Protein tyrosine kinases in Schistosoma mansoni

The identification and description of signal transduction molecules and mechanisms are essential to elucidate Schistosoma mansoni host-parasite interactions and parasite biology. This mini review focuses on recent advancements in the study of signalling molecules and transduction mechanisms in S. mansoni, drawing special attention to the recently identified and characterised protein tyrosine kinases of S. mansoni.Fiocruz Centro de Pesquisas René RachouSanta Casa de Misericórdia de Belo Horizonte Programa de Pós-Graduação e PesquisaUniversidade Federal de São Paulo (UNIFESP), Escola Paulista de Medicina (EPM)UNIFESP, EPMSciEL

Mixoma cutâneo em um pintagol

Mixomas são tumores mesenquimais benignos incomuns em aves. Este trabalho objetiva descrever os achados clínico-patológicos de um caso de mixoma em um pintagol. A ave apresentou aumento de volume na região dorsal do terceiro dígito do membro pélvico esquerdo. Macroscopicamente, notou-se um nódulo de 0,9x0,5x0,4cm, macio, esbranquiçado, com áreas amareladas e enegrecidas na superfície de corte. A histopatologia revelou população monomórfica de células fusiformes, com baixo pleomorfismo, arranjadas em meio à matriz mixóide positiva para a coloração de azul alciano. Com base nos achados histopatológicos, foi firmado o diagnóstico de mixomaMyxomas are benign mesenchymal tumors rarely described in birds. This report describes the clinical and pathological findings in a case of myxoma in a pintagol (Sporagra magellanica X Serinus canaria). The animal had a nodule on the dorsal region of the third digit on the left hindlimb. Grossly, it was a 0.9×0.5×0.4cm, soft, white nodule, with black and yellow areas on the cut surface. Microscopically, a well-differentiated monomorphic population of spindle cells arranged in an abundant Alcian blue-positive myxoid matrix was observed. The diagnosis of myxoma was based on the microscopic findings

Serological screening of the Schistosoma mansoni adult worm proteome

BackgroundNew interventions tools are a priority for schistosomiasis control and elimination, as the disease is still highly prevalent. The identification of proteins associated with active infection and protective immune response may constitute the basis for the development of a successful vaccine and could also indicate new diagnostic candidates. In this context, post-genomic technologies have been progressing, resulting in a more rational discovery of new biomarkers of resistance and antigens for diagnosis.Methodology/Principal FindingsTwo-dimensional electrophoresed Schistosoma mansoni adult worm protein extracts were probed with pooled sera of infected and non-infected (naturally resistant) individuals from a S. mansoni endemic area. A total of 47 different immunoreactive proteins were identified by mass spectrometry. Although the different pooled sera shared most of the immunoreactive protein spots, nine protein spots reacted exclusively with the serum pool of infected individuals, which correspond to annexin, major egg antigen, troponin T, filamin, disulphide-isomerase ER-60 precursor, actin and reticulocalbin. One protein spot, corresponding to eukaryotic translation elongation factor, reacted exclusively with the pooled sera of non-infected individuals living in the endemic area. Western blotting of two selected recombinant proteins, major egg antigen and hemoglobinase, showed a similar recognition pattern of that of the native protein.Concluding/SignificanceUsing a serological proteome analysis, a group of antigens related to the different infection status of the endemic area residents was identified and may be related to susceptibility or resistance to infection

Use of humanised rat basophilic leukaemia cell line RS-ATL8 for the assessment of allergenicity of Schistosoma mansoni proteins.

BACKGROUND Parasite-specific IgE is thought to correlate with protection against Schistosoma mansoni infection or re-infection. Only a few molecular targets of the IgE response in S. mansoni infection have been characterised. A better insight into the basic mechanisms of anti-parasite immunity could be gained from a genome-wide characterisation of such S. mansoni allergens. This would have repercussions on our understanding of allergy and the development of safe and efficacious vaccinations against helminthic parasites. METHODOLOGY/PRINCIPAL FINDINGS A complete medium- to high-throughput amenable workflow, including important quality controls, is described, which enables the rapid translation of S. mansoni proteins using wheat germ lysate and subsequent assessment of potential allergenicity with a humanised Rat Basophilic Leukemia (RBL) reporter cell line. Cell-free translation is completed within 90 minutes, generating sufficient amounts of parasitic protein for rapid screening of allergenicity without any need for purification. Antigenic integrity is demonstrated using Western Blotting. After overnight incubation with infected individuals' serum, the RS-ATL8 reporter cell line is challenged with the complete wheat germ translation mixture and Luciferase activity measured, reporting cellular activation by the suspected allergen. The suitability of this system for characterization of novel S. mansoni allergens is demonstrated using well characterised plant and parasitic allergens such as Par j 2, SmTAL-1 and the IgE binding factor IPSE/alpha-1, expressed in wheat germ lysates and/or E. coli. SmTAL-1, but not SmTAL2 (used as a negative control), was able to activate the basophil reporter cell line. CONCLUSION/SIGNIFICANCE This method offers an accessible way for assessment of potential allergenicity of anti-helminthic vaccine candidates and is suitable for medium- to high-throughput studies using infected individual sera. It is also suitable for the study of the basis of allergenicity of helminthic proteins

Avaliação do efeito do revestimento à base de quitosana na conservação pós-colheita do Umbu / Evaluation of the effect of chitosan-based coating on the post-harvest conservation of Umbu

Para manter a qualidade e estender o tempo de prateleira das frutas, as indústrias têm buscado novas técnicas de aprimoramento, como o uso de revestimentos comestíveis. A quitosana é um biopolímero promissor nessa área, por apresentar atividade antimicrobiana, características físico-químicas e biodegradáveis pertinentes para a utilização como revestimento comestível. O umbu é um fruto da região semiárida brasileira que tem importância econômica e nutricional contribuindo para a geração de renda da região Nordeste do Brasil. Neste estudo foi realizada a avaliação do efeito do revestimento comestível à base de quitosana sobre o tempo de prateleira dos umbus. Para tal, os frutos foram imersos numa solução de quitosana 1% (p/v) em ácido acético 1% (v/v) por 2 min à 25°C. Grupos com e sem revestimento foram estocados à 8 °C por 15 dias e as análises de perda de massa, acidez total titulável, pH, sólidos solúveis totais e parâmetros de cor (L e ºH) foram realizados. Os resultados revelaram que durante o período de armazenamento houve redução da perda de massa, aumento da acidez titulável e diminuição do pH nos frutos revestidos em comparação com o grupo sem revestimento (controle). Contudo, nenhuma diferença foi encontrada nas análises colorimétricas, nem nos sólidos solúveis totais. O estudo sugere que o revestimento de quitosana pode ser um candidato promissor para aumentar a vida de prateleira dos umbus.

Vaccination with a CD4+ and CD8+ T-cell epitopes-based recombinant chimeric protein derived from Leishmania infantum proteins confers protective immunity against visceral leishmaniasis.

Vaccination seems to be the best approach to control visceral leishmaniasis (VL). Resistance against infection is based on the development of a Th1 immune response characterized by the production of interferons-? (IFN-?), interleukin-12 (IL-12), granulocyte-macrophage-colony-stimulating factor (GM-CSF), and tumor necrosis factor-? (TNF-?), among others. A number of antigens have been tested as potential targets against the disease; few of them are able to stimulate human immune cells. In the present study, 1 prediction of MHC class I and II molecules-specific epitopes in the amino acid sequences of 3 Leishmania proteins: 1 hypothetical, prohibitin, and small glutamine-rich tetratricopeptide repeat-containing proteins, was performed using bioinformatics tools, and a T-cell epitopes-based recombinant chimeric protein was constructed, synthetized and purified to be evaluated in invitro and in vivo experiments. The purified protein was tested regarding its immunogenicity in peripheral blood mononuclear cells (PBMCs) from healthy subjects and VL patients, as well as to its immunogenicity and protective efficacy in a murine model against Leishmania infantum infection. Results showed a Th1 response based on high IFN-? and low IL-10 levels derived from in chimera-stimulated PBMCs in both healthy subjects and VL patients. In addition, chimera and/or saponin-immunized mice presented significantly lower parasite burden in distinct evaluated organs, when compared to the controls, besides higher levels of IFN-?, IL-2, IL-12, and GM-CSF, and an IgG2a isotype-based humoral response. In addition, the CD4+ and CD8+ T-cell subtypes contributed to IFN-? production in the protected animals. The results showed the immunogenicity in human cells and the protective efficacy against L. infantum in a murine model, and well indicate that this recombinant chimera can be considered as a promising strategy to be used against human disease

In vivo antileishmanial efficacy of a naphthoquinone derivate incorporated into a Pluronic? F127-based polymeric micelle system against Leishmania amazonensis infection.

New therapeutic strategies against leishmaniasis are desirable, since the treatment against disease presents problems, such as the toxicity, high cost and/or parasite resistance. As consequence, new antileishmanial compounds are necessary to be identified, as presenting high activity against Leishmania parasites, but low toxicity in mammalian hosts. Flau-A is a naphthoquinone derivative recently showed to presents an in vitro effective action against Leishmania amazonensis and L. infantum species. In the present work, the in vivo efficacy of Flau-A, which was incorporated into a Poloxamer 407-based micelle system, was evaluated in a murine model against L. amazonensis infection. Amphotericin B (AmB) and Ambisome? were used as controls. The animals were infected and later treated with the compounds. Thirty days after the treatment, parasitological and immunological parameters were evaluated. Results showed that AmB, Ambisome? , Flau-A or Flau-A/M-treated animals presented significantly lower average lesion diameter and parasite burden in tissue and organs evaluated, when compared to the control (saline and micelle) groups. Flau-A or Flau-A/M-treated mice were those presenting the most significant reductions in the parasite burden, when compared to the others. These animals developed also a more polarized antileishmanial Th1 immune response, which was based on significantly higher levels of IFN-?, IL-12, TNF-?, GM-CSF, and parasite-specific IgG2a isotype; associated with low levels of IL-4, IL10, and IgG1 antibody. The absence of toxicity was found in these animals, although mice receiving AmB have showed high levels of renal and hepatic damage markers. In conclusion, results suggested that the Flau-A/M compound may be considered as a possible therapeutic target to be evaluated against human leishmaniasis

Caracterização molecular de uma nova proteína tirosina quinase, SmFes, de Schistosoma mansoni.

Submitted by Nuzia Santos ([email protected]) on 2017-08-25T16:57:25Z No. of bitstreams: 1 Fernanda Ludolf Ribeiro.pdf: 37073099 bytes, checksum: 866dc18aba4b6a7fa2899db801df0c20 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2017-08-25T17:02:54Z (GMT) No. of bitstreams: 1 Fernanda Ludolf Ribeiro.pdf: 37073099 bytes, checksum: 866dc18aba4b6a7fa2899db801df0c20 (MD5)Made available in DSpace on 2017-08-25T17:02:54Z (GMT). No. of bitstreams: 1 Fernanda Ludolf Ribeiro.pdf: 37073099 bytes, checksum: 866dc18aba4b6a7fa2899db801df0c20 (MD5) Previous issue date: 2005Fundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brasil.A identif icação de moléculas sinalizadoras e a elucidação de processo s de transdução de sinal de Schistosoma mansoni são necessárias para o entendi mento da interação hospedeiro- parasito e de proces sos fisiológicos do parasito. Proteínas tirosinas quinase (PTK s) são molécu las importan tes na comunica ção intra e intercelu lar, tendo um papel crucial nos mecan ismos de transdução de sinal. Estas proteínas estão envolvidas nos processo s de desenvolvim ento, diferenciação e comuni cação celular entre outros. Uma nova PTKs foi identificad a em S. mansoni e nomeada, SmFes. SmFes é um gene de cópia única e o seu cDNA comple to contém uma ORF de 3.780 pb. SmFes exibe característi cas da família Fes/Fps/Fer das PTKs, com assinaturas de domín io proteína quinase, SH2 e coiled- coil característ icos. SmFes é o prime iro gene da família Fes/Fp s/Fer identificado em S. mansoni. Análise filogenétic a revelou que SmFes está relacionada a proteínas da família Fes/Fps/Fer, agrupadas mais próximas às proteínas ortólogas de organismos invertebrados. O alinham ento entre o cDNA de SmFes e seqüências genôm icas depositadas pelos bancos de dados de S. mansoni indicara m a presença de 17 introns, alguns demonstrados experi mentalmente. Análises parciais de clones de cDNA indicara m a inserção de 9 pb na posição 3’ do exon 10, formando duas populações de cDNA, comprovadas por RFLP. A análise da seqüência genômic a indicou que existem dois possíveis sítios de edição do RNA na proximidad e 5’ do intron, indicando um evento de edição alternat iva. A existência de um alelo contendo uma inserção de 15 pb na seqüência genômi ca foi observada. As amplif icaçõ es de diferentes espécies de Schistosoma com pares de iniciador es desenhados para SmFes indicara m a presença de ortólogos de SmFes em várias espécies do gênero. SmFes é transcrita em baixos níveis nas diversas fases de desenvolvi mento do parasito, com uma maior taxa de transcrição em verm es machos, como detectado por PCR em tempo real e northern- blot. A expressão da proteína SmFes foi verificada em esporocisto, mirac ídio, verme adulto e em nível mais elevado em cercári a. Uma proteína recombinan te de SmFes foi expressa para ser usada em experim entos de identif icação de parceiros de SmFes em futuros ensaios de co-precipitaç ão e també m em experim entos de duplo híbrido. Vários possíveis parceiros de SmFes foram identificados por análise compu tacion al. A compre ensão de genes envolvidos nos mecan ismos de transdução de sinal pode fornecer novas estratégias de desenvolvi mento de drogas.The identifi cation and elucida tion of signal transduction molecules and mechan isms are necessary for the understanding of the Schistosoma mansoni host-parasite interac tion and of physiologica l process of the parasite. Protein Tyrosine kinases (PTKs) are impor tant molecu les for intra and interce llular comm unication, playing a crucial role in signal transduction processes. These proteins are known to be involved in developm ental, differentiat ion and communication processes of cells, among others. A novel member of PTK was identified in S. mansoni and designated SmFes. SmFes is a single copy gene and the comple te cDNA contains a 3.780 bp ORF. SmFes exhibits the characteristi c features of Fes/Fps/Fer protein tyrosine kinases family, containing three coiled- coil regions, an SH2 and a protein tyrosine kinase catal ytic doma in signatures. SmFes is the first gene from Fes/Fps/Fer family identified in S. mansoni. Phylogene tic analyses reveled that SmFes is more closel y related to invertebrate proteins orthologues of the Fes/Fps/Fer family. The assembly of the SmFes cDNA and genomi c sequences indicated the presence of 17 introns, some experim entally demonstra ted. Analy sis of partial cDNA clones indicated the presence of a 9 pb insertion at the 3’ end of exon 10, producing two different populations of cDNA. The analysis of the genomi c sequence indicated that there are two possible splice sites at the 5’end of the intron, pointing to an alternative splicing event in the SmFes pre-mRNA. An allele of the SmFes containing a 15 pb insertion was observed in genomi c sequence. The amplification of different Schistosoma species with SmFes specific primers indicat ed that orthologues of SmFes are present in several species of the genus. SmFes is transcribed at low levels in the different develop mental stages of the parasite, with higher level in adult male worms as detected by real time PCR and northern-blot . The expression of SmFes protein was verified in sporoc yst, mirac idium, adult worm and in higher levels in cercariae. A recombin ant protein was expressed to be used in experi ments to identif y SmFes partner proteins in future pull- down assay s and also in two-h ybrid systems. Various possible partners or SmFes were identified by computa tional analy sis. The comprehension of signal transduction processes could also contribute to the identific ation of po ssible new targets for drug s