Effects of isoflavones on the coagulation and fibrinolytic system of postmenopausal women

Objective: We evaluated the effects of soy isoflavone supplementation on hemostasis in healthy postmenopausal women. Methods: In this double-blinded, placebo-controlled study, 47 postmenopausal women 47-66 y of age received 40 mg of soy isoflavone (n = 25) or 40 mg of casein placebo (n = 22) once a day for 6 mo. Levels of factors VII and X. fibrinogen, thrombin-antithrombin complex, prothrombin fragments I plus 2, antithrombin, protein C, total and free protein S, plasminogen, plasminogen activator inhibitor-1, and D-dimers were measured at baseline and 6 mo. Urinary isoflavone concentrations (genistein and daidzein) were measured as a marker of compliance and absorption using high-performance liquid chromatography. Baseline characteristics were compared by unpaired Student`s t test. Within-group changes and comparison between the isoflavone and casein placebo groups were determined by a mixed effects model. Results: The levels of hemostatic variables did not change significantly throughout the study in the isoflavone group; however, the isoflavone group showed a statistically significant reduction in plasma concentration of prothrombin fragments I plus 2; both groups showed a statistically significant reduction in antithrombin, protein C, and free protein S levels. A significant increase in D-dimers was observed only in the isoflavone group. Plasminogen activator inhibitor-l levels increased significantly in the placebo group. However, these changes were not statistically different between groups. Conclusion: The results of the present study do not support a biologically significant estrogenic effect of soy isoflavone on coagulation and fibrinolysis in postmenopausal women. However, further research will be necessary to definitively assess the safety and efficacy of isoflavone. (D 2008 Elsevier Inc. All rights reserved.

Identification and characterization of antigenic proteins potentially expressed during the infectious process of Paracoccidioides brasiliensis

Paracoccidioides brasiliensis causes paracoccidioidomycosis (PCM), a systemic mycosis presenting clinical manifestations ranging from mild to severe forms. A P. brasiliensis cDNA expression library was produced and screened with pooled sera from PCM patients adsorbed against antigens derived from in vitro-grown P. brasiliensis yeast cells. Sequencing DNA inserts from clones reactive with PCM patients sera indicated 35 open reading frames presenting homology to genes involved in metabolic pathways, transport, among other predicted functions. The complete cDNAs encoding aromatic-L-amino-acid decarboxylase (Pbddc), lumazine synthase (Pbls) and a homologue of the high affinity copper transporter (Pbctr3) were obtained. Recombinant proteins PbDDC and PbLS were obtained; a peptide was synthesized for PbCTR3. The proteins and the synthetic peptide were recognized by sera of patients with confirmed PCM and not by sera of healthy patients. Using the in vivo-induced antigen technology (IVIAT), we identified immunogenic proteins expressed at high levels during infection. Quantitative real time RTPCR demonstrated high transcript levels of Pbddc, Pbls and Pbctr3 in yeast cells infecting macrophages. Transcripts in yeast cells derived from spleen and liver of infected mice were also measured by qRT-PCR. Our results suggest a putative role for the immunogenic proteins in the infectious process of P. brasiliensis. (C) 2009 Elsevier Masson SAS. All rights reserved.

Identification and lineage genotyping of South American trypanosomes using fluorescent fragment length barcoding

Trypanosoma cruzi and Trypanosoma rangeli are human-infective blood parasites, largely restricted to Central and South America. They also infect a wide range of wild and domestic mammals and are transmitted by a numerous species of triatomine bugs. There are significant overlaps in the host and geographical ranges of both species. The two species consist of a number of distinct phylogenetic lineages. A range of PCR-based techniques have been developed to differentiate between these species and to assign their isolates into lineages. However, the existence of at least six and five lineages within T. cruzi and T. rangeli, respectively, makes identification of the full range of isolates difficult and time consuming. Here we have applied fluorescent fragment length barcoding (FFLB) to the problem of identifying and genotyping T. cruzi, T. rangeli and other South American trypanosomes. This technique discriminates species on the basis of length polymorphism of regions of the rDNA locus. FFLB was able to differentiate many trypanosome species known from South American mammals: T. cruzi cruzi. T. cruzi marinkellei, T. dionisii-like, T. evansi, T. lewisi, T. rangeli, T. theileri and T. vivax. Furthermore, all five T. rangeli lineages and many T. cruzi lineages could be identified, except the hybrid lineages TcV and TcVI that could not be distinguished from lineages III and II respectively. This method also allowed identification of mixed infections of T. cruzi and T. rangeli lineages in naturally infected triatomine bugs. The ability of FFLB to genotype multiple lineages of T. cruzi and T. rangeli together with other trypanosome species, using the same primer sets is an advantage over other currently available techniques. Overall, these results demonstrate that FFLB is a useful method for species diagnosis, genotyping and understanding the epidemiology of American trypanosomes. (C) 2010 Elsevier B.V. All rights reserved.

A real-time polymerase chain reaction assay for the identification and quantification of American Leishmania species on the basis of glucose-6-phosphate dehydrogenase

A real-time polymerase chain reaction (PCR) test was developed on the basis of the Leishmania glucose-6-phosphate dehydrogenase locus that enables identification and quantification of parasites. Using two independent pairs of primers in SYBR-Green assays, the test identified etiologic agents of cutaneous leishmaniasis belonging to both subgenera, Leishmania (Viannia) and Leishmania (Leishmania) in the Americas. Furthermore, use of TaqMan probes enables distinction between L. (V.) braziliensis or L. (V.) peruviania from the other L. (Viannia) species. All assays were negative with DNA of related trypanosomatids, humans, and mice. The parasite burden was estimated by normalizing the number of organisms per total amount of DNA in the sample or per host glyceraldehyde-3-phosphate dehydrogenase copies. The real-time PCR assay for L. (Leishmania) subgenus showed a good linear correlation with quantification on the basis of a limiting dilution assay in experimentally infected mice. The test successfully identifies and quantifies Leishmania in human biopsy specimens and represents a new tool to study leishmaniasis.

Upgrading a TCABR data analysis and acquisition system for remote participation using Java, XML, RCP and modern client/server communication/authentication

The TCABR data analysis and acquisition system has been upgraded to support a joint research programme using remote participation technologies. The architecture of the new system uses Java language as programming environment. Since application parameters and hardware in a joint experiment are complex with a large variability of components, requirements and specification solutions need to be flexible and modular, independent from operating system and computer architecture. To describe and organize the information on all the components and the connections among them, systems are developed using the extensible Markup Language (XML) technology. The communication between clients and servers uses remote procedure call (RPC) based on the XML (RPC-XML technology). The integration among Java language, XML and RPC-XML technologies allows to develop easily a standard data and communication access layer between users and laboratories using common software libraries and Web application. The libraries allow data retrieval using the same methods for all user laboratories in the joint collaboration, and the Web application allows a simple graphical user interface (GUI) access. The TCABR tokamak team in collaboration with the IPFN (Instituto de Plasmas e Fusao Nuclear, Instituto Superior Tecnico, Universidade Tecnica de Lisboa) is implementing this remote participation technologies. The first version was tested at the Joint Experiment on TCABR (TCABRJE), a Host Laboratory Experiment, organized in cooperation with the IAEA (International Atomic Energy Agency) in the framework of the IAEA Coordinated Research Project (CRP) on ""Joint Research Using Small Tokamaks"". (C) 2010 Elsevier B.V. All rights reserved.