RESUMO
Parasitic helminths can reside in humans owing to their ability to disrupt host protective immunity. Receptor for advanced glycation end products (RAGE), which is highly expressed in host skin, mediates inflammatory responses by regulating the expression of pro-inflammatory cytokines and endothelial adhesion molecules. In this study, we evaluated the effects of venestatin, an EF-hand Ca2+-binding protein secreted by the parasitic helminth Strongyloides venezuelensis, on RAGE activity and immune responses. Our results demonstrated that venestatin bound to RAGE and downregulated the host immune response. Recombinant venestatin predominantly bound to the RAGE C1 domain in a Ca2+-dependent manner. Recombinant venestatin effectively alleviated RAGE-mediated inflammation, including footpad edema in mice, and pneumonia induced by an exogenous RAGE ligand. Infection experiments using S. venezuelensis larvae and venestatin silencing via RNA interference revealed that endogenous venestatin promoted larval migration from the skin to the lungs in a RAGE-dependent manner. Moreover, endogenous venestatin suppressed macrophage and neutrophil accumulation around larvae. Although the invasion of larvae upregulated the abundance of RAGE ligands in host skin tissues, mRNA expression levels of tumor necrosis factor-α, cyclooxygenase-2, endothelial adhesion molecules vascular cell adhesion protein-1, intracellular adhesion molecule-1, and E-selectin were suppressed by endogenous venestatin. Taken together, our results indicate that venestatin suppressed RAGE-mediated immune responses in host skin induced by helminthic infection, thereby promoting larval migration. The anti-inflammatory mechanism of venestatin may be targeted for the development of anthelminthics and immunosuppressive agents for the treatment of RAGE-mediated inflammatory diseases.
Assuntos
Proteínas de Helminto/metabolismo , Interações Hospedeiro-Parasita/fisiologia , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Strongyloides/imunologia , Strongyloides/metabolismo , Estrongiloidíase/metabolismo , Animais , Larva/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Strongyloides/patogenicidadeRESUMO
Strongyloides venezuelensis is an important alternative source of antigen for the serologic diagnosis of human strongyloidiasis. Proteomics techniques applied to the analysis of the protein content of infective third stage larvae (iL3) of S. venezuelensis provide a powerful tool for the discovery of new candidates for immunodiagnosis. This study presents an overview of the protein iL3 S. venezuelensis focusing on the diagnosis of strongyloidiasis. A total of 877 proteins were identified by shotgun proteomics. Many of these proteins are involved in different cellular processes, metabolic as well as structural maintenance. Our results point to a catalog of possible diagnostic targets for human strongyloidiasis and highlight the need for evaluation of uncharacterized proteins, especially the proteins within the CAP domain, transthyretin, and BTPI inhibitor domains, as a repertoire as yet unexplored in the context of strongyloidiasis diagnostic markers. We believe that the protein profile presented in this shotgun analysis extends our understanding of the protein composition within the Strongyloides genus, opening up new perspectives for research on biomarkers that may help with the diagnosis of human strongyloidiasis. Data are available via ProteomeXchange with identifier PXD013703.
Assuntos
Biomarcadores/metabolismo , Larva/metabolismo , Proteoma , Strongyloides/metabolismo , Estrongiloidíase/diagnóstico , Animais , Catepsinas/metabolismo , Galectinas/metabolismo , Interações Hospedeiro-Parasita , Humanos , Testes Imunológicos , Metaloproteases/metabolismo , Patologia Molecular , ProteômicaRESUMO
Infection with Strongyloides sp. induces a host immune response, predominantly the Th2 type, that is able to eliminate the parasite. However, little is known about the role of the nitric oxide (NO) mediator, induced by the enzyme nitric oxide synthase (NOS), in strongyloidiasis. Therefore, in this study, we investigated the immune response of mice genetically deficient in the enzyme inducible nitric oxide synthase (iNOS-/- ), infected with Strongyloides venezuelensis. C57BL/6 wild-type (WT) and iNOS-/- mice were individually inoculated by subcutaneous injection of 3000 S. venezuelensis L3 larvae. In the absence of iNOS, mice were more susceptible to the infection than WT animals, in which the parasite was completely eliminated. The overall production of cytokines and specific IgG, IgG1 or IgE antibodies against the parasite was significantly lowered in infected iNOS-/- mice. The expression of iNOS was observed in the intestine of WT hosts but mainly in the wall of the parasite, despite the presence of iNOS in mice. Altogether, we concluded that iNOS expression may play an important role in the control of S. venezuelensis infection.
Assuntos
Anticorpos Antiprotozoários/imunologia , Mucosa Intestinal/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico/metabolismo , Strongyloides/metabolismo , Estrongiloidíase/imunologia , Animais , Anticorpos Antiprotozoários/biossíntese , Arvicolinae/parasitologia , Citocinas/biossíntese , Citocinas/imunologia , Imunoglobulina E/imunologia , Imunoglobulina G/imunologia , Mucosa Intestinal/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Strongyloides/citologia , Strongyloides/isolamento & purificação , Estrongiloidíase/parasitologia , Células Th2/imunologiaRESUMO
The secretory EF-hand Ca++-binding proteins act as calcium signaling molecules for control of cell functions, but those proteins from parasitic helminths are poorly understood. Here, we have identified and characterized an EF-hand Ca++-binding protein from the rodent nematode, Strongyloides venezuelensis, termed 'venestatin', which is highly conserved in Strongyloides spp. Canonical two EF-hand domains and a signal peptide are present in venestatin. A gel mobility shift assay and Ruthenium red staining indicated that the recombinant venestatin possesses binding ability with Ca++ ions. Endogenous venestatin was seemingly localized in the hypodermis and gut of the worms and was found in the excretory-secretory products. Quantitative reverse transcription-PCR data showed that venestatin-specific transcript was upregulated in the parasitic stages of S. venezuelensis, and the upregulation occurred promptly after larval invasion through the host's skin, but not in the case of in vitro incubation. Immunization of mice with recombinant venestatin caused a 55% reduction in larval migration to the lungs, and lung hemorrhaging was mild compared with non-immunized groups, suggesting that anti-venestatin sera may interfere with larval migration from skin to lung. Our results suggest that venestatin is secreted from the hypodermis and gut of S. venezuelensis, and has pivotal roles in larval migration.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Helminto/metabolismo , Strongyloides/metabolismo , Estrongiloidíase/parasitologia , Animais , Proteínas de Ligação ao Cálcio/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas de Helminto/genética , Larva/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Nematodes of the genus Strongyloides are important parasites of vertebrates including man. Currently, little is known about their germline organization or reproductive biology and how this influences their parasitic life strategies. Here, we analyze the structure of the germline in several Strongyloides and closely related species and uncover striking differences in the development, germline organization, and fluid dynamics compared to the model organism Caenorhabditis elegans. With a focus on Strongyloides ratti, we reveal that the proliferation of germ cells is restricted to early and mid-larval development, thus limiting the number of progeny. In order to understand key germline events (specifically germ cell progression and the transcriptional status of the germline), we monitored conserved histone modifications, in particular H3Pser10 and H3K4me3. The evolutionary significance of these events is subsequently highlighted through comparisons with six other nematode species, revealing underlying complexities and variations in the development of the germline among nematodes.
Assuntos
Células Germinativas/metabolismo , Histonas/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Processos de Determinação Sexual/fisiologia , Strongyloides/metabolismo , Animais , Caenorhabditis elegans/citologia , Caenorhabditis elegans/metabolismo , Diferenciação Celular , Proliferação de Células , Metilação , Reprodução/fisiologia , Strongyloides/citologia , Strongyloides/genéticaRESUMO
Phage display is a powerful technology that selects specific proteins or peptides to a target. We have used Phage Display to select scFv (single-chain variable fragment) clones from a combinatorial library against total proteins of Strongyloides venezuelensis. After scFv characterization, further analysis demonstrated that this recombinant fragment of antibody was able to bind to an S. venezuelensis antigenic fraction of ~65 kDa, present in the body periphery and digestive system of infective larvae (L3), as demonstrated by immunofluorescence. Mass spectrometry results followed by bioinformatics analysis showed that this antigenic fraction was a heat shock protein 60 (HSP60) of Strongyloides sp. The selected scFv was applied in serodiagnosis by immune complexes detection in serum samples from individuals with strongyloidiasis using a sandwich enzyme-linked immunosorbent assay (ELISA), showing sensitivity of 97.5% (86.84-99.94), specificity of 98.81 (93.54-99.97), positive likelihood ratio of 81.60 and an area under the curve of 0.9993 (0.9973-1.000). Our study provided a novel monoclonal scFv antibody fragment which specifically bound to HSP60 of Strongyloides sp. and was applied in the development of an innovative serodiagnosis method for the human strongyloidiasis.
Assuntos
Antígenos de Helmintos/imunologia , Chaperonina 60/imunologia , Anticorpos de Cadeia Única/metabolismo , Strongyloides/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Sequência de Aminoácidos , Animais , Antígenos de Helmintos/sangue , Feminino , Humanos , Larva/metabolismo , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Ratos , Ratos Wistar , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Anticorpos de Cadeia Única/química , Anticorpos de Cadeia Única/genética , Strongyloides/crescimento & desenvolvimento , Adulto JovemRESUMO
BACKGROUND: Strongyloidiasis is commonly a clinically unapparent, chronic infection, but immuno suppressed subjects can develop fatal disease. We carried out a review of literature on hyperinfection syndrome (HS) and disseminated strongyloidiasis (DS), in order to describe the most challenging aspects of severe strongyloidiasis. METHODS: We conducted a structured search using PubMed to collect case reports and short case series on HS/DS published from 1991 to 2011. We restricted search to papers in English, Spanish, Italian and French. Case reports were classified as HS/DS according to given definitions. RESULTS: Records screened were 821, and 311 were excluded through titles and abstract evaluation. Of 510 full-text articles assessed for eligibility, 213 were included in qualitative analysis. As some of them were short case series, eventually the number of cases analyzed was 244.Steroids represented the main trigger predisposing to HS and DS (67% cases): they were mostly administered to treat underlying conditions (e.g. lymphomas, rheumatic diseases). However, sometimes steroids were empirically prescribed to treat signs and symptoms caused by unsuspected/unrecognized strongyloidiasis. Diagnosis was obtained by microscopy examination in 100% cases, while serology was done in a few cases (6.5%). Only in 3/29 cases of solid organ/bone marrow transplantation there is mention of pre-transplant serological screening. Therapeutic regimens were different in terms of drugs selection and combination, administration route and duration. Similar fatality rate was observed between patients with DS (68.5%) and HS (60%). CONCLUSIONS: Proper screening (which must include serology) is mandatory in high - risk patients, for instance candidates to immunosuppressive medications, currently or previously living in endemic countries. In some cases, presumptive treatment might be justified. Ivermectin is the gold standard for treatment, although the optimal dosage is not clearly defined in case of HS/DS.
Assuntos
Estrongiloidíase/diagnóstico , Animais , Antinematódeos/uso terapêutico , Humanos , Strongyloides/metabolismo , Estrongiloidíase/tratamento farmacológico , Estrongiloidíase/metabolismoRESUMO
Nematode parasites from the genus Strongyloides spp. are important pathogens of the intestinal mucosa of animals and humans. Their complex life cycles involve alternating developmental adaptations between larvae stages and the adult parthenogenetic female. Here, we report, primarily through homology-based searching, the existence of the major components of the ubiquitin-proteasome system in this genus, using the available EST data from S. ratti, S. stercoralis, and Parastrongyloides trichosuri. In this study, S. venezuelensis was used as our model organism for detection of proteasome activity and ubiquitinated substrates in cytosolic preparations from the L3 larvae and the adult female. Marked differences in proteasome capabilities were found when these two stages were compared. A preference for degradation of chymotryptic synthetic peptides was found in both stages with the adult exhibiting a higher rate of hydrolysis compared to the larvae. Due to the high evolutionary conservation of proteasome alpha subunits, an anti-human proteasome antibody was able to recognize proteasome subunits in these preparations by Western blotting, supporting the proposal that the activity of the ubiqutin-proteasome system is developmentally regulated in this nematode.
Assuntos
Complexo de Endopeptidases do Proteassoma/metabolismo , Strongyloides/metabolismo , Ubiquitina/metabolismo , Animais , DNA de Helmintos/genética , Etiquetas de Sequências Expressas , Feminino , Genômica , Larva/metabolismo , Análise de Sequência , Strongyloides/genéticaRESUMO
Complementary DNA (cDNA) sequencing can be used to sample an organism's transcriptome, and the generated EST sequences can be used for a variety of purposes. They are especially important for enhancing the utility of a genome sequence or for providing a gene catalog for a genome that has not or will not be sequenced. In planning and executing a cDNA project, several criteria must be considered. One should clearly define the project purpose, including organism tissue(s) choice, whether those tissues should be pooled, ability to acquire adequate amounts of clean and well-preserved tissue, choice of type(s) of library, and construction of a library (or libraries) that is compatible with project goals. In addition, one must possess the skills to construct the library (or libraries), keeping in mind the number of clones that will be necessary to meet the project requirements. If one is inexperienced in cDNA library construction, it might be wise to outsource the library production and/or sequence and analysis to a sequencing center or to a company that specializes in those activities. One should also be aware that new sequencing platforms are being marketed that may offer simpler protocols that can produce cDNA data in a more rapid and economical manner. Of course, the bioinformatics tools will have to be in place to de-convolute and aid in data analysis for these newer technologies. Possible funding sources for these projects include well-justified grant proposals, private funding, and/or collaborators with available funds.
Assuntos
Etiquetas de Sequências Expressas , Técnicas Genéticas , Algoritmos , Ancylostoma/metabolismo , Animais , Clonagem Molecular , DNA Complementar/metabolismo , Evolução Molecular , Biblioteca Gênica , Genoma , Hibridização de Ácido Nucleico , Análise de Sequência com Séries de Oligonucleotídeos , Polimorfismo de Nucleotídeo Único , RNA Mensageiro/metabolismo , Strongyloides/metabolismoRESUMO
Parastrongyloides trichosuri is a nematode parasite of Australian brushtail possums that has an alternative free-living life cycle which can be readily maintained indefinitely in a laboratory setting. The ability to maintain this parasite in a free-living cycle and induce it to parasitism at the free-living L1 stage makes this an excellent model for the study of genes associated with parasitism. A 70kD protein from infective larvae of P. trichosuri that appears to be immunogenic in infected possums has been identified as a heat shock protein (Hsp)70 homologue. The complete gene for Pt-Hsp70 was cloned and sequenced. The protein encoded by the Pt-Hsp70 gene is the likely orthologue of the Caenorhabditis elegans protein, Hsp70A, also known as hsp-1. Reverse transcriptase-PCR data indicate that Pt-Hsp70 (designated Pt-hsp-1) is expressed at readily detectable levels in all developmental stages of both the parasitic and free-living P. trichosuri life cycles and the promoter is mildly inducible by heat shock. Bioinformatic analysis of expressed sequence tag databases indicates that C. eleganshsp-1 homologues, together with C. eleganshsp-3 homologues, are the predominant members of the Hsp70 superfamily that are normally expressed in parasitic stages of the Strongyloididae family. Promoter fusions to a beta-galactosidase coding sequence were prepared and introduced into wild type C. elegans to produce transgenic nematodes. Reporter gene expression was clearly present within embryonic cells and within intestinal cells of larval and adult stages. Thus, the expression of the Pt-hsp-1 promoter within P. trichosuri and transgenic C. elegans appears similar to the known expression of C. elegans hsp-1. This promoter should be of value in efforts to develop genetic manipulation tools for P. trichosuri.
Assuntos
Genes de Helmintos , Proteínas de Choque Térmico HSP70/genética , Proteínas de Helminto/genética , Strongyloides/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Caenorhabditis elegans/genética , Biologia Computacional , Regulação da Expressão Gênica no Desenvolvimento , Proteínas de Choque Térmico HSP70/isolamento & purificação , Proteínas de Helminto/isolamento & purificação , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Strongyloides/crescimento & desenvolvimento , Strongyloides/metabolismo , Estrongiloidíase/parasitologia , Transformação Genética , Trichosurus/parasitologiaRESUMO
The parasitic female of Strongyloides venezuelensis keeps invading the epithelial layer of the host intestinal mucosa. Upon invasion, it adheres to the surface of the intestinal epithelial cells with adhesion molecules secreted from the mouth. It has been demonstrated that S. venezuelensis are expelled from the intestine because mucosal mast cells inhibit the attachment of adult worms to the mucosal surface. In the present study, we generated specific antibodies against secreted adhesion molecules to investigate their function in vivo, because these molecules have been demonstrated only in vitro in spite of the importance in the infection processes. A mouse monoclonal antibody specific to S. venezuelensis adhesion molecules inhibited the attachment of adult worms to plastic dishes and the binding of adhesion molecules to rat intestinal epithelial cells. Immunohistochemical study revealed that adhesion molecules were produced by oesophageal glands and were continuously secreted in vivo to line the wall of the tunnels formed by adult worms in the intestinal mucosa. Our findings indicate that adhesion molecules play essential roles in the infection processes of S. venezuelensis in the host intestine.
Assuntos
Moléculas de Adesão Celular/biossíntese , Mucosa Intestinal/parasitologia , Strongyloides/patogenicidade , Estrongiloidíase/parasitologia , Animais , Anticorpos Monoclonais , Especificidade de Anticorpos , Proteínas de Transporte/metabolismo , Moléculas de Adesão Celular/imunologia , Moléculas de Adesão Celular/metabolismo , Células Cultivadas , Esôfago/anatomia & histologia , Esôfago/citologia , Esôfago/metabolismo , Gerbillinae , Interações Hospedeiro-Parasita , Imunização , Masculino , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Ratos , Strongyloides/anatomia & histologia , Strongyloides/metabolismoRESUMO
Immunologically damaged Strongyloides venezuelensis adult worms were examined for their mucosal invasion ability and secretion of heparin-binding adhesion substances. S. venezuelensis was expelled from male Wistar rats 4 to 5 weeks after infection. Four-week-old adult worms were smaller and had fewer eggs than 1-week-old adult worms. One-week-old, 4-week-old, and 5-week-old adult worms equally established in the recipient mouse intestine when surgically implanted. Adult worms of 4 and 5 weeks of age secreted adhesion substances as much as 1-week-old adult worms. There was no difference in the heparin-binding activities and the lectin-binding profile of adhesion substances among adult worms of different ages. The rate of secretion of adhesion substances from the mouth was also identical. Heparin-binding activities were detected in crude adult worm proteins; however, proteins of 5-week-old adult worms had weaker heparin-binding activities than those of 1-week-old adult worms. Western blotting revealed that a number of heparin-binding proteins were lost in 5-week-old adult worms. A heparin-binding protein of 42. 0 kDa, which was consistently expressed in adult worms, was a possible component of heparin-binding adhesion substances which are secreted from the mouth.
Assuntos
Duodeno/parasitologia , Proteínas de Helminto/metabolismo , Heparina/metabolismo , Enteropatias Parasitárias/parasitologia , Strongyloides/metabolismo , Estrongiloidíase/parasitologia , Animais , Western Blotting , Adesão Celular/fisiologia , Duodeno/imunologia , Enteropatias Parasitárias/imunologia , Larva/imunologia , Larva/metabolismo , Lectinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Ratos Wistar , Strongyloides/imunologia , Estrongiloidíase/imunologiaRESUMO
Adhesion substances produced by adult worms of Strongyloides venezuelensis bound strongly to hepin-Sepharose beads after incubation at 37 degrees C for 1 h. This binding was completely inhibited by highly sulfated carbohydrates such as soluble heparin, dextran surfate, fucoidan, and pentosan polysulfate. Chondroitin sulfate E and chondroitin sulfate A inhibited to a lesser degree and chondroitin sulfate C and dextran did not inhibit significantly. Carbohydrate moieties as well as the number and position of negatively charged sulfate groups of sulfated glycans were important determinants for the interaction between sulfated carbohydrates and adhesion substances. Adhesion substances of S. venezuelensis adult worms also bound to negatively charged rat red blood cells. The binding was significantly inhibited by heparin but not by mono- or disaccharides. Thus the intraction between red cells and adhesion substances was electrostatic in nature, but did not involve lectin-sugar interactions.
Assuntos
Metabolismo dos Carboidratos , Moléculas de Adesão Celular/metabolismo , Strongyloides/metabolismo , Animais , Sulfatos de Condroitina/metabolismo , Sulfato de Dextrana/metabolismo , Dextranos/metabolismo , Eritrócitos/metabolismo , Heparina/metabolismo , Masculino , Microesferas , Poliéster Sulfúrico de Pentosana/metabolismo , Polissacarídeos/metabolismo , Ratos , Ratos WistarRESUMO
Adults worms of Strongyloides venezuelensis were cultured in vitro. After overnight incubation, about 60% of the worms adhered firmly to the bottom of culture vessels by secreting adhesive substances from the mouth. A single worm produced 24.5 +/- 10.1 of the adhesion spots overnight. When they were transferred to new culture vessels, they still produced new spots comparable to those produced for first 24 hr. The adhesion spots were positively stained with Coomassie brilliant blue and also with mucicarmine, periodic acid-Schiff, and alcian blue, pH 2.5, but not with alcian blue, pH 0.3, indicating their glycoprotein nature. The substances were amorphous and did not contain cells or nuclei. Histologic staining with a panel of lectins showed that the adhesive substances were rich in mannose, N-acetyl galactosamine, and N-acetyl glucosamine, but devoid of sialic acid. These characteristics were distinct from those of jejunal goblet cell mucins of rats. Adhesive substances contained antigenic components recognized by sera from infected rats. Thus, the adhesive substances secreted from the mouth of S. venezuelensis were clearly of parasite origin. We consider the production/secretion of the adhesive substances by S. venezuelensis adult worms a key step for the parasites to invade and establish the host epithelial layer.
Assuntos
Antígenos de Helmintos/metabolismo , Glicoproteínas/metabolismo , Proteínas de Helminto/metabolismo , Strongyloides/metabolismo , Adesividade , Animais , Anticorpos Anti-Helmínticos/biossíntese , Antígenos de Helmintos/química , Antígenos de Helmintos/imunologia , Vidro , Glicoproteínas/química , Glicoproteínas/imunologia , Proteínas de Helminto/química , Proteínas de Helminto/imunologia , Soros Imunes/imunologia , Masculino , Plásticos , Ratos , Ratos WistarRESUMO
The in vitro transformation system of Strongyloides venezuelensis has been established which induces free-living infective larvae to transform into the parasitic stage by a temperature shift from 25 to 37 degrees C. Comparison of the profiles of proteins labeled with [35S]-methionine in infective larvae at 25 and 37 degrees C with marked morphological transformation revealed an increase in the 70-kDa protein, thus showing these proteins to be related to the family of heat-shock proteins. In addition, the new appearance of two complexes between 16-kDa and 22-kDa proteins was observed. The 70-kDa protein cross-reacted with monoclonal antibody against human heat-shock protein 70, which was constitutively expressed. These proteins, synthesized during the transformation of S. venezuelensis from the infective larval stage to the parasitic stage, might be crucial for biochemical events that regulate the infectivity of the parasite for the host.
Assuntos
Proteínas de Choque Térmico HSP70/biossíntese , Proteínas de Helminto/biossíntese , Strongyloides/crescimento & desenvolvimento , Strongyloides/metabolismo , Animais , Anticorpos Anti-Helmínticos , Western Blotting , Reações Cruzadas , Eletroforese em Gel Bidimensional , Eletroforese em Gel de Poliacrilamida , Proteínas de Choque Térmico HSP70/imunologia , Proteínas de Helminto/imunologia , Humanos , MorfogêneseRESUMO
Strongyloidosis was induced by percutaneous injection of rabbits; then organ changes were tested. It has been found that Strongyloides papillosus evoked changes in intestine, liver, kidney and lung. No notable changes were found in heart, spleen and suprarenal gland. These changes may be provoked not only directly by the presence of these parasites but also by the products of metabolism of S. papillosus.
Assuntos
Strongyloides/metabolismo , Estrongiloidíase/patologia , Animais , Intestinos/patologia , Rim/patologia , Fígado/patologia , Pulmão/patologia , CoelhosRESUMO
Live, intact third-stage larvae (L3s) of Strongyloides ratti in the absence of exogenous substrates consumed oxygen at a rate (E-QO2) of 181.8 +/- 12.4 ng atoms min-1 mg dry weight-1 at 35 degrees C. Respiratory electron transport (RET) Complex I inhibitor rotenone (2 microM) produced 33 +/- 6.5% inhibition of the E-QO2. Unusually the rotenone-induced inhibition was not relieved by 5 mM-succinate. The E-QO2 of intact L3s was refractory to RET Complex III inhibitor antimycin A at 2 microM; 4 microM-antimycin inhibited less than or equal to 10% of the E-QO2. The electron donor couple ascorbate/TMPD augmented the E-QO2 in the presence of rotenone (2 microM) and antimycin A (4 microM) by 110%. Azide (1 mM) stimulated the antimycin A refractory QO2 by 36.6 +/- 7.2% which was only partially inhibited by 1.0 mM-KCN (IC50 = 0.8 mM). The data suggest the presence of classical (CPW) and alternate (APW) electron transport pathways in S. ratti L3s.
Assuntos
Rotenona/farmacologia , Strongyloides/metabolismo , Animais , Antimicina A/farmacologia , Azidas/farmacologia , Transporte de Elétrons/efeitos dos fármacos , Larva/metabolismo , Consumo de Oxigênio/efeitos dos fármacos , Cianeto de Potássio/farmacologia , Strongyloides/efeitos dos fármacosRESUMO
A technique is described for radiolabeling Strongyloides stercoralis larvae with [75Se]selenomethionine. Cultures of an auxotrophic methionine-dependent stain of Escherichia coli were grown in a medium containing Dulbecco's modified Eagle's medium supplemented with 5% nutrient broth, amino acids, and [75Se]selenomethionine. When the 75Se-labeled bacterial populations were in the stationary phase of growth, cultures were harvested and the bacteria dispersed on agar plates to serve as food for S. stercoralis larvae. Use of nondividing bacteria is important for successful labeling because the isotope is not diluted by cell division and death of larvae attributable to overgrowth by bacteria is prevented. First-stage S. stercoralis larvae were recovered from feces of infected dogs and reared in humid air at 30 C on agar plates seeded with bacteria. After 7 days, infective third-stage larvae were harvested. The mean specific activity of 6 different batches of larvae ranged from 75 to 330 counts per min/larva with 91.8 +/- 9.5% of the population labeled sufficiently to produce an autoradiographic focus during a practicable, 6-wk period of exposure. Labeled infective larvae penetrated the skin of 10-day-old puppies and migrated to the small intestine, where the developed to adulthood.
Assuntos
Selênio/metabolismo , Selenometionina/metabolismo , Strongyloides/metabolismo , Estrongiloidíase/parasitologia , Animais , Cães , Escherichia coli , Intestino Delgado/parasitologia , Larva/metabolismo , Larva/fisiologia , Radioisótopos de Selênio , Pele/parasitologia , Strongyloides/fisiologiaRESUMO
The effect of fatty acids was studied on the developmental direction of Strongyloides ratti first-stage larvae (L1). The proportion of third-stage infective larvae increased markedly when L1 were cultured in faeces with added fatty acids such as palmitic (C16), stearic (C18), oleic (C18:1) and linoleic (C18:2) acids. Unsaturated fatty acids were more effective than saturated ones. Moreover, the proportion of infective larvae increased with quantity of linoleic acid but the triacylglycerols of any fatty acid had no effect. These results suggest that these free fatty acids cause physiological changes that determine the developmental course of L1 of S. ratti in nature.