Endovascular management of chronic disabling ilio-caval obstructive lesions: long-term results.

OBJECTIVE: To report the long-term results of stenting for chronic ilio-caval obstructive lesions. MATERIAL AND METHODS: From January 1996 to January 2008, 89 patients (72 women, 17 men; median age 43 years) were admitted for endovascular treatment of chronic disabling non-malignant obstructive ilio-caval lesions. Patients were classified as C2 in 15 cases, C3 in 59, C4 in seven, C5 in two and C6 in six. Median preoperative venous disability score (VDS) and venous clinical severity score (VCSS) were 2 and 9, respectively. Aetiology was primary in 52 patients, secondary in 35 and congenital in two. Lesions were bilateral in seven cases, eight patients had inferior vena cava (IVC) involvement and 18 had common femoral vein (CFV) obstructive lesions. Complete occlusion was found in 30 cases. RESULTS: Technical success was achieved in 98%. The median hospital stay was 2 days. During a median follow-up of 38 months (range: 1-144 months), one patient died and five cases of thromboses occurred. Iterative stenting was performed for restenosis in six cases. Primary, assisted-primary and secondary patency rates, in terms of intention to treat, were 83%, 89% and 93%, respectively, at 3 and 10 years, with a median VDS of 1. Univariate analysis found that significant factors affecting patency were CFV involvement for primary patency and history of deep venous thrombosis (DVT) and CFV involvement for secondary patency. The last 46 patients had statistically more severe lesions than the first 43 (higher VDS, more secondary lesions, more occlusions, more stented segments, higher length of stented vein), and in spite of which patency rates are not different. CONCLUSION: Endovenous angioplasty, combined with stenting, is a sure, safe, effective and very minimally invasive technique which provides good long-term patency rates. Currently, it is recognised as the technique of choice for the treatment of ilio-caval obstructive lesions. Surgery should be proposed only in case of failure.

In vitro generation of endothelial microparticles and possible prothrombotic activity in patients with lupus anticoagulant.

Microparticles (MPs) resulting from vesiculation of platelets and other blood cells have been extensively documented in vitro and have been found in increased numbers in several vascular diseases, but little is known about MPs of endothelial origin. The aim of this study was to analyze morphological, immunological, and functional characteristics of MPs derived from human umbilical vein endothelial cells (HUVECs) stimulated by TNF, and to investigate whether these MPs are detectable in healthy individuals and in patients with a prothrombotic coagulation abnormality. Electron microscopy evidenced bleb formation on the membrane of TNF-stimulated HUVECs, leading to increased numbers of MPs released in the supernatant. These endothelial microparticles (EMPs) expressed the same antigenic determinants as the corresponding cell surface, both in resting and activated conditions. MPs derived from TNF-stimulated cells induced coagulation in vitro, via a tissue factor/factor VII-dependent pathway. The expression of E-selectin, ICAM-1, alphavbeta3, and PECAM-1 suggests that MPs have an adhesion potential in addition to their procoagulant activity. In patients, labeling with alphavbeta3 was selected to discriminate EMPs from those of other origins. We provide evidence that endothelial-derived MPs are detectable in normal human blood and are increased in patients with a coagulation abnormality characterized by the presence of lupus anticoagulant. Thus, MPs can be induced by TNF in vitro, and may participate in vivo in the dissemination of proadhesive and procoagulant activities in thrombotic disorders.

Two miscarriages, consecutive or non-consecutive, does it change something?

OBJECTIVES: To assess the rate of anomalies in the etiological evaluation of patients presenting recurrent early miscarriages (RM) according to miscarriage chronology (number of miscarriages, history of live birth and succession of RM). METHODS: Retrospective single centre study including RM, defined as at least 2 miscarriages at less than 14 weeks of gestation (WG) between the 1st January 2012 and the 31st December 2015. Clinical data and etiological evaluation include blood glucose levels, screening for antiphospholipid syndrome (APS), endocrine assessment, vitamin levels, pelvic imaging, karyotyping of both partners, chronic endometritis and thrombophilia screening. RESULTS: Two hundred and eighty-eight patients were included over this period, 118 (41%) patients had no history of live birth. Two hundred and twenty-three (77%) patients had consecutive RM and 65 (22%) patients had non-consecutive RM. For consecutive RM, 62,8% had thrombophilic disorders versus 69,8% for non-consecutive RM (P>0,05); 44,7% had endocrine disorders or vitamin deficiencies versus 39,7%; 34,6% of patients with consecutive RM had uterine anomalies versus 45,5% respectively. No difference was found depending on the recurrence of RM or the history of live birth (P>0.05) apart from the age of the patient. Fifty-nine (17.4%) patients had uterine anomalies. There are 24 chronic endometritis on 31 biospsies performed. Seventy-eight (27%) patients were offered treatment. Ninety-four (90%) patients showed good therapy compliance. Eighty-one (78%) patients became pregnant. CONCLUSION: An etiological evaluation provides, for over half of the cases, an etiology or the identification of risk factors responsible for RM, as well as in some cases offering an adapted, efficient, therapeutic approach. This evaluation should be offered regardless of the obstetric history of the patient.

Modulation of plasminogen activator systems by matrix components in two breast cancer cell lines: MCF-7 and MDA-MB-231.

We have analyzed the plasminogen activator (PA) systems of two metastatic breast adenocarcinoma cell lines, MCF-7 and MDA-MB-231, as a function of 17 beta-estradiol stimulation when the cells were cultured on purified components of extracellular matrix. Laminin enhanced PA levels in both cell lines, but this enhancement seemed to occur via different mechanisms, including dissociation of inhibitor complexes. The major effect was the marked increase in cell-associated urokinase-type PA (u-PA); the increase was independent of estrogen in hormone-insensitive MDA-MB-231 cells grown on laminin-coated surfaces. In estrogen-sensitive MCF-7 cells, 17 beta-estradiol stimulated u-PA secretion in a similar fashion on plastic, laminin, fibronectin, or collagen but acted in synergy with laminin in the production and release of tissue-type PA.

Cytofluorometric detection of human endothelial cells in whole blood using S-Endo 1 monoclonal antibody.

It has long been debated whether endothelial cells are present at very low frequency in peripheral blood. Elevated concentrations of such circulating cells may represent a good marker of vascular injury. We have therefore designed an immunocytometric assay for the detection of rare endothelial cells in whole blood. This assay is based on a new monoclonal antibody (MAb) S-Endo 1, made against human umbilical vein endothelial cells (HUVEC) and specific for endothelial cells of various origins without detectable reactivity with blood cells. First, the sensitivity of the assay was established by using normal blood samples with admixed HUVEC as an in vitro model. A good correlation was obtained between added and counted endothelial cells; the recovery was greater than 90% and the minimum detectable concentration of HUVEC was about 0.2 cells/microliters whole blood. Using this rapid counting technique, no detectable levels of endothelial cells were found in the blood of normal individuals (CE less than or equal to 0.1 cells/microliters) while elevated concentrations (up to 8 cells/microliters) were detected in a human model of vascular injury corresponding to a traumatic venepuncture. Thus, this new whole blood immunocytometric assay using S-Endo 1 MAb may be useful in determining the levels of circulating endothelial cells in vascular disorders.

Plasma thrombomodulin in orthotopic liver transplantation.

Plasma thrombomodulin (TM), a specific marker of vascular endothelial injury was measured pre-, per-, and postoperatively in 16 consecutive patients undergoing orthotopic liver transplantation (OLT). The TM level, which was already elevated preoperatively, remained unchanged during OLT, except for an acute and transitory spike at the time of graft reperfusion. This TM peak is probably attributable to an acute release from the patient's endothelium because the TM level in the last saline rinse of the graft before implantation was low. This TM spike was not correlated with the progressive tissue-type plasminogen activator (t-PA) increase, plasminogen activator inhibitor 1 (PAI-1), or von Willebrand factor (vWF) values. The absence of accumulation of TM in plasma, unlike that of t-PA, suggests that the liver does not play a major role in TM clearance in humans. At the end of surgery, individual TM values returned to preoperative levels and remained unchanged during the 7 days following surgery. This observation suggests that the high (or very high) TM levels measured in these patients might be due to an indirect rather than a direct effect of liver dysfunction on the vascular endothelium which remained damaged during the postoperative period. The possibility that TM might be a predictive marker for thrombotic OLT complications remains to be investigated in a postoperative follow-up study.

Release pattern of the vascular plasminogen activator and its inhibitor in human postvenous occlusion plasma as assessed by a spectrophotometric solid-phase fibrin-tPA activity assay.

Vascular or tissue-type plasminogen activator (plasma t-PA) is the circulating physiological fibrinolytic enzyme of endothelial cell origin which function is regulated by fibrin and a specific inhibitor (PAI). To study the pattern of release of t-PA and the behavior of t-PA-PAI complexes in plasma we determined t-PA activity in 44 healthy subjects before and after 10 min of forearm venous occlusion using a new spectrophotometric solid-phase fibrin-tPA activity assay. The assay is based on 1) the high affinity binding of t-PA to fibrin, and 2) the detection of fibrin-bound t-PA by measuring the release of pNA from a chromogenic substrate in the presence of plasminogen. Values at rest were rather undetectable in plasma (0.05 +/- 0.03 IU/ml, in 23 out of 44 samples) but were positively detected in all the euglobulins: 0.88 +/- 0.68 IU/ml. After venous occlusion the majority of plasmas (36 out of 44) showed a slight increase in t-PA activity (0.65 +/- 0.63 IU/ml) as compared to the important level observed in all the euglobulins (9.78 +/- 9.58 IU/ml). So, the ratio plasma/euglobulin t-PA activity was very low (0.06) and remained identical in both pre- and postocclusion samples. However, when diluted plasmas were tested the inhibitory effect disappeared and t-PA activity increased indicating that although t-PA circulates in a neutralized state it can be available for fibrinolysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Concomitant secretion by A431 cells of tissue plasminogen activator and a specific inhibitor masks EGF modulation of tPA activity.

It has previously been reported that EGF enhances uPA but not tPA in the A431 squamous carcinoma cell line. To determine whether the absence of tPA modulation by EGF reflected steady levels or the action of an anti-activator, we assayed tPA, PAI-1 and tPA/PAI-1 complexes by zymography and immunological assays. Under conditions in which EGF had no effect on tPA activity, tPA antigen paradoxically increased with a concomitant rise of tPA/PAI-1 complexes. This indicated that tPA was rapidly inactivated through the formation of a complex, immunologically and electrophoretically related to tPA/PAI-1. tPA antigen and tPA/PAI-1 complexes were modulated by EGF in a time and concentration dependent manner. PAI-1 antigen was secreted into A431 medium (CM) after a lag phase of 16 h in both control and EGF-treated cultures. Evidence is presented here that two forms of PAI-1 are present in A431 CM: an inactive form and an active form which neutralizes the tPA secreted, masking its enhancement by EGF in functional assays.

Rapid isolation of human endothelial cells from whole blood using S-Endo1 monoclonal antibody coupled to immuno-magnetic beads: demonstration of endothelial injury after angioplasty.

The presence in whole blood of circulating endothelial cells (EC) has been a subject of debate for many years. It could represent a good marker of vessel injury. We demonstrate here that human endothelial cells can be directly isolated and identified in circulating blood by means of an endothelial cell specific monoclonal antibody, S-Endo1, coupled to micromagnetic beads. The specificity and efficacy of the assay were established using normal blood samples with cultured EC added. Specific rosettes formed between EC and beads could subsequently be isolated with a magnet. The rosetted cells were recovered with a yield greater than 80%. Their endothelial origin was confirmed by the positive labelling of von Willebrand factor and thrombomodulin, as well as the presence of Weibel-Palade bodies. We applied this method to demonstrate significantly increased levels of EC in venous and arterial human blood samples in patients undergoing heart catheterization. This new whole blood immuno-separation method may be useful in determining endothelial cell injury in vascular disorders.

Quantification of lupus anticoagulants in clinical samples using anti-beta2GP1 and anti-prothrombin monoclonal antibodies.

Quantification of lupus anticoagulant (LA) in clinical samples is hampered by the lack of a suitable standard of activity. We evaluated the use of mAbs displaying LA activity for this purpose. As most patient samples contain both beta2Glycoprotein I (beta2GP1) and prothrombin dependent LA, a combination of two mAbs, one of each specificity, was added to normal plasma in a concentration from 0 to 60 microg/ml. Eight assay systems using different reagents and instruments were used. The calibration curves were linear for all but one, with marked differences between the responsiveness to each mAb. A panel of plasmas from 69 patients with persistent LA diagnosed using the SSC-ISTH criteria was tested. An antiphospholipid syndrome (APS) was present in 40, whereas 29 were asymptomatic. LA activities of individual plasmas varied between assays (p < 10(-4)), but homogeneous subgroups were identified. In a majority of samples, LA activity displayed a prothrombin-dependent profile, with a variable contribution of beta2GP1-dependent activity. The latter was associated to beta2GP1 antibodies detected by solid-phase immunoassay. By using 3 dilute Russell viper venom time assays, higher LA titers were found in APS, compared to asymptomatic patients (p <0.05).

Experimental infection of human erythrocytes from alcoholic patients with Bartonella quintana.

Bartonella spp. are found in the erythrocytes of their specific natural hosts and B. quintana bacteremia is associated epidemiologically with lice, alcoholism, and homelessness. The aim of our study was to compare the growth and the number of bacteria per erythrocyte in vitro in laboratory-infected red blood cells from alcoholic patients versus normal blood donor erythrocytes. Enumeration of bacteria was performed either with plate counting or with a real-time PCR quantitative assay. Number of bacteria per cell was determined using immunofluorescence assay and laser confocal microscopy. Although the number of bacteria after 4 days of incubation was similar in the two groups of erythrocytes, we found that the distribution of bacteria per erythrocyte in the two groups was different. Erythrocytes from alcoholics contain significantly more bacteria per cell than erythrocytes from blood donors. Our results suggest that there is a link between alcoholism and infections of B. quintana that may be due to the macrocytosis of erythrocytes.

Intraoperative evolution of coagulation parameters and t-PA/PAI balance in orthotopic liver transplantation.

The changes in coagulation and fibrinolysis were investigated in 10 patients undergoing orthotopic liver transplantation (OLT) which is known to be frequently associated with perturbations of haemostasis. The coagulation profile, already deteriorated before surgery in most patients, showed no appreciable further alteration. On the other hand, important modifications of fibrinolytic parameters occurred, essentially concerning tissue-type plasminogen activator (t-PA) and its specific inhibitor (PAI). t-PA activity constantly increased in the course of transplantation, reaching a maximum at the end of anhepaty. Large interindividual variations were noted in the level of t-PA activity (7.5 to 135 IU/ml). Free PAI activity followed a reverse kinetics, remaining low during the anhepatic stage, and dramatically increasing after allograft reperfusion. Despite the fibrinolytic potential related to high circulating t-PA levels, no biologic nor clinical evidence of systemic fibrinolysis was observed peroperatively. These findings suggest that PAI release could represent an early process making the use of antifibrinolytic drugs during OLT unnecessary.

Modulation of tPA, PAI-1 and PAI-2 antigen and mRNA levels by EGF in the A431 cell line.

It has been reported that EGF treatment enhances uPA but not tPA in the A431 epidermoid carcinoma cell line. To determine whether the absence of tPA modulation by EGF could be due to the action of inhibitors, we assayed tPA, PAI-1, PAI-2 and tPA/PAI-1 complexes by immunological assays and zymography in A431 serum-free medium. We found that, under conditions in which EGF had no effect on tPA activity, tPA antigen increased with a concomitant rise of tPA/PAI-1 complexes, indicating the action of an inhibitor. Both tPA antigen and tPA/PAI-1 complexes were modulated by EGF in a time and concentration dependent manner. tPA/PAI-1 complex levels were lower than tPA levels, suggesting the presence of other inhibitors. Immunological assays detected PAI-2 in addition to PAI-1 and showed a time and dose response to EGF. Modulation of tPA and the anti-activators by the growth factor was confirmed by identification of the corresponding transcripts with cDNA probes. We conclude that the net plasminogen activator activity in A431 cells is the result of a balance between activators and inhibitors.

Antiphospholipid syndrome and factor V Leiden. Three cases with recurrent venous thrombosis.

Recurrent thrombosis is a common complication of various rheumatic disorders and is part of the definition of antiphospholipid syndrome. We report three cases of recurrent venous thrombosis due not only to antiphospholipid syndrome with a normal activated partial thromboplastin time but also to resistance to activated protein C caused by the factor V Leiden mutation. These three cases confirm that thrombotic disease is frequently multifactorial and suggest that resistance to activated protein C should be looked for routinely in patients with suggestive clinical manifestations, particularly when standard clotting tests are normal.

[Antiphospholipid antibodies: clinical significance and biological diagnosis]. / Les anticorps "antiphospholipides": intérêt clinique et diagnostic biologique.

The term "antiphospholipids" (aPLs) refers to an heterogeneous family of antibodies diagnosed either by clotting tests: the lupus anticoagulants or by Elisa: anticardiolipin (aCL) and anti-beta2-glycoprotein I (anti-beta2GP1) especially. aPLS recognize phospholipids, alone or bound to plasma protein cofactor(s), or the cofactors themselves. aPLs have long been described in autoimmune diseases such as SLE, but may also be found in other clinical settings including infections, malignancies and drug administration. Their persistent presence can be associated with venous and/or arterial thrombotic complications and/or recurrent miscarriage, thus defining the "antiphospholipid syndrome" (APS). The heterogeneity of aPLs makes a comprehensive approach to laboratory investigation essential. Detection of lupus anticoagulants relies on increased clotting times in phospholipid-dependent tests. Their 4 step diagnosis includes: 1) screening (by at least two different tests); 2) demonstration of an inhibitory activity; 3) evidence of its phospholipid dependence; 4) exclusion of an associated coagulopathy. Among the aPLs detected by Elisa, IgG aCL are the most frequently investigated. However, other antibodies may represent useful biological tools. Among them, anti-beta2GP1 are thought to be more closely associated with a history of thrombosis than aCL and testing for anti-beta2 GP1 should now be systematically included in the biological diagnosis of APS. The Elisa used for aCL and anti-beta2GP1 are not fully standardized, and a number of methodological parameters may account for the interlaboratory discrepancies often observed. The clinical importance of other antibodies such as antiphosphatidylethanolamine, antiprothrombin or antiannexin V is being evaluated. An appropriate laboratory investigation of APS should, in all cases, combine the use of clotting and immunological assays, and assess the persistence of autoantibodies over time.

[Antiphospholipid antibodies]. / Les anticorps antiphospholipides.

The term 'antiphospholipids' (APL) refers to heterogeneous auto-antibodies, including anticardiolipins detected by immunological methods and lupus anticoagulants detected by clotting tests. APL are currently of considerable interest, both from a clinical and a biological point of view, since their presence is associated with thromboembolic events. In this review, the authors emphasize the diversity of the clinical settings where APL are diagnosed and investigate the relationship between APL and thrombosis. The heterogeneity of APL and the lack of standard techniques make their laboratory diagnosis difficult and require the use of various types of tests. Several pathogenic mechanisms, all related to a possible effect of APL on the antithrombotic functions of vascular endothelium, have been proposed: decrease in prostacyclin synthesis, induction of procoagulant activity, inhibition of the endothelial anticoagulant functions, and impairment of fibrinolysis. Given the heterogeneity of these antibodies, it is unlikely that a single mechanism can account for their prothrombotic effect.

[Determination of factor X Ag by laser immunonephelemetry]. / Dosage du facteur X Ag par immunonéphélémétrie laser.

A method of dosage of X Ag factor by laser immunonephelemetry is proposed. This technique, carried out with a rabbit antiserum in the presence of PEG and EDTA, presents the advantage to be sensitive, reproducible and mostly faster than the other immunological methods, such as Electroimmunodiffusion (EID) and Enzyme Linked Immuno-Sorbent Assay (ELISA). It constitutes a simple tool, applicable in the demonstration of an enzyme dysfunction or liver pathology. Finally, it could bring a supplement of information in monitoring antivitamin K therapy.

[Acquired factor V inhibitor: etiology, bleeding risk and therapeutic management with regard to three cases]. / Anticoagulant acquis antifacteur V: nosologie, risque hémorragique et prise en charge thérapeutique à propos de trois observations.

INTRODUCTION: Acquired factor V inhibitor is rare and clinical symptoms are quite variable. Bleeding is the leading symptom but some patients are asymptomatic. Several diseases or conditions are associated with factor V inhibitors. Various treatments have been attempted but randomized or prospective trials are not available. EXEGESIS: Here we report three cases of acquired factor V inhibitor. These reports highlight the clinical variability of this disorder. Pathogenesis and therapy with reference to the literature are discussed. CONCLUSION: Factor V inhibitors are rare and associated to several diseases or conditions. Pathogenesis is still unclear except in patients exposed to bovine thrombin. The majority of the cases developed after surgery. In a few cases there is an association to a malignant or autoimmune disease. Plasmapheresis and platelet transfusions might be the best treatment in case of severe bleeding. High-dose intravenous immunoglobulin infusions have been used successfully in some cases and we report here their efficacy in two cases.

[Fibrinolytic activity in patients undergoing hepatic transplantation]. / Activité fibrinolytique chez les patients opérés pour transplantation hépatique.

Bleeding complications during liver transplantation have been attributed to accelerated fibrinolysis. In order to determine its cause, 11 adults (mean age: 38.9 +/- 13.2 yr) undergoing liver transplantation were studied. There were three groups of patients: cirrhosis (n = 4), fulminating hepatitis (n = 4) and one group including a primary biliary cirrhosis, a hepatic metastasis and a hepatoma. The following factors were studied in the immediate preoperative period, at different surgical times throughout the procedure and 2-3 h after the end of the abdominal sutures: platelet count, prothrombin concentration, fibrinogen, activated kephalin time, factors II, V, VII + X and VIIIc, antithrombin III, protein C, D-dimers, fibrinogen and fibrin degradation products (PDF), plasma plasminogen, tissue plasminogen activator (tPA) and the fast tPA inhibitor (PAi). Preoperatively, only the two patients with hepatic cancer had a normal haemostatic profile. Throughout the procedure, all patients had only moderate changes in platelets, coagulation factors and their inhibitors, and plasminogen, because platelet concentrates and fresh frozen plasma were transfused. Levels of tPA rose, becoming very high during the anhepatic period and just after graft reperfusion. An abrupt fall occurred at the end of surgery. There were important individual differences in tPA activity. PAi activity was low during the preanhepatic and anhepatic stages, rising rapidly after revascularization.(ABSTRACT TRUNCATED AT 250 WORDS)