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1.
Int J Mol Med ; 43(3): 1531-1541, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30664181

RESUMO

Dogs are a major source of indoor allergens. However, the prevalence of dog allergies in China remains unclear, especially in children. In the present study, Can f 7, a canine allergen belonging to the Niemann pick type C2 protein family, was selected to study its sensitization rate in Chinese children with dog allergies. The Can f 7 gene was subcloned into a pET­28a vector and expressed in Escherichia coli BL21 (DE3) cells. Recombinant Can f 7 was purified by nickel affinity chromatography, identified by SDS­PAGE electrophoresis, and had its allergenicity assessed by western blot, ELISA and basophil activation tests. Through a series of bioinformatical approaches, B­cell epitopes, secondary structures, and 3 dimensional (3D) homology modeling of Can f 7 were predicted. The activity of the B cell epitopes was verified by ELISA. The recombinant Can f 7 showed a distinct band with a molecular weight of 14 kDa. Six of 20 sera from dog­allergic children reacted positively to the Can f 7. Can f 7 induced an ~4.0­fold increase in cluster of differentiation 63 and C­C motif chemokine receptor R3 expression in basophils sensitized with the serum of dog­allergic children compared with those of non­allergic controls. The secondary structure analysis showed that Can f 7 contains 6 ß­sheets. Five B cell epitopes of Can f 7 were predicted, and two of these were confirmed by ELISA. These results indicate that Can f 7 is an important canine allergen in Chinese children and provide novel data for further research concerning the use of Can f 7 in the diagnosis and treatment of Chinese children with canine allergy symptoms.


Assuntos
Alérgenos/genética , Alérgenos/imunologia , Epitopos de Linfócito B/genética , Epitopos de Linfócito B/imunologia , Expressão Gênica , Lipocalinas/genética , Lipocalinas/imunologia , Adolescente , Alérgenos/química , Alérgenos/isolamento & purificação , Sequência de Aminoácidos , Animais , Criança , Pré-Escolar , Códon , Cães , Ensaio de Imunoadsorção Enzimática , Epitopos de Linfócito B/química , Feminino , Humanos , Hipersensibilidade/imunologia , Imunoglobulina E/imunologia , Lactente , Lipocalinas/química , Lipocalinas/isolamento & purificação , Masculino , Modelos Moleculares , Conformação Proteica , Proteínas Recombinantes
2.
Elife ; 72018 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-30398151

RESUMO

Stable mutualism between a host and its resident bacteria requires a moderated immune response to control bacterial population size without eliciting excessive inflammation that could harm both partners. Little is known about the specific molecular mechanisms utilized by bacterial mutualists to temper their hosts' responses and protect themselves from aggressive immune attack. Using a gnotobiotic larval zebrafish model, we identified an Aeromonas secreted immunomodulatory protein, AimA. AimA is required during colonization to prevent intestinal inflammation that simultaneously compromises both bacterial and host survival. Administration of exogenous AimA prevents excessive intestinal neutrophil accumulation and protects against septic shock in models of both bacterially and chemically induced intestinal inflammation. We determined the molecular structure of AimA, which revealed two related calycin-like domains with structural similarity to the mammalian immune modulatory protein, lipocalin-2. As a secreted bacterial protein required by both partners for optimal fitness, AimA is an exemplar bacterial mutualism factor.


Assuntos
Aeromonas/genética , Interações Hospedeiro-Patógeno/genética , Imunidade Inata/genética , Simbiose/genética , Animais , Interações Hospedeiro-Patógeno/imunologia , Intestinos/imunologia , Intestinos/microbiologia , Larva/imunologia , Larva/microbiologia , Lipocalinas/genética , Lipocalinas/imunologia , Domínios Proteicos/genética , Simbiose/imunologia , Peixe-Zebra/imunologia , Peixe-Zebra/microbiologia
4.
J Allergy Clin Immunol ; 142(4): 1113-1120.e9, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29852259

RESUMO

BACKGROUND: Sensitization to dog dander is an important risk factor for rhinoconjunctivitis and asthma but is not sufficient for diagnosing dog allergy. Molecular allergy diagnostics offer new opportunities for refined characterization. OBJECTIVES: We sought to study the association between sensitization to all presently known dog allergen components and clinical symptoms of dog allergy in children evaluated by using nasal provocation tests (NPTs). METHODS: Sixty children (age, 10-18 years) sensitized to dog dander extract underwent NPTs with dog dander extract. Measurement of IgE levels to dog dander and to Can f 1, Can f 2, Can f 3, and Can f 5 was performed with ImmunoCAP, and measurement of IgE levels to Can f 4 and Can f 6 was performed with streptavidin ImmunoCAP. An IgE level of 0.1 kUA/L or greater was considered positive. RESULTS: There was an association between sensitization to an increasing number of dog allergen components and a positive nasal challenge result (P = .01). Sensitization to lipocalins (odds ratio [OR], 6.0; 95% CI, 1.04-34.5), in particular Can f 4 (OR, 6.80; 95% CI 1.84-25.2) and Can f 6 (OR, 5.69; 95% CI, 1.59-20.8), was associated with a positive NPT result. Monosensitization to Can f 5 was related to a negative NPT result (OR, 5.78; 95% CI, 1.01-33.0). CONCLUSION: Sensitization to an increasing number of dog allergen components and to lipocalins is associated with dog allergy. Monosensitization to Can f 5 should not be regarded primarily as a marker for dog allergy.


Assuntos
Alérgenos/administração & dosagem , Alérgenos Animais/imunologia , Hipersensibilidade/diagnóstico , Lipocalinas/administração & dosagem , Antígeno Prostático Específico/administração & dosagem , Adolescente , Alérgenos/imunologia , Animais , Criança , Dessensibilização Imunológica , Cães , Feminino , Humanos , Hipersensibilidade/imunologia , Imunoglobulina E/imunologia , Lipocalinas/imunologia , Masculino , Antígeno Prostático Específico/imunologia
5.
Int Immunopharmacol ; 61: 126-131, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29859469

RESUMO

BACKGROUND: Increasing dairy consumption in China has been accompanied by rising incidence of milk allergy. Here we analyzed profiles of specific immunoglobulin E (sIgE) against cow's milk proteins, and assessed their value for milk allergy diagnosis among infants and young children from northern China. METHODS: Sera collected from 48 patients with milk allergy and 27 negative control subjects was analyzed by enzyme-linked immunosorbent assay to measure sIgE to α-lactalbumin (Bos d 4), ß-lactoglobulin (Bos d 5), α-casein (Bos d 9), ß-casein (Bos d 11), and κ-casein (Bos d 12). RESULTS: Among milk-allergic individuals, most were sensitized to at least one milk protein; about half were sensitized to Bos d 5, Bos d 9, Bos d 11 and Bos d 12, respectively, while few had positive serum sIgE against Bos d 4. Bos d 12 sIgE had the largest area under curve (AUC) (0.878; 95% CI, 0.800-0.957) and thus showed the best diagnostic performance in discriminating between milk-allergic and non-milk allergic patients, with a sensitivity of 92.6% and specificity of 72.9% using a statistically optimal cut-off value (OD450nm, 0.191). The combinations of Bos d 5 + Bos d 12 showed an AUC of 0.926, which was larger than for any individual components. CONCLUSIONS: Our results revealed inter-individual variation in the sensitization to different milk allergen component. Bos d 12 sIgE showed best performance in diagnosing milk allergy. Milk allergy diagnostic accuracy was further improved using combinations of milk allergen components by application of ROC curves based on logistic regression.


Assuntos
Alérgenos/imunologia , Lipocalinas/imunologia , Hipersensibilidade a Leite/diagnóstico , Animais , Bovinos , Pré-Escolar , China , Diagnóstico Diferencial , Feminino , Humanos , Imunoglobulina E/sangue , Lactente , Recém-Nascido , Masculino , Valor Preditivo dos Testes , Prognóstico , Padrões de Referência , Sensibilidade e Especificidade
6.
Sci Rep ; 8(1): 1598, 2018 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-29371615

RESUMO

The major cow's milk allergen Bos d 5 belongs to the lipocalin protein family, with an intramolecular pocket for hydrophobic ligands. We investigated whether Bos d 5 when loaded with the active vitamin A metabolite retinoic acid (RA), would elicit differential immune responses compared to the unloaded state. By in silico docking an affinity energy of -7.8 kcal/mol was calculated for RA into Bos d 5. Loading of RA to Bos d 5 could be achieved in vitro, as demonstrated by ANS displacement assay, but had no effect on serum IgE binding in tolerant or challenge-positive milk allergic children. Bioinformatic analysis revealed that RA binds to the immunodominant T-cell epitope region of Bos d 5. In accordance, Bos d 5 significantly suppressed the CD3+ CD4+ cell numbers, proliferative response and IL-10, IL-13 and IFN-γ secretion from stimulated human PBMCs only when complexed with RA. This phenomenon was neither associated with apoptosis of T-cells nor with the activation of Foxp3+ T-cells, but correlated likely with enhanced stability to lysosomal digestion due to a predicted overlap of Cathepsin S cleavage sites with the RA binding site. Taken together, proper loading of Bos d 5 with RA may suppress its immunogenicity and prevent its allergenicity.


Assuntos
Alérgenos/imunologia , Alérgenos/metabolismo , Epitopos de Linfócito T/metabolismo , Fatores Imunológicos/metabolismo , Lipocalinas/imunologia , Lipocalinas/metabolismo , Tretinoína/metabolismo , Animais , Bovinos , Proliferação de Células/efeitos dos fármacos , Humanos , Imunoglobulina E/metabolismo , Interferon gama/metabolismo , Interleucina-10/metabolismo , Interleucina-13/metabolismo , Leucócitos Mononucleares/imunologia , Lisossomos/metabolismo , Simulação de Acoplamento Molecular , Ligação Proteica , Proteólise
8.
Monoclon Antib Immunodiagn Immunother ; 36(4): 185-191, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28806153

RESUMO

Human lipocalin 6 (hLCN6) is a member of the lipocalin family, which is a group of structurally conserved hydrophobic ligand binding proteins, and widely distributed in animal, plant, and bacteria. Specific expression of hLCN6 in the epididymis and localization of this protein on the surface of spermatozoa suggest a role played by hLCN6, which may function as a transporter to carry ligands in the epididymal channel. However, the role of hLCN6 in sperm maturation has been largely unknown due to the lack of effective antibodies. In this study, we report the prokaryotic expression, purification, and refolding of recombinant hLCN6. Purified hLCN6 protein was used to generate monoclonal antibody (mAb) against this protein using conventional hybridoma techniques. The sensitivity and specificity of the anti-hLCN6 mAb were determined based on their activities in enzyme-linked immunosorbent assay and Western blotting analysis using various human tissues. The results showed that the antibody induced by recombinant hLCN6 protein had high sensitivity and specificity. Taken together, the recombinant hLCN6 protein and mAb against this protein obtained from our study provided useful tools for further exploration of the biological functions and molecular mechanism, as well as pathological significance of LCN6 in human.


Assuntos
Anticorpos Monoclonais Murinos/química , Lipocalinas/biossíntese , Animais , Anticorpos Monoclonais Murinos/imunologia , Anticorpos Monoclonais Murinos/isolamento & purificação , Especificidade de Anticorpos , Western Blotting , Epididimo/metabolismo , Escherichia coli , Feminino , Humanos , Imunoglobulina G/química , Imunoglobulina G/imunologia , Imunoglobulina G/isolamento & purificação , Lipocalinas/química , Lipocalinas/imunologia , Masculino , Camundongos Endogâmicos BALB C , Redobramento de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/imunologia
9.
Sci Rep ; 7: 43642, 2017 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-28276430

RESUMO

Quinolinic acid, a macrophage/microglia-derived excitotoxin fulfills a plethora of functions such as neurotoxin, gliotoxin, and proinflammatory mediator, and it alters the integrity and cohesion of the blood-brain barrier in several pathophysiological states. Beta-trace protein (BTP), a monomeric glycoprotein, is known to indicate cerebrospinal fluid leakage. Thus, the prior aim of this study was to investigate whether BTP might non-invasively indicate quinolinic acid-induced impaired blood-brain barrier integrity. The research hypotheses were tested in three subsamples with different states of immune activation (patients with HCV-infection and interferon-α, patients with major depression, and healthy controls). BTP has also been described as a sensitive marker in detecting impaired renal function. Thus, the renal function has been considered. Our study results revealed highest quinolinic acid and highest BTP- levels in the subsample of patients with HCV in comparison with the other subsamples with lower or no immune activation (quinolinic acid: F = 21.027, p < 0.001 [ANOVA]; BTP: F = 6.792, p < 0.01 [ANOVA]). In addition, a two-step hierarchical linear regression model showed that significant predictors of BTP levels are quinolinic acid, glomerular filtration rate and age. The neurotoxin quinolinic acid may impair blood-brain barrier integrity. BTP might be a new non-invasive biomarker to indicate quinolinic acid-induced impaired blood-brain barrier integrity.


Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Oxirredutases Intramoleculares/metabolismo , Lipocalinas/metabolismo , Ácido Quinolínico/efeitos adversos , Adulto , Biomarcadores , Barreira Hematoencefálica/imunologia , Feminino , Taxa de Filtração Glomerular , Humanos , Oxirredutases Intramoleculares/sangue , Oxirredutases Intramoleculares/imunologia , Lipocalinas/sangue , Lipocalinas/imunologia , Masculino , Pessoa de Meia-Idade , Neurotoxinas/efeitos adversos , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/tratamento farmacológico , Insuficiência Renal Crônica/etiologia , Insuficiência Renal Crônica/metabolismo
10.
Eur Ann Allergy Clin Immunol ; 49(2): 92-96, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28294591

RESUMO

Summary: It has been shown that allergen immunotherapy (AIT) is effective in reducing symptoms of allergic asthma and rhinitis. Data on the efficacy are less convincing with regard to AIT for allergens of common pets (cats/dogs). We describe a case of dog allergy in which we explored if dog AIT (DAI) could reduce a concomitant allergic sensitization to other allergens of furry animals. Our case demonstrates the efficacy of sublingual DAI on SPTs, symptom score, and spirometric responses despite persistent exposure to dog allergens at home in a patient sensitized, but not exposed, to several other furry animals. Moreover, this is the first report suggesting that DAI is able to reduce SPTs responses not only to dog, but also to other furry animals such as rabbit, horse, mouse, rat, hamster, cow. We recommend an accurate anamnesis and diagnosis of dog allergy before prescribing DAI. In particular, the use of ImmunoCAP ISAC is essential to verify the presence of IgE to lipocalins / albumins belonging to other furry animals. Obviously further studies carried out by using different DAI schedules, allergen amount and time of re-evaluation, laboratory procedure should be performed to confirm our findings.


Assuntos
Alérgenos/administração & dosagem , Asma/terapia , Cães/imunologia , Cabelo/imunologia , Imunoterapia Sublingual/métodos , Administração Sublingual , Adulto , Albuminas/imunologia , Alérgenos/imunologia , Animais , Asma/diagnóstico , Asma/imunologia , Biomarcadores/sangue , Humanos , Imunoglobulina E/sangue , Testes Intradérmicos , Lipocalinas/imunologia , Masculino , Especificidade da Espécie , Espirometria , Resultado do Tratamento
11.
Allergy ; 71(10): 1490-5, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-27289080

RESUMO

We investigated the prevalence of sensitization to the cat lipocalin Fel d 7 among 140 cat-sensitized Swedish patients and elucidated its allergenic activity and cross-reactivity with the dog lipocalin Can f 1. Sixty-five of 140 patients had IgE to rFel d 7 whereof 60 also had IgE to rCan f 1. A moderate correlation between IgE levels to rFel d 7 and rCan f 1 was found. rFel d 7 activated basophils in vitro and inhibited IgE binding to rCan f 1 in 4 of 13 patients, whereas rCan f 1 inhibited IgE binding to rFel d 7 in 7 of 13 patients. Fel d 7 and Can f 1 showed high similarities in protein structure and epitopes in common were found using cross-reactive antisera. Fel d 7 is a common allergen in a Swedish cat-sensitized population that cross-reacts with Can f 1, and may contribute to symptoms in cat- but also in dog-allergic patients.


Assuntos
Alérgenos/imunologia , Reações Cruzadas/imunologia , Lipocalinas/imunologia , Alérgenos/química , Animais , Basófilos/imunologia , Gatos , Cães , Epitopos/imunologia , Hipersensibilidade/sangue , Hipersensibilidade/imunologia , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Lipocalinas/química , Modelos Moleculares , Peptídeos/química , Peptídeos/imunologia , Conformação Proteica , Suécia
12.
Rev Alerg Mex ; 63(1): 1-10, 2016.
Artigo em Espanhol | MEDLINE | ID: mdl-26943824

RESUMO

BACKGROUND: Lipocalins seem to explain the cross-reactivity between some pets such as cat and dog. However, its role in other animals and its possible clinical impact in allergy diseases have been scarcely studied. OBJECTIVE: To analyze by bioinformatics techniques, the identity between lipocalin of some animals and to explore the clinical impact on allergic diseases. MATERIAL AND METHOD: An in silico study was done to search for lipocalin sequences using the BLAST program of NCBI Database. The protein sequences were aligned with CLUSTAL Omega UniProt version 1.2.1 software. The base sequences for alignments were lipocalins dogs and cats. The defined percentage identity was compared with the frequency of sensitization to animals exposed in a population of 284 patients with suspected allergic diseases. RESULTS: Identities between sequences were 10% to 70%. The highest values were found with Can f 6-Fel d 4 (68%) and Fel d 4-Equ c 1 (68%). The lower identity was found with lipocalin porpurin and retinol binding (<20%). We observed a relationship between sensitization and the percent identity between the species studied. CONCLUSION: Lipocalins as Can f 6, Fel de 4 and Equ c 1 seem to play an important role in the cross-reactivity to cat, horse and dog but not for the co-sensitization to hamster, cow or birds. Fel de 4 and Equ c 1 could be a prevalent allergen for horse and cat. These results come from predictive analysis and must be confirmed by in vitro and in vivo studies.


Assuntos
Alérgenos/imunologia , Simulação por Computador , Reações Cruzadas/imunologia , Hipersensibilidade/veterinária , Lipocalinas/imunologia , Alérgenos/genética , Animais , Gatos , Bovinos , Galinhas , Cricetinae , Reações Cruzadas/genética , Cães , Feminino , Cavalos , Humanos , Hipersensibilidade/genética , Hipersensibilidade/imunologia , Lipocalinas/genética , Análise de Sequência de DNA , Especificidade da Espécie
13.
J Immunol Methods ; 432: 82-6, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26899825

RESUMO

UNLABELLED: The distinction between acute infections of bacterial or viral causes is clinically important, but often very difficult even for experienced doctors. Previous studies indicated that serum measurements of HNL (Human Neutrophil Lipocalin) might be a superior diagnostic means in this regard, but also indicated that the antibody conformation of the HNL assay might have an impact on the diagnostic performance. The aim of the present report was to examine this further. METHODS: Several different (n=24) HNL ELISA assays were developed using different combinations of monoclonal and polyclonal HNL antibodies. Sera were collected from healthy persons (n=188) and from 155 patients with acute infections before any antibiotics treatment. The patients were diagnosed as having bacterial (n=69) or viral causes (n=86) of their infections. Plasma and serum were also examined by Western blotting using HNL-specific polyclonal antibodies. RESULTS: The optimal assay format for the distinction between bacterial and viral infection resulted in an area under the receiver operating characteristics curve (AuROC) for S-HNL of 0.98. (95% CI 0.94-1.00) as compared to 0.83 (0.76-0.88) for blood neutrophil counts and 0.69 (0.61-0.76) for S-CRP. Results also showed that different assay formats of HNL identified monomeric and dimeric HNL differently, the monomeric HNL being elevated in viral infections and the dimeric HNL being elevated in bacterial infections. CONCLUSION: We conclude that serum measurement of HNL is a superior diagnostic means to distinguish between acute infections caused by bacteria or virus. For optimal clinical performance the immunoassay should address conformational epitopes in the dimeric HNL.


Assuntos
Infecções Bacterianas/diagnóstico , Ensaio de Imunoadsorção Enzimática , Lipocalinas/sangue , Viroses/diagnóstico , Adulto , Idoso , Anticorpos Monoclonais/imunologia , Área Sob a Curva , Infecções Bacterianas/sangue , Infecções Bacterianas/microbiologia , Biomarcadores/sangue , Western Blotting , Estudos de Casos e Controles , Diagnóstico Diferencial , Epitopos , Feminino , Humanos , Lipocalinas/imunologia , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Multimerização Proteica , Curva ROC , Regulação para Cima , Viroses/sangue , Viroses/virologia , Adulto Jovem
14.
Am J Reprod Immunol ; 75(2): 126-33, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26773532

RESUMO

PROBLEM: A wide variety of mediators are involved in inflammatory processes. However, the identity of those participating in vaginal immune responses has not been established. We correlated extracellular matrix metalloproteinase inducer (EMMPRIN), matrix metalloproteinase-8 (MMP-8), hyaluronan (HA), hyaluronidase-1 (Hyal-1), human ß-defensin-2 (hBD2), and neutrophil gelatinase-associated lipocalin (NGAL) concentrations with the extent of leukocyte infiltration into the vagina and suggest their participation in vaginal inflammation. METHODS OF STUDY: Vaginal fluid was obtained from 233 women seen at the outpatient clinic in the Department of Obstetrics and Gynecology at Campinas University, Brazil. The magnitude of vaginal inflammation was determined by the leukocyte count on vaginal smears and categorized as no inflammation (0 leukocytes/field), moderate inflammation (1-4 leukocytes/field), and intense inflammation (>4 leukocytes/field). Concentrations of EMMPRIN, MMP-8, HA, Hyal-1, hBD2, and NGAL were determined in vaginal fluid by ELISA. RESULTS: EMMPRIN, MMP-8, HA, hBD2, and NGAL concentration increased with elevated leukocyte numbers (P < 0.05), while Hyal-1 did not. EMMPRIN concentrations were correlated with HA and MMP-8 levels. CONCLUSION: EMMPRIN, MMP-8, HA, ß-defensin, and NGAL are elevated in women with vaginal inflammation.


Assuntos
Proteínas da Fase Aguda/imunologia , Basigina/imunologia , Ácido Hialurônico/imunologia , Lipocalinas/imunologia , Metaloproteinase 8 da Matriz/imunologia , Proteínas Proto-Oncogênicas/imunologia , Vaginite/imunologia , beta-Defensinas/imunologia , Adulto , Feminino , Humanos , Hialuronoglucosaminidase/imunologia , Leucócitos/imunologia , Lipocalina-2 , Adulto Jovem
15.
Allergy ; 71(3): 286-94, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26497994

RESUMO

Owners and their domestic animals via skin shedding and secretions, mutually exchange microbiomes, potential pathogens and innate immune molecules. Among the latter especially lipocalins are multifaceted: they may have an immunomodulatory function and, furthermore, they represent one of the most important animal allergen families. The amino acid identities, as well as their structures by superposition modeling were compared among human lipocalins, hLCN1 and hLCN2, and most important animal lipocalin allergens, such as Can f 1, Can f 2 and Can f 4 from dog, Fel d 4 from cats, Bos d 5 from cow's milk, Equ c 1 from horses, and Mus m 1 from mice, all of them representing major allergens. The ß-barrel fold with a central molecular pocket is similar among human and animal lipocalins. Thereby, lipocalins are able to transport a variety of biological ligands in their highly conserved calyx-like cavity, among them siderophores with the strongest known capability to complex iron (Fe(3+) ). Levels of human lipocalins are elevated in nonallergic inflammation and cancer, associated with innate immunoregulatory functions that critically depend on ligand load. Accordingly, deficient loading of lipocalin allergens establishes their capacity to induce Th2 hypersensitivity. Our similarity analysis of human and mammalian lipocalins highlights their function in innate immunity and allergy.


Assuntos
Alérgenos/química , Alérgenos/imunologia , Hipersensibilidade/imunologia , Imunidade Inata , Lipocalinas/química , Lipocalinas/imunologia , Conformação Proteica , Alérgenos/metabolismo , Animais , Humanos , Hipersensibilidade/metabolismo , Tolerância Imunológica , Imunoglobulina E/imunologia , Imunomodulação , Lipocalinas/metabolismo , Relação Estrutura-Atividade , Células Th2/imunologia , Células Th2/metabolismo
16.
Int J Cancer ; 138(5): 1269-80, 2016 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-26421425

RESUMO

The standard of care for diagnosis and therapy monitoring of gliomas is magnetic resonance imaging (MRI), which however, provides only an indirect and incomplete representation of the tumor mass, offers limited information for patient stratification according to WHO-grades and may insufficiently indicate tumor relapse after antiangiogenic therapy. Anticalins are alternative binding proteins obtained via combinatorial protein design from the human lipocalin scaffold that offer novel diagnostic reagents for histology and imaging applications. Here, the Anticalins N7A, N7E and N9B, which possess exquisite specificity and affinity for oncofetal fibronectin carrying the extra domain B (ED-B), a well-known proangiogenic extracellular matrix protein, were applied for immunohistochemical studies. When investigating ED-B expression in biopsies from 41 patients with confirmed gliomas of WHO grades I to IV, or in non-neoplastic brain samples, we found that Anticalins specifically detect ED-B in primary glioblastoma multiforme (GBM; WHO IV) but not in tumors of lower histopathological grade or in tumor-free brain. In primary GBM samples, ED-B specific Anticalins locate to fibronectin-rich perivascular areas that are associated with angiogenesis. Anticalins specifically detect ED-B both in fixed tumor specimen and on vital cells, as evidenced by cytofluorometry. Beyond that, we labeled an Anticalin with the γ-emitter (123) I and demonstrated specific binding to GBM-tissue samples using in vitro autoradiography. Overall, our data indicate that ED-B specific Anticalins are useful tools for the diagnosis of primary GBM and related angiogenic sites, presenting them as promising tracers for molecular tumor imaging.


Assuntos
Anticorpos/metabolismo , Neoplasias Encefálicas/diagnóstico , Fibronectinas/análise , Glioblastoma/diagnóstico , Lipocalinas/imunologia , Imagem Molecular/métodos , Neoplasias Encefálicas/química , Linhagem Celular Tumoral , Fibronectinas/metabolismo , Glioblastoma/química , Humanos , Imuno-Histoquímica , Lipocalinas/metabolismo , Biblioteca de Peptídeos , Molécula-1 de Adesão Celular Endotelial a Plaquetas/análise , Ligação Proteica , Estrutura Terciária de Proteína
17.
Eur Ann Allergy Clin Immunol ; 47(5): 163-7, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26357003

RESUMO

Although this highly refined diagnostic approach has been used in several fields of allergy diagnosis, we noticed the scarcity of data on the role of CDR in detecting current sensitization to the allergens of common pets (cat / dog) and, especially, its potential usefulness in predicting the risk of sensitization to other furry animals. Reported data suggest that cross-reacting mechanisms might play an important role in a significant proportion of allergic sensitizations to furry animals (common pets and unusual / exotic mammals) especially in the absence of any possible direct / indirect contact. In this context an evaluation of specific IgE by using the micro-array technique ImmunoCAP ISAC (Thermofisher Scientific - Immuno-Diagnostics, Sweden) for lipocalins (Can f 1, Can f 2, Equ c 1, Fel d 4, Mus m 1) and albumins (Bos d 6, Can f 3, Equ c 3, Fel d 2) might be very useful to evaluate the possibility of cross-reactions between the allergens of different animals. In fact, allergic sensitization without animal exposure is a relevant risk for patients, because they are not aware about the possibility that even severe respiratory symptoms may develop after an occasional animal contact. This aspect should be taken into account by susceptible individuals before acquiring new pets, after removal of common pets or beginning a contact for working / leisure activity with a common as well as uncommon animal.


Assuntos
Gatos/imunologia , Cães/imunologia , Hipersensibilidade/diagnóstico , Animais de Estimação/imunologia , Animais , Humanos , Lipocalinas/imunologia , Risco , Albumina Sérica/fisiologia
18.
Immunity ; 43(3): 475-87, 2015 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-26320658

RESUMO

Interleukin-17 (IL-17) induces pathology in autoimmunity and infections; therefore, constraint of this pathway is an essential component of its regulation. We demonstrate that the signaling intermediate MCPIP1 (also termed Regnase-1, encoded by Zc3h12a) is a feedback inhibitor of IL-17 receptor signal transduction. MCPIP1 knockdown enhanced IL-17-mediated signaling, requiring MCPIP1's endoribonuclease but not deubiquitinase domain. MCPIP1 haploinsufficient mice showed enhanced resistance to disseminated Candida albicans infection, which was reversed in an Il17ra(-/-) background. Conversely, IL-17-dependent pathology in Zc3h12a(+/-) mice was exacerbated in both EAE and pulmonary inflammation. MCPIP1 degraded Il6 mRNA directly but only modestly downregulated the IL-6 promoter. However, MCPIP1 strongly inhibited the Lcn2 promoter by regulating the mRNA stability of Nfkbiz, encoding the IκBζ transcription factor. Unexpectedly, MCPIP1 degraded Il17ra and Il17rc mRNA, independently of the 3' UTR. The cumulative impact of MCPIP1 on IL-6, IκBζ, and possibly IL-17R subunits results in a biologically relevant inhibition of IL-17 signaling.


Assuntos
Inflamação/imunologia , Interleucina-17/imunologia , Ribonucleases/imunologia , Transdução de Sinais/imunologia , Proteínas da Fase Aguda/genética , Proteínas da Fase Aguda/imunologia , Proteínas da Fase Aguda/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Candida albicans/imunologia , Candida albicans/fisiologia , Candidíase/genética , Candidíase/imunologia , Candidíase/microbiologia , Linhagem Celular , Células Cultivadas , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Feminino , Células HEK293 , Interações Hospedeiro-Patógeno/imunologia , Humanos , Immunoblotting , Inflamação/genética , Inflamação/metabolismo , Interleucina-17/metabolismo , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-6/metabolismo , Lipocalina-2 , Lipocalinas/genética , Lipocalinas/imunologia , Lipocalinas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Nucleares/genética , Proteínas Nucleares/imunologia , Proteínas Nucleares/metabolismo , Proteínas Oncogênicas/genética , Proteínas Oncogênicas/imunologia , Proteínas Oncogênicas/metabolismo , Pneumonia/genética , Pneumonia/imunologia , Pneumonia/metabolismo , Receptores de Interleucina-17/genética , Receptores de Interleucina-17/imunologia , Receptores de Interleucina-17/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ribonucleases/genética , Ribonucleases/metabolismo
19.
Eur J Immunol ; 45(11): 3073-86, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26332507

RESUMO

Lipocalin-2 (Lcn2) is an innate immune peptide with pleiotropic effects. Lcn2 binds iron-laden bacterial siderophores, chemo-attracts neutrophils and has immunomodulatory and apoptosis-regulating effects. In this study, we show that upon infection with Salmonella enterica serovar Typhimurium, Lcn2 promotes iron export from Salmonella-infected macrophages, which reduces cellular iron content and enhances the generation of pro-inflammatory cytokines. Lcn2 represses IL-10 production while augmenting Nos2, TNF-α, and IL-6 expression. Lcn2(-/-) macrophages have elevated IL-10 levels as a consequence of increased iron content. The crucial role of Lcn-2/IL-10 interactions was further demonstrated by the greater ability of Lcn2(-/-) IL-10(-/-) macrophages and mice to control intracellular Salmonella proliferation in comparison to Lcn2(-/-) counterparts. Overexpression of the iron exporter ferroportin-1 in Lcn2(-/-) macrophages represses IL-10 and restores TNF-α and IL-6 production to the levels found in wild-type macrophages, so that killing and clearance of intracellular Salmonella is promoted. Our observations suggest that Lcn2 promotes host resistance to Salmonella Typhimurium infection by binding bacterial siderophores and suppressing IL-10 production, and that both functions are linked to its ability to shuttle iron from macrophages.


Assuntos
Proteínas da Fase Aguda/imunologia , Homeostase/imunologia , Ferro/metabolismo , Lipocalinas/imunologia , Macrófagos/metabolismo , Proteínas Oncogênicas/imunologia , Salmonelose Animal/imunologia , Proteínas da Fase Aguda/metabolismo , Animais , Western Blotting , Lipocalina-2 , Lipocalinas/metabolismo , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Oncogênicas/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Salmonelose Animal/metabolismo , Salmonella typhimurium , Transfecção
20.
Sci Rep ; 5: 13841, 2015 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-26346541

RESUMO

Lipocalins are one of the most important groups of inhalant animal allergens. The analysis of structural features of these proteins is important to get insights into their allergenicity. We have determined two different dimeric crystal structures for bovine dander lipocalin Bos d 2, which was earlier described as a monomeric allergen. The crystal structure analysis of all other determined lipocalin allergens also revealed oligomeric structures which broadly utilize inherent structural features of the ß-sheet in dimer formation. According to the moderate size of monomer-monomer interfaces, most of these dimers would be transient in solution. Native mass spectrometry was employed to characterize quantitatively transient dimerization of two lipocalin allergens, Bos d 2 and Bos d 5, in solution.


Assuntos
Alérgenos/química , Lipocalinas/química , Multimerização Proteica , Alérgenos/imunologia , Lipocalinas/imunologia , Espectrometria de Massas , Modelos Moleculares , Conformação Proteica
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