5 (peak 1) and the remainder was obtained using 0.1 M glycine/0. 5 M NaCl at pH 2.5 (peak 2). In an enzyme-linked immunosorbent assay (ELISA), we then showed that each patient had developed antibodies to PF4 displaying different characteristics. In patient 1, peak 1 IgG reacted almost exclusively with H-PF4 complexes, whereas peak 2 IgG had similar reactivity with PF4 whether or not heparin was present. Patient 2 expressed a mixture of IgA, IgM and IgG and both fractions bound to PF4 alone or to H-PF4 complexes. Finally, IgG in patient 3 only bound to H-PF4 and was unreactive with PF4 alone. Using [14C]-serotonin release assays, the antibodies developed in the three patients and exhibiting the strongest ability to activate platelets with heparin were those having the highest affinity to H-PF4. These results strongly support the hypothesis that HIT antibodies to PF4 are heterogeneous regarding their affinity and specificity for target antigens and this may greatly influence their ability to activate platelets and their pathogenicity.     

Comparison of fondaparinux, low molecular-weight heparin and unfractionated heparin in preventing thrombus formation on mechanical heart valves: results of an in-vitro study

BACKGROUND AND AIM OF THE STUDY: The study aim was to investigate the efficacy of three different anticoagulants in preventing thrombus formation on mechanical heart valves, using an in-vitro system. METHODS: Blood samples (470 ml) were taken from young and healthy male volunteers and anticoagulated with unfractionated heparin (UFH; n=18), low molecular-weight heparin (LMWH; n=18) or fondaparinux (n=16). Bileaflet mechanical heart valves were placed in a new device--the 'Thrombosis Tester'--and exposed in a continuous circulation at a rate of 80 beats per min to the anticoagulated blood samples for a total exposure time of 60 min. Results for thrombus weight were skewed distributed and presented as log-transformed values. RESULTS: The weight of each valve was measured before and after 1 h of perfusion; subsequent mean (+/-SD) thrombus weights were 0.739 +/- 0.573 g for UFH, 0.789 +/- 0.099 g for LMWH, and 0.934 +/- 0.145 g for fondaparinux (p = 0.397 for comparison of all groups by ANOVA). Electron microscopy showed concordant results with regard to thrombus formation on heart valve surfaces. CONCLUSION: Fondaparinux and LMWH were as effective as UFH in preventing thrombus formation on mechanical heart valves in vitro. The 'Thrombosis Tester' proved to be a new, unique instrument for investigating the ability of anticoagulants to prevent valve thrombosis on mechanical heart valves under in-vitro conditions.     

普通肝素、达肝素及依诺肝素在冠状动脉造影术中对血小板激活的对比研究

目的:了解普通肝素(UFH)、达肝素及依诺肝素对血小板激活的差别.方法:58例行冠状动脉造影术的心脏瓣膜病患者,分为UFH组、达肝素组和依诺肝素组.术前测体重,动脉穿刺成功后,按50 U/kg经动脉鞘管内注射UFH(UFH组)、达肝素(达肝素组)或依诺肝素(依诺肝素组).分别于给药前、给药后10 min、20 min取血,测定血小板活化标记物GPⅡb/Ⅲa、sCD40L、P-选择素(CD62P)及血友病因子(vWF).结果:UFH组给药后10 minGPⅡb/Ⅲa、血浆sCD40L及CD62P水平明显高于给药前(P<0.05),给药后20 min GPⅡb/Ⅲa、sCD40L水平仍高于给药前(P<0.05),但CD62P水平与给药前差异无统计学意义(P>0.05);达肝素组和依诺肝素组给药后10 min、20 minGPⅡb/Ⅲa均明显高于给药前(P<0.05),但血浆sCD40L及CD62P水平变化差异无统计学意义(P>0.05);达肝素组给药后20 min、依诺肝素组给药后10 min血浆vWF较给药前显著降低(P<0.05).结论:冠状动脉造影术中鞘内注射UFH、达肝素及依诺肝素均可激活血小板;与UFH相比,达肝素和依诺肝素对血小板的激活作用较小;而达肝素和依诺肝素对血小板的激活作用无明显差异.     

Spontaneous heparin-induced thrombocytopenia syndrome without any proximate heparin exposure,infection, or inflammatory condition: Atypical clinical features with heparin-dependent platelet activating antibodies

Recent studies suggest that a thromboembolic disorder resembling heparin-induced thrombocytopenia (HIT), so-called spontaneous HIT syndrome, can occur in patients without any history of heparin exposure. It is likely due to anti-platelet factor 4 (PF4)/polyanion antibodies induced by other polyanions, such as bacterial surfaces and nucleic acids. We describe an atypical case of spontaneous HIT syndrome. A 70-year-old man suddenly presented with acute cerebral sinus thrombosis (CST). Soon after the initiation of unfractionated heparin (UFH) for the treatment of CST, his platelet count fell precipitously and he developed deep vein thrombosis, a clinical picture consistent with rapid-onset HIT but without any proximate episodes of heparin exposure, infection, trauma, surgery, or other acute illness. Antigen assays and a washed platelet activation assay indicated that the patient already possessed anti-PF4/heparin IgG antibodies with heparin-dependent platelet activation properties on admission. Cessation of UFH and initiation of argatroban resulted in prompt recovery of his platelet count without further thromboembolic events. We identified two similar cases in the literature. However, these patients do not meet the recently proposed criteria for spontaneous HIT syndrome. Even in atypical cases, however, inappropriate or delayed diagnosis of HIT appears to be associated with worse outcomes. We propose that these atypical cases should be included in the category of spontaneous HIT syndrome.     

Low molecular weight heparin versus unfractionated heparin in the colonoscopy peri-procedure period: a cost modeling study

OBJECTIVE: The aim of this study was to compare the economic outcomes of peri-procedure anticoagulation approaches for elective colonoscopy. METHODS: Decision analysis was used to model the economic outcomes of five peri-procedure anticoagulation options: outpatient low molecular weight heparin (LMWH), inpatient unfractionated heparin infusion (UFHi), continuous warfarin (with probability of a repeat procedure using LMWH or UFHi), and discontinuation of anticoagulation therapy. The model's base-case scenario assumed drug therapy options for high-risk patients were equally effective in preventing a thromboembolic event (0.1% risk), with a higher probability for the no anticoagulation strategy (0.4%); event costs were based on published data and adjusted to 1997 dollars. Drug costs reflected 1997 average wholesale price. Medical costs for other variables were estimated based on local hospital charges. Indirect costs were not considered. Risk probabilities and LMWH drug cost were tested in sensitivity analysis. RESULTS: In the base-case scenario, costs for the options evaluated were $1436/patient, $1792/patient, $1848/patient, $2629/patient, and $5196/patient for no anticoagulation, continuous warfarin/repeat LMWH, LMWH as outpatient, continuous warfarin/repeat UFHi, and UFHi as inpatient respectively ($1997). Discontinuing anticoagulation was the least costly approach but involved the greatest thromboembolic risk. The cost of continued warfarin anticoagulation/repeat LMWH was minimally less than the LMWH option, but assumes 25% of patients would require a second procedure. The traditional approach (UFHi) requires an extended hospitalization and is the most costly option. Varying risk category or LMWH cost in sensitivity analysis had a negligible impact on overall costs. CONCLUSION: Within the model's assumptions, LMWH offers a novel, convenient, and economical solution to the problem of peri-procedure anticoagulation for elective colonoscopy.     

Changes in platelet count after cardiac surgery can effectively predict the development of pathogenic heparin-dependent antibodies

Cardiopulmonary bypass (CPB) induces the release of platelet factor 4 (PF4) and patients are at risk of heparin-induced thrombocytopenia (HIT). This study was aimed to determine whether an abnormal evolution in platelet count (PC) after CPB is predictive of the development of HIT antibodies. Two abnormal PC patterns were defined: pattern P1, characterized by a decrease in PC following previous correction of thrombocytopenia occurring during CPB, and pattern P2, defined as a persistent low PC in the days following CPB. PC was evaluated for 10 d in 305 consecutive patients before and after CPB. Serotonin release assay (SRA) was carried out between days 8 and 10 to detect pathogenic heparin-dependent antibodies. Moreover, antibodies to heparin-PF4 (H-PF4) complexes were assayed by enzyme-linked immunosorbent assay. PC evolution after CPB was normal in 300 patients although antibodies to H-PF4 were frequently present (53.4%). Changes in PC were abnormal in five patients with pattern P1 (n = 4) or P2 (n = 1). As SRA was positive in four of the five cases, the positive predictive value of abnormal PC pattern for pathogenic HIT antibodies was 80%. Careful follow-up of PC after CPB makes it possible to predict with high specificity (99%) for those patients who develop pathogenic HIT antibodies.     

低分子肝素与普通肝素在血液透析中对血脂的影响

目的探讨长期应用低分子肝素(LMWH)与普通肝素(UFH)抗凝治疗对血液透析患者脂质代谢的影响.方法选择病情稳定血液透析半年以上的尿毒症患者31例,分成UFH组(15例)和LMWH组(16例)两组,第1年2组均按个体化使用肝素抗凝;第2年前组继续使用肝素抗凝,而后组改用LMWH,于血透前动脉端1次注入LMWH.2组分别于实验前、实验第1年和实验第2年结束时检测血浆胆固醇(CH)、甘油三脂(TG)、高密度脂蛋白(HDL)、低密度脂蛋白(LDL)、脂蛋白LP(a)及载脂蛋白(ApoA1、ApoB)水平.结果 UFH组患者随着透析时间的延长,血浆CH、TG、LDL、ApoB水平明显升高,HDL、ApoA1明显降低,而LMWH组患者前1年使用UFH,血脂变化与同期UFH组相似,改用LMWH 1年后CH、TG、LDL和Apo-B水平明显降低,HDL、ApoA1水平升高.结论长期使用肝素可引起脂质代谢异常,低分子肝素在一定程度上可以缓解高脂血症和改善脂质代谢.     

Antibodies to platelet factor 4-heparin after cardiopulmonary bypass in patients anticoagulated with unfractionated heparin or a low-molecular-weight heparin : clinical implications for heparin-induced thrombocytopenia.

BACKGROUND: Cardiopulmonary bypass (CPB) induces platelet activation with release of platelet factor 4 (PF4), and patients are exposed to high doses of heparin (H). We investigated whether this contributes to the development of antibodies to H-PF4 and heparin-induced thrombocytopenia (HIT). METHODS AND RESULTS: CPB was performed with unfractionated heparin (UFH) in 328 patients. After surgery, patients received UFH (calcium heparin, 200 IU. kg-1. d-1) (group 1, n=157) or low-molecular-weight heparin (LMWH, Dalteparin, 5000 IU once daily) (group 2, n=171). Eight days after surgery, antibodies to H-PF4 were present in 83 patients (25.3%), 46 in group 1 and 37 in group 2 (P=0.12). Most patients (61%) had IgG1 to H-PF4, but only 8 samples with antibodies induced platelet activation with positive results on serotonin release assay. HIT occurred in 6 patients in group 1, but no thrombocytopenia was observed in subjects receiving LMWH, although 2 had high levels of antibodies with positive serotonin release assay results. When antibodies to H-PF4 were present, mean platelet counts were lower only in patients with FcgammaRIIA R/R131 platelets. CONCLUSIONS: These results provide evidence that the development of antibodies to H-PF4 after CPB performed with UFH is not influenced by the postoperative heparin treatment. The antibodies associated with high risk of HIT are mainly IgG1, which is present at high titers in the plasma of patients continuously treated with UFH.     

Intracellular events in platelet activation induced by antiphospholipid antibodies in the presence of low doses of thrombin

OBJECTIVE: Thrombosis and thrombocytopenia are features of the antiphospholipid syndrome (APS), suggesting that antiphospholipid antibodies (aPL) may bind platelets, causing activation and aggregation of platelets and thrombosis. The intracellular events involved in aPL-mediated platelet activation are not fully understood and are therefore the subject of this study. METHODS: IgG fractions and their F(ab')(2) fragments were purified from the sera of 7 patients with APS and from the pooled sera of 10 healthy subjects (as controls). Phosphorylation of p38 MAPK, ERK-1/2, and [Ca(2+)]-dependent cytosolic phospholipase A(2) (cPLA(2)) was determined in lysates of washed platelets pretreated with low doses of thrombin and aPL or control IgG or their F(ab')(2) fragments, by immunoblot. The effects of aPL on platelet aggregation in the presence or absence of a p38 MAPK inhibitor, SB203580, were examined. Thromboxane B(2) (TXB(2)) production was detected by enzyme-linked immunosorbent assay on gel-filtered platelets treated with aPL and thrombin, with or without SB203580. Calcium mobilization studies were done utilizing a fluorometric assay. RESULTS: Treatment of platelets with IgG aPL, or their F(ab')(2) fragments, in conjunction with subactivating doses of thrombin resulted in a significant increase in phosphorylation of p38 MAPK. Neither the IgG aPL nor their F(ab')(2) fragments increased significantly the phosphorylation of ERK-1/2 MAPKs. Furthermore, pretreatment of platelets with SB203580 completely abrogated the aPL-mediated enhanced aggregation of the platelets. Platelets treated with F(ab')(2) aPL and thrombin produced significantly larger amounts of TXB(2) when compared with controls, and this effect was completely abrogated by treatment with SB203580. In addition, cPLA(2) was also significantly phosphorylated in platelets treated with thrombin and F(ab')(2) aPL. There were no significant changes in intracellular [Ca(2+)] when platelets were treated with aPL and low doses of thrombin. CONCLUSION: The data strongly indicate that aPL in the presence of subactivating doses of thrombin induce the production of TXB(2) mainly through the activation of p38 MAPK and subsequent phosphorylation of cPLA(2). The ERK-1/2 pathway does not seem to be involved in this process, at least in the early stages of aPL-mediated platelet activation.     

Low-molecular-weight heparin versus unfractionated heparin in the treatment of patients with acute pulmonary thromboembolism

BACKGROUND: Low-molecular-weight heparin (LMWH) appears to be as effective as unfractionated heparin (UFH) for both treatment and prophylaxis of deep vein thrombosis (DVT), but limited data are available for its use in acute pulmonary thromboembolism (PTE). OBJECTIVE: To determine whether enoxaparin, a LMWH, was clinically as efficient and safe as UFH in patients with a diagnosis of acute PTE. MATERIAL AND METHODS: After exclusion of those with massive forms, 59 patients with acute PTE were randomly assigned to either subcutaneous enoxaparin given twice daily (1 mg/kg/dose) or adjusted dose intravenous UFH. Oral anticoagulant treatment was begun on the second day and was given for at least 6 months. We compared the treatment regimens at day 8 and day 90 with respect to a combined end point of major bleeding, recurrent venous thromboembolism (VTE), and death. RESULTS: In the first 8 days of treatment, 1 of 30 patients assigned to receive UFH (3.3%) reached one of the end points (recurrence), as compared with none of 29 patients assigned to enoxaparin. Statistically this difference was not significant (p = 0.508). By day 90, 3 patients assigned to UFH (10%) had symptomatic recurrent VTE, as compared with 1 patient assigned to enoxaparin (3.4%). There was neither major bleeding nor death in the study groups. There was an absolute difference of 6.4 percentage points between the two treatment groups, but the difference was statistically not significant (p = 0.318). CONCLUSION: Initial subcutaneous treatment with enoxaparin appeared to be as effective and safe as UFH in acute PTE.     

Increased circulating platelet-neutrophil, platelet-monocyte complexes, and platelet activation in patients with ulcerative colitis: a comparative study

It is reported that the incidence of thromboembolism is increased in ulcerative colitis (UC), and hypercoagulability persists even when patients are in remission. We evaluated the association of inflammatory response parameters with UC activity, and activation parameters of the platelets, endothelium, and the coagulation system in UC. Eighteen UC patients and 19 healthy subjects were included in the study. The patients' clinical features were recorded down; whole blood counts and acute phase parameters were evaluated. UC patients were divided into two as active (9 patients) and inactive (9 patients) according to combined clinical activity index (CAI) and endoscopic activity index (EAI) scores. In all subjects, platelet CD62P expression, platelet-monocyte complexes (PMC), platelet-neutrophil complexes (PNC), and platelet microparticles (PMP) were determined by flow cytometry. E-selectin, thrombin-antithrombin complex (TAT) levels in plasma, and sCD40L levels in serum were determined by ELISA. In both active and inactive UC patients, platelet CD62P expression, the percentages of PMC, and PNC were significantly higher than those in the control group (P< 0.01). PMP level was higher in the control group than in inactive UC patients (P = 0.001). sCD40L level was significantly higher in active UC group than in the control group (P < 0.01). EAI score correlated significantly with PMP (r = 0.5, P = 0.04) and sCD40L (r = 0.48, P = 0.044); CAI score had a negative correlation (r = -0.68, P = 0.002) with sE-selectin level. In addition to increased CD62P expression and sCD40L, increased formation of PMC and PNC suggests a role for platelet-leukocyte complex formation together with platelet activation in thromboembolic events observed in UC.