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1.
Parasite Immunol ; 32(4): 221-31, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-20398222

RESUMO

Deposition of Schistosoma mansoni eggs in the intestinal mucosa is associated with recruitment of mucosal mast cells (MMC) expressing mouse mast cell protease-1 (mMCP-1). We investigated the involvement of mMCP-1 in intestinal barrier disruption and egg excretion by examining BALB/c mice lacking mMCP-1 (Mcpt-1(-/-)). Tissue and faecal egg counts from 6 weeks until 12 weeks post-infection (w p.i.) revealed no differences between wild type (WT) and Mcpt-1(-/-)mice. Using chamber experiments on ileal tissue revealed that at 8 w p.i., the epithelial barrier and secretory capacity were severely impaired, whereas no difference was found between WT and Mcpt-1(-/-)mice in this respect. However, a fragmented distribution of the tight junction (TJ) protein occludin, but not of claudin-3 or ZO-1, was observed in WT mice at 8 w p.i., while no changes in TJ integrity were seen in Mcpt-1(-/-)mice. Therefore, we conclude that in contrast to the situation in Trichinella spiralis-infected mice, in schistosomiasis, mMCP-1 is not a key mediator in egg excretion or impairment of the intestinal barrier. The marked decrease in ileal secretory capacity during S. mansoni egg excretion suggests that the mechanisms facilitating the passage of schistosoma eggs through the gut wall are directed more particularly at the epithelial cells.


Assuntos
Quimases/metabolismo , Mucosa Intestinal/patologia , Mucosa Intestinal/parasitologia , Mastócitos/imunologia , Schistosoma mansoni/patogenicidade , Esquistossomose mansoni/imunologia , Esquistossomose mansoni/patologia , Animais , Quimases/deficiência , Íleo/imunologia , Íleo/parasitologia , Íleo/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Técnicas de Cultura de Órgãos , Contagem de Ovos de Parasitas , Schistosoma mansoni/imunologia
2.
Vet Immunol Immunopathol ; 120(1-2): 47-54, 2007 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-17714792

RESUMO

Infection of sheep with Teladorsagia circumcincta triggers an immune response with predominantly type-2 (Th2) characteristics, including local eosinophila, mastocytosis and increased mucus production. In order to better understand the protective immune responses elicited, we used RT-PCR assays to define the changes in expression levels of a range of cytokine transcripts in lymph nodes draining the ovine abomasum following a challenge infection with T. circumcincta. This study compared the changes in cytokine expression in the abomasal lymph node following challenge with T. circumcincta in naïve sheep (Group 2) and sheep immunised by a previous trickle infection (Group 3), in comparison to unchallenged naive sheep (Group 1). There was a significant up-regulation of interleukin-4 (IL-4), IL-5 and IL-13 in both the challenged groups compared to naïve individuals. There was also an up-regulation of IL-1beta, IL-6, IL-10, IL-18, transforming growth factor-beta1 (TGFbeta1) and tumour necrosis factor-alpha (TNFalpha) by day 5 after infection. IL-12p40 was found to be increased in the previously infected Group 3 animals by day 5 following challenge. By contrast, transcription of this cytokine was found to be reduced by day 10 following infection of Group 2 animals. Expression of IL-2 and Interferon-gamma (IFNgamma) did not significantly differ between the three groups.


Assuntos
Citocinas/metabolismo , Regulação da Expressão Gênica/imunologia , Doenças dos Ovinos/imunologia , Trichostrongyloidea/fisiologia , Tricostrongiloidíase/veterinária , Animais , Citocinas/genética , Ovinos , Doenças dos Ovinos/metabolismo , Doenças dos Ovinos/parasitologia , Tricostrongiloidíase/imunologia , Tricostrongiloidíase/metabolismo
3.
Vet Immunol Immunopathol ; 120(1-2): 55-60, 2007 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-17709146

RESUMO

Mammalian chitinases and chitinase-like proteins are a group of molecules known to be upregulated and secreted in Th2-induced inflammatory responses, such as asthma, allergy and nematode infection. As part of an investigation of potential components of the innate immune response to Teladorsagia circumcincta, a gastrointestinal nematode that colonises the abomasum in sheep, we carried out RT-PCR analysis of two members of the mammalian chitinase family of molecules, acidic chitinase (ChiA) and chitinase-3 like 1 (Chi3L1) using primers to homologous bovine/human sequences. Both sets of primers detected transcripts in the abomasum which were confirmed to be ovine ChiA and Chi3L1 by sequence analysis. Chi3L1 transcripts were found to be significantly upregulated in both the abomasum and gastric lymph nodes in response to T. circumcincta challenge of previously infected animals.


Assuntos
Abomaso/imunologia , Quitinases/metabolismo , Doenças dos Ovinos/imunologia , Trichostrongyloidea/fisiologia , Tricostrongiloidíase/veterinária , Regulação para Cima/imunologia , Animais , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Ovinos , Doenças dos Ovinos/metabolismo , Tricostrongiloidíase/imunologia , Tricostrongiloidíase/metabolismo
4.
Vet Immunol Immunopathol ; 110(1-2): 187-91, 2006 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-16216338

RESUMO

The high-affinity receptor for IgE (FcepsilonRI) plays a central role in IgE-mediated allergic reactions. Cross-linking of FcepsilonRI by IgE-antigen complexes results in the activation of mast cells and basophils and is thought to contribute to the immunopathology of Heaves, a chronic obstructive pulmonary disease of horses. Recombinant protein corresponding to the extra-cellular portion of the FcepsilonRI alpha subunit, cloned and sequenced previously, was expressed using both mammalian cells and insect cells. The yield of expressed protein was considerably greater using insect cells and the baculovirus expression system. The recombinant proteins differed in size between the two systems, presumably due to differences in the extent of glycosylation. However, recombinant protein from both cell systems bound equine IgE present in bronchoalveolar lavage fluid from horses with Heaves. These results suggest that the recombinant extra-cellular part of FcepsilonRI should be a useful tool with which to study equine IgE responses.


Assuntos
Doenças dos Cavalos/imunologia , Imunoglobulina E/imunologia , Doença Pulmonar Obstrutiva Crônica/veterinária , Receptores de IgE/genética , Animais , Baculoviridae/genética , Western Blotting/veterinária , Líquido da Lavagem Broncoalveolar/imunologia , Células COS , Chlorocebus aethiops , Doenças dos Cavalos/diagnóstico , Cavalos , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Doença Pulmonar Obstrutiva Crônica/diagnóstico , Doença Pulmonar Obstrutiva Crônica/imunologia , Receptores de IgE/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia
5.
Vet Immunol Immunopathol ; 105(1-2): 141-50, 2005 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-15797483

RESUMO

Earlier studies of cattle and sheep have demonstrated that Psoroptes ovis infestations provoke an intense immunoinflammatory response dominated by eosinophils accompanied by a substantial infiltrate of lymphocytes. However, the kinetics of the lymphocyte response and the subtypes involved have not been characterised. We employed two groups of sheep to investigate the early (1-21 days) and later (21-63 days) infiltration of lymphocyte subpopulations and dendritic cells in primary infestations of sheep with P. ovis. Immunohistochemistry indicated that by 4 days after infestation numbers of CD4+ and CD45RA+ cells in lesional skin had increased significantly (P<0.03 and P<0.005, respectively) and that a significant increase in gammadelta T cells and dendritic cells (CD1b+) had occurred by 8 days (P<0.02 and P<0.01, respectively). Numbers of lymphocyte and dendritic cells declined from 49 to 63 days after infestation. Our observations suggest that mite-derived products exert a profound influence on the early recruitment of lymphocytes that may significantly influence the genesis of the adaptive immune response.


Assuntos
Células Dendríticas/imunologia , Infestações por Ácaros/imunologia , Psoroptidae/imunologia , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/parasitologia , Dermatopatias Parasitárias/veterinária , Subpopulações de Linfócitos T/imunologia , Animais , Biópsia/veterinária , Células Dendríticas/parasitologia , Imuno-Histoquímica/veterinária , Contagem de Linfócitos/veterinária , Infestações por Ácaros/parasitologia , Ovinos , Dermatopatias Parasitárias/imunologia , Dermatopatias Parasitárias/parasitologia , Subpopulações de Linfócitos T/parasitologia , Linfócitos T/imunologia , Linfócitos T/parasitologia
6.
J Comp Pathol ; 131(4): 318-29, 2004 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-15511540

RESUMO

Clinical observation has indicated that Psoroptes ovis mites provoke cutaneous inflammation within hours of experimental infestation, but the nature of this reaction has not been described. After infestation of naive sheep with ovigerous P. ovis mites, significant influxes of eosinophils (P<0.004) and neutrophils (P<0.001) were detected within 24 h. A significant (P<0.001) increase in mast cell numbers was observed by 96 h post-infestation. In addition, marked degenerative and proliferative epidermal lesions were evident 24 and 96 h, respectively, after infestation. The influence of the later, adaptive response on the cellular infiltrate at the advancing margin of the lesion and the original site of infestation was also monitored. Mast cell numbers were greatest at 21 days while recruitment of eosinophils and neutrophils was maximal 63 days after infestation. Lesional severity was particularly pronounced from 42 to 63 days after infestation, but significant resolution had occurred by 84 days. Pathological changes at the advancing margin of the lesion were more severe than at the initial site of infestation, and this was reflected by the numbers of mites present. These data suggest that P. ovis elicits an early innate cutaneous response that is subsequently augmented by the development of an adaptive immune response, the intensity of which corresponds to the local population density of mites.


Assuntos
Imunidade Celular/imunologia , Infestações por Ácaros/veterinária , Doenças Parasitárias em Animais/imunologia , Psoroptidae/imunologia , Doenças dos Ovinos/imunologia , Pele/imunologia , Animais , Contagem de Células , Interações Hospedeiro-Parasita/imunologia , Infestações por Ácaros/imunologia , Infestações por Ácaros/patologia , Doenças Parasitárias em Animais/parasitologia , Doenças Parasitárias em Animais/patologia , Ovinos , Doenças dos Ovinos/parasitologia , Doenças dos Ovinos/patologia , Pele/parasitologia , Pele/patologia
7.
Am J Physiol Gastrointest Liver Physiol ; 287(1): G178-91, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15016615

RESUMO

Upregulation of CGRP-immunoreactive (IR) primary afferent nerve fibers accompanied by mastocytosis is characteristic for the Schistosoma mansoni-infected murine ileum. These mucosal mast cells (MMC) and CGRP-IR fibers, which originate from dorsal root (DRG) and nodose ganglia, are found in close apposition. We examined interactions between primary cultured MMC and CGRP-IR DRG neurons in vitro by confocal recording of intracellular Ca(2+) concentration ([Ca(2+)](i)). The degranulatory EC(50) for the mast cell secretagogue compound 48/80 (C48/80; 10 microg/ml) and the neuropeptides CGRP (2.10(-8) M) and substance P (SP; 3.10(-8) M) were determined by measurement of extracellular release of the granule chymase, mouse mast cell protease-1. Application of C48/80 (10 microg/ml) and CGRP and SP (both 10(-7) M) to Fluo-4-loaded MMC induced a transient rise in [Ca(2+)](i) after a lag time, indicative of mast cell degranulation and/or secretion. The CGRP response could be completely blocked by pertussis toxin (2 microg/ml), indicating involvement of G(i) proteins. Application of MMC juice, obtained by C48/80 degranulation of MMC, to Fluo-4-loaded DRG neurons induced in all neurons a rise in [Ca(2+)](i), indicative of activation. Degranulation of MMC by C48/80 in culture dishes containing Fluo-4-loaded DRG neurons also caused activation of the DRG neurons. In conclusion, these results demonstrate a bidirectional cross-talk between cultured MMC and CGRP-IR DRG neurons in vitro. This indicates that such a communication may be the functional relevance for the close apposition between MMC and CGRP-IR nerve fibers in vivo.


Assuntos
Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Degranulação Celular/fisiologia , Gânglios Espinais/fisiologia , Mucosa Intestinal/fisiologia , Mastócitos/fisiologia , Neurônios/fisiologia , Animais , Líquidos Corporais/fisiologia , Peptídeo Relacionado com Gene de Calcitonina/farmacologia , Cálcio/metabolismo , Células Cultivadas , Gânglios Espinais/citologia , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Membranas Intracelulares/metabolismo , Masculino , Mastócitos/metabolismo , Camundongos , Concentração Osmolar , Substância P/farmacologia , p-Metoxi-N-metilfenetilamina/farmacologia
8.
Clin Exp Allergy ; 33(7): 1005-12, 2003 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-12859460

RESUMO

BACKGROUND: Gastrointestinal nematode infection is associated with mucosal mast cell (MMC) hyperplasia. In the mouse, this is accompanied by the release of substantial quantities of the chymase mouse mast cell proteinase-1 (mMCP-1) into the gut lumen and peripheral bloodstream. Expression of mMCP-1 is largely restricted to intraepithelial MMC and is thought to play a role in the regulation of epithelial permeability. MMCs also express mouse mast cell proteinase-2 (mMCP-2), but less is known about the expression or biological function of this proteinase. OBJECTIVES: (1) To purify and characterize mMCP-2. (2) To compare the expression and release of mMCP-2 and mMCP-1 in vivo using specific antibodies. METHODS: Bone marrow-derived mast cells (mBMMCs) were generated from mMCP-1(-/-) BALB/c mice. mMCP-2 was purified, characterized and used to generate rat and sheep polyclonal antibodies. The expression and systemic release of mMCP-1 and -2 were compared in vivo by immunohistochemistry and ELISA. RESULTS: mMCP-2 was successfully purified from mMCP-1(-/-) mBMMC and its identity confirmed by N-terminal amino acid sequencing. mMCP-2 bound [3H]-labelled DFP, indicating the presence of an active serine proteinase catalytic site, but showed little evidence of chymotryptic activity. MMC expressed comparable levels of mMCP-1 and -2 in the jejunum but not in the gastric mucosa, where mMCP-2 was more abundant. Expression of both proteinases increased substantially during primary Nippostrongylus brasiliensis infection and this was accompanied by a substantial increase in peripheral blood levels of mMCP-1 (70 microg/mL on day 12). By contrast, mMCP-2 was not detected in the serum of uninfected mice and only increased to approximately 25 ng/mL on day 12. CONCLUSION: As in the case of mMCP-1, mMCP-2 expression is restricted to MMC. However, mMCP-2 lacks chymase activity, is expressed at higher levels in gastric MMC and appears to be differentially released into the peripheral bloodstream.


Assuntos
Serina Endopeptidases/isolamento & purificação , Animais , Quimases , Ensaio de Imunoadsorção Enzimática , Feminino , Imuno-Histoquímica , Mucosa Intestinal/metabolismo , Doenças do Jejuno/metabolismo , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Nippostrongylus , Serina Endopeptidases/metabolismo , Gastropatias/metabolismo , Infecções por Strongylida/metabolismo , Especificidade por Substrato
9.
J Helminthol ; 77(2): 155-61, 2003 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-12756069

RESUMO

Mucosal mast cells (MMC) play an important role in the immune response against selected species of intestinal nematode. The kinetics with which different strains of inbred mice resolve infection with Trichinella spiralis correlates with their ability to mount MMC responses in the intestinal mucosa. Homologues of MMC that express and constitutively secrete abundant amounts of the granule chymase, mouse mast cell protease-1 (mMCP-1), can be generated in vitro from bone marrow cultures supplemented with interleukins-3 and -9, stem cell factor and transforming growth factor-beta1. Using the enhanced growth characteristics of these MMC homologues, a novel limiting dilution assay for mast cell precursor (MCp) frequency has been developed. The assay is highly specific, in that cultures containing mast cells are identified with mMCP-1 specific antibody, and almost three-fold more sensitive than previously published systems. MCp frequencies were compared in BALB/c and C57/BL10 strains of mice that, respectively, respond rapidly and slowly to infection with T. spiralis. MCp frequency (1/378 bone marrow cells) was significantly greater in BALB/c than C57/BL10 mice (frequency: 1/751). Similarly the rate of growth of MMC homologues and the production of mMCP-1 was significantly greater in BALB/c than in C57/BL10 bone marrow cultures.


Assuntos
Enteropatias Parasitárias/imunologia , Mucosa Intestinal/imunologia , Mastócitos/imunologia , Trichinella spiralis , Triquinelose/imunologia , Animais , Anticorpos/análise , Células da Medula Óssea , Diferenciação Celular , Células Cultivadas , Quimases , Meios de Cultura , Imunofluorescência , Interleucina-3 , Interleucina-9 , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Serina Endopeptidases/imunologia , Especificidade da Espécie , Fator de Células-Tronco , Fator de Crescimento Transformador beta
10.
Immunogenetics ; 55(2): 122-5, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12709836

RESUMO

The high-affinity receptor for IgE is expressed on the surface of mast cells and basophils. It is a transmembrane protein with one alpha, one beta and two gamma subunits. The cDNA sequences for the alpha subunit have already been determined. We report here the cDNA sequences for the beta and gamma subunits. The cytoplasmic domains of these subunits are important for intracellular signalling and the deduced amino acid sequences show the expected immunoreceptor tyrosine-based activation motifs. The gamma subunit is highly conserved between species but more variation is seen with the beta subunit. Near the C terminus of the equine beta chain there is a two-base deletion, which changes the reading frame: residue 237 (human numbering) becomes Asp instead of Glu and the chain is three amino acids shorter than the other known mammalian and rodent sequences.


Assuntos
DNA Complementar , Cavalos/genética , Receptores de IgE/genética , Ovinos/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Dados de Sequência Molecular
11.
Clin Exp Allergy ; 33(1): 132-46, 2003 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-12534561

RESUMO

BACKGROUND: The mucosal mast cell (MMC) granule-specific beta-chymase, mouse mast cell protease-1 (mMCP-1), is released systemically into the bloodstream early in nematode infection before parasite-specific IgE responses develop and TGF-beta1 induces constitutive release of mMCP-1 by homologues of MMC in vitro. Intraepithelial MMC may also express the chemokine CCL2 (monocyte chemotactic protein-1) during nematode infection but the expression of this chemokine by MMC homologues has not been investigated. OBJECTIVE: To investigate the expression and to compare the mechanisms of constitutive release of the chymase, mMCP-1, and the chemokine, CCL2. METHODS: MMC homologues were generated by culturing bone marrow cells in the presence of TGF-beta1, IL-3, IL-9 and stem cell factor (SCF). The intracellular distribution of mMCP-1 and CCL2 was examined by confocal microscopy. The involvement of the Golgi complex and of protein synthesis in the constitutive release of mMCP-1 and CCL2 was investigated using the Golgi-disrupting agent brefeldin A and cycloheximide to block protein synthesis. Secreted analytes were quantified by ELISA. RESULTS: mMCP-1 colocalized with Golgi matrix protein 130 but was most abundant in the granules, whereas CCL2 was not found in the granules but appeared to be located uniquely in the Golgi complex. Extracellular release of mMCP-1 was significantly inhibited ( approximately 40%) by cycloheximide and by the Golgi-disrupting agent brefeldin A, indicating both continuous protein synthesis and transportation via the Golgi complex are required for optimal mMCP-1 secretion. A similar but more marked inhibitory effect with both compounds was demonstrated on the constitutive secretion of CCL2. CONCLUSION: The culture conditions that promote mMCP-1 expression and release by MMC homologues also promote the expression and release of CCL2. Constitutive release involves de novo protein synthesis and requires a functional Golgi complex, suggesting that similar mechanisms of extracellular secretion operate for both mediators.


Assuntos
Proteínas de Bactérias/metabolismo , Complexo de Golgi/química , Mastócitos/enzimologia , Proteínas de Membrana/metabolismo , Mucosa Respiratória/imunologia , Serina Endopeptidases/metabolismo , Animais , Apoptose , Células da Medula Óssea , Brefeldina A/farmacologia , Técnicas de Cultura de Células , Células Cultivadas , Quimases , Cicloeximida/farmacologia , Complexo de Golgi/efeitos dos fármacos , Masculino , Mastócitos/ultraestrutura , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Confocal , Inibidores da Síntese de Proteínas/farmacologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Serina Endopeptidases/análise , Serina Endopeptidases/genética
12.
Vet Immunol Immunopathol ; 91(2): 105-17, 2003 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-12543547

RESUMO

We have previously shown that infestation with Psoroptes ovis induces an IgE response and intense tissue eosinophilia, typical of a Type I hypersensitivity response [Parasite Immunol. 22 (2000) 407]. Intradermal tests (IDSTs) suggest that there are also delayed and Arthus-type responses to this parasite. In order to study the nature of ovine cutaneous reactions to P. ovis, naïve controls and experimentally infested sheep (n = 5) were challenged intradermally with mite antigen. Challenge elicited immediate (P < 0.001) and delayed (P < 0.005) wheal reactions in sensitised sheep. At 6 (P < 0.02) and 30 h (P < 0.001) the predominant infiltrating cells were eosinophils. To explore the role of circulating antibodies, naïve sheep (n = 5) were subjected to Prausnitz-Kustner (PK) tests. These elicited immediate (P < 0.02) but not delayed wheal reactions. At 6 h eosinophils (P < 0.001) dominated the infiltrate. These results suggest that P. ovis allergens provoke an IgE-dependent immediate and late phase response and a cell-mediated eosinophil-rich delayed-type hypersensitivity response (ER-DTH).


Assuntos
Antígenos de Dermatophagoides/imunologia , Hipersensibilidade/imunologia , Hipersensibilidade/veterinária , Psoroptidae/imunologia , Carneiro Doméstico/imunologia , Carneiro Doméstico/parasitologia , Pele/imunologia , Animais , Anticorpos/imunologia , Antígenos/administração & dosagem , Antígenos/imunologia , Proteínas de Artrópodes , Cisteína Endopeptidases , Granulócitos/citologia , Granulócitos/imunologia , Contagem de Leucócitos , Mastócitos/citologia , Mastócitos/imunologia , Infestações por Ácaros/imunologia , Infestações por Ácaros/veterinária , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/parasitologia , Fatores de Tempo
13.
Vet Parasitol ; 111(2-3): 217-30, 2003 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-12531296

RESUMO

In sheep Psoroptes ovis provokes an allergic dermatitis with significant P. ovis antigen-specific IgE responses. The kinetics of the IgE response to primary and challenge infestations of P. ovis were reported earlier [Parasite Immunol. 22 (2000) 407]. The present study examines IgG, IgM and IgA responses to primary and challenge infestations of P. ovis and the profile of antigens/allergens reacting with IgG and IgE antibodies. Antigen-specific enzyme-linked immunosorbent assays (ELISAs) demonstrate that primary infestations elicited significant increases in levels of IgG and IgM but not IgA antibodies. IgG and IgM responses to primary and challenge infestations were not significantly different. Western blots of reduced P. ovis proteins indicate that IgG antibodies reacted with five major antigens following primary infestation and only three of these after challenge infestation. IgE antibodies bound to three major and five minor allergens after primary infestation and two additional minor allergens after challenge infestation. Immunodominant antigens >100 and <15 kDa and allergens >100 kDa were most consistent in stimulating substantial IgG and IgE antibody responses, respectively. These antigens/allergens may be exploited in immunodiagnosis and modulation of the host immune response.


Assuntos
Isotipos de Imunoglobulinas/biossíntese , Infestações por Ácaros/veterinária , Psoroptidae/imunologia , Doenças dos Ovinos/parasitologia , Alérgenos/imunologia , Animais , Western Blotting/veterinária , Ensaio de Imunoadsorção Enzimática/veterinária , Epitopos Imunodominantes/imunologia , Isotipos de Imunoglobulinas/imunologia , Infestações por Ácaros/imunologia , Infestações por Ácaros/parasitologia , Ovinos , Doenças dos Ovinos/imunologia
14.
Parasite Immunol ; 24(8): 413-22, 2002 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12406195

RESUMO

Infestation of sheep with the ectoparasitic mite Psoroptes ovis, results in a severe allergic dermatitis. Currently, little is known about the allergens/antigens that stimulate the allergic response. We have isolated an 836-bp cDNA from a P. ovis cDNA library which displays strong homology to cysteine proteases and, in particular, to the group I house dust mite allergens Der p 1, Der f 1 and Eur m 1. The cDNA was expressed in Escherchia coli, fused to a hexahistidine tag and the recombinant protein (Pso o 1) purified using a nickel-affinity column. The recombinant Pso o 1 was tested for recognition by immunoglobulin (Ig)G and IgE in serum from P. ovis naïve and P. ovis infested sheep. Using Western blots, both classes of antibody to Pso o 1 were detected in postinfestation serum. In enzyme-linked immunosorbent assays, a pronounced IgG-antibody response to Pso o 1 was detected in five of five sheep 3 weeks postinfestation. The IgE-antibody response to whole mite extract was poor in four of five animals. However, a marked IgE response occurred in the fifth animal, and IgE anti Pso o 1 was detected in the serum.


Assuntos
Alérgenos/imunologia , Antígenos/imunologia , Infestações por Ácaros/imunologia , Psoroptidae/imunologia , Alérgenos/genética , Sequência de Aminoácidos , Animais , Antígenos/genética , Antígenos de Dermatophagoides/imunologia , Sequência de Bases , Clonagem Molecular , Imunoglobulina E/análise , Imunoglobulina E/imunologia , Imunoglobulina G/análise , Imunoglobulina G/imunologia , Infestações por Ácaros/diagnóstico , Dados de Sequência Molecular , Psoroptidae/citologia , Psoroptidae/patogenicidade , Pyroglyphidae/genética , Pyroglyphidae/imunologia , Proteínas Recombinantes/imunologia , Alinhamento de Sequência , Análise de Sequência de DNA , Análise de Sequência de Proteína , Ovinos
15.
Clin Exp Allergy ; 32(2): 315-24, 2002 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-11929499

RESUMO

BACKGROUND: Mucosal mast cells (MMC) play a central role in gut hypersensitivities and inflammation. They are morphologically, biochemically and functionally distinct from their connective tissue counterparts. Massive hyperplasia of MMC occurs 7-10 days after intestinal infection with nematodes but it has never been possible to replicate this phenomenon in vitro. OBJECTIVE: (1) To determine whether mouse bone marrow-derived mast cells (mBMMC) grown in the presence of transforming growth factor (TGF)-beta1 could develop over the same time frame (7-10 days) as MMC in parasitized mice. (2) To compare the early expression of surface receptors (integrins alphaE and beta7, c-kit and FcepsilonR) with that of the MMC-specific granule chymase mouse mast cell protease-1 (mMCP-1). METHODS: Mouse bone marrow cells were cultured in the presence of IL-9, IL-3 and Stem Cell Factor (SCF) with or without TGF-beta1. mBMMC were quantified after toluidine blue or Leishmans' staining. Expression of MMC-specific mouse mast cell proteases was analysed by ELISA, immunohistochemistry and RT-PCR. Surface antigen expression was characterized by flow cytometry and confocal microscopy. RESULTS: TGF-beta1 promotes the development of abundant MMC-like mBMMC from bone marrow progenitor cells with kinetics, which closely parallel that seen in vivo. mRNA transcripts encoding mMCP-1 and -2 are readily detectable by day 4 ex vivo in cultures grown in the presence of TGF-beta1. Between 30 and 40% and 75-90% of the cells in these cultures on days 4 and 7, respectively, have typical mast cell morphology, are c-kit+, FcepsilonR+, integrin alphaEbeta7+, and express and secrete abundant mMCP-1. The integrin alphaE subunit is coexpressed with mMCP-1. CONCLUSION: The kinetics of mMCP-1+/alphaE+ mBMMC development, regulated by TGF-beta1, are consistent with that seen in vivo in the parasitized intestine. The normally down-regulatory functions of TGF-beta1 in haematopoiesis are superseded in this culture system by its ability to promote the early expression of alphaE and mMCP-1.


Assuntos
Antígenos CD/metabolismo , Células da Medula Óssea/citologia , Cadeias alfa de Integrinas , Mastócitos/citologia , Serina Endopeptidases/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Animais , Células da Medula Óssea/efeitos dos fármacos , Diferenciação Celular/fisiologia , Células Cultivadas , Quimases , Interleucina-3/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mucosa/citologia , Serina Endopeptidases/genética , Fixação de Tecidos , Transcrição Gênica , Fator de Crescimento Transformador beta1
16.
Vet Dermatol ; 7(1): 11-20, 1996 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-34644992

RESUMO

Abstract The in vivo dynamics of differentiated cells and interleukin (IL)-lß, IL-8, granulocyte-macrophage colony-stimulating factor (GM-CSF) and interferon (IFN)-γ titres in afferent lymph were compared following orf virus reinfection and inactivated virus injection of previously infected sheep. The biphasic lymphoblast and cytokine response in the lymph to virus reinfection is consistent with a response initially to orf virus as recall antigen followed by a response to viral replication. CD4 T cells increased in output over other cell types in the lymph in both groups. A rapid immune/inflammatory response was detected in lymph plasma as an increase in cytokine titres within 24 h of virus reinfection or injection. Lymph cells producing IL-1ß and IL-8 appeared prior to those producing GM-CSF in both groups. In spite of variations in the concentration of individual cytokines in lymph following reinfection, both the size of the orf lesion and the time to resolve were similar in all cases. Résumé- Les dynamiques in vivo des cellules différenciées et des taux d'IL-1ß, de GM-CSF et d'interferon γ dans la lymphe afférente furent comparés chez des moutons antéricurement infectés après une réinfection par le virus de l'ecthyma contagieux et après injection d'un virus inactivé. La réponse biphasique lymphoblastique et des cytokines à la réinfection virale est compatibles avec une réponse primaire au virus de l'ecthyma contagieux comme antigène mémoire suivie par une réponse secondaire à la réplication virale. Dans les 2 groupes, le nombre de TCD4 est plus élevé que les autres populations cellulaires mises en évidence dans la lymphe. Une réponse de type immune/inflammatoire est révélée dans le plasma par une élévation des taux de cytokines dans les 24 heures qui suivent la réinfection virale ou l'injection de virus inactivé. Les lymphocytes producteurs dTL-1ß et d'IL-8 apparaissent avant les lymphocytes producteurs de GM-CSF dans les deux groupes. En dépit des variations de concentration des cytokines individuelles dans le lymphe après reinfection, à la fois la taille des lesions d'ecthyma et les délais de guérison sont identiques dans tous les cas. [Haig, D., Deane, D., Percival, A., Myatt, N., Thomson, J., Inglis, L., Rothel, J., Heng-Fong Seow, Wood, P., Miller, H. R. P., Reid, H. W. The cytokine response of afferent lymph following orf virus reinfection of sheep (Effet sur les cytokines de la lymphe afferente d'une reinfection par les virus de l'ecthyma contagieux chez le mouton.) Veterinary Dermatology 1996; 7: 11-20.] Resumen Se comparó la actividad de células diferenciadas y los titulos de interleuquina (IL)-1ß, IL-8, factor de estimulación de colonias de granulocitos y macrófagos (GM-CSF) e interferon (IFN)-y entre reinfecciones por el virus del ectima contagioso e inyección de virus inactivado de ovinos previamente infectados. La respuesta a la reinfección en forma bifásica linfoblástica y de citoquinas en la linfa está de acuerdo con una respuesta inicial al virus del ectima por estimulación antigénica, seguida por una respuesta a la replicación viral. Las células T CD4 aumentaron respecto a otros tipos celulares en la linfa de ambos grupos. Se detectó una respuesta inmune/inflamatoria rápida en el linfa-plasma en forma de aumento de los titulos de citoquinas dentro de las 24 h de reinfección o inyección del virus. Las células linfáticas productoras de IL-1ß e IL-8 aparecicron antes que las productoras de GM-CSF en ambos grupos. A pesar de las variaciones en la concentración de citoquinas individuates en la linfa después de la reinfección, tanto el tamaño de la lesión por el virus del ectima contagioso como el tiempo de resolución fueron similares en todos los casos. [Haig, D., Deane, D., Percival, A., Myatt, N., Thomson, J., Inglis, L., Rothel, J., Heng-Fong Seow, Wood, P., Miller, H. R. P., Reid, H. W. The cytokine response of afferent lymph following orf virus reinfection of sheep (Produccion de citoquinas en el ganglio linfatico afferente tras la reinfección por el virus del ectima contagioso.) Veterinary Dermatology 1996; 7: 11-20.] Zusammenfassung- Es wurde die in-vivo-Dynamik der Titer differenzierter Zellen und Interleukin (1L)-1ß, IL-8, Granulozytenmakrophagenkolonien-stimulierender Faktor (GM-CSF) und Interferon (IFN)-γ in afferenter Lymphe nach einer Reinfektion mit ORF-Virus und einer Injektion inaktivierten Virusmaterials von früher infizierten Schafen verglichen. Die biphasische Lymphoblasten- und Zytokin-Reaktion in der Lymphe auf die Virusreinfektion stimmte mit einer initialen Reaktion gegenüber ORF-Virus überein, da der Wiederabruf von Antigen von einer Reaktion auf die Virusreplikation gefolgt wird. CD4-T-Zellen vermehrten sich im Output stärker als andere Zelltypen in der Lymphe beider Gruppen. Es wurde eine rasche immunologische/entzündliche Reaktion im Lymphplasma in Form eines Anstieges von Zytokin-Titern innerhalb von 24 Stunden nach Virusreinfektion oder -injektion festgestellt. Lymphatische Zellen, die IL-1ß und IL-8 produzieren, erschienen vor dem Auftreten von solchen, die GM-CSF produzieren, in beiden Gruppen. Trotz des Schwankens der Konzentration der individuellen Zytokine in der Lymphe nach Reinfektion, waren sowohl die Größe der ORF-Veränderungen und die Zeit der Heilung ähnlich in alien Fällen. [Haig, D., Deane, D., Percival, A., Myatt, N., Thomson, J., Inglis, L., Rothel, J., Heng-Fong Seow, Wood, P., Miller, H. R. P., Reid, H. W. The cytokine response of afferent lymph following orf virus reinfection of sheep (Die ZytokinReaktion afferenter Lymphe nach einer Reinfektion mit orf-virus beim schaf.) Veterinary Dermatology 1996; 7: 11-20.].

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