Nadia Montani

Study of three patients with monoclonal gammopathies and ‘lupus‐like’ anticoagulants

In three patients with monoclonal gammopathies: a case of multiple myeloma, a case of monoclonal gammopathy of uncertain significance (MGUS) and a case of monoclonal gammopathy associated with lymphocytic lymphoma, we found the presence of a circulating lupus‐like anticoagulant. Coagulative studies showed that the paraproteins: an IgG3k, an IgG1k and an IgMλ, were responsible for the anticoagulant activity by interacting with the thromboplastin phospholipids. Using isoelectrofocusing we demonstrated that the three monoclonal immunoglobulins had a strong basic charge which may have contributed to determining their interaction with the acidic thromboplastin phospholipids. The binding of various phospholipids to the monoclonal proteins was assessed by the fluorescence quenching method which showed heterogeneous specificity. In order to establish whether the electrical charge is also relevant in cases with polyclonal lupus anticoagulant, the polyclonal immunoglobulins were fractionated according to their charge. The strongest inhibitor activity was found in the most basic immunoglobulins. Monoclonal lupus‐like anticoagulants represent useful tools for investigating the heterogeneous world of polyclonal lupus‐like anticoagulants.

Bleeding Tendency of Unknown Origin and Protein Z Levels

known origin showed low levels of protein Z, suggesting that a protein Z assay must be included in

Platelet function after in vivo and in vitro treatment with thrombolytic agents

Whereas in vitro studies showed that plasmin may induce both inhibition and activation of platelets, in vivo and ex vivo investigations suggested that thrombolytic agents are responsible for platelet stimulation. To gain further information on this topic, ex vivo platelet function was studied in 24 subjects with acute myocardial infarction treated with streptokinase or recombinant tissue-type plasminogen activator (rt-PA). Ten patients with acute myocardial infarction who did not receive thrombolytic treatment were also investigated. The data shows that at the end of thrombolytic infusion, the maximal extent of platelet aggregation and adenosine triphosphate release was reduced in treated patients compared with that in untreated ones. In subjects treated with streptokinase, the defect in platelet aggregation derived from both cellular and plasmatic defects. Plasmatic beta-thromboglobulin concentration was significantly reduced after streptokinase, but unchanged after rt-PA. Three days after thrombolytic treatment, platelet aggregation of patients receiving streptokinase or rt-PA was not significantly different from that of untreated subjects. A similar defect in platelet function was obtained in vitro, incubating normal platelet-rich plasma with pharmacologic concentrations of streptokinase. Again, platelet function defect derived from both cellular and plasmatic damages. It cannot be excluded that platelet activation occurs in patients with acute myocardial infarction during the very early phases of thrombolytic treatment. However, it is suggested that a transient defect in platelet function follows both streptokinase and rt-PA infusion.

Abnormalities in thrombin-antithrombin pathway in AL amyloidosis.

Various pathogenic factors have been proposed to explain the abnormal hemostasis observed in AL amyloidosis. Since imbalance between clotting factors and inhibitors could play a pathogenic role in both hemorrhagic and thrombotic manifestations, we investigated the thrombin-antithrombin pathway in 35 patients with AL amyloidosis. Ten patients suffered from bleeding while 3 patients experienced deep venous thrombosis. Thrombin time was prolonged in 29 subjects, the mean values of antithrombin III activity (ATIII Act) were significantly lower than those of antithrombin III antigen (ATIII Ag) with loss of relationship between these two different techniques of ATIII detection, normally observed in healthy controls. In 19 patients increased levels of thrombin-antithrombin (TAT) complexes were present. Crossed immunoelectrophoresis of ATIII, performed in presence of heparin, evidenced ATIII forms with reduced binding capacity to heparin and TAT complexes of various electrophoretic mobilities. In conclusion, the impairment of the thrombin-antithrombin pathway, in association with the low ATIII biological activity, might play a pathogenic role in the hypercoagulable state reported in AL amyloidosis, despite the higher frequency of bleeding manifestations.

Plasma levels of protease inhibitors in acute myeloid leukemia at the onset of the disease and during antiblastic therapy

Abstract The levels of some plasma protease inhibitors during acute myeloid leukemia were investigated in two groups of patients: one with DIC and one without DIC. Our results indicate that plasma levels of α 1 -P.I. and α 1 -Achy, two specific inhibitors of leucocyte neutral proteases, were constantly high in all subjects and showed further increases after chemotherapy. α 2 -M, the third specific inhibitor, was generally low and did not seem to be univocally affected by the therapy. In patients with DIC a strong discrepancy between ATIII R: AG and ATIII activity was observed, probably indicating the presence of ATIII-thrombin complexes with partial inactivation of its biological function.

Haemostatic variables, serum lipid abnormalities and vascular complications in diabetes mellitus: a 5 year follow-up study.

SummaryThe relationship among blood lipids, haemostatic and fibrinolytic parameters have been evaluated, during a follow-up study, in 27 non-insulin dependent (type II) diabetic patients. Upon recruitment, and in periodical controls, we observed that plasma triglycerides and VLDL levels correlated inversely, and HDL directly, with the fibrinolytic activity of plasma and euglobulin precipitate. Furthermore triglycerides and VLDL correlated directly with factor VIII antigen (vWFAg). After 5 years in the study, 12 patients (44%) had macroangiopathic complications, and 9 of these subjects showed persistently high levels of triglycerides (above 2.36 mmol/l). These haemostatic and lipid components, however, do not influence the progression of diabetic retinopathy and nephropathy. The alterations of lipid, haemostatic and fibrinolytic parameters and their possible relationships seem to play an important role in the occurrence of diabetic macroangiopathy.

Activated prothrombin complex concentrate (FEIBA®) for the treatment and prevention of bleeding in patients with acquired haemophilia: A sequential study

Despite anti-haemorrhagic therapy with proper doses of activated prothrombin complex concentrate (aPCC, Feiba®), patients with acquired haemophilia A (AHA) have a considerable risk of recurrent bleeding complications. Evidence in support of the benefit-to-risk ratio of prevention strategies with the use of lower doses of aPCC following the end of the initial treatment period is scarce and inconclusive. We report our experience in the management of 18 consecutive patients with AHA admitted to two Haemophilia centres in Italy. We managed the first 11 according to current guidelines (e.g., with conventional aPCC doses until bleeding resolution). Then, we decided to prolong the treatment beyond bleeding resolution with lower doses of the same concentrate (short-term prophylaxis) in the 7 additional patients. In these patients, the treatment was continued for as long as the titre of FVIII inhibitor was found to decrease by at least 50% when compared to the baseline one. We observed six relapses of bleeding in patients in whom aPCC was confined to the treatment of the qualifying bleeding episode, and none in patients to whom lower doses were administered until the pre-specified decrease in the titre of FVIII inhibitor was achieved. No patients experienced thrombotic complications during the study period. Prolonging the treatment with lower doses of aPCC beyond the initial phase in patients with AHA in whom the titre of FVIII inhibitor is still high is likely to safely prevent further bleeding complications.

Protocollo per l’identificazione di pazienti con difetti congeniti dell’emostasi non noti: screening pre-operatorio

Congenital Hemorrhagic Diseases (CHD) are often complicated by intra- and post-surgical bleeding that is the cause of diagnostic procedures. This study presents a protocol applied to the patients with indication of elective surgery, aimed to detected CHD and the consequent surgical hemorrhagic risk, on the basis of structured history concerning bleeding tendency (Bleeding Score-BS) and/or laboratory clotting screening tests. 1100 consecutive patients have been enrolled. The abnormal BS and/or alterations of screening clotting tests, have been further evaluate by medical staff experienced on hemostasis to make the diagnosis of CHD and to suggest hemostatic prophylaxis during and after surgery. In conclusion these preliminary data do not support the hypothesis that mild CHD are under diagnosed in general populations. A case-control study will be planned to register eventual differences on bleeding surgical complications between patients considered as care or as control in our protocol.

Prevalenza di trombosi arteriose e venose in presenza delle mutazioni G20210A del gene della protrombina e R506Q del gene del fattore V (Leiden)

Arterial and venous thrombosis is becoming a pathology of primary importance because of other concomitant pathologies (metabolic, cardiac, neoplastic diseases) which often acerbate the prognosis. Therefore, our research aims to clarify in particular the function of congenital prothrombotic risk factors, that is the R506Q gene muta-tion of the FV (Leiden) and the G20210A gene mutation of the FII. We analyzed 351 subjects in treatment at the Hemostasis Diseases Unit of IRCCS Policlinico San Matteo Foundation, Pavia. We collected each personal and family anamnesis and carried out the thrombophilia screening test. Among this population, 63 subjects resulted positive to, at least, one of the two mutations we studied. Comparing the number of subjects having at least a mutation with the ones without mutations, we found out that the first ones had a higher VTE percentage than the second ones. We also examined risk factors. All the results were supporting the thrombosis disease. Processing people to the thrombophilia screening test means to recognise and manage subjects at risk before the thrombosis event happens.

Modalità di diagnosi e caratteristiche cliniche della Malattia di von Willebrand Acquisita

Acquired von Willebrand’s disease (AVWD) is a rare disorder of hemostasis, with pathogenesis variable, associated with underlying diseases like lymphoproliferative, myeloproliferative disorders and cardiovascular diseases. Unlike the congenital form, in AVWD the reduction of von Willebrand Factor (VWF) is due to the increased clearance attributed to several mechanisms. From 2006 to 2012, have been diagnosed at the unit of Haemostasis Diseases of the Medical Clinic III 13 clinical cases with the following conditions: lymphoproliferative (4 cases), myeloproliferative disorders (7 cases), chronic inflammatory diseases (2 cases). The diagnosis was made for the appearance of severe hemorrhagic diathesis in 3 patients, moderate in 5, mild in 2, and in 3 patients in the absence of hemorrhagic manifestations, as affected by diseases often associated with AVWD. Six of the 13 patients required an invasive procedure for the treatment of the hemorrhages, and only 3 of 13 patients were treated with FVIII concentrates / VWF for bleeding complications. The outcome of AVWD is variable. In our study we obtained a stable remission of AVWD in the patient with Waldenstrom’s disease, following the chemotherapy.