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Biomedical subjects

M Furlan

Publications and source records attributed to M Furlan.

At least 19 recordsLinked to original sources

Antioxidant activity of Maytenus ilicifolia root bark.

Maytenus ilicifolia is an important plant with potential on cancer treatment and has been largely used in Brazil and other countries. We have evaluated the crude ethanolic extract of M. ilicifolia as a potential antioxidant source using an assay based on the bleaching of the radical monocation 2,2'-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS(*+)) and by HOCl scavenger capacity. Trolox and uric acid were used as positive controls. The results indicated M. ilicifolia root bark as a great source of antioxidants based on its potential as scavenger of radicals.

Antioxidants↗

A Phase II trial of docetaxel and cisplatin in patients with recurrent or metastatic nasopharyngeal carcinoma.

UNLABELLED: A Phase II study was conducted to determine the efficacy and toxicity of the combination of docetaxel and cisplatin, in patients with recurrent or metastatic nasopharyngeal carcinoma (NPC). Nine patients (median age 39 years) with NPC were enrolled, none had prior chemotherapy for their recurrent or metastatic disease. Docetaxel was administered as a 1-h intravenous infusion at a dose of 75 mg/m(2) on day 1; cisplatin was administered at a dose of 75 mg/m(2) on day 1, immediately after docetaxel. Treatment was repeated every 3 weeks. The primary endpoint was objective response rate and the secondary endpoints were duration of response, time to progression, and overall survival. A total of 45 chemotherapy cycles were administered. In an intention-to-treat analysis two patients (22%, 95% confidence interval (CI): 3-60%) achieved a partial response. The median duration of response was 4.1 months, the median time to progression 8.4 months and the overall survival at 1 year from start of treatment was 76%. Grade 3-4 neutropenia was observed in all (100%) patients over 93% of the treatment cycles, and in three cases this was complicated by fever. Other toxicities were mild. CONCLUSIONS: The combination of docetaxel and cisplatin has manageable toxicity but little efficacy as first-line treatment in patients with recurrent or metastatic NPC. In view of the low response rate, accrual was terminated and the trial was aborted.

Adult↗

Remission of thrombotic thrombocytopenic purpura in a patient with compound heterozygous deficiency of von Willebrand factor-cleaving protease by infusion of solvent/detergent plasma.

UNLABELLED: Plasma exchange or plasma infusion is considered to be the therapy of choice in patients with thrombotic thrombocytopenic purpura (TTP) who are deficient in von Willebrand factor-cleaving protease (VWF-CP). Recently, mutations in the ADAMTS 13 gene were identified as being responsible for VWF-CP deficiency in patients with familial TTP (VWF-CP deficiency in the absence of an inhibitor). Here we report on a girl who presented with recurrent thrombocytopenia and anaemia since birth, developing the full pentad of characteristic TTP at the age of 16 y. Congenital TTP was confirmed on the basis of severe VWF-CP deficiency in the absence of an acquired inhibitor. The patient was found to be compound heterozygous for two hitherto undescribed mutations in the ADAMTS 13 gene: a truncating frame shift mutation, 4143insA in exon 29, and the nonsense mutation 3100A >T in exon 24 (R1034X). After infusion of solvent/detergent plasma, the patient went into remission and remained asymptomatic under regular plasma therapy at 2-wk intervals for over two years. CONCLUSION: TTP in childhood may be mild and oligosymptomatic. Determination of VWF-CP activity is helpful in the differential diagnosis of thrombocytopenia.

ADAM Proteins↗

Partial amino acid sequence of purified von Willebrand factor-cleaving protease.

von Willebrand factor-cleaving protease (vWF-cp) is responsible for the continuous degradation of plasma vWF multimers released from endothelial cells. It is deficient in patients with thrombotic thrombocytopenic purpura, who show unusually large vWF multimers in plasma. Purified vWF-cp may be useful for replacement in these patients, who are now treated by plasma therapy. In this study, vWF-cp was purified from normal human plasma by affinity chromatography on the IgG fraction from a patient with autoantibodies to vWF-cp and by a series of further chromatographic procedures, including affinity chromatography on Protein G, Ig-TheraSorb, lentil lectin, and heparin. Four single-chain protein bands, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions, showed M(r) of 150, 140, 130, and 110 kd and were found to share the same N-terminal amino acid sequence, suggesting that they were derived from the same polypeptide chain that had been partially degraded at the carboxy-terminal end. A hydrophobic sequence (Ala-Ala-Gly-Gly-Ile-Leu-His-Leu-Glu-Leu-Leu-Val-Ala-Val-Gly) of the first 15 residues was established. The protease migrates in gel filtration as a high-molecular-weight complex with clusterin, a 70-kd protein with chaperonelike activity. vWF-cp bound to clusterin is dissociated by the use of concentrated chaotropic salts. vWF-cp in normal human plasma or serum is not associated with clusterin, suggesting that the observed complex is due to vWF-cp denaturation during the purification procedure. Activity of vWF-cp is unusually stable during incubation at 37 degrees C; its in vitro half-life in citrated human plasma, heparin plasma, or serum is longer than 1 week. There was even a temporary increase in protease activity during the first 3 days of incubation.

ADAM Proteins↗

Fibrinogen Milano XII: a dysfunctional variant containing 2 amino acid substitutions, Aalpha R16C and gamma G165R.

Fibrinogen Milano XII was detected in an asymptomatic Italian woman, whose routine coagulation test results revealed a prolonged thrombin time. Fibrinogen levels in functional assays were considerably lower than levels in immunologic assays. Polymerization of purified fibrinogen was strongly impaired in the presence of calcium or ethylenediaminetetraacetic acid (EDTA). Two heterozygous structural defects were detected by DNA analysis: Aalpha R16C and gamma G165R. As seen previously with other heterozygous Aalpha R16C variants, thrombin-catalyzed release of fibrinopeptide A was 50% of normal. Additionally, the release of fibrinopeptide B was delayed. Immunoblotting analysis with antibodies to human serum albumin indicated that albumin is bound to Aalpha 16 C. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of plasmin digests of fibrinogen Milano XII in the presence of calcium or EDTA showed both normal and novel D1 and D3 fragments. Further digestion of abnormal D3 fragments by chymotrypsin resulted in degradation products of the same size as the fragments derived from normal fibrinogen. SDS-PAGE analysis under reducing conditions showed no difference between normal fibrinogen and fibrinogen Milano XII or between their plasmic fragments. Circular dichroism analysis revealed a shift in the mean residual ellipticity and a significant reduction of the alpha-helix content in the variant D3 fragment. It is concluded that the Aalpha-chain substitution is mainly responsible for the coagulation abnormalities, whereas the substitution in the gamma-chain induced a conformational change in the D3 fragment.

Aged↗

Microangiopathic haemolytic anaemia in metastasizing malignant tumours is not associated with a severe deficiency of the von Willebrand factor-cleaving protease.

Complete deficiency of von Willebrand factor-cleaving protease (VWF-cp) has recently been identified as a pathogenetically important factor for thrombotic thrombocytopenic purpura (TTP). Microangiopathic haemolytic anaemia (MAHA) with thrombocytopenia in patients with metastasizing neoplasms is clinically similar to TTP, however, the pathogenesis of the condition is unclear. Partial deficiency of VWF-cp in metastasizing malignancy has recently been reported in patients without MAHA. Our study shows normal or subnormal VWF-cp activity in four patients with metastasizing neoplasia-associated MAHA but, in contrast to classical TTP, no complete deficiency of VWF-cp despite the full clinical picture of MAHA.

ADAM Proteins↗

Ten years of prophylactic treatment with fresh-frozen plasma in a child with chronic relapsing thrombotic thrombocytopenic purpura as a result of a congenital deficiency of von Willebrand factor-cleaving protease.

We report the results of 10 years of prophylactic fresh-frozen plasma (FFP) infusion therapy in a 14-year-old girl with chronic relapsing thrombotic thrombocytopenic purpura (TTP), in whom a severe congenital von Willebrand factor (VWF)-cleaving protease deficiency has been documented. Severe haemolytic crises triggered by infections were prevented and her present renal and neurological functions have been preserved. Sequential measurements of protease activity and platelet count after FFP infusion led us to conclude tentatively that 5% may be sufficient to degrade very large and adhesive VWF multimers.

ADAM Proteins↗

Aetiology and pathogenesis of thrombotic thrombocytopenic purpura and haemolytic uraemic syndrome: the role of von Willebrand factor-cleaving protease.

Thrombotic thrombocytopenic purpura (TTP) and haemolytic uraemic syndrome (HUS) are today often regarded as variants of one syndrome denoted as TTP/HUS, characterized by thrombocytopenia caused by intravascular platelet clumping, microangiopathic haemolytic anaemia, fever, renal abnormalities and neurological disturbances. Unusually large von Willebrand factor multimers have been observed in plasma from patients with chronic relapsing forms of TTP. Their appearance in patients with classic TTP is caused by deficiency of a specific von Willebrand factor-cleaving protease. A constitutional deficiency of this protease has consistently been found in familial cases of TTP, whereas in acquired TTP the protease deficiency is caused by the presence of an inhibiting autoantibody. A normal activity of von Willebrand factor-cleaving protease has been established in patients with HUS. In this chapter, we report 23 cases with severe constitutional protease deficiency: about one half of these patients had their first acute episode as children, whereas the other half had their first TTP event at an adult age, several of them during their first pregnancy. Two of these 23 individuals with congenital protease deficiency, both older than 35 years, have never had an acute TTP event. These results indicate that a deficiency of von Willebrand factor-cleaving protease alone is not sufficient to cause acute TTP. Patients with long-lasting dormant protease deficiency have been found to experience multiple relapses of TTP after having had their first acute episode. In one protease-deficient, plasma-dependent patient with chronic relapsing TTP, we estimated that 5% of normal protease activity is sufficient to remove the most adhesive von Willebrand factor multimers and prevent the formation of platelet microthrombi. The deficiency of von Willebrand factor-cleaving protease is a very strong risk factor for TTP, but the development of an acute bout requires a trigger, possibly causing the activation or apoptosis of endothelial cells in the microcirculation. It is unclear whether anti-endothelial cell antibodies, cytokines or other agents are involved in triggering thrombotic microangiopathy. The release of platelet calpain (and/or other proteases), leading to a degradation of von Willebrand factor and to platelet aggregation, has been reported in patients during their acute TTP episode. It is unknown whether calpain directly triggers an acute event or whether it merely reflects its release during the aggregation of platelets by the unusually large von Willebrand factor multimers. With regard to the heterogeneous aetiology of thrombotic microangiopathies, requiring distinct therapeutic measures, a new classification of thrombotic microangiopathy should replace the current, frequently inappropriate clinical discrimination between TTP and haemolytic uraemic syndrome.

ADAM Proteins↗

von Willebrand factor-cleaving protease in childhood diarrhoea-associated haemolytic uraemic syndrome.

A deficiency of von Willebrand factor (vWF)-cleaving protease, either due to a congenital deficiency or to the presence of a protease inhibitor of vWF-cleaving protease has been associated with thrombotic thrombocytopenic purpura (TTP). We have studied vWF-cleaving protease in diarrhoea-associated haemolytic uraemic syndrome (D+ HUS), which shares clinical features with TTP. 29 children with acute D+ HUS and 13 control children were studied. vWF-cleaving protease activity was normal (range 50-150%) in 39 of 42 plasma samples. Levels of protease activity between 25 and 50% were noted in plasma from two D+ HUS patients. One D+HUS patient, who had clinical features of TTP, had a vWF-cleaving protease inhibitor producing a severe deficiency of vWF-cleaving protease. Thus a deficiency of vWF-cleaving protease appears to be atypical in D+HUS. The detection of a vWF-cleaving protease inhibitor in one patient suggests it may be associated with infection such as E. coli O157.

ADAM Proteins↗

Similarities and differences between cobalamins and cobaloximes. Accurate structural determination of methylcobalamin and of LiCl- and KCl-containing cyanocobalamins by synchrotron radiation.

The accurate crystal structure determinations of MeCbl (1), CNCbl.2LiCl (2), and CNCbl.KCl (3), based on synchrotron diffraction data collected at 100 K and using high-quality single crystals, are reported. Refinements gave R1 indices of 0.0834 (1), 0.0434 (2), and 0.0773 (3). The influence of the water of crystallization and ion content on the crystal packing of these and other cobalamins (XCbl) is discussed, and a relationship between the crystal packing and the corrin side chain conformations is presented. An analysis of the bond lengths within the corrin moiety, based on 13 accurate structures with several X groups, shows that the trend of the C-C and C-N distances can be interpreted in terms of electronic and steric factors. The variation in structural, NMR and IR spectroscopic, and electrochemical properties are compared with those of cobaloximes, the B12 model, when X is varied. This comparison indicates that the pi-back-donation from metal to the CN axial ligand and the transmission of the trans influence of the X ligand are more effective in cobalamins than in cobaloximes. These findings are consistent with a significantly greater availability of electron charge on Co in cobalamins, and, hence, a semiquantitative evaluation of the electronic difference between the cobalt centers in the two systems is allowed.

Journal Article↗

Biochemical properties of the fibrinogen component of a fibrin glue before and after severe dry heat treatment.

Functional biochemical properties of 5 batches of the fibrinogen component of a fibrin glue produced by the ZLB Central Laboratory, Bern, each consisting of 4 different in-process samples (taken after the first and second precipitation step, lyophilization, and dry-heat treatment) were studied in vitro. We focused our attention on the effect of the anti-viral treatment of the lyophilized product by dry heat for 1 h at 100 degrees C. A slight reduction in maximal turbidity of all heat-treated samples was observed during the clotting assay compared to nontreated samples. Treatment with dry heat did not result in generation of fibrinogen fragments that might accelerate tissue-plasminogen-activator (t-PA)-enhanced plasminogen to plasmin conversion. The time course of fibrin cross-linking by factor XIII showed no differences between heated and unheated samples. This result indicates that exposure of the fibrinogen component to severe heat neither reduced activity of factor XIIIa nor affected the correct alignment of cross-linking sites in polymerized fibrin. Incubation of fibrinogen with thrombin, plasminogen, and t-PA resulted in a slightly enhanced degradation of fibrin derived from the heat-treated samples. The amount of residual moisture, determined to be within the range of 0.6-2.1% before heat treatment, did not influence clotting, cross-linking, and fibrinolysis parameters. In conclusion, the virus inactivation treatment by dry heat for 1 h at 100 degrees C induces no significant alterations of the in vitro biochemical properties of the fibrinogen component of this fibrin glue.

Cross-Linking Reagents↗

Should whole-body red cell mass be measured or calculated?

The whole-body volume of red blood cells must be known for correct diagnosis of polycythemia vera. Since the venous hematocrit may not correctly reflect the absolute amount of red blood cells, the red cell mass (RCM) is usually determined by radioisotope labeling of red blood cells according to recommendations of the International Committee for Standardization in Haematology. We examined whether the radioisotope labeling procedure can be replaced by a simple calculation of RCM from the values of venous blood hematocrit and plasma volume, using an empirical factor (Ratio f) equal to the mean ratio between whole-body and venous hematocrit. A retrospective study was performed of 264 cases in which the RCM was assayed using (51)Cr or (99m)Tc for red cell labeling, and (125)I-labeled albumin was used for estimation of plasma volume. The results showed wide scattering of Ratio f (mean value 0.911; range from 0.76 to 1.15) that was responsible for substantial differences between the measured and calculated values of RCM. We also tested whether the calculated RCM may be used as an appropriate marker for polycythemia vera according to recommendations of the International Council for Standardization in Haematology. Our data showed that 146 patients had measured RCM values more than 25% above the mean normal predicted value. Using the calculated RCM, 17 of these patients would be lost from the polycythemia vera group, whereas 29 subjects with measured RCM levels equal to or lower than 125% of predicted values would incorrectly meet the RCM criterion for polycythemia vera. Thus, a total of 46 patients would be misclassified by using the calculated RCM. We conclude that the radioisotope labeling of red blood cells remains mandatory for correct determination of the whole-body red cell volume.

Adolescent↗

Autosomal recessive inheritance of von Willebrand factor-cleaving protease deficiency.

A child with chronic relapsing thrombotic thrombocytopenic purpura (TTP/HUS) had recurrent thrombocytopenia, microangiopathic hemolytic anemia with fragmented erythrocytes, microthrombi in the lung vessels, and renal dysfunction. Assay of von Willebrand factor (vWF)-cleaving protease showed a complete protease deficiency in the patient and subnormal activities in the mother and in two asymptomatic siblings. No inhibitor of vWF-cleaving protease was detected in the patient's plasma. Periodic transfusions of fresh-frozen plasma prevented further acute episodes of TTP/HUS. Specific diagnosis of the constitutional deficiency of vWF-cleaving protease helps to provide successful prophylactic therapy.

ADAM Proteins↗

Antifungal amides from Piper hispidum and Piper tuberculatum.

Piper hispidum and Piper tuberculatum accumulate amides bearing isobutyl, pyrrolidine, dihydropyridone and piperidine moieties. The isolation and characterization of several representatives including two hitherto unreported amides were performed by chromatographic techniques and by analysis of spectroscopic data. The antifungal activity of each amide was determined by direct bioautography against Cladosporium sphaerospermum.

Amides↗

Biosynthesis of friedelane and quinonemethide triterpenoids is compartmentalized in Maytenus aquifolium and Salacia campestris.

Maytenus aquifolium (Celastraceae) and Salacia campestris (Hippocrateaceae) species accumulate friedelane and quinonemethide triterpenoids in their leaves and root bark, respectively. Enzymatic extracts obtained from leaves displayed cyclase activity with conversion of the substrate oxidosqualene to the triterpenes, 3beta-friedelanol and friedelin. In addition, administration of (+/-)5-(3)H mevalonolactone in leaves of M. aquifolium seedlings produced radio labelled friedelin in the leaves, twigs and stems, while the root bark accumulated labelled maytenin and pristimerin. These experiments indicated that the triterpenes once biosynthesized in the leaves are translocated to the root bark and further transformed to the antitumoral quinonemethide triterpenoids.

Chromatography, High Pressure Liquid↗