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1.
Gene ; 744: 144616, 2020 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-32222531

RESUMO

AIM: The purpose of this study was to investigate the possible effects of Myrtus communis subsp. communis (MC) on cognitive impairment in ovariectomized diabetic rats. MATERIAL AND METHOD: Female Sprague-Dawley rats were divided into 5 groups consisting of 15 rats each; Control (C), Diabetes (D), Ovariectomy and diabetes (OVX + D), Ovariectomy, diabetes and donepezil (OVX + D + Don), Ovariectomy, diabetes and Myrtus communis subsp. communis (OVX + D + MC). Blood glucose measurements were made at the beginning and end of the experiments. The animals underwent the novel object recognition test (NORT) and their performance was evaluated. In hippocampal tissues; amyloid beta (Aß) and neprilysin levels, acetylcholinesterase (AChE), and choline acetyltransferase (ChAT) activities, polysialylated neural cell adhesion molecule (PSA-NCAM), α7 subunit of neuronal nicotinic acetylcholine receptor (α7-nAChR) and brain derived neurotrophic factor (BDNF) gene expressions were examined. RESULTS: Animals with ovariectomy and diabetes showed increased levels of blood glucose, AChE activity and Aß levels, and decreased neprilysin levels, ChAT activity, α7-nAChR, PSA-NCAM and BDNF gene expressions in parallel with a decrease in NORT performance score. On the other hand, in the MC-treated OVX + D group, there was a significant decrease observed in blood glucose levels and AChE activities while there was improvement in NORT performances and an increase in hippocampal ChAT activity, neprilysin levels, α7-nAChR, PSA-NCAM and BDNF expressions. CONCLUSION: These results suggest that MC extract could improve cognitive and neuronal functions with its anticholinesterase and antihyperglycemic properties.


Assuntos
Cognição/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Myrtus , Fitoterapia , Acetilcolinesterase/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Glicemia/metabolismo , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Colina O-Acetiltransferase/metabolismo , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/psicologia , Feminino , Hipocampo/metabolismo , Neprilisina/metabolismo , Molécula L1 de Adesão de Célula Nervosa/genética , Molécula L1 de Adesão de Célula Nervosa/metabolismo , Ovariectomia , Extratos Vegetais/uso terapêutico , Ratos Sprague-Dawley , Ácidos Siálicos/genética , Ácidos Siálicos/metabolismo , Receptor Nicotínico de Acetilcolina alfa7/genética , Receptor Nicotínico de Acetilcolina alfa7/metabolismo
2.
Am J Physiol Lung Cell Mol Physiol ; 318(1): L165-L179, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31617733

RESUMO

Pulmonary fibrosis involves the formation of inappropriate scar tissue in the lungs, but what drives fibrosis is unclear. Sialidases (also called neuraminidases) cleave terminal sialic acids from glycoconjugates. In humans and mice, pulmonary fibrosis is associated with desialylation of glycoconjugates and upregulation of sialidases. Of the four mammalian sialidases, we previously detected only NEU3 in the bronchoalveolar lavage fluid from mice with bleomycin-induced pulmonary fibrosis. In this report, we show that NEU3 upregulates extracellular accumulation of the profibrotic cytokines IL-6 and IL-1ß, and IL-6 upregulates NEU3 in human peripheral blood mononuclear cells, suggesting that NEU3 may be part of a positive feedback loop potentiating fibrosis. To further elucidate the role of NEU3 in fibrosis, we used bleomycin to induce lung fibrosis in wild-type C57BL/6 and Neu3-/- mice. At 21 days after bleomycin, compared with male and female C57BL/6 mice, male and female Neu3-/- mice had significantly less inflammation, less upregulation of other sialidases and the profibrotic cytokine active transforming growth factor ß1, and less fibrosis in the lungs. Our results suggest that NEU3 participates in fibrosis and that NEU3 could be a target to develop treatments for fibrosis.


Assuntos
Neuraminidase/metabolismo , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/fisiopatologia , Animais , Bleomicina/farmacologia , Líquido da Lavagem Broncoalveolar , Linhagem Celular , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Inflamação/metabolismo , Inflamação/fisiopatologia , Interleucina-1beta/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/fisiologia , Pulmão/metabolismo , Pulmão/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ácidos Siálicos/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia
3.
Sci Total Environ ; 699: 134380, 2020 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-31678878

RESUMO

The oral health of preschool children in an electronic waste (e-waste) area is susceptible to lead (Pb) exposure increasing the risk of dental caries and causing periodontitis and other oral diseases. The aim of the present study is to investigate the relationship between chronic exposure to Pb and oral anti-inflammatory potential of preschool children. For this analysis, 574 preschool children from 2.5 to 6 years of age were recruited between November and December 2017, in which 357 preschool children were from Guiyu (n = 357), an e-waste-contaminated town, and 217 from Haojiang Shantou. We measured the levels of child blood Pb, salivary sialic acid, serum interleukin-6 (IL-6) and serum tumor necrosis factor-α (TNF-α), and investigated the prevalence of dental caries in deciduous teeth. The medians of blood Pb levels, serum IL-6 and TNF-α were significantly higher in the Guiyu children than in Haojiang children. Concomitantly, salivary sialic acids were lower in the Guiyu children [9.58 (3.97, 18.42) mg/dL] than in Haojiang [17.57 (5.95, 24.23) mg/dL]. Additionally, the prevalence of dental caries in deciduous teeth was significantly higher in the Guiyu children than in Haojiang (62.5% vs. 53.9%). Blood Pb levels were negatively correlated with salivary sialic acids, in which IL-6 played as a mediator of the association between blood Pb levels and saliva sialic acid concentrations according to the mediation model. To our knowledge, this is the first report on the potential association between chronic Pb exposure and the anti-inflammatory ability of oral sialic acids among preschool children. These results suggest that the chronic Pb exposure can reduce salivary sialic acid levels, attenuate oral anti-inflammatory potential and increase the potential risk of dental caries in deciduous teeth among preschool children in an e-waste site.


Assuntos
Resíduo Eletrônico , Exposição Ambiental/análise , Poluentes Ambientais/metabolismo , Chumbo/metabolismo , Ácidos Siálicos/metabolismo , Anti-Inflamatórios , Pré-Escolar , Exposição Ambiental/estatística & dados numéricos , Humanos
4.
Nat Struct Mol Biol ; 26(12): 1151-1157, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31792450

RESUMO

The Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe and often lethal respiratory illness in humans, and no vaccines or specific treatments are available. Infections are initiated via binding of the MERS-CoV spike (S) glycoprotein to sialosides and dipeptidyl-peptidase 4 (the attachment and entry receptors, respectively). To understand MERS-CoV engagement of sialylated receptors, we determined the cryo-EM structures of S in complex with 5-N-acetyl neuraminic acid, 5-N-glycolyl neuraminic acid, sialyl-LewisX, α2,3-sialyl-N-acetyl-lactosamine and α2,6-sialyl-N-acetyl-lactosamine at 2.7-3.0 Å resolution. We show that recognition occurs via a conserved groove that is essential for MERS-CoV S-mediated attachment to sialosides and entry into human airway epithelial cells. Our data illuminate MERS-CoV S sialoside specificity and suggest that selectivity for α2,3-linked over α2,6-linked receptors results from enhanced interactions with the former class of oligosaccharides. This study provides a structural framework explaining MERS-CoV attachment to sialoside receptors and identifies a site of potential vulnerability to inhibitors of viral entry.


Assuntos
Coronavírus da Síndrome Respiratória do Oriente Médio/química , Ácidos Siálicos/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Sítios de Ligação , Configuração de Carboidratos , Microscopia Crioeletrônica , Dipeptidil Peptidase 4/química , Dipeptidil Peptidase 4/metabolismo , Dipeptidil Peptidase 4/ultraestrutura , Hemaglutinação por Vírus , Humanos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Mapeamento de Interação de Proteínas , Ácidos Siálicos/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/ultraestrutura , Relação Estrutura-Atividade
5.
Nat Commun ; 10(1): 5404, 2019 11 27.
Artigo em Inglês | MEDLINE | ID: mdl-31776339

RESUMO

Glycosylation plays important roles in cellular function and endows protein therapeutics with beneficial properties. However, constructing biosynthetic pathways to study and engineer precise glycan structures on proteins remains a bottleneck. Here, we report a modular, versatile cell-free platform for glycosylation pathway assembly by rapid in vitro mixing and expression (GlycoPRIME). In GlycoPRIME, glycosylation pathways are assembled by mixing-and-matching cell-free synthesized glycosyltransferases that can elaborate a glucose primer installed onto protein targets by an N-glycosyltransferase. We demonstrate GlycoPRIME by constructing 37 putative protein glycosylation pathways, creating 23 unique glycan motifs, 18 of which have not yet been synthesized on proteins. We use selected pathways to synthesize a protein vaccine candidate with an α-galactose adjuvant motif in a one-pot cell-free system and human antibody constant regions with minimal sialic acid motifs in glycoengineered Escherichia coli. We anticipate that these methods and pathways will facilitate glycoscience and make possible new glycoengineering applications.


Assuntos
Sistema Livre de Células/metabolismo , Engenharia de Proteínas/métodos , Proteínas/metabolismo , Antígenos CD/metabolismo , Escherichia coli/genética , Glicoproteínas/biossíntese , Glicosilação , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Humanos , Redes e Vias Metabólicas , Oligossacarídeos/metabolismo , Polissacarídeos/metabolismo , Proteínas/genética , Ácidos Siálicos/química , Ácidos Siálicos/metabolismo , Sialiltransferases/metabolismo
6.
Analyst ; 144(24): 7378-7389, 2019 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-31670365

RESUMO

The development of carbohydrate-binding ligands is crucial for expanding knowledge on the glycocode and for achieving systematic carbohydrate targeting. Amongst such ligands, carbohydrate-binding peptides (CBPs) are attractive for use in bioanalytical and biomedical systems due to their biochemical and physicochemical properties; moreover, given the biological significance of lectin-carbohydrate interactions, these ligands offer an opportunity to study peptide sequence and binding characteristics to inform on natural target/ligand interactions. Here, a high-throughput microarray screening technique is described for the identification and study of CBPs, with a focus on polysialic acid (PSA), a polysaccharide found on neural stem cells. The chemical and biological uniqueness of PSA suggests that an ability to exclusively target this glycan may promote a number of diagnostic and therapeutic applications. PSA-binding peptides from phage display screening and from epitope mapping of an scFv for oligosialic acid were screened in an optimized microarray format with three ligand density conditions. Hypothesis-driven mutations were additionally applied to select peptides to modulate peptide affinity and selectivity to PSA. Peptide compositional and positional analyses revealed the significance of various residues for PSA binding and suggested the importance of basic residue positioning for PSA recognition. Furthermore, selectivity studies performed directly on microarrays with chondroitin sulfate A (CS-A) demonstrated the value of screening for both affinity and selectivity in the development of CBPs. Thus, the integrated approach described, with attention to design strategy, screening, and peptide characterization, successfully identified novel PSA-binding ligands and offers a platform for the identification and study of additional polysaccharide-binding peptides.


Assuntos
Lectinas/análise , Peptídeos/análise , Sequência de Aminoácidos , Ensaios de Triagem em Larga Escala/métodos , Lectinas/química , Lectinas/metabolismo , Ligantes , Análise em Microsséries/métodos , Biblioteca de Peptídeos , Peptídeos/química , Peptídeos/metabolismo , Ligação Proteica , Ácidos Siálicos/metabolismo
7.
Epidemiol Infect ; 147: e297, 2019 11 04.
Artigo em Inglês | MEDLINE | ID: mdl-31679542

RESUMO

In the human population, influenza A viruses are associated with acute respiratory illness and are responsible for millions of deaths annually. Avian and human influenza viruses typically have a different α2-3- and α2-6-linked sialic acid (SA) binding preference. Only a few amino acid changes in the haemagglutinin on the surface of avian influenza viruses (AIV) can cause a switch from avian to human receptor specificity, and the individuals with pathognostic chronic diseases might be more susceptible to AIV due to the decreased expression level of terminal α2-3-linked SA in their saliva. Here, using lectin and virus histochemical staining, we observed the higher expression levels of α2-3/6-linked SA influenza virus receptors in the airway of HBV-transgenic mice compared with that of control mice due to the significant decrease in control mice during ageing, which imply that this is also a risk factor for individuals with pathognostic chronic diseases susceptible to influenza viruses. Our findings will help understand the impact on influenza virus pathogenesis and transmission.


Assuntos
Vírus da Influenza A/imunologia , Pulmão/metabolismo , Ácidos Siálicos/metabolismo , Traqueia/metabolismo , Animais , Biomarcadores/metabolismo , Imuno-Histoquímica , Pulmão/imunologia , Pulmão/virologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Traqueia/imunologia , Traqueia/virologia
8.
Nat Commun ; 10(1): 4816, 2019 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-31645552

RESUMO

Sialic acids are a family of related sugars that play essential roles in many biological events intimately linked to cellular recognition in both health and disease. Sialidases are therefore orchestrators of cellular biology and important therapeutic targets for viral infection. Here, we sought to define if uncharacterized sialidases would provide distinct paradigms in sialic acid biochemistry. We show that a recently discovered sialidase family, whose first member EnvSia156 was isolated from hot spring metagenomes, defines an unusual structural fold and active centre constellation, not previously described in sialidases. Consistent with an inverting mechanism, EnvSia156 reveals a His/Asp active center in which the His acts as a Brønsted acid and Asp as a Brønsted base in a single-displacement mechanism. A predominantly hydrophobic aglycone site facilitates accommodation of a variety of 2-linked sialosides; a versatility that offers the potential for glycan hydrolysis across a range of biological and technological platforms.


Assuntos
Domínio Catalítico , Neuraminidase/metabolismo , Ácidos Siálicos/metabolismo , Cristalografia por Raios X , Glicocálix/metabolismo , Neuraminidase/ultraestrutura , Estrutura Terciária de Proteína
9.
ACS Chem Biol ; 14(10): 2141-2147, 2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31584261

RESUMO

Neu5Ac, Neu5Gc, and KDN are three forms of sialic acids in vertebrates that possess distinct biological functions. Herein, we report the synthesis and metabolic incorporation of the 9-azido analogues of three sialic acid forms in mammalian cells. The incorporated sialic acid analogues enable fluorescent imaging of cell-surface sialoglycans and proteomic profiling of sialoglycoproteins. Furthermore, we apply them to metabolically engineer cell surfaces with sialoglycans terminated with distinct sialic acids or their 9-azido analogues. The remodeled cells expressing specific cell-surface sialoglycoforms show distinct binding affinity toward subtilase cytotoxin (SubAB), a toxin secreted by Shiga toxigenic Escherichia coli. The 9-azido analogues of sialic acid forms developed in this work provide a versatile tool for metabolic remodeling of cell-surface properties and modulating pathogen-host interactions.


Assuntos
Azidas/metabolismo , Glicoproteínas de Membrana/metabolismo , Ácidos Siálicos/metabolismo , Sialoglicoproteínas/metabolismo , Animais , Células CHO , Engenharia Celular/métodos , Linhagem Celular Tumoral , Chlorocebus aethiops , Cricetulus , Proteínas de Escherichia coli/metabolismo , Humanos , Glicoproteínas de Membrana/química , Proteômica , Sialoglicoproteínas/química , Subtilisinas/metabolismo , Células Vero
10.
Nat Commun ; 10(1): 4512, 2019 10 04.
Artigo em Inglês | MEDLINE | ID: mdl-31586047

RESUMO

Plasmodium species are frequently host-specific, but little is currently known about the molecular factors restricting host switching. This is particularly relevant for P. falciparum, the only known human-infective species of the Laverania sub-genus, all other members of which infect African apes. Here we show that all tested P. falciparum isolates contain an inactivating mutation in an erythrocyte invasion associated gene, PfEBA165, the homologues of which are intact in all ape-infective Laverania species. Recombinant EBA165 proteins only bind ape, not human, erythrocytes, and this specificity is due to differences in erythrocyte surface sialic acids. Correction of PfEBA165 inactivating mutations by genome editing yields viable parasites, but is associated with down regulation of both PfEBA165 and an adjacent invasion ligand, which suggests that PfEBA165 expression is incompatible with parasite growth in human erythrocytes. Pseudogenization of PfEBA165 may represent a key step in the emergence and evolution of P. falciparum.


Assuntos
Eritrócitos/parasitologia , Especificidade de Hospedeiro/genética , Malária Falciparum/parasitologia , Plasmodium falciparum/genética , Proteínas de Protozoários/genética , Animais , Sistemas CRISPR-Cas/genética , Engenharia Celular , Eritrócitos/metabolismo , Evolução Molecular , Mutação da Fase de Leitura , Edição de Genes , Células HEK293 , Humanos , Mutação com Perda de Função , Pan troglodytes/parasitologia , Plasmodium falciparum/isolamento & purificação , Plasmodium falciparum/patogenicidade , Ácidos Siálicos/metabolismo
11.
Curr Top Med Chem ; 19(25): 2271-2282, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31648641

RESUMO

Polysialic acid (polySia) is a novel glycan that posttranslationally modifies neural cell adhesion molecules (NCAMs) in mammalian cells. Up-regulation of polySia-NCAM expression or NCAM polysialylation is associated with tumor cell migration and progression in many metastatic cancers and neurocognition. It has been known that two highly homologous mammalian polysialyltransferases (polySTs), ST8Sia II (STX) and ST8Sia IV (PST), can catalyze polysialylation of NCAM, and two polybasic domains, polybasic region (PBR) and polysialyltransferase domain (PSTD) in polySTs play key roles in affecting polyST activity or NCAM polysialylation. However, the molecular mechanisms of NCAM polysialylation and cell migration are still not entirely clear. In this minireview, the recent research results about the intermolecular interactions between the PBR and NCAM, the PSTD and cytidine monophosphate-sialic acid (CMP-Sia), the PSTD and polySia, and as well as the intramolecular interaction between the PBR and the PSTD within the polyST, are summarized. Based on these cooperative interactions, we have built a novel model of NCAM polysialylation and cell migration mechanisms, which may be helpful to design and develop new polysialyltransferase inhibitors.


Assuntos
Movimento Celular , Moléculas de Adesão de Célula Nervosa/metabolismo , Ácidos Siálicos/metabolismo , Animais , Humanos , Moléculas de Adesão de Célula Nervosa/química , Ácidos Siálicos/química
12.
Fish Shellfish Immunol ; 94: 72-80, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31472263

RESUMO

In the present study, a sialic acid-binding lectin was cloned and characterized from Manila clam Ruditapes philippinarum (designed as RpSabl). The open reading frame of RpSabl encoded a polypeptide of 162 amino acids with a calculated molecular mass of 17.7 kDa. Analysis of the conserved domain suggested that RpSabl was a new member of the sialic acid-binding lectins family. In non-stimulated clams, RpSabl transcripts were constitutively expressed in all five tested tissues, especially in hepatopancreas. After Vibrio anguillarum challenge, the expression of RpSabl mRNA in hepatopancreas was significantly up-regulated at 3 h (3.8-fold, P < 0.05), 6 h (4.9-fold, P < 0.05), 12 h (12.3-fold, P < 0.01) and 24 h (9.7-fold, P < 0.01), while RpSabl transcripts in hemocytes was only significantly up-regulated at 6 h (8.5-Fold, P < 0.01). RNAi-mediated knockdown of RpSabl transcripts affected the survival rates of Manila clam against V. anguillarum, perhaps mainly due to the inhibited expression of antibacterial effectors (e.g. lysozyme and defensin). Moreover, recombinant protein of RpSabl (rRpSabl) possessed binding activities towards lipopolysaccharides (LPS), peptidoglycan (PGN) and glucan in vitro. Coinciding with the Pathogen-associated molecular patterns (PAMPs) binding assay, rRpSabl displayed broad bacterial-agglutination properties towards Vibrio harveyi, Vibrio splendidus, V. anguillarum, Enterobacter cloacae and Aeromonas hydrophila. Meanwhile, the phagocytosis and encapsulation ability of hemocytes could be significantly enhanced by rRpSabl incubation. All these results showed that RpSabl could function as a versatile molecule involved in the innate immune responses of R. philippinarum.


Assuntos
Antibacterianos/farmacologia , Bivalves/genética , Bivalves/imunologia , Lectinas/genética , Proteínas Opsonizantes/farmacologia , Ácidos Siálicos/metabolismo , Aeromonas hydrophila/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Enterobacter cloacae/efeitos dos fármacos , Lectinas/química , Lectinas/metabolismo , Alinhamento de Sequência , Vibrio/efeitos dos fármacos
13.
Fish Shellfish Immunol ; 94: 230-238, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499201

RESUMO

In the study, two c-type lectins were identified and characterized from the manila clam Venerupis philippinarum (designed as VpClec-1 and VpClec-2, respectively). Multiple alignments and phylogenetic analysis strongly suggested that they were new members of the c-type lectin superfamily. In normal tissue of clams, both VpClec-1 and VpClec-2 transcripts were highly expressed in the tissue of hepatopancreas. After Vibrio anguillarum challenge, the temporal expression of both VpClec-1 and VpClec-2 transcripts was up-regulated in the hemocytes of manila clams. The recombinant protein VpClec-1 (rVpClec-1) showed obvious binding activities to lipopolysaccharide (LPS), peptidoglycan (PGN), glucan and zymosan in vitro, while the recombinant protein VpClec-2 (rVpClec-2) could only bind LPS, glucan and zymosan. Coinciding with the PAMPs binding assay, both rVpClec-1 and rVpClec-2 displayed broad agglutination and antibacterial activities towards Vibrio harveyi, Vibrio splendidus, Vibrio anguillarum, Enterobacter cloacae and Aeromonas hydrophila. Moreover, the phagocytosis and encapsulation ability of hemocytes could be significantly enhanced by rVpClec-1 and rVpClec-2. Notably, the rVpClec-1 but not rVpClec-2 elicited a chemotactic response from hemocytes. All the results showed that VpClec-1 and VpClec-2 functioned as pattern recognition receptors (PRRs) with distinct recognition spectrum, and involved in the innate immune responses of manila clams.


Assuntos
Antibacterianos/farmacologia , Bivalves/genética , Bivalves/imunologia , Imunidade Inata/genética , Lectinas Tipo C/genética , Proteínas Opsonizantes/genética , Ácidos Siálicos/metabolismo , Aeromonas hydrophila/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Enterobacter cloacae/efeitos dos fármacos , Lectinas Tipo C/química , Lectinas Tipo C/metabolismo , Receptores de Reconhecimento de Padrão/genética , Receptores de Reconhecimento de Padrão/imunologia , Alinhamento de Sequência , Transcriptoma , Vibrio/efeitos dos fármacos
14.
Artigo em Inglês | MEDLINE | ID: mdl-31476363

RESUMO

Understanding variation in physiological traits across taxa is a central question in evolutionary biology that has wide-ranging implications in biomedicine, disease ecology, and environmental protection. Sialic acid (Sia), and in particular, 5-N-acetylneuraminic acid (Neu5Ac), is chemically bound to galactose and the underlying glycan via α2-3 or α2-6 glycosidic linkage (i.e., Siaα2-3Galactose or Siaα2-6Galactose), conferring two different cell surface structures that affects cell to cell communication and interactions with foreign agents including microparasites and toxins. As an initial step towards understanding variation of Sia across the class Aves, we collected red blood cells (RBCs or erythrocytes) and measured Sia quantity in 76 species and 340 individuals using HPLC-MS/MS and glycosidic linkage type in 24 species and 105 individuals using hemagglutination assay. Although Sia quantity did not, α2-6 glycosidic linkage did exhibit a discernable phylogenetic pattern as evaluated by a phylogenetic signal (λ) value of 0.7. Sia quantity appeared to be higher in after hatch year birds than hatch year birds (P < 0.05); moreover, ~80% of the measured Sia across all individuals or species was expressed by ~20% of the individuals or species. Lastly, as expected, we detected a minimal presence of 5-N-glycolylneuraminic acid in the avian RBCs tested. These data provide novel insights and a large baseline dataset for further study on the variability of Sia in the class Aves which might be useful for understanding Sia dependent processes in birds.


Assuntos
Aves/metabolismo , Eritrócitos/metabolismo , Ácidos Siálicos/metabolismo , Animais , Aves/classificação , Eritrócitos/química , Ácidos Siálicos/química , Especificidade da Espécie
15.
Colloids Surf B Biointerfaces ; 183: 110413, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31401461

RESUMO

A novel bioinspired nanoplatform capable of fast endocytosis, lysosomal pH-triggered drug release, and reduced drug efflux based on PBA-PEG-b-P(Glu-co-GluDA) copolymer was developed in this study. The synthesized copolymer could facilitate doxorubicin encapsulation with relatively high drug-loading content and efficiency. Inspired by mussel byssal threads, a core crosslinking strategy based on the coordination between catechol and ferric ions was introduced to improve the stability of nanomicelles and realize lysosomal pH-controlled drug release. This nanoplatform could maintain integrity even after being dissolved in a good solvent, demonstrating its the potential to withstand infinite dilution of plasma after intravenous injection. Moreover, this nanoplatform demonstrated lysosomal pH-triggered drug release, and the cumulative release amount of doxorubicin under a simulated lysosomal condition was 13 times higher than that under a simulated plasma condition. Moreover, as a result of the high binding capacity between phenylboronic acid (PBA) and sialic acid on the surface of human hepatoma cell line (HepG2), the fast and enhanced endocytosis in addition to lysosomal pH-triggered release property and significantly low efflux, this nanoplatform exhibits improved delivery efficiency of doxorubicin into the nucleus and notably outstanding antiproliferative effects compared with doxorubicin. Furthermore, the PBA modification remarkably increased the mean fluorescence intensity of this nanoplatform endocytosed by HepG2 cells to twice that of doxorubicin after one hour of incubation. The nanoplatform exhibited an inhibition rate of 70% against tumor growth. Thus, this novel nanoplatform based on PBA-PEG-b-P(Glu-co-GluDA) copolymer displayed multifunctionality and exhibited great potential as an intelligent nanoplatform for antitumor drug delivery.


Assuntos
Antibióticos Antineoplásicos/farmacologia , Núcleo Celular/efeitos dos fármacos , Preparações de Ação Retardada , Doxorrubicina/farmacologia , Lisossomos/efeitos dos fármacos , Nanopartículas/química , Antibióticos Antineoplásicos/metabolismo , Materiais Biomiméticos/química , Ácidos Borônicos/química , Ácidos Borônicos/metabolismo , Catecóis/química , Catecóis/metabolismo , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Doxorrubicina/metabolismo , Composição de Medicamentos/métodos , Endocitose , Células Hep G2 , Humanos , Concentração de Íons de Hidrogênio , Ferro/química , Ferro/metabolismo , Lisossomos/metabolismo , Micelas , Nanopartículas/ultraestrutura , Polietilenoglicóis/química , Polietilenoglicóis/metabolismo , Ácidos Siálicos/química , Ácidos Siálicos/metabolismo
16.
Glycobiology ; 29(11): 750-754, 2019 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-31361010

RESUMO

Glycosylation is a common modification found on numerous proteins and lipids. However, direct detection of glycans on these intact biomolecules has been challenge. Here, utilizing enzymatic incorporation of fluorophore-conjugated sialic acids, dubbed as direct fluorescent glycan labeling, we report the labeling and detection of N- and O-glycans on glycoproteins. The method allows detection of specific glycans without the laborious gel blotting and chemiluminescence reactions used in Western blotting. The method can also be used with a variety of fluorescent dyes.


Assuntos
Fluorescência , Polissacarídeos/análise , Sialiltransferases/química , Animais , Bovinos , Clostridium perfringens/enzimologia , Corantes Fluorescentes/química , Glicosilação , Humanos , Polissacarídeos/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Ácidos Siálicos/química , Ácidos Siálicos/metabolismo , Sialiltransferases/metabolismo
17.
Ann Rheum Dis ; 78(11): 1488-1496, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31300460

RESUMO

OBJECTIVE: An increased proportion of circulating follicular helper T (Tfh) cells was reported in rheumatoid arthritis (RA), but it remains uncertain how Tfh cells affect antibody hyposialylation. We investigated the regulation of autoantibody hyposialylation by Tfh cells in RA using murine model. METHODS: Behaviours of Tfh cells and their function on B cell promotion were analysed. Change of arthritogenicity and sialylation of autoantibodies during the course of arthritis was examined by mass spectrometry. Tfh-mediated regulation of hyposialylation was investigated, and the responsible cell surface molecule was specified both in vitro and in vivo. The relation between circulating Tfh cells and hyposialylation was analysed in patients with RA. RESULTS: An increase in Tfh, particularly interleukin-17 producing Tfh (Tfh17) cells, at the onset of arthritis and their enhancement of autoantibody production were found. Autoantibodies at the onset phase demonstrated stronger inflammatory properties than those at the resolution phase, and mass spectrometric analysis revealed their difference in sialylation. In vitro coculture showed enhanced hyposialylation by the Tfh cells via OX40, which was highly expressed in the Tfh and Tfh17 cells. Blockade of OX40 prevented the development of arthritis with reduction in Tfh17 cells and recovery of autoantibody sialylation. Analysis of patients with RA showed abundance of OX40-overexpressing Tfh17 cells, and their proportion correlated negatively with the expression of α2,6-sialyltransferase 1, an enzyme responsible for sialylation. CONCLUSIONS: OX40 expressed on Tfh cells can regulate autoantibody sialylation and play a crucial role in the development of autoimmune arthritis.


Assuntos
Artrite Reumatoide/imunologia , Autoanticorpos/metabolismo , Receptores OX40/metabolismo , Ácidos Siálicos/metabolismo , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Modelos Animais de Doenças , Imunidade Celular/genética , Masculino , Camundongos , Camundongos Endogâmicos DBA
18.
Infect Immun ; 87(10)2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31308084

RESUMO

Our studies reveal that the oral colonizer and cause of infective endocarditis Streptococcus oralis subsp. dentisani displays a striking monolateral distribution of surface fibrils. Furthermore, our data suggest that these fibrils impact the structure of adherent bacterial chains. Mutagenesis studies indicate that these fibrils are dependent on three serine-rich repeat proteins (SRRPs), here named fibril-associated protein A (FapA), FapB, and FapC, and that each SRRP forms a different fibril with a distinct distribution. SRRPs are a family of bacterial adhesins that have diverse roles in adhesion and that can bind to different receptors through modular nonrepeat region domains. Amino acid sequence and predicted structural similarity searches using the nonrepeat regions suggested that FapA may contribute to interspecies interactions, that FapA and FapB may contribute to intraspecies interactions, and that FapC may contribute to sialic acid binding. We demonstrate that a fapC mutant was significantly reduced in binding to saliva. We confirmed a role for FapC in sialic acid binding by demonstrating that the parental strain was significantly reduced in adhesion upon addition of a recombinantly expressed, sialic acid-specific, carbohydrate binding module, while the fapC mutant was not reduced. However, mutation of a residue previously shown to be essential for sialic acid binding did not decrease bacterial adhesion, leaving the precise mechanism of FapC-mediated adhesion to sialic acid to be defined. We also demonstrate that the presence of any one of the SRRPs is sufficient for efficient biofilm formation. Similar structures were observed on all infective endocarditis isolates examined, suggesting that this distribution is a conserved feature of this S. oralis subspecies.


Assuntos
Proteínas de Bactérias/ultraestrutura , Biofilmes/crescimento & desenvolvimento , Saliva/metabolismo , Ácidos Siálicos/metabolismo , Streptococcus oralis/genética , Sequência de Aminoácidos , Aderência Bacteriana , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Endocardite Bacteriana/microbiologia , Endocardite Bacteriana/patologia , Expressão Gênica , Humanos , Mutação , Ligação Proteica , Domínios Proteicos , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/ultraestrutura , Saliva/química , Ácidos Siálicos/química , Streptococcus oralis/química , Streptococcus oralis/metabolismo
19.
Proc Natl Acad Sci U S A ; 116(32): 16036-16045, 2019 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-31332008

RESUMO

Cardiovascular disease (CVD) events due to atherosclerosis cause one-third of worldwide deaths and risk factors include physical inactivity, age, dyslipidemia, hypertension, diabetes, obesity, smoking, and red meat consumption. However, ∼15% of first-time events occur without such factors. In contrast, coronary events are extremely rare even in closely related chimpanzees in captivity, despite human-like CVD-risk-prone blood lipid profiles, hypertension, and mild atherosclerosis. Similarly, red meat-associated enhancement of CVD event risk does not seem to occur in other carnivorous mammals. Thus, heightened CVD risk may be intrinsic to humans, and genetic changes during our evolution need consideration. Humans exhibit a species-specific deficiency of the sialic acid N-glycolylneuraminic acid (Neu5Gc), due to pseudogenization of cytidine monophosphate-N-acetylneuraminic acid (Neu5Ac) hydroxylase (CMAH), which occurred in hominin ancestors ∼2 to 3 Mya. Ldlr -/- mice with human-like Cmah deficiency fed a sialic acids (Sias)-free high-fat diet (HFD) showed ∼1.9-fold increased atherogenesis over Cmah wild-type Ldlr -/- mice, associated with elevated macrophage cytokine expression and enhanced hyperglycemia. Human consumption of Neu5Gc (from red meat) acts as a "xeno-autoantigen" via metabolic incorporation into endogenous glycoconjugates, as interactions with circulating anti-Neu5Gc "xeno-autoantibodies" potentiate chronic inflammation ("xenosialitis"). Cmah -/- Ldlr -/- mice immunized with Neu5Gc-bearing antigens to generate human-like anti-Neu5Gc antibodies suffered a ∼2.4-fold increased atherosclerosis on a Neu5Gc-rich HFD, compared with Neu5Ac-rich or Sias-free HFD. Lesions in Neu5Gc-immunized and Neu5Gc-rich HFD-fed Cmah -/- Ldlr -/- mice were more advanced but unexplained by lipoprotein or glucose changes. Human evolutionary loss of CMAH likely contributes to atherosclerosis predisposition via multiple intrinsic and extrinsic mechanisms, and future studies could consider this more human-like model.


Assuntos
Aterosclerose/enzimologia , Oxigenases de Função Mista/deficiência , Animais , Bovinos , Citocinas/metabolismo , Dieta Hiperlipídica , Feminino , Humanos , Hiperglicemia/patologia , Inflamação/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos Endogâmicos C57BL , Oxigenases de Função Mista/metabolismo , Modelos Biológicos , Fenótipo , Receptores de LDL/deficiência , Receptores de LDL/metabolismo , Ácidos Siálicos/metabolismo , Especificidade da Espécie
20.
J Biol Chem ; 294(31): 11910-11919, 2019 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-31201275

RESUMO

A sperm that fertilizes an egg has successfully survived multiple checkpoints within the female reproductive tract, termed pre-fertilization events. The leukocytic response is a pre-fertilization event in which sperm trigger an immune response that promotes homing of circulating leukocytes to the uterine lumen to destroy most sperm. Various glycoconjugates decorate the sperm surface, including sialic acids, which are abundant at the sperm surface where they cap most glycan chains and regulate sperm migration through cervical mucus, formation of the sperm oviductal reservoir, and sperm capacitation. However, the role of sperm-associated sialic acids in the leukocytic reaction remains unknown. The cognate endogenous binding partners of sialic acids, sialic acid-binding immunoglobulin-like lectins (Siglecs) play a pivotal role in regulating many immune responses. Here we investigated whether sperm-associated sialic acids inhibit activation of neutrophils, one of the major immune cells involved in the leukocytic reaction. We used in vitro interactions between sperm and neutrophils as well as binding assays between sperm and recombinant Siglec-Fc chimeric proteins to measure interactions. Moreover, we examined whether Siglecs are expressed on human and mouse endometria, which have a role in initiating the leukocytic reaction. Surprisingly less sialylated, capacitated, sperm did not increase neutrophil activation in vitro However, we observed expression of several Siglecs on the endometrium and that these receptors interact with sialylated sperm. Our results indicate that sperm sialic acids may interact with endometrial Siglecs and that these interactions facilitate sperm survival in the face of female immunity.


Assuntos
Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/metabolismo , Espermatozoides/metabolismo , Animais , Endométrio/metabolismo , Feminino , Fragmentos Fc das Imunoglobulinas/genética , Fragmentos Fc das Imunoglobulinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neuraminidase/metabolismo , Ativação de Neutrófilo , Neutrófilos/metabolismo , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/isolamento & purificação , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/genética , Ácidos Siálicos/metabolismo
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