RESUMO
GalNAc-glycopeptides derived from mucin MUC1 are an important class of tumor-associated antigens. α- O-glycosylation forces the peptide to adopt an extended conformation in solution, which is far from the structure observed in complexes with a model anti-MUC1 antibody. Herein, we propose a new strategy for designing potent antigen mimics based on modulating peptide/carbohydrate interactions by means of O â S/Se replacement at the glycosidic linkage. These minimal chemical modifications bring about two key structural changes to the glycopeptide. They increase the carbohydrate-peptide distance and change the orientation and dynamics of the glycosidic linkage. As a result, the peptide acquires a preorganized and optimal structure suited for antibody binding. Accordingly, these new glycopeptides display improved binding toward a representative anti-MUC1 antibody relative to the native antigens. To prove the potential of these glycopeptides as tumor-associated MUC1 antigen mimics, the derivative bearing the S-glycosidic linkage was conjugated to gold nanoparticles and tested as an immunogenic formulation in mice without any adjuvant, which resulted in a significant humoral immune response. Importantly, the mice antisera recognize cancer cells in biopsies of breast cancer patients with high selectivity. This finding demonstrates that the antibodies elicited against the mimetic antigen indeed recognize the naturally occurring antigen in its physiological context. Clinically, the exploitation of tumor-associated antigen mimics may contribute to the development of cancer vaccines and to the improvement of cancer diagnosis based on anti-MUC1 antibodies. The methodology presented here is of general interest for applications because it may be extended to modulate the affinity of biologically relevant glycopeptides toward their receptors.
Assuntos
Anticorpos Monoclonais/imunologia , Antígenos de Neoplasias/imunologia , Neoplasias da Mama/microbiologia , Carboidratos/imunologia , Glicopeptídeos/imunologia , Oxigênio/imunologia , Animais , Anticorpos Monoclonais/química , Neoplasias da Mama/patologia , Neoplasias da Mama/terapia , Carboidratos/química , Desenho de Fármacos , Feminino , Glicopeptídeos/química , Glicosídeos/química , Glicosídeos/imunologia , Glicosilação , Humanos , Neoplasias Mamárias Experimentais/imunologia , Neoplasias Mamárias Experimentais/patologia , Neoplasias Mamárias Experimentais/terapia , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Oxigênio/química , Selênio/química , Selênio/imunologia , Enxofre/química , Enxofre/imunologiaRESUMO
BACKGROUND: Recurrent tonsillitis might reduce the immunological capability of fighting against the infection of tonsil tissue. Polypodium leucotomos (Anapsos) immunomodulating effect has been subject of research in the last years. The aim of this research is to test the in vitro immunomodulating capacity of Anapsos in a child palatine tonsil explants model. METHODS: Palatine tonsils explants of children undergoing amigdalectomy were stimulated with mononuclear cells obtained from their own blood by density gradient centrifugation. Some were then treated with Anapsos while others rest untreated. Cytokines were measured by ELISA, immune cells activation was measured by flow cytometry and activation of immunoglobulins was appreciated by indirect immunofluorescence in tonsils tissue. RESULTS: Anapsos activates Natural Killers cells. It increases IL-2 and IFN-γ levels by the activation of Th2 lymphocytes, and IL-10, by the Th1 lymphocytes. Anapsos also increases immunoglobulins IgM, IgD and IgG4 by B-lymphocyte activation in tonsils tissue. CONCLUSION: Anapsos has an immunomodulating effect, both in humoral and cellular responses, which might benefit children suffering of recurrent tonsillitis as it could enhance their immune system. This effect might reduce the number of episodes suffered and therefore the number of children undergoing surgery.
Assuntos
Citocinas/metabolismo , Glicosídeos/imunologia , Imunoglobulinas/metabolismo , Fatores Imunológicos/uso terapêutico , Leucócitos Mononucleares/imunologia , Tonsila Palatina/efeitos dos fármacos , Tonsilite/tratamento farmacológico , Técnicas de Cultura de Células , Criança , Pré-Escolar , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Imunofluorescência , Humanos , Leucócitos Mononucleares/metabolismo , Tonsila Palatina/imunologia , Tonsila Palatina/metabolismo , Polypodium , Tonsilectomia , Tonsilite/imunologia , Tonsilite/cirurgiaRESUMO
Flavonoid-rich diets are expected to decrease the risk of cardiovascular diseases. The localization and target sites of flavonoids underlying the protective mechanism in vivo have not been fully investigated because the methods for detection of flavonoids have been limited to chemical analysis such as high-performance liquid chromatography. To further understand the actions of flavonoids in vivo, we developed a novel methodology that immunochemically evaluates flavonoids using specific antibodies. Quercetin-3-glucuronide (Q3GA), a major metabolite in human plasma, was coupled with keyhole limpet hemocyanin. Alternatively, the sugar moiety of quercetin-3-glucoside (Q3G) was succinylated and then coupled with a carrier protein. Using these two immunogens, we finally obtained two monoclonal antibodies, mAb14A2 and mAb11G6, from the immunogen using Q3GA and Q3G, respectively. Competitive enzyme-linked immunosorbent assay showed the unique difference in the specificity between the two similar antibodies: mAb14A2 recognized several quercetin-3-glycosides including Q3G and rutin but mAb11G6 was highly specific to the Q3G structure. The macrophage-derived foam cells in human atherosclerotic lesions were significantly stained with mAb14A2 but scarcely with mAb11G6. These results showed that the anti-flavonoid glycoside antibodies are useful tools for evaluating their localization in tissues and that the specificities strongly depend on the immunogen design for synthesizing the hapten-protein conjugates.
Assuntos
Anticorpos Monoclonais/química , Flavonoides/imunologia , Glicosídeos/imunologia , Animais , Aterosclerose/sangue , Aterosclerose/patologia , Ensaio de Imunoadsorção Enzimática , Células Espumosas/metabolismo , Células Espumosas/patologia , Glucuronídeos/sangue , Glucuronídeos/farmacologia , Hemocianinas/metabolismo , Humanos , Macrófagos/patologia , Moluscos , Quercetina/sangue , Quercetina/metabolismo , Sensibilidade e EspecificidadeRESUMO
OBJECTIVE: Several other immunosuppressive agents still need to be found for rejection as alternatives to Tacrolimus in lung transplantation. We tried to elucidate the treatment effect of Multiglycosidorum tripterygii on tracheal allografts in comparison to that of Tacrolimus. METHODS: Treatment effect of agents on tracheal allografts, undergoing incomplete immunosuppression for 12 weeks after transplantation, was investigated using a heterotopic rat tracheal transplantation model. Treatments with Tacrolimus (1.0 or 1.5mg/kg per day), Multiglycosidorum tripterygii (150 or 225mg/kg per day) and a combination of Tacrolimus (1.0mg/kg per day) and Multiglycosidorum tripterygii (150mg/kg per day) were applied as a therapy for allografts. Four weeks after administering this therapy, the effect of each treatment was investigated by the morphologic assessment of transplants. RESULTS: Treatment group with high doses of Multiglycosidorum tripterygii demonstrated a significantly better graft patency and lower cartilage dislocation than that without any treatment and tended to show better morphological findings than the other treatment groups, in addition to being safe. Some of allografts with high doses of Tacrolimus or Multiglycosidorum tripterygii therapy had a viable epithelium and viable tracheal glands in part, whereas the allografts with other treatments showed almost a completely denuded epithelium. High doses of Multiglycosidorum tripterygii therapy demonstrated less infiltration of mononuclear cells into the allografts, whereas other therapies showed a higher infiltration of such cells. CONCLUSIONS: We conclude that high doses of Multiglycosidorum tripterygii may be a useful alternative to Tacrolimus as an immunosuppressant for rat tracheal allografts.
Assuntos
Glicosídeos/uso terapêutico , Imunossupressores/uso terapêutico , Tacrolimo/uso terapêutico , Traqueia/transplante , Animais , Peso Corporal/imunologia , Cartilagem/imunologia , Cartilagem/patologia , Epitélio/imunologia , Epitélio/patologia , Glicosídeos/imunologia , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/prevenção & controle , Tolerância Imunológica/imunologia , Imunossupressores/imunologia , Masculino , Modelos Animais , Ratos , Ratos Endogâmicos BN , Ratos Endogâmicos Lew , Tacrolimo/imunologia , Traqueia/imunologia , Traqueia/patologiaRESUMO
We evaluate the ability of a Fasciola hepatica FABP native antigen (Fh12) with a new vaccination system called ADAD to protect mice and sheep against an experimental F. hepatica infection. The vaccination protocol consists of a set of two injections. The first injection contains a micelle in which two components are included, saponin from Quillaja saponaria (Qs) and/or Anapsos (A) a Polypodium leucotomos hydroalcoholic extract, both emulsified in a non-mineral oil (Montanide) in a water/oil emulsion (30/70). This is subcutaneously injected to achieve the "adaptation" of the immune system to subsequent stimuli. The second injection contains in addition the Fh12 antigen. Two different experiments were carried out using two mouse strains (BALB/c and CD-1). Mice vaccinated with Qs+A+Fh12 presented a survival rate of 40%, when compared with control groups. Furthermore, we evaluated the efficiency of the vaccination in sheep against an experimental F. hepatica challenge. The vaccinated sheep presented lower fluke recovery (24.5%), number of eggs in bile fluid (58.1%) and faeces (40.3%) than control groups. The recovered flukes were shorter (32.7%), immature (34.0%) and with lower body mass (31.6%) than non-complete vaccinated sheep. Thus, the new ADAD system could be a good alternative for future vaccination experiments against fasciolosis.
Assuntos
Adjuvantes Imunológicos , Proteínas de Transporte/imunologia , Fasciola hepatica/imunologia , Fasciolíase/veterinária , Imunização/veterinária , Doenças dos Ovinos/prevenção & controle , Animais , Anticorpos Anti-Helmínticos/biossíntese , Antígenos de Helmintos/imunologia , Emulsões , Fasciolíase/prevenção & controle , Proteínas de Ligação a Ácido Graxo , Feminino , Glicosídeos/imunologia , Imunização/métodos , Imunização/normas , Magnoliopsida , Camundongos , Camundongos Endogâmicos BALB C , Micelas , Extratos Vegetais/imunologia , Polypodium , Distribuição Aleatória , Saponinas/imunologia , Ovinos , Vacinação/veterináriaRESUMO
The immunomodulating effects of Anapsos, an aqueous hydrosoluble extract obtained from the rhizomes of the fern Polypodium leucotomos, on both pathogenicity and cytokine levels in serum (IFN-gamma/IL-4) were assayed in a Trichomonas vaginalis experimental model (BALB/c mice infected with 10(7) trichomonads and examined at day 15 after infection). Doses of 20 mg/kg/day administered for 10 days before the infection with the parasite induced a decrease of the experimental pathogenicity approximately 10-20% compared to controls. Gross histopathologic changes at abdominal organs and mortality rate, as a consequence of pathogenicity of the protozoa and the immune response of the host, were evaluated. IFN-gamma and IL-4 cytokines were determined on days -5, 0, 5, 10, and 15 postinfection by indirect ELISA. Treatment with PAL before infection modulates and downregulates the IFN-gamma concentration, while anticipates and upregulates the IL-4 level. The assays performed have showed the utility of the murine model of experimental trichomoniasis for the evaluation of immunomodulatory activity of synthetic or natural products.
Assuntos
Adjuvantes Imunológicos/farmacologia , Citocinas/biossíntese , Glicosídeos/farmacologia , Extratos Vegetais/farmacologia , Tricomoníase/imunologia , Trichomonas vaginalis/imunologia , Animais , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Glicosídeos/imunologia , Interferon gama/biossíntese , Interleucina-4/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Polypodium/química , Distribuição Aleatória , Fatores de Tempo , Tricomoníase/parasitologia , Tricomoníase/patologia , Trichomonas vaginalis/patogenicidadeRESUMO
The crystal structure of the verotoxin 1 (VT1) B subunit complexed with a globotriaosylceramide (Gb(3)) analogue showed the presence of three receptor binding sites per monomer. We wished to study the effects of altering the three sites, singly or in combination, on animal toxicity and cytokine induction in vitro. We found that while the site 1 and 2 mutants were modestly (two- to sevenfold) reduced in their ability to cause disease in BALB/c mice, the site 3 mutant, W34A, was as toxic as VT1. However, all the double-mutant proteins, irrespective of which two sites were mutated, exhibited approximately a 100-fold reduction in their 50% lethal doses for mice. These results suggest that multivalent receptor binding is important in vivo and that all three binding sites make a similar contribution to the latter process. The triple-mutant holotoxin, F30A G62T W34A, administered intraperitoneally without adjuvant, stimulated a strong antibody response in BALB/c mice, and the immune sera neutralized the activity of VT1 in vitro. Induction of tumor neurosis factor alpha release from differentiated human monocytes (THP-1 cells) was relatively impaired for site 1 and site 2 but not site 3 mutants, suggesting an auxiliary role for the latter site in mediation of cytokine release in vitro. Cytotoxicity assays on undifferentiated THP-1 cells have also demonstrated the importance of sites 1 and 2 and the relatively small role played by site 3 in causing cell death. These data suggest an association between the cytotoxicity of the protein and its ability to induce cytokine release.
Assuntos
Escherichia coli/patogenicidade , Toxinas Shiga/toxicidade , Fator de Necrose Tumoral alfa/biossíntese , Animais , Linhagem Celular , Feminino , Glicosídeos/imunologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Mutação , Subunidades Proteicas , Toxinas Shiga/imunologia , Toxinas Shiga/metabolismo , Triexosilceramidas/metabolismo , Triterpenos/imunologiaRESUMO
Human sera contain high levels of natural antibody (Ab) to Galalpha1-3Gal, a terminal glycosidic structure expressed on the surface of cells of mammals other than Old World primates. Incorporation of this determinant onto retroviral membranes by passage of viruses in cells encoding alpha-1-3-galactosyltransferase (GT) renders retroviruses sensitive to lysis by natural Ab and complement in normal human serum (NHS). Plasma membrane-budding viruses representing four additional virus groups were examined for their sensitivities to serum inactivation after passage through human cell lines that lack a functional GT or human cells expressing recombinant porcine GT. The inactivation of lymphocytic choriomeningitis virus (LCMV) by NHS directly correlated with host modification of the virus via expression of Galalpha1-3Gal and was blocked by incorporation of soluble Galalpha1-3Gal disaccharide into the inactivation assay. GT-deficient mice immunized to make high levels of Ab to Galalpha1-3Gal (anti-Gal Ab) were tested for resistance to LCMV passaged in GT-expressing cells. Resistance was not observed, but in vitro analyses of the mouse immune sera revealed that the antiviral activity of the sera was insufficient to eliminate LCMV infectivity on its natural targets of infection, macrophages, which express receptors for Ab and complement. Newcastle disease virus and vesicular stomatitis virus (VSV) were inactivated by NHS regardless of cell passage history, whereas Sindbis virus (SV) passaged in human cells resisted inactivation. Both VSV and SV passaged in Galalpha1-3Gal-expressing human cells incorporated this sugar moiety onto their major envelope glycoproteins. SV passaged in mouse cells expressing Galalpha1-3Gal was moderately sensitive to inactivation by NHS. These results indicate that enveloped viruses expressing Galalpha1-3Gal differ in their sensitivities to NHS and that a potent complement source, such as that in NHS, is required for efficient inactivation of sensitive viruses in vitro and in vivo.
Assuntos
Anticorpos Antivirais/imunologia , Formação de Anticorpos , Antígenos Virais/imunologia , Glicosídeos/imunologia , Herpesvirus Humano 1/imunologia , Vírus da Estomatite Vesicular Indiana/imunologia , Animais , Especificidade de Anticorpos , Linhagem Celular , Epitopos/imunologia , Humanos , Camundongos , Proteínas do Envelope Viral/imunologiaRESUMO
Analysis of cell surface glycosylation not only provides information about cell properties such as their state of differentiation or histogenetic lineage. The carbohydrate chains also provide potentially functional binding sites to endogenous carbohydrate-binding proteins. This interaction can elicit consequent signalling processes. Because of the importance of neutrophils in the host defence system, we monitored the effect of the binding of such sugar receptors to their cell surface on the release of the enzymatic activities of lysozyme, elastase, and myeloperoxidase. Besides the mannose-binding lectin concanavalin A and the immunomodulatory alpha/beta-galactoside-binding lectin from Viscum album L., three preparations of human sugar receptors - beta-galactoside-binding lectin (M(r) 14 kDa) and two affinity-purified polyclonal IgG fractions from serum with the capacity to recognize alpha- or beta-galactosides, respectively - were used. Two animal lectins from chicken liver and intestine that bind beta-galactosides, as well as the lectin-like human serum amyloid P component, were included in order to assess the importance of slight differences in ligand recognition. Cytochalasin B-enhanced enzyme release was invariably seen with the two plant lectins and the chicken liver beta-galactoside-binding lectin, but the related intestinal lectin did not increase enzyme release. The mammalian homologue of these avian lectins triggered lysozyme secretion, and the lactoside-binding IgG fraction enhanced the amount of extracellular elastase activity slightly but significantly. Thus, the actual lectin, not the nominal specificity of sugar receptors, is crucial for elucidation of responses. Due to the highly stimulatory activity of the two plant lectins, neutrophils from patients with non-cancerous diseases and from patients with lung cancer were monitored for the extent of lectin-mediated enzyme release. Only the concanavalin A-mediated reactivity of the neutrophils was associated with the type of disease.
Assuntos
Autoanticorpos/metabolismo , Lectinas/metabolismo , Neutrófilos/enzimologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Aglutininas/metabolismo , Metabolismo dos Carboidratos , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma de Células Pequenas/imunologia , Carcinoma de Células Pequenas/metabolismo , Concanavalina A/farmacologia , Feminino , Galactosídeos/imunologia , Galactosídeos/metabolismo , Glicosídeos/imunologia , Glicosídeos/metabolismo , Humanos , Imunoglobulina G/imunologia , Imunoglobulina G/isolamento & purificação , Elastase de Leucócito , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/metabolismo , Masculino , Pessoa de Meia-Idade , Erva-de-Passarinho/química , Muramidase/imunologia , Muramidase/metabolismo , Neutrófilos/metabolismo , Elastase Pancreática/imunologia , Elastase Pancreática/metabolismo , Peroxidase/imunologia , Peroxidase/metabolismo , Lectinas de Plantas , Proteínas de Plantas/metabolismo , Plantas Medicinais , Ligação Proteica/fisiologia , Componente Amiloide P Sérico/metabolismoRESUMO
Proprietary extract of mistletoe (Iscador) that has federal approval for clinical application can exhibit immunomodulatory capacity. However, the nature of this responsible substance has still remained elusive. To validate the hypothesis that specific lectin-carbohydrate interactions at least participate in eliciting immunomodulation, the modulatory efficiency of the major beta-galactoside-specific mistletoe lectin (ML I) from the clinically applied extract on selected immunological parameters was monitored "in vivo" in rabbits. Injections of nontoxic doses of the purified lectin or even only of its carbohydrate-binding subunit (0.25-1.0 ng/kg) into rabbits yielded significant increases in natural killer cytotoxicity, frequency of large granular lymphocytes, and phagocytic activity of granulocytes. In the clinically relevant situation, changes in these parameters were also determined in cancer patients after extract (Iscador) injection s.c. as well as i.v., emphasizing the potential relevance of the lectin. Comparative analyses of the changes in the selected parameters following injection of extract with normal lectin content as well as of extract, selectively depleted of lectin, into healthy volunteers corroborated this inference. Lectin depletion by affinity chromatography was highly specific and did not affect any other substance in the extract. Remarkably, contamination by endotoxin has been rigorously excluded in each applied specimen. These results encourage detailed elucidation of lectin action on various parts of the tumor defense system "in vitro" with the long range goal of achieving progress in the treatment of cancer through immunological strategies, exploring selective mediatory lectin-carbohydrate interactions.
Assuntos
Adjuvantes Imunológicos , Galactosídeos/imunologia , Glicosídeos/imunologia , Erva-de-Passarinho , Extratos Vegetais/farmacologia , Preparações de Plantas , Proteínas de Plantas , Plantas Medicinais , Toxinas Biológicas/farmacologia , Adulto , Idoso , Animais , Temperatura Corporal/efeitos dos fármacos , Neoplasias da Mama/imunologia , Citotoxicidade Imunológica , Feminino , Humanos , Células Matadoras Naturais/imunologia , Masculino , Pessoa de Meia-Idade , Erva-de-Passarinho/análise , Neutrófilos/imunologia , Fagocitose , Extratos Vegetais/análise , Coelhos , Proteínas Inativadoras de Ribossomos Tipo 2 , Toxinas Biológicas/isolamento & purificaçãoRESUMO
Expression of Gal alpha 1-3Gal cell surface residues has been correlated with the metastatic potential of murine tumor cells. We report that Gal alpha 1-3Gal residues are expressed at the cell surface of malignant human cancer cells, including four cell lines and 50% of the malignant breast specimens obtained by aspiration biopsy. In contrast, all benign breast biopsies and normal cells were Gal alpha 1-3Gal negative. Affinity-purified anti-alpha-galactosyl IgG (anti-Gal) antibody, which specifically recognizes Gal alpha 1-3Gal residues, significantly inhibited cell attachment in two in vitro assays thought to indicate tumor cell extravasation of the circulatory system during the metastatic process: attachment to perfused human umbilical vein endothelium, and attachment to isolated laminin. Since anti-Gal antibody is a natural component of all human sera, we propose that it may be part of the natural antitumor defense system in humans.
Assuntos
Antígenos Glicosídicos Associados a Tumores/imunologia , Galactosídeos/imunologia , Glicosídeos/imunologia , Células Tumorais Cultivadas/imunologia , Antígenos de Superfície/imunologia , Adesão Celular , Imunofluorescência , Humanos , Metástase Neoplásica , Células Tumorais Cultivadas/citologiaRESUMO
Specific multimer complexes of glycoproteins of enveloped viruses were prepared by treatment of suspensions of purified concentrated virus with a non-ionic detergent MECK mixed with 0.2% glycoside Quil A. Mixed complexes of glycoproteins with glycoside Quil A were formed upon removal of the detergent from the mixture of solubilized glycoproteins and glycoside Quil A. According to the results of electron microscopy, the formed complexes differed morphologically from the conventional micelles of glycoproteins and presented spherical reticular structures 20-30 nm in diameter. The immunogenic potency of the complexes was much higher than that of virus particles and micelles of purified glycoproteins and was comparable to the immunogenic potency of glycoproteins mixed with complete Freund adjuvant. The protective activity of the complex of protein G of rabies virus with glycoside Quil A was higher than that of the subunit rabies vaccine adsorbed on aluminium hydroxide.
Assuntos
Adjuvantes Imunológicos , Glicoproteínas/isolamento & purificação , Glicosídeos/farmacologia , Polímeros/isolamento & purificação , Saponinas/farmacologia , Proteínas do Envelope Viral/isolamento & purificação , Animais , Anticorpos Antivirais/análise , Relação Dose-Resposta Imunológica , Eletroforese em Gel de Poliacrilamida , Glicoproteínas/análise , Glicoproteínas/imunologia , Glicosídeos/imunologia , Imunização , Vírus da Influenza A/efeitos dos fármacos , Vírus da Influenza A/imunologia , Vírus da Influenza A/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C , Micelas , Vírus da Parainfluenza 1 Humana/efeitos dos fármacos , Vírus da Parainfluenza 1 Humana/imunologia , Vírus da Parainfluenza 1 Humana/isolamento & purificação , Polímeros/análise , Polímeros/imunologia , Saponinas de Quilaia , Vírus da Raiva/efeitos dos fármacos , Vírus da Raiva/imunologia , Vírus da Raiva/isolamento & purificação , Árvores , Proteínas do Envelope Viral/análise , Proteínas do Envelope Viral/imunologiaAssuntos
Glicolipídeos/metabolismo , Proteínas de Membrana/metabolismo , Fosfatidilinositóis/metabolismo , Glicolipídeos/imunologia , Glicosídeos/imunologia , Glicosídeos/metabolismo , Glicosilfosfatidilinositóis , Inositol/análise , Proteínas de Membrana/imunologia , Modelos Químicos , Peptídeos/metabolismo , Fosfatidilinositóis/imunologia , Conformação Proteica , Precursores de Proteínas/metabolismoRESUMO
The phenyl-beta-galactoside (phi-beta-gal)-specific monoclonal antibody (mAb) 49H.8 cross-reacts with the terminal disaccharide structure of the asialo GM1 (AGM1) molecule. It was found to react with phi-beta-gal determinants on murine and rat splenic natural killer (NK) cells, as measured by complement depletion studies. Flow cytometric analysis identified the antigen on two IL 2-dependent cloned murine NK cell lines and the rat large granular lymphocyte leukemia RNK. We have compared the 49H.8 reactivity to that of anti-AGM1 antisera (alpha-AGM1) on NK cells and a panel of NK related killer cells, including bone marrow-derived killer cells, lymphokine-activated killer cells (LAK), and anomalous killer cells (AK). We found that the 49H.8 specificity closely paralleled that of alpha-AGM1. When tested against Con A-reactive T cells, the 49H.8 mAb was less reactive than the alpha-AGM1, indicating that it may be a more specific marker for splenic NK populations than the alpha-AGM1.
Assuntos
Anticorpos Monoclonais/imunologia , Gangliosídeo G(M1) , Galactosídeos/imunologia , Glicosídeos/imunologia , Células Matadoras Naturais/imunologia , Animais , Anticorpos Monoclonais/fisiologia , Reações Antígeno-Anticorpo , Soro Antilinfocitário/imunologia , Células Clonais/imunologia , Concanavalina A/farmacologia , Epitopos/imunologia , Glicoesfingolipídeos/imunologia , Interleucina-2/fisiologia , Leucemia Experimental/imunologia , Depleção Linfocítica , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos A , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Endogâmicos DBA , Cavidade Peritoneal , Ratos , Ratos Endogâmicos F344 , Ratos Endogâmicos , Linfócitos T/imunologiaRESUMO
Hybridomas secreting monoclonal antibodies to Nippostrongylus brasiliensis antigens were generated by hybridization of IR983F myeloma cells with spleen cells from Lou/M/Wol rats infected with living third-stage larvae. Antibodies specific either for larval or worm antigens were identified by enzyme-linked immunosorbent assays with Nippostrongylus brasiliensis fragments, homogenates and secretions as antigens. The results demonstrate that all antibodies which recognized larval antigens (38 antibodies) also reacted with worm surfaces. Ten antibodies were specific only for worm antigens. Ten antibodies reacted with worm homogenate, three antibodies recognized components of worm secretion and 17 antibodies combined with acetylcholinesterase. The epitope specificity was investigated by the capacity of various glycosides, aminoacids, N-acetylneuraminic acid and phosphorylcholine to inhibit the binding to worm fragments. The analysis revealed that alpha-methylglucoside, alpha-methylmannoside, N-acetylglucosamine, N-acetylgalactosamine, fucose and the amino acids leucine, phenylalanine, tyrosine, serine, tryptophan did not combine with the antigen-binding sites of the antibodies. Proline, arginine and histidine, however, displayed inhibitory effects. With N-acetylneuraminic acid as inhibitor three groups of antibodies could be discriminated. At a concentration of 10-20 mM, phosphorylcholine was a potent inhibitor for all antibodies.
Assuntos
Anticorpos Monoclonais , Antígenos de Helmintos/imunologia , Nippostrongylus/imunologia , Acetilcolinesterase/imunologia , Aminoácidos/imunologia , Animais , Especificidade de Anticorpos , Antígenos de Helmintos/análise , Epitopos , Feminino , Glicosídeos/imunologia , Larva/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Nippostrongylus/crescimento & desenvolvimento , Nippostrongylus/metabolismo , Fosforilcolina/imunologia , Ratos , Ratos Endogâmicos , Ácidos Siálicos/imunologiaAssuntos
Antígenos de Neoplasias/imunologia , Epitopos , Neoplasias Experimentais/imunologia , Animais , Carboidratos/imunologia , Linhagem Celular , Testes de Fixação de Complemento , Citotoxicidade Imunológica , Imunofluorescência , Glicosídeos/imunologia , Masculino , Camundongos , Papaína , Ácido PeriódicoRESUMO
The dynamics of the titers and number of the antibody forming cells was studied within 26 hours after a single injection of anthracycline to immunized animals at various phases of the immune response. The specificity of the "rapid" effect of anthracyclines suggests that the low differentiated precursors of the antibody forming cells are the main target for the rubomycin effect and the immune competent cells in the state of multiplication and differentiation under the effect of the antigen are the main target for the carminomycin effect. The study was performed with noninbred mice immunized with sheep red cells.