Fibrinogen and von Willebrand factor and susceptibility vessel sign on T2*-weighted gradient echo imaging.

BACKGROUND: We investigated the relationship between von Willebrand factor (vWF), fibrin monomers (FM), fibrinogen baseline levels and the presence of susceptibility vessel sign (SVS) on T2*-weighted gradient echo imaging in acute ischemic stroke. METHODS: SVS was assessed at admission using T2*-weighted GRE. Plasmatic levels of vWF, FM and fibrinogen were evaluated before the initiation of intravenous thrombolysis. RESULTS: Forty-four patients were enrolled in this study. SVS was noted in 26 patients. Univariate analysis revealed that vWF >160% (p = 0.02) and fibrinogen >4 g/l (p = 0.03) were associated with a significant decrease in the likelihood of SVS. Multivariate analysis confirmed that higher levels of vWF or fibrinogen predicted the absence of SVS. CONCLUSIONS: The increased activity of vWF may promote a fibrin-platelet recruitment mainly contributing to the absence of (SVS).

The natural occurrence of human fibrinogen variants disrupting inter-chain disulfide bonds (A{alpha}Cys36Gly, A{alpha}Cys36Arg and A{alpha}Cys45Tyr) confirms the role of N-terminal A{alpha} disulfide bonds in protein assembly and secretion.

Analyses of site-directed fibrinogen mutants expressed in several recombinant models have previously shown that both inter- and intra-chain disulfide bonds are critical for fibrinogen assembly and secretion. Four naturally occurring mutations on AαCys36 and AαCys45 residues are reported here to be associated with decreased fibrinogen levels. This confirms the main role of the AαCys36-BßCys65 and AαCys45-γCys23 disulfide bonds in reaching a normal fibrinogen plasma level. Decreased coagulant/antigen ratios indicate abnormal species secretion in heterozygous subjects which varies between individuals. However, in contrast to overexpression in experimental models, disruption of the AαCys36-BßCys65 disulfide bond did not result in the appearance of Aα-Bß-γ moieties in vivo. A 188 kDa molecule reacting only with anti Aα and anti Bß chains was found in the plasma of the AαCys45Tyr variant. Heterozygous carriers of Aα chain mutations usually have normal fibrinogen levels, in contrast to the AαCys36Gly, AαCys36Arg and AαCys45Tyr variants that are shown here to cause hypofibrinogenemia.

Extracorporeal membrane oxygenation with danaparoid sodium after massive pulmonary embolism.

During extracorporeal membrane oxygenation, anticoagulation therapy is usually achieved with unfractionated heparin. We report on an extracorporeal membrane oxygenation with danaparoid sodium for a patient with severe respiratory failure due to massive pulmonary embolism and suspected type 2 heparin-induced thrombocytopenia. Danaparoid, a low molecular weight heparinoid, is an alternative to heparin for patients who develop type 2 heparin-induced thrombocytopenia. Danaparoid was given at 400 IU/h with an objective of antifactor Xa activity of 0.6-0.8 U/mL, which was monitored twice a day. No excessive bleeding or clotting of the circuit was noted. The patient was weaned from extracorporeal membrane oxygenation after 9 days of treatment.

Does p16ink4a expression increase with the number of cell doublings in normal and malignant lymphocytes?

p16(ink4a) is known to be a major inhibitor of cyclin-dependent kinases of G1-phase. Its accumulation is associated with replicative senescence. We analyzed to what extent the number of cell doublings may participate to p16(ink4a) expression in normal and malignant lymphocytes. p16(ink4a) expression, not found in normal quiescent B or T-lymphocytes, was observed after stimulation of B-lymphocytes (72 h) and T-lymphocytes (2 weeks) before the occurrence of replicative senescence markers such as senescence-associated-beta-galactosidase activity. Afterwards, in lymphocyte long-term cultures, the increase in p16(ink4a) followed the expression of features of cell ageing. In acute lymphoblastic leukemia, the analysis of the individual differences between peripheral blood and blood compartments (34 cases) showed a decrease in cell proliferation (p<0.005), in telomerase activity (p<0.0005), and in hTERT expression (p<0.04), associated with an increase of p16(ink4a) (p<0.035) in blood leukemic cells. These results support the hypothesis that (i) an increase in p16(ink4a) expression in normal lymphocytes is linked, in part, to the number of cell doublings before the occurrence of replicative senescence and (ii) this process is maintained in leukemic cell populations of numerous patients.

Predictive factors for thrombosis and major bleeding in an observational study in 181 patients with heparin-induced thrombocytopenia treated with lepirudin.

The antithrombotic efficacy of lepirudin in patients with heparin-induced thrombocytopenia (HIT) is compromised by an increased risk for bleeding. A retrospective observational analysis in 181 patients (median age, 67 years) with confirmed HIT treated in routine practice with lepirudin was performed to identify predictive factors for thrombotic and bleeding complications. Lepirudin was administered at a mean (+/- SD) dose of 0.06 +/- 0.04 mg/kg/h (compared with a recommended initial dose of 0.15 mg/kg/h). Mean activated partial thromboplastin time was greater than 1.5 times baseline value in 99.4% of patients. Median treatment duration was 7.7 days. Until discharge from the hospital, 13.8% and 20.4% of patients experienced a thrombotic or a major bleeding event, respectively. On multivariate analysis, mean lepirudin dose was not a significant predictive factor for thrombosis. In contrast, mean lepirudin dose greater than 0.07 mg/kg/h, long duration of lepirudin treatment, and moderate to severe renal impairment were significant positive factors for major bleeding. Overall, these results suggest that the recommended dose of lepirudin in patients with HIT is too high; the use of reduced doses may be safer with regard to bleeding risk and does not compromise antithrombotic efficacy.

Autolysosomes accumulate during in vitro CD8+ T-lymphocyte aging and may participate in induced death sensitization of senescent cells.

As autophagic inclusions accumulate in senescent fibroblasts, we wondered whether an increase in cellular fragility during in vitro lymphocyte aging may be related to an autolysosome accumulation. We established that, during long-term cultures, repeatedly stimulated T-lymphocytes acquired characteristics of replicative senescence and became progressively intolerant to activation. Cell death following stimulations: (i) corresponded to apoptosis, associated with necrosis at the end of the culture; (ii) was not, for its main part, mediated through CD95/CD178 or TNFRII/TNF alpha interactions; and (iii) occurred in spite of bcl-2 increased expression. After 14 weeks of culture, the percentage of lymphocytes containing at least one autophagic inclusion (p<0.0001) and the lipofuscin autofluorescence in lymphocytes (p<0.0001) were significantly increased. The expression of several genes regulating autophagy did not significantly vary with the age of the culture. Forty-eight hours after each stimulation, the percentage of induced cell death rose while, in the remaining living cells, the percentage of lymphocytes with autophagic vacuoles (p<0.05), with beta-galactosidase activity and the lipofuscin autofluorescence (p<0.001) significantly decreased, suggesting the preferential death of cells with autophagy. Our data support the view that the accumulation of autolysosomes in senescent lymphocytes might aggravate cellular fragility, leading to apoptosis and necrosis mainly induced by lymphocyte activation.

Early fibrinogen degradation coagulopathy is predictive of parenchymal hematomas in cerebral rt-PA thrombolysis: a study of 157 cases.

BACKGROUND: Little is known about the coagulation factors as predictors of cerebral bleeding in rt-PA thrombolysis. The aim of this study was to determine what early coagulation parameters could predict early hemorrhagic lesions. METHODS: Consecutive patients were included in the Lyon rt-PA protocol. Early hematomas (within 24 hours), diagnosed on an anatomoradiological basis (symptomatic and not symptomatic) were considered for the study. Fibrinogen and fibrin(ogen) degradation products (FDP) were assessed at entry and at 2 and 24 hours after the beginning of thrombolysis. RESULTS: Of 157 patients, 11 had early parenchymal hematomas (7%), 31 had early hemorrhagic infarcts (19.7%), and 115 had no bleeding (73.2%). In logistic regression, FDP at 2 hours was the single predictor of parenchymal hematomas (OR: 2.5; CI: 1.09 to 5.8), whereas an increase of FDP >200 mg/L multiplied the odds of parenchymal hematoma by 4.95 (IC: 1.09 to 22.4). Early parenchymal hematomas were indicative of a poor prognosis at 3 months (P=0.001). CONCLUSIONS: Early parenchymal hematomas appear as both "malignant" and exclusively related to an explosive increase of FDP at 2 hours, ie, an early fibrinogen degradation coagulopathy (EFDC). All patients scheduled to rt-PA thrombolysis should have an assay of FDP 2 hours after the beginning of thrombolysis: patients with an established EFDC (FDP >200 mg/L) should be monitored specifically, with no antithrombotic drug during the first 72 hours. Patients with FDP >100 mg should share the same monitoring.

CDK2 is involved in the S-phase lengthening induced by glucocorticoids in normal human lymphocytes.

Involvement of CDK2 in glucocorticoid-mediated S-phase lengthening was analyzed in this work. Dexamethasone (DXM) treatment of PHA-stimulated lymphocytes induced a decrease in CDK2 mRNA expression without any change in mRNA stability. This glucocorticoid-induced decrease in CDK2 mRNA expression could be suppressed by cycloheximide treatment. These results support the hypothesis of an indirect effect of DXM at the transcriptional level. Furthermore, CDK2 protein expression also decreased while the rate of protein synthesis and stability remained unchanged, suggesting a second post-translational level of regulation with the preferential degradation of a CDK2 protein stock fraction. The analysis of co-precipitated proteins showed that glucocorticoids induced modifications of protein complex composition. We found i) a preservation of the linkage capability of CDK2 for cyclin E and A, ii) a relative increase in p27kip1 linkage, and iii) a decrease in p21waf1 complexed with CDK2. As a consequence of CDK2 decrease and modifications of protein complex composition, pRb and histone H1 kinase activity of CDK2 was profoundly decreased. All these results pinpoint the role of CDK2 in glucocorticoid-induced S-phase lengthening and the potential activator role of p21waf1 for CDK2 in human lymphocytes.

A splicing donor site point mutation in intron 6 of the plasmin inhibitor (alpha2 antiplasmin) gene with heterozygous deficiency and a bleeding tendency.

A new case of familial plasmin inhibitor (alpha2 antiplasmin) deficiency is reported. The bleeding symptoms are moderate, happening after surgery or trauma or consisting of abnormal uterine bleeding induced by hormone replacement therapy. It is easily corrected with tranexamic acid. Gene sequencing makes it possible to find a splicing donor site mutation of intron 6, leading to exon 6 skipping. Neither a shortened variant nor an abnormal plasmin interaction was found in plasma by immunoblotting, and fibrin binding is unaffected. The mutation is heterozygous, associated with an intermediate decrease of both antiplasmin activity and antigen levels, and was found in four other family members out of five tested. It is different from the five mutations previously reported. At the time of diagnosis, the patient was living in Artas, France, allowing the defect to be named plasmin inhibitor (alpha2 antiplasmin) Artas.

In vitro tests for assessing heparin-induced thrombocytopenia in patients after elective hip replacement. A medico-economical evaluation.

OBJECTIVES: Considering the previously published incidences of heparin-induced thrombocytopenia (HIT) in patients receiving a thromboprophylactic therapy, the role of the hemostasis laboratory is essential in making a clinical decision. The purpose of this project was to compare the strategies of diagnosis and associated care of patients with suspected HIT after elective hip replacement using platelet aggregation assay, carbon 14-serotonin release, and "doing nothing." METHODS: The authors used an incremental cost-effectiveness analysis based on data extracted from the literature. The effectiveness of the strategies was represented by the number of deep venous thromboses prevented. Cost data were collected from the observation of biological and medical practice at Edouard Herriot University Hospital, Lyon, France, in 1999. RESULTS: In comparison with the strategies of doing nothing using no biological test for diagnosis, and clinical care of HIT-suspected patients, the strategy using platelet aggregation test was more expensive and less effective. With respect to the strategy using carbon 14-serotonin release assay, the incremental cost-effectiveness ratio, expressed as U.S. dollars per deep venous thrombosis prevented, reached $200,000, with a marginal effectiveness of eight deep venous thromboses prevented for 10,000 HIT-suspected patients. CONCLUSION: This study suggests that clinical hemostasis laboratories might consider replacing the platelet aggregation test with the carbon 14-serotonin release assay or should use another functional assay such as the flow cytometric assay for the diagnosis and care of patients with suspected HIT.