Análise da expressão do gene TcNTPDase-1 em diferentes subpopulações e formas evolutivas de Trypanosoma cruzi

Made available in DSpace on 2016-04-04T12:35:26Z (GMT). No. of bitstreams: 2 natalia_gomes_ioc_mest_2014.pdf: 3321973 bytes, checksum: 59b3c4b51a637c1e44229ae143291433 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2014Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, BrasilA doença de Chagas é uma doença negligenciada causada pelo protozoário parasita Trypanosoma cruzi, o qual afeta cerca de 6-8 milhões de pessoas em áreas endêmicas da América Latina. As diferentes cepas envolvidas na infecção, e sua distribuição regional, tem sido sugeridas como causa das diferentes manifestações clínicas, resposta ao diagnóstico e eficácia terapêutica. A ecto-NTPDase é uma enzima localizada na superfície externa da membrana plasmática do T.cruzi, que hidrolisa nucleotídeos tri e difosfatados. Estudos anteriores têm relacionado esta enzima à infectividade e virulência de tripanosomatídeos, além de sugerir um papel importante na captação de purinas pelos parasitos. Neste trabalho avaliamos os níveis de mRNA para a TcNTPDase-1 por RT-PCR, em diferentes isolados do parasito (Dm28c- T.cruzi I; Y- T.cruzi II; 3663- T.cruzi III; 4167- T.cruzi IV; LL014- T.cruzi V; CL-14- T.cruzi VI- avirulenta), em formas não-infectantes (epimastigotas) e infectantes (tripomastigotas e amastigotas). Os ensaios de PCR em Tempo Real foram realizados com oligonucleotídeos desenhados para amplificar uma sequência de 111pb do gene TcNTPDase-1 e, como controles endógenos, uma sequência de 268pb do gene da TcCalmodulina e uma sequência de 100pb do gene TcGAPDH foram utilizados. Nós observamos que epimastigotas das cepas Y, 3663, 4167 e LL014 apresentaram os mesmos níveis de expressão do gene TcNTPDase-1 que o clone CL-14, avirulento Por outro lado, o clone Dm28c expressou 7,2\F0B11,5 vezes mais o gene desta enzima que o clone CL-14. Além disso, observamos que as formas infectantes tripomastigota e amastigota apresentaram, respectivamente, uma expressão 22,5\F0B15,6 e 16,3\F0B13,8 vezes maior que a forma epimastigota. Observamos também que durante a curva de crescimento de formas epimastigotas a 28°C e 37°C a expressão deste gene aumenta gradativamente com o tempo de cultivo, sendo mais pronunciado a 37°C, sugerindo que o choque-térmico e o longo período de cultivo pode aumentar a expressão do gene da TcNTPDase-1. Em conjunto, nossos dados sugerem que a TcNTPDase-1 estaria envolvida nos processos de infectividade do T.cruzi, bem como adaptação do parasita a temperatura do hospedeiro vertebrado. Devido sua importância para o T.cruzi, a TcNTPDase-1 apresenta potencial para ser avaliada como possível alvo para quimioterapia da Doença de ChagasChagas disease is a negl ected illness caused by the parasite protozoan Trypanosoma cruzi , which affects 6 - 8 million people in endemic areas of Latin America . The distinct strains involved in infection, in conjunction with the regional distribution, have been suggested to cause di fferent clinical manifestations and therapeutic efficiency in Chagas Disease. The ecto - NTPDase is an apyrase located on the outer surface of T. cruzi plasma membrane - that hydrolyzes tri - and/or di - phosphate nucleotides . Previous studies related this enzyme to the infectivity and virulence of the parasite , and suggest an important role in the uptake of purines . In this study, we evaluate the mRNA levels for the e cto - NTPDase I by RT - PCR in distinct isolated parasites (Dm28c - T. cruzi I; Y - T. cruzi II; 3663 - T. cruzi III; 4167 - T. cruzi IV; LL014 - T.cruzi V ; CL - 14 - T. cruzi VI - avirulent ) , in non - infective forms (epimastigotes) and infective forms (amastigotes and trypomastigotes) . The Real Time PCR assays were performed with primers designed to amplify a 111b p seq uence in the TcNTPDase - 1 gene and, as endogenous controls, a 268bp sequence in the TcCalmoduline gene and a 100bp sequence in the Tc GAPDH gene were used. We observed that the epimastigotes of strain s Y, 3663, 4167 e LL014 expressed the same levels of the g ene of TcNTPDase - 1 that the CL - 14 clone, avirulent. On the other hand, the Dm28c clone expressed 7 . 2 1 . 5 times higher the gene of this enzyme that the C L - 14 clone. Surprisingly, w e observed that the trypomastigote and amastigote presented expression levels , respectively, 22 . 5 5 . 6 and 16 . 3 3 . 8 time s higher than the epimastigote form. Moreover, following the epimastigotes growth curve at 28°C e 37°C , we also observed that the expression of TcNTPDase - 1 increases with the days of growth, and this increase is mo re pronounced at 37°C , suggesting that heat shock and long - term cultivation could increase TcNTPDase - 1 gene expression . In conjunction, our results suggest the role of TcNTPDase - 1 on T. cruzi infectivity and adjustment to stress conditions s uch as nutrients starvation and heat shock. Due to its importance for T. cruzi , TcNTPD ase - 1 has the potential to be e valuated as a possible target for chemotherapy of Chagas Disease

Overexpression of TcNTPDase-1 Gene Increases Infectivity in Mice Infected with Trypanosoma cruzi

Ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) are enzymes located on the surface of the T. cruzi plasma membrane, which hydrolyze a wide range of tri-/-diphosphate nucleosides. In this work, we used previously developed genetically modified strains of Trypanosoma cruzi (T. cruzi), hemi-knockout (KO +/−) and overexpressing (OE) the TcNTPDase-1 gene to evaluate the parasite infectivity profile in a mouse model of acute infection (n = 6 mice per group). Our results showed significantly higher parasitemia and mortality, and lower weight in animals infected with parasites OE TcNTPDase-1, as compared to the infection with the wild type (WT) parasites. On the other hand, animals infected with (KO +/−) parasites showed no mortality during the 30-day trial and mouse weight was more similar to the non-infected (NI) animals. In addition, they had low parasitemia (45.7 times lower) when compared with parasites overexpressing TcNTPDase-1 from the hemi-knockout (OE KO +/−) group. The hearts of animals infected with the OE KO +/− and OE parasites showed significantly larger regions of cardiac inflammation than those infected with the WT parasites (p < 0.001). Only animals infected with KO +/− did not show individual electrocardiographic changes during the period of experimentation. Together, our results expand the knowledge on the role of NTPDases in T. cruzi infectivity, reenforcing the potential of this enzyme as a chemotherapy target to treat Chagas disease (CD)

FIRST RECORD ABOUT EFFICIENCY AND HUMIDITY OF COBIA SUBMMITED TO SALTING PROCESS

With the cultivation of cobia Rachycentron canadum via off-shore system in Brazil, the development of salting methods that enable elaboration of top quality products, may be a manner to aggregate value to the fish. This research evaluated the efficiency and humidity for the R. canadum, when it was submitted to different procedures and salinity concentrations during the mixed salting process at two forms of presentation: split in half and sliced-refrigerated. The major efficiency was 49.79% for the sliced fishes with 10% of salinity, while on the treatment of the split in half fish at 40% of salt, the efficiency was 45.95%. The split in half fish, treated with 20% of salinity, suffered deteriorative process. The fishes presented initial humidity of 70.53%, being reduced to 34.74% on the sliced form with 40% of salt, and 66.41% and 64.67% on the split in half form at 10% and 15% of salt, respectively. This research verified that the mixed salting process of cobia is efficient for the sliced products, such as for those ones split in half on a traditional manner, with 40% of salinity, being an alternative to entrepreneurs and communities

TGF-β inhibitor therapy decreases fibrosis and stimulates cardiac improvement in a pre-clinical study of chronic Chagas’ heart disease

International audienceTGF-β involvement in Chagas disease cardiomyopathy has been clearly demonstrated. The TGF-β signaling pathway is activated in the cardiac tissue of chronic phase patients and is associated with an increase in extracellular matrix protein expression. The aim of this study was to investigate the effect of GW788388, a selective inhibitor of TβR1/ALK5, on cardiac function in an experimental model of chronic Chagas’ heart disease. To this end, C57BL/6 mice were infected with Trypanosoma cruzi (102 parasites from the Colombian strain) and treated orally with 3mg/kg GW788388 starting at 120 days post-infection (dpi), when 100% of the infected mice show cardiac damage, and following three distinct treatment schedules: i) single dose; ii) one dose per week; or iii) three doses per week during 30 days. The treatment with GW788388 improved several cardiac parameters: reduced the prolonged PR and QTc intervals, increased heart rate, and reversed sinus arrhythmia, and atrial and atrioventricular conduction disorders. At 180 dpi, 30 days after treatment interruption, the GW3x-treated group remained in a better cardiac functional condition. Further, GW788388 treatment reversed the loss of connexin-43 enriched intercellular plaques and reduced fibrosis of the cardiac tissue. Inhibition of the TGF-β signaling pathway reduced TGF-β/pSmad2/3, increased MMP-9 and Sca-1, reduced TIMP-1/TIMP-2/TIMP-4, and partially restored GATA-6 and Tbox-5 transcription, supporting cardiac recovery. Moreover, GW788388 administration did not modify cardiac parasite load during the infection but reduced the migration of CD3+ cells to the heart tissue. Altogether, our data suggested that the single dose schedule was not as effective as the others and treatment three times per week during 30 days seems to be the most effective strategy. The therapeutic effects of GW788388 are promising and suggest a new possibility to treat cardiac fibrosis in the chronic phase of Chagas’ heart disease by TGF-β inhibitors

Divergent cerebrospinal fluid cytokine network induced by non-viral and different viral infections on the central nervous system.

Submitted by Nuzia Santos ([email protected]) on 2016-04-06T12:50:09Z No. of bitstreams: 1 Divergent cerebrospinal fluid cytokine .pdf: 6666541 bytes, checksum: b39732a64c755314d74c20e896a13597 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2016-04-06T13:24:06Z (GMT) No. of bitstreams: 1 Divergent cerebrospinal fluid cytokine .pdf: 6666541 bytes, checksum: b39732a64c755314d74c20e896a13597 (MD5)Made available in DSpace on 2016-04-06T13:24:06Z (GMT). No. of bitstreams: 1 Divergent cerebrospinal fluid cytokine .pdf: 6666541 bytes, checksum: b39732a64c755314d74c20e896a13597 (MD5) Previous issue date: 2015Fundação de Medicina Tropical Dr. Heitor Vieira Dourado. Manaus, AM, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratorio de Biomarcadores para Diagnostico e Monitoramento. Belo Horizonte, MG, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratorio de Biomarcadores para Diagnostico e Monitoraramento. Belo Horizonte, MG, Brasil/Instituto Hermes Pardini. Belo Horizonte, MG, BrasilFundação Oswaldo Cruz. Instituto Leônidas e Maria Deane. Manaus, AM, Brasil.Fundação de MedicinaTropical Dr. Heitor Vieira Dourado. Manaus, AM, Brasil.Fundação de Hematologia e Hemoterapia do Amazonas. Manaus, AM, Brasil.Fundação de Medicina Tropical Dr. Heitor Vieira Dourado. Manaus, AM, Brasil.Fundação de Medicina Tropical Dr. Heitor Vieira Dourado. Manaus, AM, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratorio de Biomarcadores para Diagnostico e Monitoramento. Belo Horizonte, MG, Brasil.Universidade Federal de Minas Gerais. Faculdade de Medicina. Departamento de Patologia Anatomica. Belo Horizonte, MG, Brasil.Fundação de Hematologia e Hemoterapia do Amazonas. Manaus, AM, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratorio de Biomarcadores para Diagnostico e Monitoramento. Belo Horizonte, MG, Brasil.Fundação de Hematologia e Hemoterapia do Amazonas. Manaus, AM, Brasil.Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratorio de Biomarcadores para Diagnostico e Monitoramento. Belo Horizonte, MG, Brasil.Fundação de Medicina Tropical Dr. Heitor Vieira Dourado. Manaus, AM, Brasil/Universidade Estadual do Amazonas. Manaus, AM, Brasil.BACKGROUND: Meningoencephalitis is one of the most common disorders of the central nervous system (CNS) worldwide. Viral meningoencephalitis differs from bacterial meningitis in several aspects. In some developing countries, bacterial meningitis has appropriate clinical management and chemotherapy is available. Virus-associated and virus not detected meningoencephalitis are treatable, however, they may cause death in a few cases. The knowledge of how mediators of inflammation can induce disease would contribute for the design of affordable therapeutic strategies, as well as to the diagnosis of virus not detected and viral meningoencephalitis. Cytokine-induced inflammation to CNS requires several factors that are not fully understood yet. METHODS: Considering this, several cytokines were measured in the cerebrospinal fluid (CSF) of patients with undiagnosed and viral meningoencephalitis, and these were correlated with cellularity in the CSF. RESULTS: The results demonstrate that an altered biochemical profile alongside increased cellularity in the cerebrospinal fluid is a feature of patients with meningoencephalitis that are not associated with the detection of virus in the CNS (P < 0.05). Moreover, HIV-positive patients (n = 10) that evolve with meningoencephalitis display a distinct biochemical/cytological profile (P < 0.05) in the cerebrospinal fluid. Meningoencephalitis brings about a prominent intrathecal cytokine storm regardless of the detection of virus as presumable etiological agent. In the case of Enterovirus infection (n = 13), meningoencephalitis elicits robust intrathecal pro-inflammatory cytokine pattern and elevated cellularity when compared to herpesvirus (n = 15) and Arbovirus (n = 5) viral infections (P < 0.05). CONCLUSION: Differences in the cytokine profile of the CSF may be unique if distinct, viral or presumably non-viral pathways initially trigger the inflammatory response in the CNS

Effects of Cholinergic Stimulation with Pyridostigmine Bromide on Chronic Chagasic Cardiomyopathic Mice

Made available in DSpace on 2015-05-04T16:34:37Z (GMT). No. of bitstreams: 2 license.txt: 1914 bytes, checksum: 7d48279ffeed55da8dfe2f8e81f3b81f (MD5) constança_brittoetal_IOC_2014.pdf: 3857100 bytes, checksum: 42bae205f256c445117f5bd53964bde3 (MD5) Previous issue date: 2014Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, BrasilUniversidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, BrasilFundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, BrasilFundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, BrasilUniversidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.University of Milan. L Sacco Hospital. Department of Clinical Sciences, Internal Medicine II. Milan, Italy.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.Universidade Federal do Triângulo Mineiro. Instituto de Ciências Biológicas e Naturais. Uberaba, MG, Brasil.The aim of the present study was to assess the effects of an anticholinesterase agent, pyridostigmine bromide (Pyrido), on experimental chronic Chagas heart disease in mice. To this end, male C57BL/6J mice noninfected (control:Con) or chronically infected (5 months) with Trypanosoma cruzi (chagasic:Chg) were treated or not (NT) with Pyrido for one month. At the end of this period, electrocardiogram (ECG); cardiac autonomic function; heart histopathology; serum cytokines; and the presence of blood and tissue parasites by means of immunohistochemistry and PCR were assessed. In NT-Chg mice, significant changes in the electrocardiographic, autonomic, and cardiac histopathological profiles were observed confirming a chronic inflammatory response. Treatment with Pyrido in Chagasic mice caused a significant reduction of myocardial inflammatory infiltration, fibrosis, and hypertrophy, which was accompanied by a decrease in serum levels of IFN with no change in IL-10 levels, suggesting a shift of immune response toward an anti-inflammatory profile. Lower nondifferent numbers of parasite DNA copies were observed in both treated and nontreated chagasic mice. In conclusion, our findings confirmthe marked neuroimmunomodulatory role played by the parasympathetic autonomic nervous systemin the evolution of the inflammatory-immune response to T. cruzi during experimental chronic Chagas heart disease in mice

Perfil epidemiológico de hanseníase no sertão Pernambucano, Brasil / Epidemiological profile of hanseníase in sertão Pernambucano, Brazil

A Hanseníase é uma doença infecciosa crônica causada por Mycobacterium leprae, considerada um grave problema saúde pública mundial. O presente trabalho tem como objetivo traçar um perfil epidemiológico da Hanseníase no sertão pernambucano, tomando por base os dados eletrônicos do SINAN e DATASUS (ambos ligados ao Ministério da Saúde). A partir dessas fontes, realizou-se um estudo de série histórica observacional do tipo transversal dos casos notificados de Hanseníase entre os anos de 2006 a 2017. De acordo com os dados, observa-se que esse agravo apresentou um decréscimo no número de novos casos, na medida em que se constatava 43.642 casos no ano de 2006 contra 26.875 novos casos em 2017. Essa realidade, embora evidencie uma notável redução no número de novos casos de hanseníase, evoca a responsabilidade dos órgãos públicos de saúde no que tange à manutenção do controle desse agravo, assim como a criação de novas medidas que busquem a prevenção e a erradicação desse enfermo no território nacional.