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1.
Front Immunol ; 12: 632482, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34276644

RESUMO

Recent evidence suggests that inflammation was participated in the pathogenesis of PD, thus, to understand the potential mechanism of gut microbiota in the pathogenesis of Parkinson's disease (PD), we performed a metagenomic analysis of fecal samples from PD patient and controls. Using a two-stage metagenome-wide association strategy, fecal DNA samples from 69 PD patients and 244 controls in three groups (comprising 66 spouses, 97 age-matched, and 81 normal samples, respectively) were analyzed, and differences between candidate gut microbiota and microbiota-associated epitopes (MEs) were compared. In the study, 27 candidate bacterial biomarkers and twenty-eight candidate epitope peptides were significantly different between the PD patients and control groups. Further, enriched 4 and 13 MEs in PD were positively associated with abnormal inflammatory indicators [neutrophil percentage (NEUT.1), monocyte count/percentage (MONO/MONO.1), white blood cell count (WBC)] and five candidate bacterial biomarkers (c_Actinobacteria, f_Bifidobacteriaceae, g_Bifidobacterium, o_Bifidobacteriales, p_Actinobacteria) from Actinobacteria phylum, and they were also positively associated with histidine degradation and proline biosynthesis pathways, respectively. Additionally, enriched 2 MEs and 1 ME in PD were positively associated with above inflammatory indicators and two bacteria (f_Lactobacillaceae, g_Lactobacillus) from Firmicutes phylum, and they were also positively associated with pyruvate fermentation to propanoate I and negatively associated with isopropanol biosynthesis, respectively. Of these MEs, two MEs from GROEL2, RPSC were derived from Mycobacterium tuberculosis, triggered the T cell immune response, as previously reported. Additionally, other candidate epitope peptides derived from Mycobacterium tuberculosis and Mycobacterium leprae may also have potential immune effects in PD. In all, the altered MEs in PD may relate to abnormalities in immunity and glutamate and propionate metabolism, which furthers our understanding of the pathogenesis of PD.


Assuntos
Actinobacteria/imunologia , Epitopos/imunologia , Firmicutes/imunologia , Doença de Parkinson/microbiologia , Actinobacteria/classificação , Actinobacteria/genética , Actinobacteria/metabolismo , Idoso , Biomarcadores , Vias Biossintéticas , Citocinas/sangue , Fezes/microbiologia , Feminino , Firmicutes/classificação , Firmicutes/genética , Firmicutes/metabolismo , Microbioma Gastrointestinal/genética , Microbioma Gastrointestinal/imunologia , Humanos , Inflamação , Masculino , Pessoa de Meia-Idade , Doença de Parkinson/imunologia
2.
J Org Chem ; 85(16): 10973-10979, 2020 08 21.
Artigo em Inglês | MEDLINE | ID: mdl-32806098

RESUMO

PGL-1 epitope 1 bearing a p-aminoethylphenol group was efficiently synthesized by using linear synthetic routes. A method for efficient synthesis of oligosaccharides containing rhamnose rings was developed. The chemistry is flexible and could be used for the synthesis of other PGLs antigens. A biotinylated PGL-1 antigen 23 was synthesized and could be used as a probe for early detection of leprosy.


Assuntos
Glicolipídeos , Mycobacterium leprae , Anticorpos Antibacterianos , Antígenos de Bactérias , Antígenos de Superfície , Epitopos , Trissacarídeos
3.
Biosens Bioelectron ; 145: 111698, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31539652

RESUMO

Mycobacterium leprae causes endemic disease leprosy which becomes chronic if not treated timely. To expedite this 'timely diagnosis', and that also at an early stage, here an attempt is made to fabricate an epitope-imprinted sensor. A molecularly imprinted polymer nanoparticles modified electrochemical quartz crystal microbalance sensor was developed for sensing of Mycobacterium leprae bacteria through its epitope sequence. Multiple monomers, 3-sulphopropyl methacrylate potassium salt, benzyl methacrylate and 4-aminothiophenol were utilized to imprint this bacterial epitope. Imprinted nanoparticles were electropolymerized on gold coated quartz electrode. The sensor was able to show specific binding towards the blood samples of infected patients, even in the presence of 'matrix' and other plasma proteins such as albumin and globulin. Even other peptide sequences, similar to epitope sequences only with two amino acid mismatches were also unable to show any binding. Sensor withstood analytical tests viz. selectivity, specificity, matrix effect, detection limit (0.161 nM), quantification limit (and 0.536 nM), reproducibility (RSD 2.01%). Hence a diagnostic tool for bacterium causing leprosy is successfully fabricated in a facile manner which will broaden the clinical access and efficient population screening can be made feasible.


Assuntos
Técnicas Biossensoriais , Hanseníase/diagnóstico , Mycobacterium leprae/isolamento & purificação , Técnicas de Microbalança de Cristal de Quartzo , Epitopos/química , Epitopos/imunologia , Ouro/química , Humanos , Hanseníase/microbiologia , Impressão Molecular , Mycobacterium leprae/imunologia , Mycobacterium leprae/patogenicidade , Nanopartículas/química , Polímeros/química
4.
Sci Rep ; 8(1): 11407, 2018 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-30061618

RESUMO

More than 100 counties, mainly in southwest China, report incidence rates of leprosy >1/100,000. The current study analysed the epidemiology of leprosy in southwest China to improve our understanding of the transmission pattern and improve control programs. 207 counties were selected in southwest China. Leprosy patients and their household contacts were recruited. The data from the medical interview and the serological antileprosy antibody of the leprosy patients were analysed. A total of 2,353 new cases of leprosy were interviewed. The distribution of leprosy patients was partly associated with local natural and economic conditions, especially several pocket areas. A total of 53 from 6643 household contacts developed leprosy, and the incidence rate of leprosy in the household contacts was 364/100,000 person-years. We found that NDO-BSA attained higher positive rates than MMP-II and LID-1 regardless of clinical types, disability and infection time in leprosy patients. By means of combination of antigens, 88.4% patients of multibacillary leprosy were detected, in contrast to 59.9% in paucibacillary leprosy. Household contacts should be given close attention for the early diagnosis, disruption of disease transmission and precise control. Applications of serology for multi-antigens were recommended for effective coverage and monitoring in leprosy control.


Assuntos
Hanseníase/diagnóstico , Adolescente , Adulto , Formação de Anticorpos/imunologia , Antígenos de Bactérias/imunologia , China/epidemiologia , Epitopos/imunologia , Feminino , Geografia , Humanos , Incidência , Hanseníase/economia , Hanseníase/epidemiologia , Hanseníase/imunologia , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Fatores Socioeconômicos , Adulto Jovem
5.
BMC Infect Dis ; 13: 42, 2013 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-23351151

RESUMO

BACKGROUND: An early diagnostic test for detecting infection in leprosy is fundamental for reducing patients' sequelae. The currently used lepromin is not adequate for disease diagnosis and, so far, no antigen to be used in intradermoreaction has proved to be sensitive and specific for that purpose. Aiming at identifying new reagents to be used in skin tests, candidate antigens were investigated. METHODS: Random peptide phage display libraries were screened by using antibodies from leprosy patients in order to identify peptides as diagnostic reagents. RESULTS: Seven different phage clones were identified using purified antibodies pooled from sera of leprosy patients. When the clones were tested with serum samples by ELISA, three of them, 5A, 6A and 1B, allowed detecting a larger number of leprosy patients when compared to controls. The corresponding peptides expressed by selected phage clones were chemically synthesized. A pilot study was undertaken to assess the use of peptides in skin tests. The intradermal challenge with peptides in animals previously sensitized with Mycobacterium leprae induced a delayed-type hypersensitivity with peptide 5A (2/5) and peptide 1B (1/5). In positive controls, there was a 3/5 reactivity for lepromin and a 4/5 reactivity of the sensitized animals with soluble extract of M. leprae. CONCLUSIONS: The preliminary data suggest that may be possible to develop reagents with diagnostic potential based on peptide mimotopes selected by phage display using polyclonal human antibodies.


Assuntos
Antígenos de Bactérias/imunologia , Hanseníase/diagnóstico , Mycobacterium leprae/imunologia , Animais , Técnicas de Visualização da Superfície Celular , Epitopos/imunologia , Feminino , Cobaias , Humanos , Hipersensibilidade Tardia/imunologia , Antígeno de Mitsuda/imunologia , Biblioteca de Peptídeos , Peptídeos/imunologia
6.
PLoS Negl Trop Dis ; 6(4): e1616, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22545169

RESUMO

During recent years, comparative genomic analysis has allowed the identification of Mycobacterium leprae-specific genes with potential application for the diagnosis of leprosy. In a previous study, 58 synthetic peptides derived from these sequences were tested for their ability to induce production of IFN-γ in PBMC from endemic controls (EC) with unknown exposure to M. leprae, household contacts of leprosy patients and patients, indicating the potential of these synthetic peptides for the diagnosis of sub- or preclinical forms of leprosy. In the present study, the patterns of IFN-γ release of the individuals exposed or non-exposed to M. leprae were compared using an Artificial Neural Network algorithm, and the most promising M. leprae peptides for the identification of exposed people were selected. This subset of M. leprae-specific peptides allowed the differentiation of groups of individuals from sites hyperendemic for leprosy versus those from areas with lower level detection rates. A progressive reduction in the IFN-γ levels in response to the peptides was seen when contacts of multibacillary (MB) patients were compared to other less exposed groups, suggesting a down modulation of IFN-γ production with an increase in bacillary load or exposure to M. leprae. The data generated indicate that an IFN-γ assay based on these peptides applied individually or as a pool can be used as a new tool for predicting the magnitude of M. leprae transmission in a given population.


Assuntos
Antígenos de Bactérias , Epitopos/imunologia , Testes de Liberação de Interferon-gama/métodos , Hanseníase/diagnóstico , Hanseníase/transmissão , Mycobacterium leprae/imunologia , Adulto , Idoso , Antígenos de Bactérias/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Redes Neurais de Computação
7.
Clin Vaccine Immunol ; 17(2): 298-303, 2010 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20016045

RESUMO

Despite the reduction in the number of leprosy cases registered worldwide as a result of the widespread use of multidrug therapy, the number of new cases detected each year remains stable in many countries. This indicates that Mycobacterium leprae, the causative agent of leprosy, is still being transmitted and that, without an earlier diagnosis, transmission will continue and infection will remain a health problem. The current means of diagnosis of leprosy is based on the appearance of clinical symptoms, which in many cases occur after significant and irreversible nerve damage has occurred. Our recent work identified several recombinant antigens that are specifically recognized by leprosy patients. The goal of the present study was to produce and validate the reactivity of a chimeric fusion protein that possesses the antibody binding properties of several of these proteins. The availability of such a chimeric fusion protein will simplify future test development and reduce production costs. We first identified the antibody binding regions within our top five antigen candidates by performing enzyme-linked immunosorbent assays with overlapping peptides representing the amino acid sequences of each protein. Having identified these regions, we generated a fusion construct of these components (protein advances diagnostic of leprosy [PADL]) and demonstrated that the PADL protein retains the antibody reactivity of the component antigens. PADL was able to complement a protein that we previously produced (the leprosy IDRI [Infectious Disease Research Institute] diagnostic 1 [LID-1] protein) to permit the improved diagnosis of multibacillary leprosy and that had a good ability to discriminate patients with multibacillary leprosy from control individuals. A serological diagnostic test consisting of these antigens could be applied within leprosy control programs to reduce transmission and to limit the appearance of leprosy-associated disabilities and stigmatizing deformities by directing treatment.


Assuntos
Antígenos de Bactérias , Técnicas de Laboratório Clínico/métodos , Hanseníase/diagnóstico , Proteínas Recombinantes de Fusão , Adolescente , Adulto , Idoso , Animais , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Mapeamento de Epitopos , Epitopos/genética , Epitopos/imunologia , Feminino , Humanos , Imunoensaio/métodos , Masculino , Pessoa de Meia-Idade , Mycobacterium leprae/genética , Mycobacterium leprae/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Adulto Jovem
8.
AIDS Res Hum Retroviruses ; 24(7): 941-6, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18593340

RESUMO

In many countries, HIV testing among tuberculosis (TB) patients is recommended so that both infections are appropriately treated. Cross-reacting antibodies to HIV antigens have been reported for several conditions, including TB, leprosy, malaria, and rheumatoid arthritis. To study the pattern and prevalence of cross-reacting antibodies to HIV antigens, we examined sera from 153 HIV-negative TB patients and 40 healthy individuals in Chennai, south India. We also studied the differences in cross-reactivity of various HIV antigens using two different Western blot kits. Of the 153 samples studied, 80 were tested using HIV Western blot and 73 were tested using INNOLIA. Most patients in the study had concordantly negative ELISA and rapid tests, and no subject had a positive Western blot. However, seven TB patients had antibodies that cross-reacted with HIV antigens, giving rise to an indeterminate result. While p51/55 was the most frequently recognized antigen in the Western blot assay, antibodies to sgp120 was most frequently identified in INNOLIA. Sequence similarities between the two organisms could be responsible for eliciting cross-reacting antibodies, since a few related epitopes were identified in HIV and Mycobacterium. These findings could have potential implications for the development of diagnostics and vaccines.


Assuntos
Anticorpos Antibacterianos/sangue , Anticorpos Antibacterianos/imunologia , Epitopos/genética , Antígenos HIV/imunologia , HIV/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculose/sangue , Western Blotting , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Antígenos HIV/genética , Proteína gp120 do Envelope de HIV/imunologia , Transcriptase Reversa do HIV/imunologia , Humanos , Mycobacterium tuberculosis/genética , Precursores de Proteínas/imunologia , Kit de Reagentes para Diagnóstico , Análise de Sequência de Proteína
9.
Artigo em Inglês | MEDLINE | ID: mdl-17050928

RESUMO

BACKGROUND: Current evidence suggests that lichen planus is an immunological disease. Cytotoxic CD8+ cells in the lesional epidermis recognize a unique antigen called lichen planus specific antigen. This antigen could be demonstrated by indirect immunofluorescence using the patient's serum and autologous lesional skin. AIM: To study indirect immunofluorescence pattern in lichen planus, among Indian patients. METHODS: Twenty-five consecutive patients with the clinical diagnosis of lichen planus were enrolled in the study. Direct immunofluorescence was done in all patients. Indirect immunofluorescence using lesional skin as substrate was done in all 25 patients and five patients with other dermatoses. RESULTS: A specific fluorescence pattern corresponding to the distribution of lichen planus specific antigen was observed in the stratum spinosum and granulosum in 22 (88%) patients. It was absent from other parts of the epidermis, dermis and in patients with other dermatoses. CONCLUSION: Indirect immunofluorescence is a useful adjuvant test in lichen planus, particularly in atypical cases.


Assuntos
Antígenos/análise , Epitopos/análise , Técnica Indireta de Fluorescência para Anticorpo/métodos , Líquen Plano/imunologia , Adolescente , Adulto , Antígenos/imunologia , Epitopos/imunologia , Feminino , Humanos , Líquen Plano/diagnóstico , Líquen Plano/patologia , Masculino , Pessoa de Meia-Idade
10.
Immunol Lett ; 88(1): 71-6, 2003 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-12853165

RESUMO

The objective of the study was to identify Mycobacterium leprae-specific immunogenic peptides for the development of a skin test reagent. Such a reagent is required for the detection of M. leprae infection and possibly for the diagnosis of patients with active leprosy. For this purpose, we analyzed the in vitro responses of human peripheral blood mononuclear cell (PBMCs) to peptides from the 35 kDa protein of M. leprae. This protein is of interest since it has no homologue within the Mycobacterium tuberculosis complex, although it has a homologue in Mycobacterium avium. The subjects enrolled in the study were paucibacillary (PB) and multibacillary (MB) leprosy patients, healthy contacts, and non-contacts. Seventy-three PB and 124 MB leprosy patients were recruited from four leprosy clinics in Thailand. Fifty-seven healthy contacts were household contacts. Twenty non-leprosy contacts had no family history of or exposure to leprosy. PBMCs from individuals were tested for stimulation with 12 overlapping peptides from the M. leprae 35 kDa protein using the lymphocyte proliferation assay. These peptides were located in four areas containing three to six residues which were distinct for the M. leprae product in comparison to that from M. avium. Four peptides (p60-76, p132-151, p206-224 and p267-286), which were the most permissive from each region and recognized by non-contacts with significantly lower frequencies than other subject groups, were identified. From this preliminary result, we conclude that these four peptides were likely to be M. leprae-specific.


Assuntos
Proteínas de Bactérias/imunologia , Epitopos , Antígenos HLA-DR/genética , Hanseníase/imunologia , Mycobacterium leprae/imunologia , Peptídeos/imunologia , Adulto , Proteínas de Bactérias/química , Feminino , Genes MHC da Classe II , Antígenos HLA-DR/imunologia , Humanos , Hanseníase/diagnóstico , Ativação Linfocitária , Masculino , Dados de Sequência Molecular , Peso Molecular , Mycobacterium avium/imunologia , Peptídeos/síntese química , Peptídeos/química , Especificidade da Espécie , Tailândia
11.
J Immunol ; 169(9): 5300-7, 2002 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-12391250

RESUMO

Microbial heat shock proteins (hsp) have been associated with the generation and induction of Th1-type immune responses. We tested the effects of treatment with five different microbial hsp (Mycobacterium leprae, Streptococcus pneumoniae, Helicobacter pylori, bacillus Calmette-Guérin, and Mycobacterium tuberculosis) in a murine model of allergic airway inflammation and airway hyperresponsiveness (AHR). Mice were sensitized to OVA by i.p. injection and then challenged by OVA inhalation. Hsp were administered to each group by i.p. injection before sensitization and challenge. Sensitized and challenged mice developed increased serum levels of OVA-specific IgE with significant airway eosinophilia and heightened responsiveness to methacholine when compared with nonsensitized animals. Administration of M. leprae hsp prevented both development of AHR as well as bronchoalveolar lavage fluid eosinophilia in a dose-dependent manner. Treatment with M. leprae hsp also resulted in suppression of IL-4 and IL-5 production in bronchoalveolar lavage fluid, while IL-10 and IFN-gamma production were increased. Furthermore, M. leprae hsp treatment significantly suppressed OVA-specific IgE production and goblet cell hyperplasia/mucin hyperproduction. In contrast, treatment with the other hsp failed to prevent changes in airway responsiveness, lung eosinophilia, or cytokine production. Depletion of gamma/delta T lymphocytes before sensitization and challenge abolished the effect of M. leprae hsp treatment on AHR. These results indicate selective and distinctive properties among the hsp, and that M. leprae hsp may have a potential therapeutic role in the treatment of allergic airway inflammation and altered airway function.


Assuntos
Proteínas de Bactérias/farmacologia , Hiper-Reatividade Brônquica/imunologia , Hiper-Reatividade Brônquica/microbiologia , Proteínas de Choque Térmico/farmacologia , Pulmão/patologia , Animais , Brônquios/imunologia , Brônquios/microbiologia , Brônquios/patologia , Hiper-Reatividade Brônquica/prevenção & controle , Líquido da Lavagem Broncoalveolar/imunologia , Líquido da Lavagem Broncoalveolar/microbiologia , Movimento Celular/imunologia , Citocinas/antagonistas & inibidores , Citocinas/biossíntese , Regulação para Baixo/imunologia , Epitopos/biossíntese , Feminino , Células Caliciformes/imunologia , Células Caliciformes/patologia , Hiperplasia , Imunoglobulina E/biossíntese , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Inflamação/imunologia , Inflamação/microbiologia , Interleucina-4/antagonistas & inibidores , Interleucina-4/biossíntese , Interleucina-5/antagonistas & inibidores , Interleucina-5/biossíntese , Pulmão/imunologia , Pulmão/microbiologia , Linfonodos/imunologia , Linfonodos/metabolismo , Depleção Linfocítica , Camundongos , Mucinas/antagonistas & inibidores , Mucinas/biossíntese , Mycobacterium leprae/fisiologia , Ovalbumina/farmacologia , Eosinofilia Pulmonar/microbiologia , Eosinofilia Pulmonar/prevenção & controle , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Subpopulações de Linfócitos T/imunologia
12.
Microbiology (Reading) ; 148(Pt 10): 3049-3057, 2002 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12368438

RESUMO

mAb CS-35 is representative of a large group of antibodies with similar binding specificities that were generated against the Mycobacterium leprae lipopolysaccharide, lipoarabinomannan (LAM), and which cross-reacted extensively with LAMs from Mycobacterium tuberculosis and other mycobacteria. That this antibody also cross-reacts with the arabinogalactan (AG) of the mycobacterial cell wall, suggesting that it recognizes a common arabinofuranosyl (Araf)-containing sequence in AG and LAM, is demonstrated. The antibody reacted more avidly with 'AraLAM' (LAM with naked Araf termini) compared to 'ManLAM' (in which many Araf termini are capped with mannose residues) and mycolylarabinogalactan-peptidoglycan complex (in which the terminal Araf units are substituted with mycolic acids). Neither did the antibody bind to AG from emb knock-out mutants deficient in the branched hexa-Araf termini of AG. These results indicate that the terminal Araf residues of mycobacterial arabinan are essential for binding. Competitive ELISA using synthetic oligosaccharides showed that the branched hexa-Araf methyl glycoside [beta-D-Araf-(1-->2)-alpha-D-Araf-(1-)(2)-(3 and 5)-alpha-D-Araf-(1-->5)-alpha-D-Araf-OCH(3)] was the best competitor among those tested. The related linear methyl glycoside, beta-D-Araf-(1-->2)-alpha-D-Araf-(1-->5)-alpha-D-Araf-(1-->5)-alpha-D-Araf-OCH(3), representing one linear segment of the branched hexa-Araf, was less effective and the other linear tetrasaccharide, beta-D-Araf-(1-->2)-alpha-D-Araf-(1-->3)-alpha-D-Araf-(1-->5)-alpha-D-Araf-OCH(3), was ineffective. The combined results suggest that the minimal epitope recognized by antibody CS-35 encompasses the beta-D-Araf-(1-->2)-alpha-D-Araf-(1-->5)-alpha-D-Araf-(1-->5)-alpha-D-Araf within the branched hexa-Araf motif of mycobacterial arabinans, whether present in LAM or AG.


Assuntos
Anticorpos Antibacterianos/imunologia , Arabinose/análogos & derivados , Epitopos/química , Lipopolissacarídeos/química , Lipopolissacarídeos/imunologia , Polissacarídeos/química , Anticorpos Monoclonais/imunologia , Arabinose/química , Configuração de Carboidratos , Sequência de Carboidratos , Mapeamento de Epitopos , Galactanos/química , Galactanos/imunologia , Mycobacterium smegmatis/genética , Mycobacterium smegmatis/imunologia , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/imunologia
13.
Immunol Cell Biol ; 80(6): 574-83, 2002 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-12406392

RESUMO

The 65 kilodalton heat shock protein (Hsp65) from mycobacterial species elicits immune responses and in some cases protective immunity. Here we have used a DNA sublibrary approach to identify antigenic fragments of Mycobacterium avium Hsp65 and a synthetic peptide approach to delineate CD4+ T cell determinants. A panel of Hsp65 reactive CD4+ T cell clones was established from lymph node cells obtained from BALB/c mice immunized with recombinant Hsp65. The clones were tested for proliferative reactivity against the products of the DNA sublibrary of the hsp65 gene. A T cell epitope, restricted by the I-Ad molecule, was identified within the C-terminal region of Hsp65 and the minimal epitope (amino acid residues 489-503) delineated using overlapping peptides spanning the C-terminal fragment. Additionally, the CD4+ T cell clone recognizing this epitope also responded to native Hsp65 present in M. avium lysates by both proliferation and cytokine production, indicating that the epitope was present and processed similarly both in the native and the recombinant forms of Hsp65. This sequence identified in BALB/c mice (Hsp65 489-503) is identical in other mycobacteria, notably M. tuberculosis, M. bovis and M. leprae, suggesting the epitope may have wider application in murine models of other mycobacterial infections.


Assuntos
Proteínas de Bactérias , Chaperoninas/metabolismo , Antígenos de Histocompatibilidade Classe II/metabolismo , Mycobacterium avium/metabolismo , Peptídeos/metabolismo , Motivos de Aminoácidos , Sequência de Aminoácidos , Linfócitos T CD4-Positivos/imunologia , Chaperonina 60 , Chaperoninas/genética , Chaperoninas/imunologia , Epitopos , Dados de Sequência Molecular , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia
14.
Immunopharmacol Immunotoxicol ; 24(2): 255-63, 2002 May.
Artigo em Inglês | MEDLINE | ID: mdl-12066851

RESUMO

Diagnosis of tuberculosis a problem, specially in the regions harboring an abundance of both pathogenic and non-pathogenic mycobacteria. This study was undertaken to assess in such a situation the predictive value of proliferative T cell response to a peptide epitope ('38G') of the 38 kDa membrane protein of Mycobacterium tuberculosis. 3[H]-thymidine incorporation assays were done with peripheral blood mononuclear cells of tuberculoid leprosy and pulmonary tuberculosis patients. The donors were also classified as PPD responders (Stimulation Index, SI> 3) or non-responders (SI < or = 3) on the basis of their T cell response to the 'Purified Protein Derivative (PPD)' of M. tuberculosis. 38G peptide was used in either free or liposome-associated form prepared by the technique of 'Dehydration-rehydration Vesicles' (Kirby and Gregoriadis, 1984). While free peptide failed to induce a positive response in study subjects, its liposomal form was T cell stimulatory and distinguished, to certain extent, between PPD responders (corresponding SI > 3 in 54% subjects) and non-responders (SI > 3 in 29% subjects). However, it did not differentiate between leprosy and tuberculosis. The study supports use of liposomes as adjuvant vehicles for antigenic peptides designed to activate human T cells.


Assuntos
Antígenos de Bactérias/administração & dosagem , Mycobacterium tuberculosis/imunologia , Linfócitos T/imunologia , Sequência de Aminoácidos , Antígenos de Bactérias/química , Antígenos de Bactérias/genética , Diagnóstico Diferencial , Epitopos/administração & dosagem , Epitopos/química , Epitopos/genética , Humanos , Técnicas In Vitro , Hanseníase Tuberculoide/diagnóstico , Hanseníase Tuberculoide/imunologia , Lipossomos , Ativação Linfocitária , Dados de Sequência Molecular , Peso Molecular , Mycobacterium tuberculosis/química , Mycobacterium tuberculosis/genética , Teste Tuberculínico , Tuberculose Pulmonar/diagnóstico , Tuberculose Pulmonar/imunologia
15.
Thromb Haemost ; 87(4): 599-605, 2002 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-12008941

RESUMO

Anticardiolipin (ACA), anti-beta2 glycoprotein I (beta2GPI), and antiprothrombin antibodies of IgG and IgM classes were quantitated by enzyme-linked immunosorbent assays in 176 untreated leprosy patients across the histopathological spectrum. Positivity rates ranged from 21% (IgG ACA) to 30% (IgM anti-prothrombin) versus 4% in healthy controls (p <10(-2) to 10(-3)). Levels of IgM anti-beta2GPI and IgG ACA were significantly higher in lepromatous leprosy and multibacillary patient subgroups. IgG3 was the most common subclass reactive to both beta2GPI and prothrombin in selected high-titer leprosy sera, unlike antibodies from patients with the antiphospholipid syndrome (APS) largely restricted to IgG2. In leprosy patients, but not in the APS control group, there was no statistical correlation between ACA and anti-beta2GPI antibody levels. Likewise, a large fraction of anti-beta2GPI positive sera (36/45 and 28/44 for IgG and IgM, respectively) were unreactive in the standard ACA assay. Most assayed anti-beta2GPI antibodies from leprosy patients showed (i) ability to recognize both human and bovine beta2GPI immobilized on non-irradiated polystyrene plates, (ii) concentration-dependent inhibition of binding by cardiolipin, and (iii) relatively high avidity binding to fluid-phase beta2GPI, thereby differing from those found in APS. Finally, the location of the major epitopic region on the beta2GPI molecule targeted by autoantibodies was different in leprosy and APS, as assessed by direct binding to domain I- and V-deleted mutants and competition with the mouse monoclonal antibody 8C3, directed at domain I. Thus, leprosy-related antiphospholipid antibodies comprise persistent IgG and IgM anti-beta2GPI that differ from APS-related ones with respect to IgG subclass, avidity and epitope specificity, possibly reflecting distinct pathophysiological significance.


Assuntos
Síndrome Antifosfolipídica/imunologia , Autoanticorpos/imunologia , Glicoproteínas/imunologia , Hanseníase/imunologia , Adolescente , Adulto , Anticorpos Anticardiolipina/imunologia , Afinidade de Anticorpos , Especificidade de Anticorpos , Síndrome Antifosfolipídica/complicações , Ensaio de Imunoadsorção Enzimática , Epitopos/imunologia , Feminino , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Masculino , Pessoa de Meia-Idade , Protrombina/imunologia , Senegal , Trombose/etiologia , Trombose/imunologia , beta 2-Glicoproteína I
16.
s.l; s.n; 2002. 9 p. graf.
Não convencional em Inglês | SES-SP, SES-SP, HANSEN, HANSENIASE, SESSP-ILSLACERVO, SES-SP | ID: biblio-1240981

RESUMO

Diagnosis of tuberculosis a problem, specially in the regions harboring an abundance of both pathogenic and non-pathogenic mycobacteria. This study was undertaken to assess in such a situation the predictive value of proliferative T cell response to a peptide epitope ('38G') of the 38 kDa membrane protein of Mycobacterium tuberculosis. 3[H]-thymidine incorporation assays were done with peripheral blood mononuclear cells of tuberculoid leprosy and pulmonary tuberculosis patients. The donors were also classified as PPD responders (Stimulation Index, SI> 3) or non-responders (SI 3 in 54 per cent subjects) and non-responders (SI > 3 in 29 per cent subjects). However, it did not differentiate between leprosy and tuberculosis. The study supports use of liposomes as adjuvant vehicles for antigenic peptides designed to activate human T cells.


Assuntos
Humanos , Antígenos de Bactérias/administração & dosagem , Antígenos de Bactérias/genética , Antígenos de Bactérias/química , Ativação Linfocitária , Dados de Sequência Molecular , Diagnóstico Diferencial , Epitopos/administração & dosagem , Epitopos/genética , Epitopos/química , Hanseníase Tuberculoide/diagnóstico , Hanseníase Tuberculoide/imunologia , Linfócitos T/imunologia , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/química , Sequência de Aminoácidos , Teste Tuberculínico , Tuberculose Pulmonar/diagnóstico , Tuberculose Pulmonar/imunologia , Lipossomos , Peso Molecular
17.
Vaccine ; 20(5-6): 731-6, 2001 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-11738736

RESUMO

Expression vectors containing rabies virus nucleoprotein B-cell and T-cell epitopes in Mycobacterium bovis BCG were constructed. The epitopes were subcloned into the M. leprae 18-kDa gene to ensure correct presentation to the host immune system. Expression of the 18-kDa::B+T epitope fusion protein was driven by either the hsp60 promoter, which is constitutively activated at a high level in M. bovis BCG, or the 18-kDa promoter, which is strongly induced in vivo. Mice were immunised intra-peritoneally with the recombinant BCG cultures and compared to a control group vaccinated with the commercial rabies vaccine Rai-SAD. Both of the expression vectors elicited a higher antibody titre than that of the rabies vaccine, with the highest response shown by M. bovis BCG (pUP203), expression controlled by the 18-kDa promoter. Immunisation with M. bovis BCG (pUP202), expression controlled by the hsp60 promoter, resulted in a continuously increasing antibody titre up to 60 days post immunisation. The mice antibodies were also capable of recognising the whole rabies virus and not only the synthetic peptide epitopes.


Assuntos
Antígenos Virais/genética , Mycobacterium bovis/genética , Mycobacterium bovis/imunologia , Nucleocapsídeo/genética , Nucleocapsídeo/imunologia , Vírus da Raiva/genética , Vírus da Raiva/imunologia , Animais , Anticorpos Antivirais/biossíntese , Linfócitos B/imunologia , Sequência de Bases , Epitopos/genética , Expressão Gênica , Vetores Genéticos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas do Nucleocapsídeo , Plasmídeos/genética , Vacinas Antirrábicas/genética , Vacinas Antirrábicas/imunologia , Linfócitos T/imunologia , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia
19.
Microbiology (Reading) ; 147(Pt 6): 1557-1564, 2001 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-11390686

RESUMO

Expression of a gene encoding a novel protein antigen of 40 kDa (p40) was detected in IS901(+) strains of Mycobacterium avium, but not in any other species or subspecies of Mycobacterium tested, including IS901(-) M. avium and the other members of the M. avium complex. Although Southern hybridization revealed that the p40 gene is widely distributed within the genus, expression of the antigen could not be detected on Western blots of mycobacterial cell lysates. Nucleotide sequence analysis of the cloned p40 gene, and a database search, revealed high levels of sequence identity with a homologous gene in IS901(-) M. avium, M. avium subsp. paratuberculosis, Mycobacterium bovis, Mycobacterium leprae, Mycobacterium smegmatis and Mycobacterium tuberculosis. Further analysis of upstream sequences identified a putative promoter region. The p40 gene is the first example of a gene that is widely distributed within the genus Mycobacterium but expressed only in association with the presence of a genomic insertion element, in this case IS901, in strains of M. avium isolated from birds and domestic livestock.


Assuntos
Antígenos de Bactérias/genética , Elementos de DNA Transponíveis , Genes Bacterianos , Mycobacterium avium/genética , Animais , Antígenos de Bactérias/metabolismo , Sequência de Bases , Southern Blotting , Western Blotting , Cromatografia Líquida de Alta Pressão , Sequência Conservada , Epitopos , Regulação Bacteriana da Expressão Gênica , Humanos , Dados de Sequência Molecular , Mycobacterium avium/metabolismo , Reação em Cadeia da Polimerase , Análise de Sequência de DNA , Homologia de Sequência do Ácido Nucleico
20.
Mol Microbiol ; 39(1): 89-99, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11123691

RESUMO

Mycobacterium tuberculosis, the causative agent of tuberculosis, produces a heparin-binding haemagglutinin adhesin (HBHA), which is involved in its epithelial adherence. To ascertain whether HBHA is also present in fast-growing mycobacteria, Mycobacterium smegmatis was studied using anti-HBHA monoclonal antibodies (mAbs). A cross-reactive protein was detected by immunoblotting of M. smegmatis whole-cell lysates. However, the M. tuberculosis HBHA-encoding gene failed to hybridize with M. smegmatis chromosomal DNA in Southern blot analyses. The M. smegmatis protein recognized by the anti-HBHA mAbs was purified by heparin-Sepharose chromatography, and its amino-terminal sequence was found to be identical to that of the previously described histone-like protein, indicating that M. smegmatis does not produce HBHA. Biochemical analysis of the M. smegmatis histone-like protein shows that it is glycosylated like HBHA. Immunoelectron microscopy demonstrated that the M. smegmatis protein is present on the mycobacterial surface, a cellular localization inconsistent with a histone-like function, but compatible with an adhesin activity. In vitro protein interaction assays showed that this glycoprotein binds to laminin, a major component of basement membranes. Therefore, the protein was called M. smegmatis laminin-binding protein (MS-LBP). MS-LBP does not appear to be involved in adherence in the absence of laminin but is responsible for the laminin-mediated mycobacterial adherence to human pneumocytes and macrophages. Homologous laminin-binding adhesins are also produced by virulent mycobacteria such as M. tuberculosis and Mycobacterium leprae, suggesting that this adherence mechanism may contribute to the pathogenesis of mycobacterial diseases.


Assuntos
Mycobacterium smegmatis/imunologia , Mycobacterium tuberculosis/imunologia , Receptores de Laminina/imunologia , Anticorpos Antibacterianos/imunologia , Aderência Bacteriana , Compartimento Celular , Clonagem Molecular , Reações Cruzadas , Epitopos , Escherichia coli/genética , Genes Bacterianos , Glicosilação , Histonas/genética , Histonas/imunologia , Histonas/isolamento & purificação , Macrófagos/microbiologia , Microscopia Imunoeletrônica , Infecções por Mycobacterium/etiologia , Alvéolos Pulmonares/microbiologia , Receptores de Laminina/genética , Receptores de Laminina/isolamento & purificação
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