Danqi Piantan Jiaonang does not modify hemostasis, hematology, and biochemistry in normal subjects and stroke patients.

BACKGROUND AND OBJECTIVE: Previous studies on Danqi Piantan Jiaonang (DPJ, NeuroAid), a traditional Chinese medicine, in stroke patients showed promising results. Our aim was to determine the safety of DPJ in normal subjects and stroke patients through a series of studies assessing its immediate and long-term effects, alone and in combination with aspirin, on hematological, hemostatic, and biochemical parameters. METHODS: We conducted 3 studies from December 2004 to May 2006. Study 1 was a case series which recruited 32 healthy volunteers who were given 2 oral doses of 4 DPJ capsules (0.4 g/capsule) 6 h apart. Study 2 was a randomized controlled trial of 22 healthy volunteers who received either 1 oral dose of aspirin 300 mg alone or a combination of 1 dose of aspirin 300 mg and 2 doses of 4 DPJ capsules taken 6 h apart. For both studies 1 and 2, hemostatic parameters (prothrombin time, activated partial thromboplastin time, fibrinogen, platelet aggregation, D-dimer) were tested at baseline, and after 2 and 8 h. Study 3 was a case series which recruited 10 patients with recent ischemic stroke (within 7 days) who were given 4 DPJ capsules taken orally 3 times a day for 1 month. Blood tests for hemostatic, hematological (complete blood count), and biochemical parameters (glucose, creatinine, alanine aminotransferase, aspartate transaminase, C-reactive protein) were performed at baseline, and after 1 and 4 weeks. RESULTS: Apart from the expected changes in platelet aggregation in subjects taking aspirin, no significant differences were detected in hemostatic parameters at baseline, and 2 and 8 h after oral intake of DPJ alone or in combination with aspirin. Likewise, no significant differences were observed in hematological, hemostatic, and biochemical parameters at baseline, and after 1 and 4 weeks of oral intake of DPJ. CONCLUSION: DPJ does not significantly modify hematological, hemostatic, and biochemical parameters in normal subjects and stroke patients.

Risk stratification and venous thromboprophylaxis in hospitalized medical and cancer patients.

Acute venous thromboembolism (VTE) is a serious and potentially fatal disorder, which often complicates the course of hospitalized medical patients. This is particularly true for carriers of malignant diseases. While the introduction of thromboprophylactic measures has probably affected the occurrence of postoperative VTE, there is an increasing awareness of the importance of medical conditions in determining thromboembolic events. Simple and clinically relevant risk assessment models are available to facilitate VTE risk assessment in hospitalized medical patients. Their validation in prospective studies is in progress. Randomized controlled clinical trials have consistently documented the efficacy of heparins and fondaparinux for prevention of VTE in hospitalized medical patients with a minimal haemorrhagic risk. Recognition of the incidence and clinical importance of thrombosis will probably encourage more widespread use of antithrombotic prophylaxis in medical patients and especially in some particular types of malignancies.

Characterization of seven novel mutations causing factor XI deficiency.

BACKGROUND AND OBJECTIVES: Factor XI (FXI) deficiency is a rare autosomal recessive disorder, the main manifestation of which is injury-related bleeding. The disorder is rare in most populations, but common among Jews in whom two mutations, E117X and F283L, account for 98% of cases. Other mutations, C38R and C128X, are prevalent in French Basques and Britons, respectively. Additional sporadic mutations have been described in most parts of the world. The objective of this study was to identify the mutations in 15 unrelated FXI-deficient patients and characterize missense mutations by expression in baby hamster kidney (BHK) cells. DESIGN AND METHODS: Clinical and laboratory information and DNA samples were obtained from the patients and mutations were identified by sequencing. Missense mutations were expressed in BHK cells and their effect on FXI secretion and dimerization was assessed using enzyme-linked immunosorbent assay and immunoblotting. RESULTS: Of 16 mutations detected, seven are novel including two deletions, one splice site and four missense mutations. Expression of the four novel missense mutations (C58Y, Y427C, C527Y and V20A) in cells revealed no secretion of FXI-C58Y, Y427C and C527Y and secretion of only 22% of normal in the medium for FXI-V20A. Secretion of FXI from BHK cells harboring a previously reported E297K substitution cells was also impaired (4.5% of wild-type). Homodimerization was normal for all five mutants. INTERPRETATION AND CONCLUSIONS: Defective homodimerization of FXI was previously recognized as a major mechanism for defective secretion of FXI from producing cells. In this study, five FXI missense mutations (four novel) were associated with impaired secretion albeit normal dimerization, underscoring the existence of other mechanisms for defective secretion.

Inhibition of clot formation process by treatment with the low-molecular-weight heparin nadroparin in patients with carotid artery disease undergoing angioplasty and stenting. A thromboelastography study on whole blood.

Low-molecular-weight heparins (LMWHs) have become the corner stone of antithrombotic treatment but their administration protocol needs to be optimized for certain groups of patients. In this paper, we studied the influence of nadroparin treatment on clot formation process assessed by thromboelastography in patients with carotid artery disease undergoing angioplasty and stenting. Standard thromboelastography assays (in-TEM and ex-TEM) and minimal TF-triggered thromboelastography assay in citrated whole blood were performed in normal volunteers (n = 20), in patients with carotid artery disease receiving only antiplatelet treatment (n = 30), and in patients undergoing angioplasty receiving nadroparin 5750 anti-Xa IU s.c. twice daily (n = 60). Blood samples were collected four hours after a second injection of nadroparin. In a subgroup of LMWH-patients (n = 18) blood samples were also obtained prior to first injection of LMWH. Antiplatelet treatment had no effect on any parameter of the thromboelastographic pattern. Nadroparin treatment resulted in significant prolongation of clotting time (CT) and clot formation time (CFT) and significantly reduced a -angle in minimal TF-triggered thromboelastography and 30 - 38% of nadroparin treated patients had thromboelastographic parameters beyond the normal maximum limit. In-TEM test revealed a significant prolongation of clotting time while ex-TEM was not modified, and 20 to 30% of the patients had thromboelastographic parameters beyond the normal maximum limit. Anti factor-Xa activity in platelet-poor plasma (PPP) was also measured, and statistical analysis showed that prolongation of CFT of minimal TF-triggered TEM was significantly correlated to the levels of anti-Xa activity in patients ' plasma (p = 0.04; r (2) = 0.7). There was no statistical correlation for any other parameter in all tests. In conclusion, the present study shows that nadroparin treatment in patients with carotid artery disease undergoing endovascular procedures induces significant modification of the thrombus kinetics assessed by minimal TF-triggered whole blood thromboelastography. The clinical relevance of these findings has to be evaluated in future studies.

Determination of prothombinase activation after adding human purified prothrombin to human clot: comparison of hirudin, an activated factor II inhibitor, with DX9065a, an activated factor X inhibitor, on clot-associated thrombin and on prothrombin activation.

Clot-associated prothrombinase and thrombin activities may contribute to thrombus extension after thrombolytic and anticoagulant treatment. We studied prothrombin activation after adding human purified prothrombin to human clot. By using two different drugs with an exclusive direct anti-activated factor X activity (DX9065a) or anti-activated factor II activity (r-hirudin), we tried to determine whether clot-bound thrombin and prothrombinase could be inhibited in our experimental system when human purified prothrombin was added. Standard clots were prepared from platelet-poor human plasma after addition of calcium. We measured clot-bound thrombin or free thrombin using a direct simple chromogenic assay. In parallel, prothrombin fragment 1+2 measurement was used to monitor prothrombin activation. For this, two protocols were used. We introduced the direct inhibitors before starting the activation process (protocol A) or at the time of the activation process (protocol B). We found a direct correlation between thrombin generation and prothrombin fragment 1+2 with an increase of thrombin activity on clots and in the incubation mixtures when clots were incubated in human purified pothrombin alone. Two protocols were used: in the first, clots were pre-incubated in presence of drugs before adding prothrombin; and in the second, clots were incubated in the presence of prothrombin and drugs. Prothrombin activation was not inhibited when clots were incubated with r-hirudin and consequently thrombin generation still occurred. However, added r-hirudin blocks thrombin activity on the clots and in the incubation mixture, but does not prevent prothrombin activation, as shown by the increase of prothrombin fragment 1+2. In contrast, DX9065a did not suppress clot-bound thrombin. However, DX9065a blocks prothrombin activation whichever protocol was used. The results show that hirudin is a poor inhibitor of thrombin generation in contrast to DX9065a. On the other hand, DX9065a cannot inhibit thrombin bound to clot in contrast to hirudin.

Biological and clinical features of low-molecular-weight heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) is a common adverse effect of unfractionated heparin (UFH) therapy. In contrast, only a few patients have been reported with HIT following low-molecular-weight heparin (LMWH) therapy (LMW-HIT). To define the clinical and biological characteristics of LMW-HIT, 180 patients treated for suspected HIT at 15 French centres were investigated. Clinical history was recorded and HIT was confirmed in 59 patients with positive serotonin release assay results: 57 of them had high levels of antibodies (Abs) to heparin-platelet factor 4 complexes (H/PF4) and two had Abs to interleukin 8. Eleven patients were treated exclusively with LMWH (LMW-HIT) and 48 with UFH either alone (UF-HIT, n = 34) or combined with LMWH (UF/LMW-HIT, n = 14). The LMW-HIT and UF-HIT groups were similar with respect to sex, age, platelet count before heparin therapy, frequency of bleeding and occurrence of disseminated intravascular coagulation. The interval to onset of HIT was longer in LMW-HIT patients compared with UF-HIT patients (P = 0.03). Severe thrombocytopenia (platelets < 15 x 10(9)/l) was more frequent in the LMW-HIT group (P = 0.04). Thrombosis occurred in three of 11 LMW-HIT patients, i.e. as frequently as in UF-HIT patients. LMW-HIT is potentially severe and may be observed after longer heparin treatment compared with UF-HIT. It is highly recommended, therefore, that platelet counts be monitored carefully whenever LMWH is administered.

Protein S Gla-domain mutations causing impaired Ca(2+)-induced phospholipid binding and severe functional protein S deficiency.

We have identified 2 PROS1 missense mutations in the exon that encodes the vitamin K-dependent Gla domain of protein S (Gly11Asp and Thr37Met) in kindred with phenotypic protein S deficiency and thrombosis. In studies using recombinant proteins, substitution of Gly11Asp did not affect production of protein S but resulted in 15.2-fold reduced protein S activity in a factor Va inactivation assay. Substitution of Thr37Met reduced expression by 33.2% (P <.001) and activity by 3.6-fold. The Gly11Asp variant had 5.4-fold reduced affinity for anionic phospholipid vesicles (P <.0001) and decreased affinity for an antibody specific for the Ca(2+)-dependent conformation of the protein S Gla domain (HPS21). Examination of a molecular model suggested that this could be due to repositioning of Gla29. In contrast, the Thr37Met variant had only a modest 1.5-fold (P <.001), reduced affinities for phospholipid and HPS21. This mutation seems to disrupt the aromatic stack region. The proposita was a compound heterozygote with free protein S antigen levels just below the lower limit of the normal range, and this is now attributed to the partial expression defect of the Thr37Met mutation. The activity levels were strongly reduced to 15% of normal, probably reflecting the functional deficit of both protein S variants. Her son (who was heterozygous only for Thr37Met) had borderline levels of protein S antigen and activity, reflecting the partial secretion and functional defect associated with this mutation. This first characterization of natural protein S Gla-domain variants highlights the importance of the high affinity protein S-phospholipid interaction for its anticoagulant role.