Anticoagulant and lipolytic effects of a low molecular weight heparin fraction

The lipolytic and anticoagulant actions of a 4000 dalton low molecular weight (LMW) heparin were compared with unfractionated mucosal heparin after intravenous and various subcutaneous doses in man. I.v. injection of 100 USP units/kg body weight lipoprotein lipase (LPL) activity, and inhibition of factor Xa decreased with a half life twice as long after LMW heparin compared to normal heparin (p < 0.05). There were no differences in half lives for HTGL activity, thrombin inhibition and on aPTT. The area under the activity time curve (AUC) of LPL and factor Xa was double with LMW heparin (p < 0.05). S.c. administration showed that the AUC of LMW heparin on the factor Xa inhibition was 10 times larger compared to normal heparin. LPL activity was released comparable to normal heparin. The effects on HTGL were three times larger compared to normal heparin. There were no differences in half lives. The data show that in contrast to normal heparin LMW heparin is rapidly and completely absorbed from the subcutaneous depots. The pharmacodynamic data of LPL activity and factor Xa inhibition suggest similar release mechanisms.     

A low molecular weight heparin compared with unfractionated heparin

A 6000 daltons low molecular weight heparin (LMWH) was compared with unfractionated mucosal heparin in vitro and in vivo. Despite unimpressive specifications by clotting assays in vitro, the LMWH gave high and sustained activity in vivo by anti-Factor Xa assays, following subcutaneous injection. However, activity measured by APTT and calcium thrombin time assays was at least as high as occurred following unfractionated heparin. On the basis of clotting assays, there seems no reason to expect a lower incidence of haemorrhagic side-effects following the clinical use of this LMWH. The study also strikingly demonstrates the inadequacy of in vitro clotting assays for assessing the in vivo behaviour of LMWH.     

The effects of heparin and low molecular weight heparins on bone

Recent clinical trials have shown that the risk of developing osteoporosis is substantially lower when low molecular weight heparins (LMWHs) are used in place of unfractionated heparin. While the reason(s) for this difference has not been fully elucidated, studies with animals have suggested that heparin causes bone loss by both decreasing bone formation and increasing bone resorption. In contrast, LMWHs appear to cause less bone loss because they only decrease bone formation. Whether all LMWHs decrease bone formation and therefore cause bone loss is unknown. For example, preliminary in vitro studies with the synthetic pentasaccaride, Fondaparinux, have suggested that it may not decrease bone formation and thus, may have no deleterious effects on bone. Further studies are required in order to determine if all LMWHs cause bone loss equally.     

Anticoagulant and antithrombotic effects of heparin and low molecular weight heparin fragments in rabbits

Heparin and heparin fragments of different molecular weight and with different anti-factor X_a/APTT activity ratios were studied with respect to their ability to inhibit thrombus formation in an animal model. It is concluded that: a) Neither anti-factor X_a nor the APTT activity alone is a good reflector of the antithrombotic activity. b) Anti-factor X_a active fragments must have a minimum molecular weight in order to elicit good antithrombotic activity. c) High affinity for antithrombin III is important for good antithrombotic activity. d) A heparin fragment of molecular weight 4 000 has the same antithrombotic activity as heparin but less effect on the clotting time.     

Prevention of experimental venous thrombosis in rabbits with low molecular weight heparin, dextran and their combinations, administered before or during induction of venous endothelial trauma

An in vivo experimental venous thrombosis model based on endothelial damage and flow reduction was used to investigate the effect of low molecular weight heparin (LMWH) alone and in combination with dextran and the effect of surgical and endothelial trauma on thrombus formation, formation of occlusive thrombi and thrombus weights. Five groups with 15 rabbits in each were studied. Two groups received dalteparin (50 anti-Xa IU/kg i.v.) before surgical trauma or after, during the endothelial trauma and two groups received dalteparin (50 anti-Xa IU/kg i.v.) with dextran 70 (1 g/kg i.v.) before surgical trauma or after, during the endothelial trauma. Compared to a control group (saline) all treatment regimes reduced significantly the frequency of thrombosis and occlusive thrombi as well as thrombus weights. No significant difference was observed between the identical treatment groups when the substances were introduced before respective after surgical trauma. It is concluded from the present study that thromboprophylaxis with LMWH in this particular in vivo model, given before or after surgical trauma is equally effective. Dextran has a certain augmenting thromboprophylactic effect when added to LMWH in this model.     

Laboratory monitoring of thromboprophylaxis with low molecular weight and standard heparin

This study was made to evaluate assays for monitoring of low dose heparin thromboprophylaxis and to evaluate its efficacy in reduction of hypercoagulation. Patients with medical diseases scheduled for routine thromboprophylaxis were subcutaneously treated with either 5.000 anti XaU low molecular weight (LMW) heparin once daily (n=20) or 5.000 IU standard (ST) heparin 3 times daily (n= 19). On days 1,2,3, before, 1 and 4 hours after heparin injection APTT, TCT, anti Xa, Heptest, thrombin-antithrombin complexes (TAT), and D-Dimer levels were measured. In the LMW heparin group, median values of APTT and TCT slightly increased after heparin and the ranges of pre- and postinjection values showed extensive overlap. However, values of anti Xa and Heptest markedly increased, showing complete separation of ranges. In the ST heparin group neither APTT, TCT, anti Xa, nor Heptest were significantly different comparing pre- and postheparin values. Half of the patients in both groups had subclinical hypercoagulation at baseline (TAT>5ng/ml, D-Dimer>200ng/ml). On day 3 of prophylaxis this percentage was not significantly decreased. Moreover, several patients in both groups increased in TAT and D-Dimer. In the LMWheparin group, negative correlations between body weight and 4 h postinjection heparin levels were found (anti Xa R=−0.50, Heptest R=−0.31) and between 1 h postinjection heparin and TAT and D-Dimer levels 3 h later (TAT-anti Xa R=−0.58, TAT-Heptest R=−0.64, D-Dimer-anti Xa R=−0.32, D-Dimer-Heptest R=−0.33). These results show that low dose LMW but not ST heparin therapy can be monitored by the anti Xa test or the Heptest.     

Studies on a highly active anticoagulant fraction of high molecular weight isolated from porcine sodium heparin

We have studied heparin fractionation using gel filtration and ion-exchange chromatographic methods. The starting material was commercial grade porcine mucosal sodium heparin (PSH). The fractionation was monitored employing synthetic substrates for assaying both antithrombin (with H-D-Phe-Pip-Arg-pNA ; S-2238) and anti-FXa (with Bz-Ileu-Glu-Gly-Arg-pNA ; S-2222) activities. The resulting fractions were evaluated in different amidolytic and coagulation methods used to determine heparin potency by comparison with PSH. By gel filtration of PSH on Ultrogel AcA 54, both strong anti-FXa and antithrombin activities were associated with the fractions eluted in the high molecular weight range (MW 20 × 10³). These fractions also had potent anticoagulant action when assayed by conventional clotting methods. PSH was also subjected to fractionation by an ion-exchange technique (DEAE-Sephacel) with increasing salt molarity. The patterns for antithrombin and anti-FXa activities were again closely related, if not identical. Four fractions were usually distinguished, with respectively negligible, intermediate, high and very high activities when compared to PSH. The very highly active fraction (HAF), approximately 15% by weight, was eluted at high salt molarity (> 0.8 M NaCl). On a weight basis its anticoagulant activity was 2–3 times that of PSH as determined by amidolytic as well as clotting methods. Intravenous injection of HAF to rabbits and dogs (1.0 and 2.5 mg/kg) produced a much stronger anticoagulant response than PSH, also showing an effect which persisted for a longer duration.     

Signs of thrombin generation in pediatric cardiac catheterization with unfractionated heparin bolus or subcutaneous low molecular weight heparin for antithrombotic cover

Introduction: Thrombosis is one of the most frequent adverse events after cardiac catheterization, which can be reduced by anticoagulation with unfractionated heparin (UFH) in both children and adults. Low molecular weight heparin (LMWH) might possibly offer advantages. Laboratory signs of thrombin generation during pediatric cardiac catheterization, with unfractionated heparin (UFH) bolus or subcutaneous LMWH for thrombosis prophylaxis, were determined in a first step to investigate the potential of LMWH for antithrombotic cover. Materials and methods: Signs of thrombin generation (D-dimer and F₁₊₂), anti-Xa activity and activated clotting time (ACT) were measured in 65 patients with congenital heart disease. A total of 40 patients were treated with a UFH bolus of 100 IU/kg bodyweight and, in 25 children, enoxaparin was subcutaneously administered at a dosage of 1/1.6 mg/kg bodyweight. Results: The dose to plasma activity of enoxaparin was more consistent than in the UFH group. Only a slight elevation of F₁₊₂ was found in some patients, which was a little higher in the enoxaparin group, but no difference of incidence of increased F₁₊₂ generation was detected between the two groups. D-dimer was elevated in three children after UFH bolus application, but no such effect was observed in any child after LMWH administration. Conclusions: Application of LMWH was equally efficacious during pediatric cardiac catheterization than UFH bolus administration, as determined by plasma levels and markers of clotting activation. In contrast to UFH bolus, no further monitoring was necessary after the application of LMWH during cardiac catheterization due to a consistent dose to plasma activity.     

Heparin-induced platelet aggregation: Dose/response relationships for a low molecular weight heparin derivative (pk 10169) and its subfractions

Addition of heparin or heparin derivatives to citrate anticoagulated platelet-rich plasma caused platelet aggregation in a dose-dependent manner. Utilizing heparin, a low molecular weight heparin derivative (PK 10169) and its various subfractions, we determined dose/response relationships for platelet aggregation and found that the ability of these agents to cause platelet aggregation was dependent upon the molecular weight of the individual subfraction used. In comparison to unmodified porcine mucosal heparin, the lower molecular weight derivative (PK 10169) yielded a dose/response curve that was shifted down and to the right, and indicated that this agent was less potent in causing platelet aggregation. In addition, as the molecular weight of PK 10169 subfractions decreased, their dose/response curves were progressively shifted down and to the right. The lowest molecular weight subfraction was essentially without platelet aggregating activity. We also measured the anti IIa and anti Xa activities of these agents and concluded that these activities did not appear to correlate with platelet aggregating activity. Platelet aggregation studies with PK 10169 subfractions of high and low affinity for antithrombin III (AT III) indicated that the platelet aggregating activity of these compounds may not be related to their affinity for AT III, but results were not definitive.     

Subcutaneous administration of heparin a randomised comparison with intravenous administration of heparin to patients with deep-vein thrombosis

One-hundred and forty-one patients with clinical signs of acute deep venous thrombosis (DVT) in the legs were randomly allocated to recieve heparin either as two daily subcutaneous injections (s.c.) or as continuous intravenous infusion (i.v.). The thrombi extended into the popliteal or femoral veins in 83% of the patients. Verification of diagnosis and evaluation of therapy was performed by phlebography, plethysmography and thermography.

The results showed that heparin administered s.c. twice daily was as efficient as continuous i.v. infusion in preventing extension of the thrombus. In two patients the s.c. administration was stopped due to local haematomas at the injection sites. Retroperitoneal or intramuscular bleedings occurred in four patients, two in each group. Two major, non-fatal pulmonary emboli occurred, one in each group.     

Comparative effects of heparin and PK 10169, a low molecular weight fraction, in a canine model of arterial thrombosis

The comparative properties of heparin and PK 10169, a low molecular weight fraction, were studied using an antithrombotic test in anaesthetized dogs. The antithrombotic properties of the two compounds were evaluated by measuring inhibition of thrombus formation following transluminar stimulation of coronary artery with anodal current and by measuring anticoagulant properties, anti Xa and anti IIa activities. The results show that PK 10169 displayed significant antithrombotic activities above 0.625 mg/kg and was equipotent at 2.5 mg/kg s.c. with heparin 10 mg/kg s.c. No correlation could be observed between antithrombotic/anti Xa ratio of both compounds. Moreover it was shown that, unlike heparin, PK 10169 s.c. was devoid of obvious anticoagulant properties and induced a negligible anti IIa activity contrasting with a high anti Xa level. A similar dissociation between anti Xa and anti IIa activities was observed following i.v. administration of 2.5 mg/kg of PK 10169 but not with heparin. This low molecular weight heparin fraction might thus be regarded as a potential arterial antithrombotic agent devoid of appreciable anticoagulant effect.     

低分子肝素对进展性缺血性脑卒中预防作用的探讨

目的 探讨低分子肝素对进展性缺血性脑卒中的预防作用。方法 将起病在72小时内的203例脑梗死患者随机分成低分子肝素治疗组及常规治疗组。低分子肝素组在常规治疗的基础上加用低分子肝素0．4ml,每日2次腹部皮下注射,连续5－10天。治疗前后检测纤溶酶原和凝血因子X活性及血液流变学各项指标,同时对进展性缺血性脑卒中患者的神经功能缺损进行评分。结果 低分子肝素组11例发生进展性缺血性脑卒中,常规治疗组23例,两组比较有显著性差异（P＜0．05）;低分子肝素组发生的进展性卒中其进展的严重程度比常规治疗组轻（P＜0．01）;近期预后较常规治疗组好（P＜0．05）;低分子肝素治疗能明显降低纤溶酶原和凝血因子X活性及血液流变学各项指标。结论 低分子肝素治疗能降低进展性缺血性脑卒中发生率,有利于进展性缺血性脑卒中患者神经功能的恢复。     

Anticoagulant effects of two types of low molecular weight heparin administered subcutaneously

Two types of LMW heparin were prepared by gel filtration of standard heparin (LMW fraction) and by degradation of heparin by nitrous acid (LMW fragment), respectively. The effects on factor Xa inhibition (XaI), APTT, platelet aggregation and AT III level of these preparations were studied after subcutaneous administration to humans and compared with those of standard heparin. At a dose of 5000 IU (XaI) the LMW fraction and LMW fragment induced peak plasma XaI activity of 0.32 IU/ml and 0.41 IU/ml respectively, compared to 0.07 IU/ml for heparin. Still 11.5 h after administration both LMW preparations gave higher activities than heparin ever induced. Following administration of 10,000 IU (XaI) of the LMW fragment the plasma peak XaI activity was 0.81 IU/ml. This prolonged the APTT from 36 sec to 46 sec only. The half-lives of the XaI activity in plasma were between 3 and 4 hours. No effect on platelet aggregation or AT-III level was demonstrated.     

A whole blood flow cytometric determination of platelet activation by unfractionated and low molecular weight heparin in vitro

The influence of unfractionated (Heparin–Natrium) and low-molecular heparin (Fragmin®) on platelet activation in whole blood was investigated by FACS analysis in vitro using antibodies against glycoprotein (gp) IIb/IIIa (CD 41), GMP 140 (CD 62P), gp 53 (CD 63) and fibrinogen. Samples were also labeled with anti-gp Ib (CD 42b). Neither unfractionated heparin (UFH) nor low molecular weight heparin (LMWH) led to significant (i.e., p<0.05) changes in fluorescence intensities of platelets labeled with anti-gp IIb/IIIa or anti-gp 53. Significant platelet activation due to unfractionated heparin could be observed by labeling with anti-GMP 140 (UFH: p=0.009; LMWH: p=0.16). The proportion of platelets with surface-bound fibrinogen was significantly increased (UFH: p=0.00006; LMWH: p=0.008). After incubation with heparins, activation ability of platelets by adenosine diphosphate (ADP) was significantly increased. The potentiating action of unfractionated heparin was larger. Therefore, flow cytometric results of platelet activation in patients receiving heparin should be interpreted carefully.     

低分子肝素对大鼠大脑皮层神经细胞缺血再灌注损伤的保护作用

目的研究低分子肝素(low molecular weight heparin,LMWH)对大鼠大脑皮层神经细胞缺血再灌注损伤的保护作用及其可能机制。方法体外培养新生大鼠大脑皮层神经细胞,建立缺血再灌注模型,MTT法检测细胞活力,Annexin V-FITC、PI双染流式测细胞凋亡率,荧光分光光度计法测定细胞内钙离子浓度。结果低分子肝素可提高缺血再灌注损伤的神经细胞活力,降低细胞凋亡率和细胞内钙离子浓度。结论低分子肝素对缺血再灌注损伤的大鼠大脑皮层神经细胞有保护作用,其机制可能与LMWH降低细胞内钙离子浓度有关。     

低分子肝素钙治疗急性脑梗死的疗效观察

目的观察低分子肝素钙（LMWHCa）治疗急性脑梗死的疗效和安全性。方法选用48例急性脑梗死病人,其中24例用常规治疗为对照组,治疗组24例除常规治疗外,加用LMWHCa4100抗Xa国际单位腹部皮下注射,bid,连续10d为1疗程。治疗前后分别作神经功能评分、血液流变学观察和PT、AFTT、TT、Fg。结果治疗组神经功能恢复、血液流变学改善均明显优于对照组（P〈0．05）。治疗组用药后Fg降低,PT延长,与对照组比较差异显著（P〈0．01或P〈0．05）,两组治疗前后的TT、APTI变化不明显;治疗组有2例出现皮下淤斑。结论脑梗死急性期给予低分子肝素钙治疗安全和有效。     

Comparative study of antithrombotic effect of a low molecular weight heparin and unfractionated heparin in an ex vivo model of deep arterial injury

Thrombosis after plaque rupture triggers the onset of acute coronary events. The treatment of choice for patients with acute coronary syndromes is conventional unfractionated heparin. Low molecular weight heparin has recently been reported to be as effective and even safer than unfractionated heparin. In this study, the effects of the low molecular weight heparin reviparin and unfractionated heparin on thrombus formation were examined under dynamic conditions using an extracorporeal perfusion chamber in a porcine model. Thrombus formation was assessed by the deposition of porcine ¹²³I-fibrin(ogen) and autologous ¹¹¹In-platelets on porcine tunica media at high and low shear rates. Reviparin reduced the fibrinogen molecules deposited on injured vessels at high shear rates (252±80 molecules×10¹²/cm² for reviparine (200 U/kg/hour) vs. 624±70×10¹²/cm² for unfractionated heparin (200 U/kg/hour) (p<0.05). At low shear rates, fibrinogen deposition was also significantly reduced by reviparin (130±15 molecules×10¹²/cm²) compared to unfractionated heparin (192±40×10¹²/cm² at 200 U/kg/hour; p<0.05). No change in platelet deposition was detected after heparin administration in either treatment group. In conclusion, the low molecular weight heparin reviparin has a higher antithrombotic potential than unfractionated heparin. Reviparin may have advantages over unfractionated heparin in treatment and prevention of acute coronary syndromes.