Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 3.850
Filtrar
1.
PLoS One ; 15(12): e0244762, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33378413

RESUMO

Non-alcoholic fatty liver disease (NAFLD) is associated with obesity and type 2 diabetes and is characterized by the accumulation of fat in the liver (steatosis). NAFLD can transition into non-alcoholic steatohepatitis (NASH), with liver cell injury, inflammation, and an increased risk of fibrosis. We previously found that injections of either 1866, a synthetic ligand for the lectin receptor CD209, or DANA, a sialidase inhibitor, can inhibit inflammation and fibrosis in multiple animal models. The methionine and choline-deficient (MCD) diet is a model of NASH which results in the rapid induction of liver steatosis and inflammation. In this report, we show that for C57BL/6 mice on a MCD diet, injections of both 1866 and DANA reversed MCD diet-induced decreases in white fat, decreases in adipocyte size, and white fat inflammation. However, these effects were not observed in type 2 diabetic db/db mice on a MCD diet. In db/db mice on a MCD diet, 1866 decreased liver steatosis, but these effects were not observed in C57BL/6 mice. There was no correlation between the ability of 1866 or DANA to affect steatosis and the effects of these compounds on the density of liver macrophage cells expressing CLEC4F, CD64, F4/80, or Mac2. Together these results indicate that 1866 and DANA modulate adipocyte size and adipose tissue macrophage populations, that 1866 could be useful for modulating steatosis, and that changes in the local density of 4 different liver macrophages cell types do not correlate with effects on liver steatosis.


Assuntos
Tecido Adiposo/efeitos dos fármacos , Moléculas de Adesão Celular/agonistas , Lectinas Tipo C/agonistas , Fígado/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Ácido N-Acetilneuramínico/análogos & derivados , Neuraminidase/antagonistas & inibidores , Receptores de Superfície Celular/agonistas , Tecido Adiposo/metabolismo , Animais , Deficiência de Colina/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Fígado Gorduroso/tratamento farmacológico , Fígado Gorduroso/metabolismo , Inflamação/metabolismo , Fígado/metabolismo , Macrófagos/metabolismo , Camundongos , Ácido N-Acetilneuramínico/farmacologia , Ácido N-Acetilneuramínico/uso terapêutico , Neuraminidase/metabolismo , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Hepatopatia Gordurosa não Alcoólica/metabolismo
2.
Nat Commun ; 11(1): 5597, 2020 11 05.
Artigo em Inglês | MEDLINE | ID: mdl-33154358

RESUMO

Seasonal influenza epidemics lead to 3-5 million severe infections and 290,000-650,000 annual global deaths. With deaths from the 1918 influenza pandemic estimated at >50,000,000 and future pandemics anticipated, the need for a potent influenza treatment is critical. In this study, we design and synthesize a bifunctional small molecule by conjugating the neuraminidase inhibitor, zanamivir, with the highly immunogenic hapten, dinitrophenyl (DNP), which specifically targets the surface of free virus and viral-infected cells. We show that this leads to simultaneous inhibition of virus release, and immune-mediated elimination of both free virus and virus-infected cells. Intranasal or intraperitoneal administration of a single dose of drug to mice infected with 100x MLD50 virus is shown to eradicate advanced infections from representative strains of both influenza A and B viruses. Since treatments of severe infections remain effective up to three days post lethal inoculation, our approach may successfully treat infections refractory to current therapies.


Assuntos
Antivirais/administração & dosagem , Antivirais/farmacologia , Imunoterapia/métodos , Infecções por Orthomyxoviridae/tratamento farmacológico , 2,4-Dinitrofenol/administração & dosagem , 2,4-Dinitrofenol/química , 2,4-Dinitrofenol/imunologia , Administração Intranasal , Animais , Anticorpos/administração & dosagem , Anticorpos/imunologia , Antivirais/química , Linhagem Celular , Citotoxicidade Imunológica/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Humanos , Vírus da Influenza A/efeitos dos fármacos , Vírus da Influenza A/enzimologia , Vírus da Influenza A/fisiologia , Vírus da Influenza B/efeitos dos fármacos , Vírus da Influenza B/enzimologia , Vírus da Influenza B/fisiologia , Infusões Parenterais , Camundongos , Camundongos Endogâmicos BALB C , Neuraminidase/antagonistas & inibidores , Neuraminidase/metabolismo , Infecções por Orthomyxoviridae/patologia , Infecções por Orthomyxoviridae/virologia , Ligação Proteica , Resultado do Tratamento , Liberação de Vírus/efeitos dos fármacos , Zanamivir/administração & dosagem , Zanamivir/química , Zanamivir/farmacologia
3.
PLoS Biol ; 18(8): e3000788, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32841232

RESUMO

Women with bacterial vaginosis (BV), an imbalance of the vaginal microbiome, are more likely to be colonized by potential pathogens such as Fusobacterium nucleatum, a bacterium linked with intrauterine infection and preterm birth. However, the conditions and mechanisms supporting pathogen colonization during vaginal dysbiosis remain obscure. We demonstrate that sialidase activity, a diagnostic feature of BV, promoted F. nucleatum foraging and growth on mammalian sialoglycans, a nutrient resource that was otherwise inaccessible because of the lack of endogenous F. nucleatum sialidase. In mice with sialidase-producing vaginal microbiotas, mutant F. nucleatum unable to consume sialic acids was impaired in vaginal colonization. These experiments in mice also led to the discovery that F. nucleatum may also "give back" to the community by reinforcing sialidase activity, a biochemical feature of human dysbiosis. Using human vaginal bacterial communities, we show that F. nucleatum supported robust outgrowth of Gardnerella vaginalis, a major sialidase producer and one of the most abundant organisms in BV. These results illustrate that mutually beneficial relationships between vaginal bacteria support pathogen colonization and may help maintain features of dysbiosis. These findings challenge the simplistic dogma that the mere absence of "healthy" lactobacilli is the sole mechanism that creates a permissive environment for pathogens during vaginal dysbiosis. Given the ubiquity of F. nucleatum in the human mouth, these studies also suggest a possible mechanism underlying links between vaginal dysbiosis and oral sex.


Assuntos
Proteínas de Bactérias/genética , Disbiose/microbiologia , Fusobacterium/metabolismo , Gardnerella vaginalis/metabolismo , Neuraminidase/genética , Polissacarídeos/metabolismo , Vaginose Bacteriana/microbiologia , Animais , Proteínas de Bactérias/metabolismo , Técnicas de Tipagem Bacteriana , Disbiose/patologia , Feminino , Fusobacterium/genética , Fusobacterium/isolamento & purificação , Fusobacterium/patogenicidade , Gardnerella vaginalis/genética , Gardnerella vaginalis/isolamento & purificação , Gardnerella vaginalis/patogenicidade , Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Microbiota/genética , Neuraminidase/metabolismo , RNA Ribossômico 16S/genética , Ácidos Siálicos/metabolismo , Simbiose/genética , Vagina/microbiologia , Vaginose Bacteriana/patologia
4.
PLoS Pathog ; 16(8): e1008816, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32853241

RESUMO

Influenza A viruses (IAVs) cause seasonal epidemics and occasional pandemics. Most pandemics occurred upon adaptation of avian IAVs to humans. This adaptation includes a hallmark receptor-binding specificity switch of hemagglutinin (HA) from avian-type α2,3- to human-type α2,6-linked sialic acids. Complementary changes of the receptor-destroying neuraminidase (NA) are considered to restore the precarious, but poorly described, HA-NA-receptor balance required for virus fitness. In comparison to the detailed functional description of adaptive mutations in HA, little is known about the functional consequences of mutations in NA in relation to their effect on the HA-NA balance and host tropism. An understudied feature of NA is the presence of a second sialic acid-binding site (2SBS) in avian IAVs and absence of a 2SBS in human IAVs, which affects NA catalytic activity. Here we demonstrate that mutation of the 2SBS of avian IAV H5N1 disturbs the HA-NA balance. Passaging of a 2SBS-negative H5N1 virus on MDCK cells selected for progeny with a restored HA-NA balance. These viruses obtained mutations in NA that restored a functional 2SBS and/or in HA that reduced binding of avian-type receptors. Importantly, a particular HA mutation also resulted in increased binding of human-type receptors. Phylogenetic analyses of avian IAVs show that also in the field, mutations in the 2SBS precede mutations in HA that reduce binding of avian-type receptors and increase binding of human-type receptors. Thus, 2SBS mutations in NA can drive acquisition of mutations in HA that not only restore the HA-NA balance, but may also confer increased zoonotic potential.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A/genética , Mutação , Neuraminidase/genética , Infecções por Orthomyxoviridae/virologia , Ácidos Siálicos/metabolismo , Replicação Viral , Substituição de Aminoácidos , Animais , Sítios de Ligação , Cães , Glicoproteínas de Hemaglutininação de Vírus da Influenza/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Vírus da Influenza A/isolamento & purificação , Células Madin Darby de Rim Canino , Neuraminidase/química , Neuraminidase/metabolismo , Infecções por Orthomyxoviridae/genética , Infecções por Orthomyxoviridae/patologia , Ligação Proteica
5.
J Chem Inf Model ; 60(10): 4582-4593, 2020 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-32845150

RESUMO

Artificial intelligence and multiobjective optimization represent promising solutions to bridge chemical and biological landscapes by addressing the automated de novo design of compounds as a result of a humanlike creative process. In the present study, we conceived a novel pair-based multiobjective approach implemented in an adapted SMILES generative algorithm based on recurrent neural networks for the automated de novo design of new molecules whose overall features are optimized by finding the best trade-offs among relevant physicochemical properties (MW, logP, HBA, HBD) and additional similarity-based constraints biasing specific biological targets. In this respect, we carried out the de novo design of chemical libraries targeting neuraminidase, acetylcholinesterase, and the main protease of severe acute respiratory syndrome coronavirus 2. Several quality metrics were employed to assess drug-likeness, chemical feasibility, diversity content, and validity. Molecular docking was finally carried out to better evaluate the scoring and posing of the de novo generated molecules with respect to X-ray cognate ligands of the corresponding molecular counterparts. Our results indicate that artificial intelligence and multiobjective optimization allow us to capture the latent links joining chemical and biological aspects, thus providing easy-to-use options for customizable design strategies, which are especially effective for both lead generation and lead optimization. The algorithm is freely downloadable at https://github.com/alberdom88/moo-denovo and all of the data are available as Supporting Information.


Assuntos
Desenho de Fármacos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Redes Neurais de Computação , Acetilcolinesterase/metabolismo , Antivirais/química , Antivirais/farmacologia , Inteligência Artificial , Betacoronavirus/enzimologia , Inibidores da Colinesterase/química , Inibidores da Colinesterase/farmacologia , Infecções por Coronavirus/tratamento farmacológico , Cisteína Endopeptidases/metabolismo , Humanos , Vírus da Influenza A/enzimologia , Simulação de Acoplamento Molecular , Neuraminidase/antagonistas & inibidores , Neuraminidase/metabolismo , Pandemias , Pneumonia Viral/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas não Estruturais Virais/antagonistas & inibidores , Proteínas não Estruturais Virais/metabolismo
6.
J Virol ; 94(18)2020 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-32641475

RESUMO

Some avian influenza (AI) viruses have a deletion of up to 20 to 30 amino acids in their neuraminidase (NA) stalk. This has been associated with changes in virus replication and host range. Currently prevalent H9N2 AI viruses have only a 2- or 3-amino-acid deletion, and such deletions were detected in G1 and Y280 lineage viruses, respectively. The effect of an NA deletion on the H9N2 phenotype has not been fully elucidated. In this study, we isolated G1 mutants that carried an 8-amino-acid deletion in their NA stalk. To systematically analyze the effect of NA stalk length and concomitant (de)glycosylation on G1 replication and host range, we generated G1 viruses that had various NA stalk lengths and that were either glycosylated or not glycosylated. The stalk length was correlated with NA sialidase activity, using low-molecular-weight substrates, and with virus elution efficacy from erythrocytes. G1 virus replication in avian cells and eggs was positively correlated with the NA stalk length but was negatively correlated in human cells and mice. NA stalk length modulated G1 virus entry into host cells, with shorter stalks enabling more efficient G1 entry into human cells. However, with a hemagglutinin (HA) with a higher α2,6-linked sialylglycan affinity, the effect of NA stalk length on G1 virus infection was reversed, with shorter NA stalks reducing virus entry into human cells. These results indicate that a balance between HA binding affinity and NA sialidase activity, modulated by NA stalk length, is required for optimal G1 virus entry into human airway cells.IMPORTANCE H9N2 avian influenza (AI) virus, one of the most prevalent AI viruses, has caused repeated poultry and human infections, posing a huge public health risk. The H9N2 virus has diversified into multiple lineages, with the G1 lineage being the most prevalent worldwide. In this study, we isolated G1 variants carrying an 8-amino-acid deletion in their NA stalk, which is, to our knowledge, the longest deletion found in H9N2 viruses in the field. The NA stalk length was found to modulate G1 virus entry into host cells, with the effects being species specific and dependent on the corresponding HA binding affinity. Our results suggest that, in nature, H9N2 G1 viruses balance their HA and NA functions by the NA stalk length, leading to the possible association of host range and virulence in poultry and mammals during the evolution of G1 lineage viruses.


Assuntos
Regulação Viral da Expressão Gênica , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A Subtipo H9N2/genética , Influenza Aviária/virologia , Neuraminidase/genética , Infecções por Orthomyxoviridae/virologia , Sequência de Aminoácidos , Animais , Galinhas , Genótipo , Glicosilação , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Hemaglutininas , Especificidade de Hospedeiro , Interações Hospedeiro-Patógeno/genética , Humanos , Vírus da Influenza A Subtipo H9N2/metabolismo , Vírus da Influenza A Subtipo H9N2/patogenicidade , Influenza Aviária/genética , Influenza Aviária/metabolismo , Influenza Aviária/patologia , Camundongos , Neuraminidase/metabolismo , Infecções por Orthomyxoviridae/genética , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/patologia , Fenótipo , Filogenia , Receptores Virais , Deleção de Sequência , Relação Estrutura-Atividade , Virulência , Internalização do Vírus , Replicação Viral
7.
PLoS Pathog ; 16(6): e1008592, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32555740

RESUMO

The neuraminidase (NA) inhibitor (NAI) oseltamivir (OST) is the most widely used influenza antiviral drug. Several NA amino acid substitutions are reported to reduce viral susceptibility to OST in in vitro assays. However, whether there is a correlation between the level of reduction in susceptibility in vitro and the efficacy of OST against these viruses in vivo is not well understood. In this study, a ferret model was utilised to evaluate OST efficacy against circulating influenza A and B viruses with a range of in vitro generated 50% inhibitory concentrations (IC50) values for OST. OST efficacy against an A(H1N1)pdm09 and an A(H1N1)pdm09 virus with the H275Y substitution in neuraminidase was also tested in the macaque model. The results from this study showed that OST had a significant impact on virological parameters compared to placebo treatment of ferrets infected with wild-type influenza A viruses with normal IC50 values (~1 nM). However, this efficacy was lower against wild-type influenza B and other viruses with higher IC50 values. Differing pathogenicity of the viruses made evaluation of clinical parameters difficult, although some effect of OST in reducing clinical signs was observed with influenza A(H1N1) and A(H1N1)pdm09 (H275Y) viruses. Viral titres in macaques were too low to draw conclusive results. Analysis of the ferret data revealed a correlation between IC50 and OST efficacy in reducing viral shedding but highlighted that the current WHO guidelines/criteria for defining normal, reduced or highly reduced inhibition in influenza B viruses based on in vitro data are not well aligned with the low in vivo OST efficacy observed for both wild-type influenza B viruses and those with reduced OST susceptibility.


Assuntos
Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H3N2/metabolismo , Vírus da Influenza B/metabolismo , Infecções por Orthomyxoviridae , Oseltamivir/farmacologia , Substituição de Aminoácidos , Animais , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Feminino , Furões , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza B/genética , Macaca fascicularis , Macrolídeos , Masculino , Mutação de Sentido Incorreto , Neuraminidase/genética , Neuraminidase/metabolismo , Infecções por Orthomyxoviridae/tratamento farmacológico , Infecções por Orthomyxoviridae/genética , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/patologia
8.
Nature ; 582(7811): 265-270, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32499653

RESUMO

Approximately one-third of the world's population suffers from allergies1. Exposure to allergens crosslinks immunoglobulin E (IgE) antibodies that are bound to mast cells and basophils, triggering the release of inflammatory mediators, including histamine2. Although IgE is absolutely required for allergies, it is not understood why total and allergen-specific IgE concentrations do not reproducibly correlate with allergic disease3-5. It is well-established that glycosylation of IgG dictates its effector function and has disease-specific patterns. However, whether IgE glycans differ in disease states or affect biological activity is completely unknown6. Here we perform an unbiased examination of glycosylation patterns of total IgE from individuals with a peanut allergy and from non-atopic individuals without allergies. Our analysis reveals an increase in sialic acid content on total IgE from individuals with a peanut allergy compared with non-atopic individuals. Removal of sialic acid from IgE attenuates effector-cell degranulation and anaphylaxis in several functional models of allergic disease. Therapeutic interventions-including removing sialic acid from cell-bound IgE with a neuraminidase enzyme targeted towards the IgE receptor FcεRI, and administering asialylated IgE-markedly reduce anaphylaxis. Together, these results establish IgE glycosylation, and specifically sialylation, as an important regulator of allergic disease.


Assuntos
Imunoglobulina E/química , Imunoglobulina E/imunologia , Ácido N-Acetilneuramínico/análise , Hipersensibilidade a Amendoim/imunologia , Hipersensibilidade a Amendoim/patologia , Adolescente , Adulto , Idoso , Alérgenos/imunologia , Anafilaxia/imunologia , Animais , Estudos de Casos e Controles , Degranulação Celular/imunologia , Criança , Pré-Escolar , Feminino , Glicosilação , Humanos , Imunoglobulina E/efeitos adversos , Imunoglobulina E/farmacologia , Lactente , Recém-Nascido , Masculino , Camundongos , Pessoa de Meia-Idade , Modelos Imunológicos , Ácido N-Acetilneuramínico/química , Ácido N-Acetilneuramínico/metabolismo , Neuraminidase/metabolismo , Receptores de IgE/metabolismo , Adulto Jovem
9.
PLoS One ; 15(3): e0228461, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32160194

RESUMO

Simulating drug binding and unbinding is a challenge, as the rugged energy landscapes that separate bound and unbound states require extensive sampling that consumes significant computational resources. Here, we describe the use of interactive molecular dynamics in virtual reality (iMD-VR) as an accurate low-cost strategy for flexible protein-ligand docking. We outline an experimental protocol which enables expert iMD-VR users to guide ligands into and out of the binding pockets of trypsin, neuraminidase, and HIV-1 protease, and recreate their respective crystallographic protein-ligand binding poses within 5-10 minutes. Following a brief training phase, our studies shown that iMD-VR novices were able to generate unbinding and rebinding pathways on similar timescales as iMD-VR experts, with the majority able to recover binding poses within 2.15 Å RMSD of the crystallographic binding pose. These results indicate that iMD-VR affords sufficient control for users to carry out the detailed atomic manipulations required to dock flexible ligands into dynamic enzyme active sites and recover crystallographic poses, offering an interesting new approach for simulating drug docking and generating binding hypotheses.


Assuntos
Protease de HIV/metabolismo , Simulação de Dinâmica Molecular , Neuraminidase/metabolismo , Tripsina/metabolismo , Realidade Virtual , Benzamidinas/metabolismo , Sítios de Ligação , Carbamatos/metabolismo , Furanos , Ligantes , Oseltamivir/metabolismo , Ligação Proteica , Sulfonamidas/metabolismo , Zanamivir/metabolismo
10.
Gene ; 742: 144538, 2020 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-32184168

RESUMO

Lysosomal desialylation is the initial step in the degradation of sialo-glycopeptides that is essential for regenerating sialo-glycoconjugates. Neu1 sialidase is the enzyme responsible for the removal of sialic acid in the mammalian lysosome. Although Neu1 sialidases are conserved in fish similar to mammals, their physiological functions remain to be fully understood. Nile tilapia (Oreochromis niloticus) is known to possess two putative Neu1 sialidases (Neu1a and Neu1b) in the genome that may have arisen by gene duplication (specifically in cichlidae family members). This suggests that understanding the Neu1 sialidase in fish, particularly cichlids, could provide insights into the (novel) physiological functions of these genes. Moreover, characterization of the tilapia Neu1 sialidase is paramount to ensure clarity of the desialylation reaction performed by the fish sialidases (like the characterized tilapia sialidases Neu3 and Neu4). Therefore, this study focused on the characterization of the tilapia Neu1 sialidases. Neu1b exhibited narrow substrate specificity when compared with Neu1a, whereas the properties of these two Neu1 sialidases, such as cathepsin A-induced activation, optimal pH, and lysosomal localization, were conserved. Neu1a mRNA levels were detected in various tissues of tilapia as compared to the mRNA levels of Neu1b. Although the cloned construct of Neu1b contained an extra exon unlike tilapia Neu1a, the exon did not affect the enzymatic properties of Neu1b. This study suggests that tilapia Neu1a profiles were highly conserved with other vertebrate Neu1 isoforms, while Neu1b probably evolved independently in other members of the cichlidae family. Moreover, the expression of sialidase genes (neu1a, neu1b, neu3a, and neu4) were determined in various stages of tilapia embryogenesis using real-time PCR; sialidase gene expression is reported to be drastically and individually altered during embryogenesis in Japanese medaka (Oryzias latipes). The mRNA levels of neu1a drastically increased between 72 and 84 hpf and mildly decreased from 84 to 144 hpf. In contrast, the transcript levels of neu1b did not change between 84 and 144 hpf and the expression of neu3a gradually increased between 84 and 120 hpf and drastically decreased at 144 hpf. The highest level of the neu4 transcripts was detected at 84 hpf. These expression patterns were different from those in Japanese medaka, possibly due to the different developmental program found in the tilapia embryo accompanied with the unique profiles of the tilapia sialidases.


Assuntos
Ciclídeos/metabolismo , Proteínas de Peixes/metabolismo , Neuraminidase/metabolismo , Animais , Ciclídeos/genética , Ciclídeos/crescimento & desenvolvimento , Clonagem Molecular , Evolução Molecular , Feminino , Proteínas de Peixes/química , Proteínas de Peixes/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Neuraminidase/química , Neuraminidase/genética , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Especificidade por Substrato/genética
11.
PLoS One ; 15(3): e0230499, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32187230

RESUMO

Glycosphingolipids (GSLs) hexosylceramides and lactosylceramides are elevated in lupus mice and human patients with nephritis. Whereas other renal diseases characterized by increased GSL levels are thought to be a result of upregulated GSL synthesis, our results suggest elevated hexosylceramides and lactosylceramides in lupus nephritis is a result of increased catabolism of ganglioside GM3 due to significantly increased neuraminidase (NEU) activity. Thus, we hypothesized GM3 would be decreased in lupus nephritis kidneys and blocking NEU activity would reduce GSLs and improve disease in lupus mice. Female MRL/lpr lupus mice were treated with water or the NEU inhibitor oseltamivir phosphate at the onset of proteinuria to block GSL catabolism. Age-matched (non-nephritic) female MRL/MpJ lupus mice served as controls. Renal GM3 levels were significantly higher in the nephritic MRL/lpr water-treated mice compared to non-nephritic MRL/MpJ mice, despite significantly increased renal NEU activity. Blocking GSL catabolism increased, rather than decreased, renal and urine GSL levels and disease was not significantly impacted. A pilot study treating MRL/lpr females with GlcCer synthase inhibitor Genz-667161 to block GSL synthesis resulted in a strong significant negative correlation between Genz-667161 dose and renal GSL hexosylceramide and GM3 levels. Splenomegaly was negatively correlated and serum IgG levels were marginally correlated with increasing Genz-667161 dose. These results suggest accumulation of renal GM3 may be due to dysregulation of one or more of the GSL ganglioside pathways and inhibiting GSL synthesis, but not catabolism, may be a therapeutic approach for treating lupus nephritis.


Assuntos
Glicoesfingolipídeos/metabolismo , Nefrite Lúpica/tratamento farmacológico , Nefrite Lúpica/metabolismo , Animais , Ceramidas/metabolismo , Feminino , Gangliosídeo G(M3)/metabolismo , Rim/efeitos dos fármacos , Rim/metabolismo , Lactosilceramidas/metabolismo , Camundongos , Camundongos Endogâmicos MRL lpr , Neuraminidase/metabolismo , Oseltamivir/análogos & derivados , Oseltamivir/uso terapêutico , Ácidos Fosforosos/uso terapêutico , Projetos Piloto , Proteinúria/tratamento farmacológico , Proteinúria/metabolismo
12.
Eur J Med Chem ; 191: 112147, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32092589

RESUMO

From our research group, it was noticed that oseltamivir derivatives targeting 150-cavity of neuraminidase enzyme (NA) could significantly increase antiviral activity. Thus, we further enriched the C5-NH2 position of oseltamivir structure to obtain more potent oseltamivir derivatives. In this article a series of oseltamivir derivatives were synthesized by modifying C5-NH2 position of oseltamivir. All the compounds were evaluated for in vitro antiviral activity against H5N1 and H5N8. Encouragingly, compounds 9a and 11e were exhibited prominent activity, which is similar to oseltamivir carboxylate (OSC) and in NAs inhibitory assay, 11e showed remarkable potency against N1 (H5N1), N2 (H5N2), N6 (H5N6) and N8 (H5N8). In addition, 11e demonstrated low cytotoxicity and no obvious toxicity at the dose of 1500 mg/kg in mice. Molecular docking studies of 9a and 11e provided a plausible rationale for the high potency against group-1 NAs. This work provided new insights to design further neuraminidase inhibitors, which can help to investigate new potent inhibitors for group-1 and group-2 shortly.


Assuntos
Antivirais/farmacologia , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Vírus da Influenza A/efeitos dos fármacos , Neuraminidase/antagonistas & inibidores , Oseltamivir/farmacologia , Animais , Antivirais/síntese química , Antivirais/química , Galinhas , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Feminino , Vírus da Influenza A/enzimologia , Masculino , Camundongos , Camundongos Endogâmicos , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Estrutura Molecular , Neuraminidase/metabolismo , Oseltamivir/síntese química , Oseltamivir/química , Relação Estrutura-Atividade
13.
J Virol ; 94(9)2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32051275

RESUMO

Sialic acids (Sia) are the primary receptors for influenza viruses and are widely displayed on cell surfaces and in secreted mucus. Sia may be present in variant forms that include O-acetyl modifications at C-4, C-7, C-8, and C-9 positions and N-acetyl or N-glycolyl at C-5. They can also vary in their linkages, including α2-3 or α2-6 linkages. Here, we analyze the distribution of modified Sia in cells and tissues of wild-type mice or in mice lacking CMP-N-acetylneuraminic acid hydroxylase (CMAH) enzyme, which synthesizes N-glycolyl (Neu5Gc) modifications. We also examined the variation of Sia forms on erythrocytes and in saliva from different animals. To determine the effect of Sia modifications on influenza A virus (IAV) infection, we tested for effects on hemagglutinin (HA) binding and neuraminidase (NA) cleavage. We confirmed that 9-O-acetyl, 7,9-O-acetyl, 4-O-acetyl, and Neu5Gc modifications are widely but variably expressed in mouse tissues, with the highest levels detected in the respiratory and gastrointestinal (GI) tracts. Secreted mucins in saliva and surface proteins of erythrocytes showed a high degree of variability in display of modified Sia between different species. IAV HAs from different virus strains showed consistently reduced binding to both Neu5Gc- and O-acetyl-modified Sia; however, while IAV NAs were inhibited by Neu5Gc and O-acetyl modifications, there was significant variability between NA types. The modifications of Sia in mucus may therefore have potent effects on the functions of IAV and may affect both pathogens and the normal flora of different mucosal sites.IMPORTANCE Sialic acids (Sia) are involved in numerous different cellular functions and are receptors for many pathogens. Sia come in chemically modified forms, but we lack a clear understanding of how they alter interactions with microbes. Here, we examine the expression of modified Sia in mouse tissues, on secreted mucus in saliva, and on erythrocytes, including those from IAV host species and animals used in IAV research. These Sia forms varied considerably among different animals, and their inhibitory effects on IAV NA and HA activities and on bacterial sialidases (neuraminidases) suggest a host-variable protective role in secreted mucus.


Assuntos
Vírus da Influenza A/metabolismo , Muco/metabolismo , Ácido N-Acetilneuramínico/metabolismo , Células A549 , Animais , Cães , Eritrócitos/metabolismo , Feminino , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Hemaglutininas/metabolismo , Humanos , Vírus da Influenza A/fisiologia , Influenza Humana/metabolismo , Células Madin Darby de Rim Canino , Masculino , Camundongos , Oxigenases de Função Mista/metabolismo , Neuraminidase/metabolismo , Orthomyxoviridae/metabolismo , Receptores Virais/metabolismo , Saliva/química
14.
Sci Rep ; 10(1): 1589, 2020 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-32005959

RESUMO

Mumps virus is one of the main cause of respiratory illnesses in humans, especially children. Among the viral surface glycoproteins, the hemagglutinin - neuraminidase, MuV-HN, plays key roles in virus entry into host cells and infectivity, thus representing an ideal target for the design of novel inhibitors. Here we report the detailed analysis of the molecular recognition of host cell surface sialylated glycans by the viral glycoprotein MuV-HN. By a combined use of NMR, docking, molecular modelling and CORCEMA-ST, the structural features of sialoglycans/MuV-HN complexes were revealed. Evidence for a different enzyme activity toward longer and complex substrates compared to unbranched ligands was also examined by an accurate NMR kinetic analysis. Our results provide the basis for the structure-based design of effective drugs against mumps-induced diseases.


Assuntos
Hemaglutininas/metabolismo , Vírus da Caxumba/metabolismo , Neuraminidase/metabolismo , Polissacarídeos/metabolismo , Proteínas Estruturais Virais/metabolismo , Sítios de Ligação , Humanos , Cinética , Espectroscopia de Ressonância Magnética , Simulação de Acoplamento Molecular , Conformação Proteica
15.
Biochim Biophys Acta Mol Basis Dis ; 1866(5): 165692, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31972227

RESUMO

Many important pathogen-host interactions rely on highly specific carbohydrate binding events. In the case of the protozoan Trypanosoma cruzi, the causative agent of Chagas disease, glycointeractions involving sialic acid (SA) residues are pivotal for parasite infectivity, escape from immune surveillance and pathogenesis. Though unable to synthesize SA de novo, T. cruzi displays a unique trans-Sialidase (TS) enzyme, which is able to cleave terminal SA residues from host donor glycoconjugates and transfer them onto parasite surface mucins, thus generating protective/adhesive structures. In addition, this parasite sheds TS into the bloodstream, as a way of modifying the surface SA signature, and thereby the signaling/functional properties of mammalian host target cells on its own advantage. Here, we discuss the pathogenic aspects of T. cruzi TS: its molecular adaptations, the multiplicity of interactions in which it is involved during infections, and the array of novel and appealing targets for intervention in Chagas disease provided by TS-remodeled sialoglycophenotypes.


Assuntos
Doença de Chagas/imunologia , Glicoproteínas/metabolismo , Interações Hospedeiro-Parasita/imunologia , Neuraminidase/metabolismo , Polissacarídeos/imunologia , Proteínas de Protozoários/metabolismo , Trypanosoma cruzi/patogenicidade , Animais , Doença de Chagas/parasitologia , Glicoproteínas/imunologia , Humanos , Evasão da Resposta Imune , Neuraminidase/imunologia , Polissacarídeos/química , Polissacarídeos/metabolismo , Proteínas de Protozoários/imunologia , Ácidos Siálicos/química , Ácidos Siálicos/imunologia , Ácidos Siálicos/metabolismo , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/imunologia , Fatores de Virulência/imunologia , Fatores de Virulência/metabolismo
16.
Lett Appl Microbiol ; 70(4): 252-258, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31990997

RESUMO

Neuraminidase (NA) is an integral membrane protein of influenza A virus (IAV) and primarily aids in the release of progeny virions, following the intracellular viral replication cycle. In an attempt to discover new functions of NA, we conducted a classical yeast two-hybrid screen and found acute myeloid leukaemia marker 1 (AML1) as a novel interacting partner of IAV-NA. The interaction was further validated by co-immunoprecipitation in IAV-infected cells and in an in vitro coupled transcription/translation system. Interestingly, we found an increase in the expression of AML1 upon IAV infection in a dose-dependent manner. As expected, we also observed an increase in the IFN-ß levels, the first line of defence against viral infections. Subsequently, when AML1 was downregulated using siRNA, the IFN-ß levels were found to be remarkably reduced. Our study also shows that AML1 is induced upon IAV infection and results in the induction of IFN-ß. Thus, AML1 is proposed to be an important player in IFN induction and has a role in an antiviral response against IAV infection. SIGNIFICANCE AND IMPACT OF THE STUDY: Influenza epidemics and pandemics are constant threats to human health. Development of antiviral therapeutics has focused on important and major IAV proteins as targets. However, the rate at which this virus mutates makes the task challenging. Thus, next-generation approaches aim at host cellular proteins that aid the virus in its replication. This study reports a new host-virus interaction, of acute myeloid leukaemia marker 1 (AML1) with influenza A neuraminidase (IAV-NA). We have found that this interaction has a direct effect on the upregulation of host IFN-ß response. Further studies may lead to a greater understanding of this new innate defence pathway in infected cells.


Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Virus da Influenza A Subtipo H5N1/metabolismo , Influenza Humana/metabolismo , Interferon beta/metabolismo , Neuraminidase/metabolismo , Proteínas Virais/metabolismo , Linhagem Celular , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Interações Hospedeiro-Patógeno , Humanos , Virus da Influenza A Subtipo H5N1/genética , Influenza Humana/genética , Influenza Humana/virologia , Interferon beta/genética , Neuraminidase/genética , Ligação Proteica , Regulação para Cima , Proteínas Virais/genética
17.
Phytomedicine ; 67: 153150, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31958713

RESUMO

BACKGROUND: Influenza virus is one of the most important human pathogens, causing substantial seasonal and pandemic morbidity and mortality. Houttuynia cordata is a traditionally used medicinal plant for the treatment of pneumonia. Flavonoids are one of the major bioactive constituents of Houttuynia cordata. PURPOSE: This study was designed to investigate the therapeutic effect and mechanism of flavonoid glycosides from H. cordata on influenza A virus (IAV)-induced acute lung injury (ALI) in mice. METHODS: Flavonoids from H. cordata (HCF) were extracted from H. cordata and identified by high-performance liquid chromatography. Mice were infected intranasally with influenza virus H1N1 (A/FM/1/47). HCF (50, 100, or 200 mg/kg) or Ribavirin (100 mg/kg, the positive control) were administered intragastrically. Survival rates, life spans, weight losses, lung indexes, histological changes, inflammatory infiltration, and inflammatory markers in the lungs were measured. Lung virus titers and neuraminidase (NA) activities were detected. The expression of Toll-like receptors (TLRs) and levels of NF-κB p65 phosphorylation (NF-κB p65(p)) in the lungs were analysed. The effects of HCF on viral replication and TLR signalling were further evaluated in cells. RESULTS: HCF contained 78.5% flavonoid glycosides. The contents of rutin, hyperin, isoquercitrin, and quercitrin in HCF were 8.8%, 26.7%, 9.9% and 31.7%. HCF (50, 100 and 200 mg/kg) increased the survival rate and life span of mice infected with the lethal H1N1 virus. In H1N1-induced ALI, mice treated with HCF (50, 100 and 200 mg/kg) showed lesser weight loss and lower lung index than the model group. The lungs of HCF-treated ALI mice presented more intact lung microstructural morphology, milder inflammatory infiltration, and lower levels of monocyte chemotactic protein 1 (MCP-1), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α) and malondialdehyde (MDA) than in the model group. Further investigation revealed that HCF exerted antiviral and TLR-inhibitory effects in vivo and in vitro. HCF (50, 100 and 200 mg/kg) reduced lung H1N1 virus titers and inhibited viral NA activity in mice. HCF (100 and 200 mg/kg) elevated the levels of interferon-ß in lungs. HCF also decreased the expression of TLR3/4/7 and level of NF-κB p65(p) in lung tissues. In vitro experiments showed that HCF (50, 100 and 200 µg/ml) significantly inhibited viral proliferation and suppressed NA activity. In RAW 264.7 cells, TLR3, TLR4, and TLR7 agonist-stimulated cytokine secretion, NF-κB p65 phosphorylation, and nuclear translocation were constrained by HCF treatment. Furthermore, among the four major flavonoid glycosides in HCF, hyperin and quercitrin inhibited both viral replication and TLR signalling in cells. CONCLUSION: HCF significantly alleviated H1N1-induced ALI in mice, which were associated with its dual antiviral and anti-inflammatory effects via inhibiting influenzal NA activity and TLR signalling. among the four major flavonoid glycosides in HCF, hyperin and quercitrin played key roles in the therapeutic effect of HCF.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/virologia , Antivirais/farmacologia , Flavonoides/farmacologia , Houttuynia/química , Vírus da Influenza A Subtipo H1N1/patogenicidade , Lesão Pulmonar Aguda/metabolismo , Animais , Antivirais/química , Cães , Flavonoides/química , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/fisiologia , Células Madin Darby de Rim Canino , Camundongos Endogâmicos BALB C , Neuraminidase/metabolismo , Infecções por Orthomyxoviridae/tratamento farmacológico , Infecções por Orthomyxoviridae/metabolismo , Receptores Toll-Like/metabolismo , Fator de Transcrição RelA/metabolismo , Replicação Viral/efeitos dos fármacos
18.
Biochem Biophys Res Commun ; 523(2): 487-492, 2020 03 05.
Artigo em Inglês | MEDLINE | ID: mdl-31889533

RESUMO

Bacterial sialidases are widely used to remove sialic acid (Sia) residues from glycans. Most of them cleave the glycosides of N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) under acidic pHs; however, currently available bacterial sialidases had no activity to the glycosides of deaminoneuraminic acid (Kdn). In this study, we found a novel sialidase from Sphingobacterium sp. strain HMA12 that could cleave any of the glycosides of Neu5Ac, Neu5Gc, and Kdn. It also had a broad linkage specificity, i.e., α2,3-, α2,6-, α2,8-, and α2,9-linkages, and the optimal pH at neutral ranges, pH 6.5-7.0. These properties are particularly important when sialidases are applied for in vivo digestion of the cell surface sialosides under physiological conditions. Interestingly, 2,3-didehydro-2-deoxy-N-acetylneuraminic acid (Neu5Ac2en), which is a transition state analog-based inhibitor, competitively inhibited the enzyme-catalyzed reaction for Kdn as well as for Neu5Ac, suggesting that the active site is common to the Neu5Ac and Kdn residues. Taken together, this sialidase is versatile and useful for the in vivo research on sialo-glycoconjugates.


Assuntos
Glicosídeos/metabolismo , Neuraminidase/metabolismo , Ácidos Siálicos/metabolismo , Sphingobacterium/enzimologia , Motivos de Aminoácidos , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Células CHO , Cricetulus , Concentração de Íons de Hidrogênio , Hidrólise , Ácido N-Acetilneuramínico/análogos & derivados , Ácido N-Acetilneuramínico/metabolismo , Ácido N-Acetilneuramínico/farmacologia , Ácidos Neuramínicos , Neuraminidase/antagonistas & inibidores , Neuraminidase/química , Neuraminidase/genética , Sphingobacterium/genética , Especificidade por Substrato , Temperatura
19.
Eur J Med Chem ; 187: 111940, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31835169

RESUMO

A novel series of 1,2,3-triazole oseltamivir derivatives, which could simultaneously occupy the classical NA catalytic site and the newly reported 430-cavity, were designed, synthesized, and evaluated for their anti-influenza activities. The results demonstrated that four compounds (6g, 6l, 6y and 8c) showed robust anti-influenza potencies against H5N1, H5N2 and H5N6 strains in both enzymatic assay and cellular assay. Especially, 6l was proved to possess the most potent and broad-spectrum anti-influenza activity, with IC50 values of 0.12 µM, 0.049 µM and 0.16 µM and EC50 values of 2.45 µM, 0.43 µM and 2.8 µM against H5N1, H5N2 and H5N6 strains, respectively, which were slightly weaker than oseltamivir carboxylate. In addition, in the embryonated egg model, 6l achieved the similar protective effect against H9N2 strain with oseltamivir carboxylate in the tested concentrations. Preliminary structure-activity relationships (SARs), molecular modeling, and calculated physicochemical properties of selected compounds were also discussed.


Assuntos
Antivirais/farmacologia , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Vírus da Influenza A/efeitos dos fármacos , Neuraminidase/antagonistas & inibidores , Triazóis/farmacologia , Animais , Antivirais/síntese química , Antivirais/química , Galinhas , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Fibroblastos/efeitos dos fármacos , Fibroblastos/virologia , Testes de Sensibilidade Microbiana , Modelos Moleculares , Estrutura Molecular , Neuraminidase/metabolismo , Relação Estrutura-Atividade , Triazóis/síntese química , Triazóis/química
20.
Fish Physiol Biochem ; 46(2): 563-574, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31792756

RESUMO

Fish store triglycerides (TGs) in the liver, muscle, and adipose tissue and TGs constitute an energy source upon metabolic demand. The liver generally plays important roles in lipid metabolism. Recent studies have suggested the possibility of hepatic lipid metabolic regulation by ganglioside in mammals; however, ganglioside-mediated regulation of lipid metabolism is unclear in fish. This study aimed to clarify the role of ganglioside in fish TG metabolism, with particular reference to Neu3a, a ganglioside-specific sialidase expressed in the fish liver. Under fasting conditions, there was a decrease in hepatic TG contents, and neu3a mRNA level was significantly up-regulated in the medaka liver. To determine the role of Neu3a in hepatic lipid metabolism, Neu3a stable transfectants were generated using fish liver Hepa-T1 cells. After treating Neu3a cells with oleic acid, reduction of TG was detected in comparison with the mock cells. Furthermore, lipase activity was greater in Neu3a cells than in mock cells. To examine which ganglioside regulates these events, alterations of ganglioside composition in Neu3a cells were analyzed. Neu3a cells exhibited increased level of lactosylceramide (LacCer), a Neu3 enzymatic product originating from GM3. In addition, exposure of LacCer toward Hepa-T1 cells resulted in an increase of neutral lipase activity. The present results suggest that Neu3a up-regulation in medaka under fasting condition accelerates hepatic TG degradation for energy production via GM3 desialylation.


Assuntos
Neuraminidase/metabolismo , Oryzias/fisiologia , Triglicerídeos/metabolismo , Animais , Hepatócitos/metabolismo , Metabolismo dos Lipídeos , Neuraminidase/genética , RNA Mensageiro/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...