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1.
Arerugi ; 68(7): 851-856, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31406081

RESUMO

BACKGROUND: The present study aimed to assess the safety of a three-level stepwise oral food challenge (OFC) in patients with peanut allergy. METHODS: This retrospective analysis reviewed laboratory test results and food allergy histories (obtained from medical records) of participants who underwent OFC using peanut butter for the first time at Miyagi Children's Hospital between January 2015 and December 2017. Total food dose was classified as follows: step 1, 0.1 g peanut butter; step 2, 0.5g peanut butter; and step 3, 3g peanut butter. RESULTS: A total of 114 participants were included. OFC-positive rates were 52% (step 1), 35% (step 2), and 20% (step 3). Of the 41 OFC-positive participants, 1 required an intramuscular adrenaline injection during step 3. Peanut allergy history rates and peanut- and Ara h 2-specific immunoglobulin E (sIgE) titers were significantly higher during the low-dose step. CONCLUSION: The three-level stepwise OFC with peanut butter is safe for patients with peanut allergy, as indicated by peanut- and Ara h 2-sIgE titers.


Assuntos
Alimentos , Hipersensibilidade a Amendoim/diagnóstico , Albuminas 2S de Plantas/imunologia , Antígenos de Plantas/imunologia , Arachis , Criança , Glicoproteínas/imunologia , Humanos , Imunoglobulina E/sangue , Estudos Retrospectivos
2.
Vet Parasitol ; 271: 45-50, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31303202

RESUMO

Cryptosporidium parvum is a protozoan parasite of the phylum Apicomplexa responsible for cryptosporidiosis in calves, a disease that causes significant diarrhea and impairs gain of body weight, generating important production losses. As to now, no effective drugs or vaccines are available for the treatment or prevention of bovine cryptosporidiosis. Several reports suggest that development of a vaccine to prevent cryptosporidiosis is feasible, but relatively few vaccine candidates have been characterized and tested. The most prominent C. parvum antigen is gp60, an O-glycosylated mucin-like protein tethered to the parasite membrane by a glycosylphosphatidylinositol (GPI) anchor. Gp60 has been shown to be involved in essential mechanisms for the survival of C. parvum, such as recognition, adhesion to, and invasion of host cells. This work was aimed at expressing gp60 in Tetrahymena thermophila, a ciliated protozoon with numerous advantages for the heterologous expression of eukaryotic proteins, as a first approach for the development of a recombinant vaccine for bovine cryptosporidiosis. T. thermophila-expressed gp60 localized to the protozoon cell surface and oral apparatus, and partitioned into the Triton X-114 detergent phase. This indicates that the protein entered the reticuloendothelial system of the ciliate, and suggests it contains a GPI-anchor. Homogenates of gp60-expressing T. thermophila cells were recognized by sera from calves naturally infected with C. parvum demonstrating their immunoreactivity. In summary, the heterologous expression of gp60, a C. parvum-encoded GPI-anchored protein, has been successfully demonstrated in the ciliate T. thermophila.


Assuntos
Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Glicoproteínas/genética , Glicoproteínas/imunologia , Tetrahymena thermophila/genética , Animais , Bovinos , Doenças dos Bovinos/imunologia , Doenças dos Bovinos/prevenção & controle , Criptosporidiose/imunologia , Criptosporidiose/prevenção & controle , Cryptosporidium parvum/genética , Vacinas Sintéticas/sangue , Vacinas Sintéticas/genética
3.
PLoS Pathog ; 15(6): e1007896, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31233555

RESUMO

Streptococcus gordonii and Streptococcus sanguinis are primary colonizers of the tooth surface. Although generally non-pathogenic in the oral environment, they are a frequent cause of infective endocarditis. Both streptococcal species express a serine-rich repeat surface adhesin that mediates attachment to sialylated glycans on mucin-like glycoproteins, but the specific sialoglycan structures recognized can vary from strain to strain. Previous studies have shown that sialoglycan binding is clearly important for aortic valve infections caused by some S. gordonii, but this process did not contribute to the virulence of a strain of S. sanguinis. However, these streptococci can bind to different subsets of sialoglycan structures. Here we generated isogenic strains of S. gordonii that differ only in the type and range of sialoglycan structures to which they adhere and examined whether this rendered them more or less virulent in a rat model of endocarditis. The findings indicate that the recognition of specific sialoglycans can either enhance or diminish pathogenicity. Binding to sialyllactosamine reduces the initial colonization of mechanically-damaged aortic valves, whereas binding to the closely-related trisaccharide sialyl T-antigen promotes higher bacterial densities in valve tissue 72 hours later. A surprising finding was that the initial attachment of streptococci to aortic valves was inversely proportional to the affinity of each strain for platelets, suggesting that binding to platelets circulating in the blood may divert bacteria away from the endocardial surface. Importantly, we found that human and rat platelet GPIbα (the major receptor for S. gordonii and S. sanguinis on platelets) display similar O-glycan structures, comprised mainly of a di-sialylated core 2 hexasaccharide, although the rat GPIbα has a more heterogenous composition of modified sialic acids. The combined results suggest that streptococcal interaction with a minor O-glycan on GPIbα may be more important than the over-all affinity for GPIbα for pathogenic effects.


Assuntos
Endocardite Bacteriana/imunologia , Glicoproteínas/imunologia , Ácidos Siálicos/imunologia , Infecções Estreptocócicas/imunologia , Streptococcus gordonii/imunologia , Streptococcus sanguis/imunologia , Animais , Modelos Animais de Doenças , Endocardite Bacteriana/patologia , Feminino , Humanos , Masculino , Ratos , Ratos Sprague-Dawley , Índice de Gravidade de Doença , Infecções Estreptocócicas/patologia , Streptococcus gordonii/patogenicidade , Streptococcus sanguis/patogenicidade
4.
PLoS Pathog ; 15(6): e1007840, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31173604

RESUMO

BLyS/BAFF is recognized for its role in B-cell ontogenesis, as well as cell fate decision towards the first-line/innate marginal zone (MZ) B-cell pool. Excess BLyS/BAFF is associated with hyperglobulinemia and increased frequencies of activated precursor-like MZ B-cells. Herein, we show that HIV highly-exposed seronegative (HESN) commercial sex workers (CSWs) had lower soluble BLyS/BAFF levels and relative frequencies of BLyS/BAFF expressing cells in their genital mucosa when compared to those from HIV-infected CSWs and HIV-uninfected non-CSWs. Furthermore, we identified genital innate and/or marginal zone (MZ)-like CD1c+ B-cells that naturally bind to fully glycosylated gp120, which frequencies were lower in HESNs when compared to HIV-infected CSWs and HIV-uninfected non-CSWs. Although genital levels of total IgA were similar between groups, HESNs had lower levels of total IgG1 and IgG3. Interestingly, HIV-gp41 reactive IgG1 were found in some HESNs. Low genital levels of BLyS/BAFF observed in HESNs may allow for controlled first-line responses, contributing to natural immunity to HIV.


Assuntos
Antígenos CD1/imunologia , Fator Ativador de Células B/imunologia , Linfócitos B/imunologia , Genitália Feminina/imunologia , Glicoproteínas/imunologia , Soronegatividade para HIV/imunologia , HIV-1/imunologia , Imunidade Inata , Profissionais do Sexo , Adulto , Linfócitos B/patologia , Feminino , Anticorpos Anti-HIV/imunologia , Humanos , Imunoglobulina G/imunologia , Pessoa de Meia-Idade
5.
Nat Commun ; 10(1): 2498, 2019 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-31175312

RESUMO

Allogeneic mesenchymal stem cells (MSCs) exhibit immunoregulatory function in human autoimmune diseases such as systemic lupus erythematosus (SLE), but the underlying mechanisms remain incompletely understood. Here we show that the number of peripheral tolerogenic CD1c+ dendritic cells (DCs) and the levels of serum FLT3L are significantly decreased in SLE patients especially with lupus nephritis, compared to healthy controls. Transplantation of allogeneic umbilical cord-derived MSCs (UC-MSCs) significantly up-regulates peripheral blood CD1c+DCs and serum FLT3L. Mechanistically, UC-MSCs express FLT3L that binds to FLT3 on CD1c+DCs to promote the proliferation and inhibit the apoptosis of tolerogenic CD1c+DCs. Conversely, reduction of FLT3L with small interfering RNA in MSCs abolishes the up-regulation of tolerogenic CD1c+DCs in lupus patients treated with MSCs. Interferon-γ induces FLT3L expression in UC-MSCs through JAK/STAT signaling pathway. Thus, allogeneic MSCs might suppress inflammation in lupus through up-regulating tolerogenic DCs.


Assuntos
Antígenos CD1/imunologia , Células Dendríticas/imunologia , Glicoproteínas/imunologia , Tolerância Imunológica/imunologia , Lúpus Eritematoso Sistêmico/terapia , Proteínas de Membrana/imunologia , Transplante de Células-Tronco Mesenquimais , Adulto , Antígenos CD1/metabolismo , Estudos de Casos e Controles , Células Dendríticas/metabolismo , Feminino , Glicoproteínas/metabolismo , Humanos , Interferon gama/farmacologia , Janus Quinases/efeitos dos fármacos , Janus Quinases/metabolismo , Lúpus Eritematoso Sistêmico/imunologia , Nefrite Lúpica/imunologia , Nefrite Lúpica/terapia , Masculino , Proteínas de Membrana/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Pessoa de Meia-Idade , Fatores de Transcrição STAT/efeitos dos fármacos , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transplante Homólogo , Adulto Jovem
6.
Vet Microbiol ; 233: 113-117, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31176396

RESUMO

Bovine ephemeral fever virus (BEFV) causes an acute febrile disease in cattle and water buffalo. The disease has an impact on dairy and beef production in tropical and subtropical countries. Vaccination is used for disease prevention and control. In this study, we developed a recombinant lentivirus to produce mammalian stable cells expressing histidine-tagged BEFV G protein with a deleted transmembrane domain (GΔTM) as a secretory protein. In addition, guinea pigs were immunised with the purified GΔTM protein and booster immunised at a 3-week interval. The mammalian stable cells were able to continuously produce GΔTM protein for a minimum of 25 passages. All of the mammalian stable cells expressing GΔTM protein could react specifically with a BEFV convalescent bovine serum. Serum samples from the immunised guinea pigs could react strongly and specifically with the purified GΔTM protein. Moreover, post-immunised guinea pig sera contained antibodies that could neutralise BEFV. These results indicate that the G protein without a transmembrane domain can be used as a subunit vaccine for the prevention and control of BEFV. The availability of the mammalian stable cells, which constitutively express GΔTM protein, could facilitate the potential use of the secretory protein for BEFV diagnosis and vaccine development.


Assuntos
Anticorpos Antivirais/sangue , Febre Efêmera/prevenção & controle , Glicoproteínas/genética , Glicoproteínas/imunologia , Imunogenicidade da Vacina , Proteínas Virais/genética , Proteínas Virais/imunologia , Animais , Anticorpos Neutralizantes/sangue , Bovinos , Linhagem Celular , Febre Efêmera/imunologia , Vírus da Febre Efêmera Bovina , Feminino , Cobaias , Células HEK293 , Humanos , Transfecção , Vacinação , Vacinas Virais/imunologia
7.
APMIS ; 127(8): 588-593, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31233243

RESUMO

Microfibrillar-associated protein 4 (MFAP4) is a non-structural matrix protein with cell regulatory activities and a potential as seromarker for fibrosis. We aimed to study the occurrence of MFAP4 in the synovial membrane from patients with rheumatoid arthritis (RA) vs osteoarthritis (OA). Formaldehyde-fixed synovial tissue sections, from patients with RA (N = 6) and OA (N = 6) undergoing total hip arthroplasty, were deparaffinized and immunostained with monoclonal antibodies against MFAP4. Elastin was detected using ElastiKit. MFAP4 in serum (sMFAP4) and synovial fluid was measured by an immunoassay. MFAP4 was present in synovial biopsies from both RA and OA patients, particularly prominent in deep arterioles where it colocalized with elastin. Notably however, MFAP4 was absent from the internal elastic lamina in RA arterioles irrespective of disease duration and synovitis activity, while present although with irregular staining patterns in OA specimens. sMFAP4 was increased in RA and OA serum vs healthy controls: median (interquartile range) 29.8 (25.3-39.1) and 25.5 U/L (18.1-43.3) vs 17.7 U/L (13.7-21.2), p = 0.006 and p = 0.02, respectively The concentration of synovial fluid was lower than in serum in both RA and OA. These findings may suggest that MFAP4 is involved in adaptive vessel wall remodeling associated with chronic joint disease.


Assuntos
Artrite Reumatoide/imunologia , Proteínas de Transporte/análise , Proteínas da Matriz Extracelular/análise , Glicoproteínas/análise , Osteoartrite/imunologia , Membrana Sinovial/imunologia , Idoso , Anticorpos Monoclonais/imunologia , Biomarcadores/análise , Proteínas de Transporte/imunologia , Estudos de Casos e Controles , Proteínas da Matriz Extracelular/imunologia , Feminino , Glicoproteínas/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Líquido Sinovial/química , Membrana Sinovial/patologia
8.
Invest Ophthalmol Vis Sci ; 60(6): 2034-2037, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31067323

RESUMO

Antibodies are key reagents used in vision research, indeed across biomedical research, but they often do not reveal the whole story about a sample. It is important for researchers to be aware of aspects of antibodies that may affect or limit data interpretation. Federal agencies now require funded grants to demonstrate how they will authenticate reagents used. There is also a push for recombinant antibodies, enabled by phage display technology awarded the 2018 Nobel Prize in Chemistry, which allow for thorough validation and a fixed DNA sequence. Here, we discuss how issues surrounding antibodies are pertinent to detecting myocilin, a protein found in trabecular meshwork and associated with a portion of hereditary glaucoma. Confirmation of myocilin expression in tissues and cell culture has been adopted as validation standard in trabecular meshwork research; thus, a discussion of antibody characteristics and fidelity is critical. Further, based on our basic structural understanding of myocilin architecture and its biophysical aggregation properties, we provide a wish list for the characteristics of next-generation antibody reagents for vision researchers. In the long term, well-characterized antibodies targeting myocilin will enable new insights into its function and involvement in glaucoma pathogenesis.


Assuntos
Anticorpos/imunologia , Proteínas do Citoesqueleto/metabolismo , Proteínas do Olho/metabolismo , Glaucoma/imunologia , Glicoproteínas/metabolismo , Malha Trabecular/metabolismo , Proteínas do Citoesqueleto/imunologia , Proteínas do Olho/imunologia , Glaucoma/diagnóstico , Glaucoma/metabolismo , Glicoproteínas/imunologia , Humanos
9.
Arch Virol ; 164(7): 1793-1803, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31079211

RESUMO

Numerous studies have shown that immunostimulatory complexes containing Quil-A saponin and various antigens are effective in stimulating the immune response and can be used as vaccine preparations for animals and humans. However, Quil-A saponin possesses toxicity and haemolytic activity. In the present work, a saponin-containing preparation named "Glabilox" was isolated from the roots of a Glycyrrhiza glabra L. plant by high-performance liquid chromatography (HPLC). The results showed that Glabilox has no toxicity or haemolytic activity and can form stable immunostimulatory complexes. Subcutaneous immunization of mice with an immunostimulating complex containing Glabilox and H7N1 influenza virus antigens stimulated high levels of humoral and cellular immunity. Vaccination of chickens with the same immunostimulating complex protected 100% of the animals after experimental infection with a homologous virus. Comparative studies showed that the immunogenic and protective activity of immunostimulatory complexes containing Quil-A and immunostimulatory complexes containing Glabilox are comparable to each other. The results of these studies indicated that Glycyrrhiza glabra saponins show great promise as safe and effective adjuvants.


Assuntos
Adjuvantes Imunológicos/farmacologia , Anticorpos Antivirais/sangue , Antígenos Virais/imunologia , Glycyrrhiza/imunologia , Vírus da Influenza A Subtipo H7N1/imunologia , Influenza Aviária/prevenção & controle , Animais , Linhagem Celular , Embrião de Galinha , Galinhas , Cães , Glicoproteínas/imunologia , Imunoglobulina A/sangue , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Influenza Aviária/imunologia , Lipídeos/imunologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos BALB C , Raízes de Plantas/imunologia , Saponinas de Quilaia/imunologia , Saponinas/imunologia , Vacinação
10.
Microb Pathog ; 132: 243-253, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31075428

RESUMO

Ebola virus (EBOV), a non-segmented single-stranded RNA virus, is often-most transmitted through body fluids like sweat, tears, saliva, and nasal secretions. Till date, there is no licensed vaccine of EBOV is available in the market; however, the world is increasingly vulnerable to this emerging threat. Hence, it is the need of time to develop a vaccine for EBOV to hinder its dissemination. The current study has been designed for identification and characterization of the potential B and T-cell epitopes using the Immuno-informatics tools, and it helped in finding the potent vaccine candidates against EBOV. Prediction, antigenicity and allergenicity testing of predicted B and T cells' epitopes was done as well to identify their potential as a vaccine candidate and to measure their safety level respectively. Among B-cell epitopes "WIPAGIGVTGVIIA" showed a high antigenicity score and it would play an important role in evoking the immune response. In T-cell epitopes, peptides "AIGLAWIPY" and "IRGFPRCRY" presented high antigenicity score, which binds to MHC class-I and MHC class-II alleles respectively. All predicted epitopes were analyzed and compared with already reported peptides carefully. Comparatively, Peptides predicted in the present study showed more immunogenicity score than already reported peptides, used as positive control, and are more immunogenic as compared to them. Peptides reported in the present study do not target only Zaire EBOV (ZEBOV), as in previous studies, but also other species, i.e. Tai Forest EBOV (TAFV), Sudan EBOV (SUDV), Bundibugyo EBOV (BDBV), and Reston EBOV (RESTV) and would bring the promising results as potent vaccine candidates.


Assuntos
Vacinas contra Ebola/imunologia , Ebolavirus/imunologia , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito T/imunologia , Glicoproteínas/imunologia , Alelos , Sequência de Aminoácidos , Vacinas contra Ebola/genética , Ebolavirus/genética , Genes MHC Classe I , Genes MHC da Classe II , Glicoproteínas/química , Glicoproteínas/genética , Antígeno HLA-B7 , Imunogenicidade da Vacina , Simulação de Acoplamento Molecular , Estrutura Secundária de Proteína
11.
Fish Shellfish Immunol ; 89: 516-524, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30986537

RESUMO

Infectious hematopoietic necrosis virus (IHNV) leads to serious disease and economic losses in the salmonid aquaculture industry. The present study aimed to develop an effective and efficient vaccine to protect rainbow trout (Oncorhynchus mykiss) against IHNV infection. Administered via the immersion route, a live vector vaccine containing the regions of the IHNV glycoprotein (G) induced immune responses in rainbow trout. Use of the immersion route induced more-efficient mucosal immunity than intramuscular injection vaccination. IHNV G gene expression was detected in the spleens of rainbow trout at 3, 7 and 15 days post-vaccination (dpv). The G gene expression continuously decreased between 3 and 15 dpv. In addition, the expression of TLR-3, TLR-7 and TLR-8 was upregulated after vaccination, and the highest expression levels of IFN-1, Mx-1, Mx-3, Vig-1 and Vig-2 were observed at 3 dpv. Four markers of the adaptive immune response (CD4, CD8, IgM and IgT) gradually increased. When experimental fish were challenged with IHNV by immersion, significant differences in cumulative percentage mortality were observed in the vaccinated fish and the unvaccinated (empty-plasmid-vaccinated) fish. The relative survival rate was 92% and 6% in the vaccinated group and empty-plasmid group, respectively. Serum antibody levels gradually increased in the vaccinated fish, unlike in the unvaccinated fish, after 7 dpv. Our results suggest there was a significant increase in fish immune responses and resistance to infection with IHNV following administration of the live vector vaccine. Therefore, this live vector vaccine is a promising vaccine that may be utilized to protect rainbow trout against IHNV.


Assuntos
Imunidade Adaptativa , Doenças dos Peixes/prevenção & controle , Vírus da Necrose Hematopoética Infecciosa/fisiologia , Oncorhynchus mykiss , Infecções por Rhabdoviridae/veterinária , Vacinas Virais/imunologia , Animais , Doenças dos Peixes/imunologia , Glicoproteínas/genética , Glicoproteínas/imunologia , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/prevenção & controle , Baço/imunologia , Vacinas Atenuadas/imunologia , Proteínas Virais/genética , Proteínas Virais/imunologia
12.
Fish Shellfish Immunol ; 89: 537-547, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30991145

RESUMO

Infectious hematopoietic necrosis virus (IHNV) causes infectious hematopoietic necrosis in salmonid fish, resulting in substantial economic losses to the aquaculture industry worldwide. The G protein, which harbors the major antigenic determinants of IHNV, is an envelope glycoprotein that plays an important role in both pathogenicity and immunogenicity of IHNV. Previous studies have demonstrated that changes to viral glycosylation sites may affect replication and immunogenicity, but little is known about the specific contributions of G protein glycosylation to IHNV replication and pathogenicity. In this study, we predicted four N-linked glycosylation sites at position 56, 379, 401, and 438 Asp (N) in G protein, and using a reverse genetics system developed in our laboratory, constructed nine recombinant viruses with single, triple, or quadruple glycosylation site disruptions using alanine substitutions in the following combinations: rIHNV-N56A, rIHNV-N379A, rIHNV-N401A, rIHNV-N438A, rIHNV-N56A-N379A-N401A, rIHNV-N56A-N379A-N438A, rIHNV-N56A-N401A-N438A, rIHNV-N379A-N401A-N438A, and rIHNV-N56A-N379A-N401A-N438A. Our results confirmed that all four asparagines are sites of N-linked glycosylation, and Western blot confirmed that mutation of each predicted N-glycosylation sited impaired glycosylation. Among the nine recombinant IHNVs, replication levels decreased significantly in vitro and in vivo in the triple and quadruple mutants that combined mutation of asparagines 401 and 438, indicating the importance of glycosylation at these sites for efficient replication. Moreover, juvenile rainbow trout mortality after challenge by each of the nine mutants showed that, while eight mutants suffered almost 100% cumulative mortality over 30 days, the mutant with a single alanine substitution at position 438 resulted in cumulative mortality of less than 50% over 30 days. This mutant also elicited specific anti-IHNV IgM production earlier than other mutants, suggesting that glycosylation of asparagine 438 may be important for viral immune escape. In conclusion, our study reveals the effect of G protein glycosylation on the pathogenicity and immunogenicity of IHNV and provides a foundation for developing a live-attenuated vaccine.


Assuntos
Doenças dos Peixes/prevenção & controle , Glicoproteínas/imunologia , Vírus da Necrose Hematopoética Infecciosa/imunologia , Vírus da Necrose Hematopoética Infecciosa/patogenicidade , Oncorhynchus mykiss , Infecções por Rhabdoviridae/veterinária , Vacinas Virais/imunologia , Animais , Doenças dos Peixes/imunologia , Glicosilação , Imunogenicidade da Vacina/imunologia , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/prevenção & controle , Virulência
13.
Int Arch Allergy Immunol ; 179(1): 10-16, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30893695

RESUMO

Peanut allergy is considered to be the most common cause for food-induced anaphylaxis. Currently, no approved treatment is available. Avoidance is the only measure to prevent anaphylactic reactions to peanuts. T-helper cells are of special importance for the sensitization process and the maintenance of allergic inflammation. Identifying markers of allergen-specific T-cell responses may help to develop novel treatment approaches. Therefore, we aimed to define new T-cell target genes in Ara h 2-specific T cells and to investigate the possibility of using them as biomarkers of peanut allergy in peripheral blood mononuclear cells (PBMCs). We performed whole mRNA array analysis (whole human genome oligo microarray) of in vitro expanded Ara h 2-specific T cells (CFSElowCD3+CD4+) from 5 peanut-allergic (PA) and 5 non-peanut-sensitized individuals. Expression of selected genes as a result of a two-step bioinformatic approach was confirmed in a second cohort by quantitative PCR. TGF-ß- activated kinase 1 and MAP3K7 binding protein 3 (TAB3), calcium/calmodulin-dependent protein kinase type IV (CAMK4) and HemK methyltransferase family member 1 (HEMK1) were significantly upregulated in Ara h 2-specific T cells of PA patients. In addition, the expression of these genes was also assessed in unstimulated PBMCs from a cohort (n = 43) of PA, atopic non-PA, and nonatopic controls. Interestingly, in unstimulated PBMCs, TAB3 expression was significantly downregulated in PA patients compared to atopic non-PA individuals. Thus, TAB3 may play a significant role at the level of T-cell activation and may also be a candidate biomarker for PA.


Assuntos
Albuminas 2S de Plantas/imunologia , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Antígenos de Plantas/imunologia , Arachis/imunologia , Linfócitos T CD4-Positivos/imunologia , Glicoproteínas/imunologia , Hipersensibilidade a Amendoim/etiologia , Adolescente , Células Cultivadas , Criança , Feminino , Humanos , Ativação Linfocitária , Masculino , Metiltransferases/fisiologia , NF-kappa B/fisiologia
14.
Viruses ; 11(3)2019 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-30836694

RESUMO

Previous studies have shown that wild-type (wt) rabies virus (RABV) evades the host immune response by restricting expression of glycoprotein (G), which blocks activation of dendritic cells (DCs) and induces production of virus-neutralizing antibodies (VNAs). In the present study, wt RABVs not only restricted G expression but also reduced incorporation of G into mature virions compared with laboratory-adapted viruses. A recombinant RABV expressing triple G was used to further determine whether G expression relates to incorporation. The recombinant virus showed higher expression and incorporation of G and activated more DCs than the virus that expressed a single copy of G. Removal of G from viruses using subtilisin or Dithiothreitol (DTT)/ Nonidet P-40 (NP40) almost completely abolishes DC activation and VNA production. Consequently, these G-depleted viruses cause lethal infection in mice. Thus, wt RABVs can subvert DC-induced antiviral immune response and maintain pathogenicity by decreasing G expression in infected cells and G incorporation into virions.


Assuntos
Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Glicoproteínas/imunologia , Evasão da Resposta Imune , Vírus da Raiva/imunologia , Vírus da Raiva/patogenicidade , Proteínas do Envelope Viral/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Antígenos Virais/genética , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Modelos Animais de Doenças , Glicoproteínas/genética , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Vírus da Raiva/genética , Proteínas do Envelope Viral/genética , Vírion , Ativação Viral
15.
Future Microbiol ; 14: 397-410, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30854893

RESUMO

AIM: Sporothrix schenckii is the causative agent of sporotrichosis. A 70-kDa glycoprotein, Gp70, is a candidate for the development of prophylactic alternatives to control the disease, and its gene (GP70) is predicted to encode for a protein of 43 kDa, contrasting with the molecular weight of the native protein. MATERIALS & METHODS: The GP70 was expressed in bacteria, the recombinant protein purified, used in immunoassays and injected to Galleria mellonella. RESULTS & CONCLUSION: The recombinant protein was detected by anti-Gp70 antibodies, confirming that the Gp70 backbone is a 43-kDa peptide. This protein showed enzyme activity of cyclase and was recognized by sera of patients with sporotrichosis. Although it was not useful for serodiagnosis of sporotrichosis, it conferred protection to animals against experimental sporotrichosis.


Assuntos
Proteínas Fúngicas/genética , Proteínas Fúngicas/imunologia , Glicoproteínas/imunologia , Sporothrix/genética , Esporotricose/microbiologia , Animais , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas Fúngicas/química , Expressão Gênica , Glicoproteínas/química , Glicoproteínas/genética , Humanos , Peso Molecular , Mariposas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Sporothrix/imunologia , Esporotricose/imunologia
16.
Immun Inflamm Dis ; 7(2): 68-73, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30851084

RESUMO

BACKGROUND: Fel d1 is the most important allergen from cats. Fel d1 is produced primarily in saliva and spread to the haircoat during grooming and then transferred to the environment via hair and dander. OBJECTIVES: A novel approach to reducing allergenic Fel d1 exposure was evaluated, involving binding the Fel d1 with an anti-Fel d1 polyclonal egg IgY antibody. The hypothesis was that hair from cats who had been fed foods containing anti-Fel d1 IgY would show a significant reduction in active Fel d1 (aFel d1). METHODS: Hair collected from 105 cats completing a 12-week study was evaluated for aFel d1 via ELISA. Hair was collected four times over a 2-week baseline period, then weekly during the 10 week treatment period during which cats consumed a food containing the anti-Fel d1 IgY. RESULTS: Baseline aFel d1 (µg/g hair) varied greatly among the cats in this study. From week 3, there was a significant reduction in mean aFel d1 with an overall average decrease of 47% by week 10, ranging from a 33-71% decrease vs baseline. Cats with the highest baseline aFel d1 showed the greatest decrease in aFel d1. CONCLUSIONS & CLINICAL IMPLICATIONS: Feeding anti-Fel d1 IgY to cats successfully reduced aFel d1 on their haircoat with the greatest decreases observed in cats with initially high levels. Feeding a diet with anti Fel d1 IgY significantly reduced the active Fel d1 on the hair of cats.


Assuntos
Alérgenos/imunologia , Glicoproteínas/imunologia , Imunoglobulinas/imunologia , Animais , Gatos , Ensaio de Imunoadsorção Enzimática , Feminino , Masculino
17.
Methods Mol Biol ; 1955: 239-246, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30868532

RESUMO

The trans-sialidase (TS), a virulence factor expressed on the surface of Trypanosoma cruzi, the agent of Chagas disease, is an enzyme that transfers sialic acids between glycoconjugates. In humans and most tested mammals, the onset of the chronic phase of T. cruzi infection correlates with the elicitation of antibodies directed to the TS catalytic domain, which inhibit the sialyl residues transfer reaction in vitro and in vivo. The method described here, termed trans-sialidase inhibition assay (TIA), enables the detection of TS-neutralizing antibodies in serum samples of different mammalian species, without the use of conjugated secondary reagents. The high specificity and exquisite sensitivity displayed by the TIA allow to overcome the limitations of routinely used Chagas disease serodiagnostic assays.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antiprotozoários/imunologia , Doença de Chagas/diagnóstico , Doença de Chagas/imunologia , Glicoproteínas/imunologia , Neuraminidase/imunologia , Trypanosoma cruzi/enzimologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antiprotozoários/sangue , Doença de Chagas/sangue , Ensaios Enzimáticos/métodos , Humanos , Proteínas Recombinantes/imunologia , Trypanosoma cruzi/imunologia , Fatores de Virulência/imunologia
18.
Crit Rev Biotechnol ; 39(3): 380-394, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30720351

RESUMO

Through the discovery of monoclonal antibody (mAb) technology, profound successes in medical treatment against a wide range of diseases have been achieved. This has led antibodies to emerge as a new class of biodrugs. As the "rising star" in the pharmaceutical market, extensive research and development in antibody production has been carried out in various expression systems including bacteria, insects, plants, yeasts, and mammalian cell lines. The major benefit of eukaryotic expression systems is the ability to carry out posttranslational modifications of the antibody. Glycosylation of therapeutic antibodies is one of these important modifications, due to its influence on antibody structure, stability, serum half-life, and complement recruitment. In recent years, the protozoan parasite Leishmania tarentolae has been introduced as a new eukaryotic expression system. L. tarentolae is rich in glycoproteins with oligosaccharide structures that are very similar to humans. Therefore, it is touted as a potential alternative to mammalian expression systems for therapeutic antibody production. Here, we present a comparative review on the features of the L. tarentolae expression system with other expression platforms such as bacteria, insect cells, yeasts, transgenic plants, and mammalian cells with a focus on mAb production.


Assuntos
Anticorpos Monoclonais/biossíntese , Clonagem Molecular/métodos , Leishmania/genética , Proteínas Recombinantes/biossíntese , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Expressão Gênica/genética , Expressão Gênica/imunologia , Glicoproteínas/biossíntese , Glicoproteínas/imunologia , Glicosilação , Humanos , Leishmania/imunologia , Processamento de Proteína Pós-Traducional/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/uso terapêutico
19.
Glycobiology ; 29(4): 307-319, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30726901

RESUMO

Successful application of potent antibody-based T-cell engaging immunotherapeutic strategies is currently limited mainly to hematological cancers. One major reason is the lack of well-characterized antigens on solid tumors with sufficient cancer specific expression. Aberrantly O-glycosylated proteins contain promising cancer-specific O-glycopeptide epitopes suitable for immunotherapeutic applications, but currently only few examples of such antibody epitopes have been identified. We previously showed that chimeric antigen receptor T-cells directed towards aberrantly O-glycosylated MUC1 can control malignant growth in a mouse model. Here, we present a discovery platform for the generation of cancer-specific monoclonal antibodies targeting aberrant O-glycoproteins. The strategy is based on cancer cell lines engineered to homogeneously express the truncated Tn O-glycoform, the so-called SimpleCells. We used SimpleCells of different cancer origin to elicit monoclonal antibodies with selectivity for aberrant O-glycoproteins. For validation we selected and characterized one monoclonal antibody (6C5) directed to a Tn-glycopeptide in dysadherin (FXYD5), known to be upregulated in cancer and promote metastasis. While dysadherin is widely expressed also in normal cells, we demonstrated that the 6C5 epitope is specifically expressed in cancer.


Assuntos
Anticorpos Monoclonais/análise , Anticorpos Monoclonais/biossíntese , Glicoproteínas/metabolismo , Neoplasias/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Linhagem Celular Tumoral , Epitopos/imunologia , Epitopos/metabolismo , Glicoproteínas/imunologia , Humanos , Camundongos , Neoplasias/imunologia , Neoplasias/patologia
20.
PLoS Pathog ; 15(2): e1007375, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30707748

RESUMO

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease localized to China, Japan, and Korea that is characterized by severe hemorrhage and a high fatality rate. Currently, no specific vaccine or treatment has been approved for this disease. To develop a therapeutic agent for SFTS, we isolated antibodies from a phage-displayed antibody library that was constructed from a patient who recovered from SFTS virus (SFTSV) infection. One antibody, designated as Ab10, was reactive to the Gn envelope glycoprotein of SFTSV and protected host cells and A129 mice from infection in both in vitro and in vivo experiments. Notably, Ab10 protected 80% of mice, even when injected 5 days after inoculation with a lethal dose of SFTSV. Using cross-linker assisted mass spectrometry and alanine scanning, we located the non-linear epitope of Ab10 on the Gn glycoprotein domain II and an unstructured stem region, suggesting that Ab10 may inhibit a conformational alteration that is critical for cell membrane fusion between the virus and host cell. Ab10 reacted to recombinant Gn glycoprotein in Gangwon/Korea/2012, HB28, and SD4 strains. Additionally, based on its epitope, we predict that Ab10 binds the Gn glycoprotein in 247 of 272 SFTSV isolates previously reported. Together, these data suggest that Ab10 has potential to be developed into a therapeutic agent that could protect against more than 90% of reported SFTSV isolates.


Assuntos
Anticorpos Neutralizantes/metabolismo , Phlebovirus/imunologia , Adulto , Animais , Anticorpos Neutralizantes/fisiologia , Anticorpos Antivirais/metabolismo , Infecções por Bunyaviridae/terapia , Epitopos/imunologia , Feminino , Febre , Glutamina/imunologia , Glutamina/metabolismo , Glicoproteínas/imunologia , Células HEK293 , Humanos , Leucopenia , Masculino , Camundongos , Camundongos Knockout , Testes de Neutralização , Phlebovirus/patogenicidade , República da Coreia , Trombocitopenia/imunologia , Proteínas do Envelope Viral/imunologia
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