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1.
Immunity ; 53(4): 793-804.e9, 2020 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-32910906

RESUMO

Allergies are considered to represent mal-directed type 2 immune responses against mostly innocuous exogenous compounds. Immunoglobulin E (IgE) antibodies are a characteristic feature of allergies and mediate hypersensitivity against allergens through activation of effector cells, particularly mast cells (MCs). Although the physiological functions of this dangerous branch of immunity have remained enigmatic, recent evidence shows that allergic immune reactions can help to protect against the toxicity of venoms. Because bacteria are a potent alternative source of toxins, we assessed the possible role of allergy-like type 2 immunity in antibacterial host defense. We discovered that the adaptive immune response against Staphylococcus aureus (SA) skin infection substantially improved systemic host defense against secondary SA infections in mice. Moreover, this acquired protection depended on IgE effector mechanisms and MCs. Importantly, our results reveal a previously unknown physiological function of allergic immune responses, IgE antibodies, and MCs in host defense against a pathogenic bacterium.


Assuntos
Imunidade Adaptativa/imunologia , Imunoglobulina E/imunologia , Mastócitos/imunologia , Infecções Estafilocócicas/imunologia , Infecções Cutâneas Estafilocócicas/imunologia , Staphylococcus aureus/imunologia , Alérgenos/imunologia , Animais , Feminino , Hipersensibilidade/imunologia , Hipersensibilidade/microbiologia , Mastócitos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Pele/imunologia , Pele/microbiologia , Infecções Estafilocócicas/microbiologia , Infecções Cutâneas Estafilocócicas/microbiologia
2.
Nature ; 597(7874): 92-96, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34433968

RESUMO

Atherosclerotic cardiovascular disease causes heart attacks and strokes, which are the leading causes of mortality worldwide1. The formation of atherosclerotic plaques is initiated when low-density lipoproteins bind to heparan-sulfate proteoglycans (HSPGs)2 and become trapped in the subendothelial space of large and medium size arteries, which leads to chronic inflammation and remodelling of the artery wall2. A proliferation-inducing ligand (APRIL) is a cytokine that binds to HSPGs3, but the physiology of this interaction is largely unknown. Here we show that genetic ablation or antibody-mediated depletion of APRIL aggravates atherosclerosis in mice. Mechanistically, we demonstrate that APRIL confers atheroprotection by binding to heparan sulfate chains of heparan-sulfate proteoglycan 2 (HSPG2), which limits the retention of low-density lipoproteins, accumulation of macrophages and formation of necrotic cores. Indeed, antibody-mediated depletion of APRIL in mice expressing heparan sulfate-deficient HSPG2 had no effect on the development of atherosclerosis. Treatment with a specific anti-APRIL antibody that promotes the binding of APRIL to HSPGs reduced experimental atherosclerosis. Furthermore, the serum levels of a form of human APRIL protein that binds to HSPGs, which we termed non-canonical APRIL (nc-APRIL), are associated independently of traditional risk factors with long-term cardiovascular mortality in patients with atherosclerosis. Our data reveal properties of APRIL that have broad pathophysiological implications for vascular homeostasis.


Assuntos
Aterosclerose/metabolismo , Aterosclerose/prevenção & controle , Proteoglicanas de Heparan Sulfato/metabolismo , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/metabolismo , Animais , Antígeno de Maturação de Linfócitos B/metabolismo , Sítios de Ligação , Doenças Cardiovasculares/sangue , Doenças Cardiovasculares/mortalidade , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ligação Proteica , Proteína Transmembrana Ativadora e Interagente do CAML/metabolismo , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/sangue , Membro 13 da Superfamília de Ligantes de Fatores de Necrose Tumoral/deficiência
3.
Mol Cell ; 75(4): 849-858.e8, 2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31442425

RESUMO

Inducing protein degradation via small molecules is a transformative therapeutic paradigm. Although structural requirements of target degradation are emerging, mechanisms determining the cellular response to small-molecule degraders remain poorly understood. To systematically delineate effectors required for targeted protein degradation, we applied genome-scale CRISPR/Cas9 screens for five drugs that hijack different substrate receptors (SRs) of cullin RING ligases (CRLs) to induce target proteolysis. We found that sensitivity to small-molecule degraders is dictated by shared and drug-specific modulator networks, including the COP9 signalosome and the SR exchange factor CAND1. Genetic or pharmacologic perturbation of these effectors impairs CRL plasticity and arrests a wide array of ligases in a constitutively active state. Resulting defects in CRL decommissioning prompt widespread CRL auto-degradation that confers resistance to multiple degraders. Collectively, our study informs on regulation and architecture of CRLs amenable for targeted protein degradation and outlines biomarkers and putative resistance mechanisms for upcoming clinical investigation.


Assuntos
Complexo do Signalossomo COP9/metabolismo , Proteínas Culina/metabolismo , Proteólise , Fatores de Transcrição/metabolismo , Complexo do Signalossomo COP9/genética , Proteínas Culina/genética , Humanos , Fatores de Transcrição/genética
5.
J Neuroinflammation ; 20(1): 68, 2023 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-36906584

RESUMO

OBJECTIVES: Reactive gliosis is a common pathological hallmark of CNS pathology resulting from neurodegeneration and neuroinflammation. In this study we investigate the capability of a novel monoamine oxidase B (MAO-B) PET ligand to monitor reactive astrogliosis in a transgenic mouse model of Alzheimer`s disease (AD). Furthermore, we performed a pilot study in patients with a range of neurodegenerative and neuroinflammatory conditions. METHODS: A cross-sectional cohort of 24 transgenic (PS2APP) and 25 wild-type mice (age range: 4.3-21.0 months) underwent 60 min dynamic [18F]fluorodeprenyl-D2 ([18F]F-DED), static 18 kDa translocator protein (TSPO, [18F]GE-180) and ß-amyloid ([18F]florbetaben) PET imaging. Quantification was performed via image derived input function (IDIF, cardiac input), simplified non-invasive reference tissue modelling (SRTM2, DVR) and late-phase standardized uptake value ratios (SUVr). Immunohistochemical (IHC) analyses of glial fibrillary acidic protein (GFAP) and MAO-B were performed to validate PET imaging by gold standard assessments. Patients belonging to the Alzheimer's disease continuum (AD, n = 2), Parkinson's disease (PD, n = 2), multiple system atrophy (MSA, n = 2), autoimmune encephalitis (n = 1), oligodendroglioma (n = 1) and one healthy control underwent 60 min dynamic [18F]F-DED PET and the data were analyzed using equivalent quantification strategies. RESULTS: We selected the cerebellum as a pseudo-reference region based on the immunohistochemical comparison of age-matched PS2APP and WT mice. Subsequent PET imaging revealed that PS2APP mice showed elevated hippocampal and thalamic [18F]F-DED DVR when compared to age-matched WT mice at 5 months (thalamus: + 4.3%; p = 0.048), 13 months (hippocampus: + 7.6%, p = 0.022) and 19 months (hippocampus: + 12.3%, p < 0.0001; thalamus: + 15.2%, p < 0.0001). Specific [18F]F-DED DVR increases of PS2APP mice occurred earlier when compared to signal alterations in TSPO and ß-amyloid PET and [18F]F-DED DVR correlated with quantitative immunohistochemistry (hippocampus: R = 0.720, p < 0.001; thalamus: R = 0.727, p = 0.002). Preliminary experience in patients showed [18F]F-DED VT and SUVr patterns, matching the expected topology of reactive astrogliosis in neurodegenerative (MSA) and neuroinflammatory conditions, whereas the patient with oligodendroglioma and the healthy control indicated [18F]F-DED binding following the known physiological MAO-B expression in brain. CONCLUSIONS: [18F]F-DED PET imaging is a promising approach to assess reactive astrogliosis in AD mouse models and patients with neurological diseases.


Assuntos
Doença de Alzheimer , Doenças Neurodegenerativas , Oligodendroglioma , Animais , Humanos , Camundongos , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Encéfalo/metabolismo , Estudos Transversais , Gliose/patologia , Inflamação/metabolismo , Camundongos Transgênicos , Monoaminoxidase/metabolismo , Doenças Neurodegenerativas/metabolismo , Oligodendroglioma/metabolismo , Oligodendroglioma/patologia , Projetos Piloto , Tomografia por Emissão de Pósitrons/métodos , Receptores de GABA/metabolismo
6.
Allergy ; 77(2): 499-512, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-33840121

RESUMO

BACKGROUND: In contrast to their clearly defined roles in allergic diseases, the physiologic functions of Immunoglobulin E antibodies (IgEs) and mast cells (MCs) remain enigmatic. Recent research supports the toxin hypothesis, showing that MCs and IgE-related type 2 immune responses can enhance host defense against certain noxious substances, including honeybee venom (BV). However, the mechanisms by which MCs can interfere with BV toxicity are unknown. In this study, we assessed the role of IgE and certain MC products in MC-mediated BV detoxification. METHODS: We applied in vitro and in vivo fluorescence microscopyimaging, and flow cytometry, fibroblast-based toxicity assays and mass spectrometry to investigate IgE-mediated detoxification of BV cytotoxicity by mouse and human MCs in vitro. Pharmacologic strategies to interfere with MC-derived heparin and proteases helped to define the importance of specific detoxification mechanisms. RESULTS: Venom-specific IgE increased the degranulation and cytokine responses of MCs to BV in vitro. Passive serum sensitization enhanced MC degranulation in vivo. IgE-activated mouse or human MCs exhibited enhanced potential for detoxifying BV by both proteolytic degradation and heparin-related interference with toxicity. Mediators released by IgE-activated human MCs efficiently degraded multiple BV toxins. CONCLUSIONS: Our results both reveal that IgE sensitization enhances the MC's ability to detoxify BV and also assign efficient toxin-neutralizing activity to MC-derived heparin and proteases. Our study thus highlights the potential importance of IgE, MCs, and particular MC products in defense against BV.


Assuntos
Venenos de Abelha , Mastócitos , Alérgenos/metabolismo , Animais , Degranulação Celular , Heparina/metabolismo , Humanos , Imunoglobulina E , Camundongos , Peptídeo Hidrolases/metabolismo
8.
Drug Chem Toxicol ; 45(5): 2233-2245, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33934680

RESUMO

Statins are 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase inhibitor drugs that lead to serum-cholesterol-lowering effects. Rosuvastatin, a third-generation statin, has shown better results in reducing cholesterol concentrations when compared to other widely prescribed statins. Recent studies by our group reported that rosuvastatin impairs reproductive function in rats possibly by disrupting the reproductive-endocrine axis. In this study, we evaluated whether rosuvastatin presents estrogenic or antiestrogenic effects, by an in vivo uterotrophic assay in rats, and investigated the direct effect of this drug upon rat uterine tissue contractility both in non-gravid and gravid periods. Rosuvastatin exposure in vivo at doses of 0 (control), 3, and 10 mg/kg/d was not associated with estrogenic or antiestrogenic effects on uterine tissue. However, in vivo (doses of 0, 3, and 10 mg/kg/d) and ex vivo (concentrations of 0, 1, 10, and 100 µg/mL) exposures to this drug were related to alterations in uterine basal contraction pattern. Furthermore, in vivo and ex vivo rosuvastatin exposures potentially modulate the action of uterine contraction inducers carbachol, norepinephrine, and prostaglandin E2. Thus, rosuvastatin can affect uterine physiology not necessarily by an endocrine mechanism related to the estrogen signaling, but possibly by its pleiotropic effects, with indirect tissue and cellular interactions, since in vivo and ex vivo exposures of uterine fragments to rosuvastatin presented different responses in uterine contractile parameters, which require further studies upon the precise mechanism of action of this drug in female reproductive function.


Assuntos
Estrogênios , Inibidores de Hidroximetilglutaril-CoA Redutases , Animais , Colesterol , Estrogênios/toxicidade , Feminino , Inibidores de Hidroximetilglutaril-CoA Redutases/toxicidade , Gravidez , Ratos , Ratos Wistar , Rosuvastatina Cálcica/toxicidade
9.
Bioorg Med Chem ; 52: 116528, 2021 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-34839158

RESUMO

Tau aggregates represent a critical pathology in Alzheimer's disease (AD) and other forms of dementia. The extent of Tau neurofibrillary tangles across defined brain regions corresponds well to the observed level of cognitive decline in AD. Compound 1 (PI-2620) was recently identified as a promising Tau positron emission tomography tracer for AD and non-AD tauopathies. To evaluate the impact of the N-atom position with respect to Tau- and off-target binding, tricyclic core analogs of PI-2620 with nitrogen atoms at different positions were prepared. Affinity to aggregated Tau was evaluated using human AD brain homogenates, and their off-target binding was evaluated in a monoamine oxidase A (MAO-A) competition assay. The novel tricyclic core derivatives all displayed inferior Tau binding or MAO-A off-target selectivity, indicating PI-2620 to be the optimal design for high affinity binding to Tau and high MAO-A selectivity.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Nitrogênio/farmacologia , Piridinas , Compostos Radiofarmacêuticos/farmacologia , Proteínas tau/antagonistas & inibidores , Doença de Alzheimer/diagnóstico , Encéfalo/metabolismo , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Monoaminoxidase/metabolismo , Nitrogênio/química , Tomografia por Emissão de Pósitrons , Piridinas/administração & dosagem , Piridinas/química , Piridinas/farmacologia , Compostos Radiofarmacêuticos/química , Relação Estrutura-Atividade , Proteínas tau/análise , Proteínas tau/metabolismo
10.
Mol Cell Proteomics ; 17(3): 516-532, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29208752

RESUMO

Peritoneal dialysis (PD) is a modality of renal replacement therapy in which the high volumes of available PD effluent (PDE) represents a rich source of biomarkers for monitoring disease and therapy. Although this information could help guide the management of PD patients, little is known about the potential of PDE to define pathomechanism-associated molecular signatures in PD.We therefore subjected PDE to a high-performance multiplex proteomic analysis after depletion of highly-abundant plasma proteins and enrichment of low-abundance proteins. A combination of label-free and isobaric labeling strategies was applied to PDE samples from PD patients (n = 20) treated in an open-label, randomized, two-period, cross-over clinical trial with standard PD fluid or with a novel PD fluid supplemented with alanyl-glutamine (AlaGln).With this workflow we identified 2506 unique proteins in the PDE proteome, greatly increasing coverage beyond the 171 previously-reported proteins. The proteins identified range from high abundance plasma proteins to low abundance cellular proteins, and are linked to larger numbers of biological processes and pathways, some of which are novel for PDE. Interestingly, proteins linked to membrane remodeling and fibrosis are overrepresented in PDE compared with plasma, whereas the proteins underrepresented in PDE suggest decreases in host defense, immune-competence and response to stress. Treatment with AlaGln-supplemented PD fluid is associated with reduced activity of membrane injury-associated mechanisms and with restoration of biological processes involved in stress responses and host defense.Our study represents the first application of the PDE proteome in a randomized controlled prospective clinical trial of PD. This novel proteomic workflow allowed detection of low abundance biomarkers to define pathomechanism-associated molecular signatures in PD and their alterations by a novel therapeutic intervention.


Assuntos
Dipeptídeos/farmacologia , Diálise Peritoneal , Proteoma , Proteínas Sanguíneas/metabolismo , Estudos Cross-Over , Feminino , Humanos , Masculino
11.
J Pharmacol Exp Ther ; 369(1): 98-106, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30728250

RESUMO

Serotonin [5-hydroxytryptamine (5-HT)] exerts multiple central and peripheral functions. High concentrations of 5-HT have been found in the epididymis, a ductal organ that plays pivotal roles in sperm transport and maturation. The contraction of the epididymal smooth muscle is essential for sperm transport and emission during ejaculation. The contributions of the epididymal 5-HT system to these events are poorly understood. Here, we assessed the contractile function of 5-HT in the rat cauda epididymis (CE), pharmacologically targeting the receptor(s) and the reuptake mechanism involved in this system. Segments of CE duct from adult Wistar rats were set up in an organ bath system for isometric tension recordings, and concentration-response curves to 5-HT and norepinephrine were obtained. 5-HT elicited concentration-dependent contractions of the CE duct (pEC50 = 6.5 ± 0.1) that were potentiated with high potency by the norepinephrine transporter (NET) inhibitor desipramine and with low potency by the highly selective serotonin transporter inhibitor paroxetine, indicating that the NET is the major mediator of 5-HT reuptake in vitro. CE contractions to 5-HT were antagonized by the α 1-adrenoceptor (α 1-AR) antagonist prazosin (pA 2 ≅ 8.9), 5-HT2A/2C antagonists ketanserin (pA 2 ≅ 9.4) and fluoxetine (pA 2 ≅ 7.4), and 5-HT1A ligands WAY 100635 (pA 2 ≅ 8.9) and buspirone (pA 2 ≅ 7.3). Reverse transcriptase polymerase chain reaction analysis demonstrated that 5-HT1A and 5-HT2A transcripts are highly abundant in the cauda epididymis, whereas 5-HT2C transcript was not found. Altogether, our results reveal that contractions of the CE duct to 5-HT encompasses at least activation of α 1-ARs and 5-HT1A and 5-HT2A receptors, providing new insights into the roles of 5-HT on the epididymal function.


Assuntos
Epididimo/efeitos dos fármacos , Epididimo/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Receptores de Serotonina/metabolismo , Serotonina/farmacologia , Animais , Relação Dose-Resposta a Droga , Masculino , Músculo Liso/efeitos dos fármacos , Músculo Liso/fisiologia , Transporte Proteico/efeitos dos fármacos , Ratos , Ratos Wistar , Receptores Adrenérgicos/metabolismo , Receptores Adrenérgicos alfa 1/metabolismo
12.
Eur J Nucl Med Mol Imaging ; 46(10): 2178-2189, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31264169

RESUMO

PURPOSE: Tau deposition is a key pathological feature of Alzheimer's disease (AD) and other neurodegenerative disorders. The spreading of tau neurofibrillary tangles across defined brain regions corresponds to the observed level of cognitive decline in AD. Positron-emission tomography (PET) has proved to be an important tool for the detection of amyloid-beta (Aß) aggregates in the brain, and is currently being explored for detection of pathological misfolded tau in AD and other non-AD tauopathies. Several PET tracers targeting tau deposits have been discovered and tested in humans. Limitations have been reported, especially regarding their selectivity. METHODS: In our screening campaign we identified pyrrolo[2,3-b:4,5-c']dipyridine core structures with high affinity for aggregated tau. Further characterization showed that compounds containing this moiety had significantly reduced monoamine oxidase A (MAO-A) binding compared to pyrido[4,3-b]indole derivatives such as AV-1451. RESULTS: Here we present preclinical data of all ten fluoropyridine regioisomers attached to the pyrrolo[2,3-b:4,5-c']dipyridine scaffold, revealing compounds 4 and 7 with superior properties. The lead candidate [18F]PI-2620 (compound 7) displayed high affinity for tau deposits in AD brain homogenate competition assays. Specific binding to pathological misfolded tau was further demonstrated by autoradiography on AD brain sections (Braak I-VI), Pick's disease and progressive supranuclear palsy (PSP) pathology, whereas no specific tracer binding was detected on brain slices from non-demented donors. In addition to its high affinity binding to tau aggregates, the compound showed excellent selectivity with no off-target binding to Aß or MAO-A/B. Good brain uptake and fast washout were observed in healthy mice and non-human primates. CONCLUSIONS: Therefore, [18F]PI-2620 was selected for clinical validation.


Assuntos
Doença de Alzheimer/diagnóstico por imagem , Inibidores da Monoaminoxidase/síntese química , Tomografia por Emissão de Pósitrons/métodos , Piridinas/síntese química , Compostos Radiofarmacêuticos/síntese química , Animais , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Radioisótopos de Flúor/farmacocinética , Humanos , Macaca mulatta , Camundongos , Monoaminoxidase/metabolismo , Inibidores da Monoaminoxidase/farmacocinética , Ligação Proteica , Piridinas/farmacocinética , Compostos Radiofarmacêuticos/farmacocinética
13.
Int J Sports Med ; 38(4): 263-269, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-28219103

RESUMO

Cardiotonic drugs and exercise training promote cardiac inotropic effects, which may affect training-induced cardiac adaptations. This study investigated the effects of long-term administration of digoxin on heart structure and function, and physical performance of rats submitted to high-intensity interval training (HIIT). Male Wistar rats, 60 days old, were divided into control (C), digoxin (DIGO), trained (T), and trained with digoxin (TDIGO). Digoxin was administered by gavage (30 µg/kg/day) for 75 days. The HIIT program consisted of treadmill running 60 min/day (8 min at 80% of the maximum speed (MS) and 2 min at 20% of the MS), 5 days per week during 60 days. The main cardiac parameters were evaluated by echocardiograph and cardiomyocyte area was determined by histology. There were no group x time effects of digoxin, HIIT or interactions (digoxin and HIIT) on functional echocardiographic parameters (heart rate; ejection fraction) or in the maximum exercise test. There was a group x time interaction, as evidenced by observed cardiac hypertrophy in the TDIGO group evaluated by ratio of left ventricle weight to body weight (p<0.002) and cardiomyocyte area (p<0.000002). Long-term administration of digoxin promoted cardiac hypertrophy without affecting cardiac function and physical performance in rats submitted to HIIT.


Assuntos
Cardiomegalia/induzido quimicamente , Digoxina/efeitos adversos , Coração/efeitos dos fármacos , Condicionamento Físico Animal , Animais , Ecocardiografia , Teste de Esforço , Frequência Cardíaca , Treinamento Intervalado de Alta Intensidade , Masculino , Distribuição Aleatória , Ratos , Ratos Wistar
14.
J Biol Chem ; 289(33): 22888-22899, 2014 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-24973214

RESUMO

Hyaluronan (HA) may have proinflammatory roles in the context of CNS autoimmunity. It accumulates in demyelinated multiple sclerosis (MS) lesions, promotes antigen presentation, and enhances T-cell activation and proliferation. HA facilitates lymphocyte binding to vessels and CNS infiltration at the CNS vascular endothelium. Furthermore, HA signals through Toll-like receptors 2 and 4 to stimulate inflammatory gene expression. We assessed the role of HA in experimental autoimmune encephalomyelitis (EAE), an animal model of MS by administration of 4-methylumbelliferone (4MU), a well established inhibitor of HA synthesis. 4MU decreased hyaluronan synthesis in vitro and in vivo. It was protective in active EAE of C57Bl/6 mice, decreased spinal inflammatory infiltrates and spinal infiltration of Th1 cells, and increased differentiation of regulatory T-cells. In adoptive transfer EAE, feeding of 4MU to donor mice significantly decreased the encephalitogenicity of lymph node cells. The transfer of proteolipid protein (PLP)-stimulated lymph node cells to 4MU-fed mice resulted in a delayed EAE onset and delayed spinal T-cell infiltration. Expression of CXCL12, an anti-inflammatory chemokine, is reduced in MS patients in CSF cells and in spinal cord tissue during EAE. Hyaluronan suppressed production of CXCL12, whereas 4MU increased spinal CXCL12 in naive animals and during neuroinflammation. Neutralization of CXCR4, the most prominent receptor of CXCL12, by administration of AMD3100 diminished the protective impact of 4MU in adoptive transfer EAE. In conclusion, hyaluronan exacerbates CNS autoimmunity, enhances encephalitogenic T-cell responses, and suppresses the protective chemokine CXCL12 in CNS tissue. Inhibition of hyaluronan synthesis with 4MU protects against an animal model of MS and may represent an important therapeutic option in MS and other neuroinflammatory diseases.


Assuntos
Sistema Nervoso Central/imunologia , Quimiocina CXCL12/imunologia , Encefalomielite Autoimune Experimental/imunologia , Regulação da Expressão Gênica/imunologia , Ácido Hialurônico/imunologia , Transferência Adotiva , Animais , Fármacos Anti-HIV/farmacologia , Benzilaminas , Sistema Nervoso Central/metabolismo , Quimiocina CXCL12/biossíntese , Ciclamos , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Compostos Heterocíclicos/farmacologia , Ácido Hialurônico/biossíntese , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Camundongos , Ratos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/patologia , Células Th1/imunologia , Células Th1/metabolismo , Células Th1/patologia
15.
PLoS Pathog ; 9(2): e1003177, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23459172

RESUMO

The basidiomycete Ustilago maydis causes smut disease in maize, with large plant tumors being formed as the most prominent disease symptoms. During all steps of infection, U. maydis depends on a biotrophic interaction, which requires an efficient suppression of plant immunity. In a previous study, we identified the secreted effector protein Pit2, which is essential for maintenance of biotrophy and induction of tumors. Deletion mutants for pit2 successfully penetrate host cells but elicit various defense responses, which stops further fungal proliferation. We now show that Pit2 functions as an inhibitor of a set of apoplastic maize cysteine proteases, whose activity is directly linked with salicylic-acid-associated plant defenses. Consequently, protease inhibition by Pit2 is required for U. maydis virulence. Sequence comparisons with Pit2 orthologs from related smut fungi identified a conserved sequence motif. Mutation of this sequence caused loss of Pit2 function. Consequently, expression of the mutated protein in U. maydis could not restore virulence of the pit2 deletion mutant, indicating that the protease inhibition by Pit2 is essential for fungal virulence. Moreover, synthetic peptides of the conserved sequence motif showed full activity as protease inhibitor, which identifies this domain as a new, minimal protease inhibitor domain in plant-pathogenic fungi.


Assuntos
Cisteína Proteases/química , Cisteína Proteases/metabolismo , Proteínas Fúngicas/metabolismo , Doenças das Plantas/microbiologia , Ustilago/fisiologia , Virulência , Zea mays/enzimologia , Sequência de Aminoácidos , Cisteína Proteases/genética , Inibidores Enzimáticos/farmacologia , Proteínas Fúngicas/genética , Interações Hospedeiro-Patógeno , Immunoblotting , Dados de Sequência Molecular , Fragmentos de Peptídeos/metabolismo , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Deleção de Sequência , Homologia de Sequência de Aminoácidos , Técnicas do Sistema de Duplo-Híbrido , Ustilago/patogenicidade , Zea mays/microbiologia
16.
New Phytol ; 206(3): 1116-1126, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25628012

RESUMO

The secreted fungal effector Pep1 is essential for penetration of the host epidermis and establishment of biotrophy in the Ustilago maydis-maize pathosystem. Previously, Pep1 was found to be an inhibitor of apoplastic plant peroxidases, which suppresses the oxidative burst, a primary immune response of the host plant and enables fungal colonization. To investigate the conservation of Pep1 in other pathogens, genomes of related smut species were screened for pep1 orthologues. Pep1 proteins were produced in Escherichia coli for functional assays. The biological function of Pep1 was tested by heterologous expression in U. maydis and Hordeum vulgare. Pep1 orthologues revealed a remarkable degree of sequence conservation, indicating that this effector might play a fundamental role in virulence of biotrophic smut fungi. Pep1 function and its role in virulence are conserved in different pathogenic fungi, even across the monocot-dicot border of host plants. The findings described in this study classify Pep1 as a phylogenetically conserved fungal core effector. Furthermore, we documented the influence of Pep1 on the disease caused by Blumeria graminis f. sp. hordei which is a non-smut-related pathosystem.


Assuntos
Proteínas Fúngicas/genética , Fungos/genética , Sequência de Aminoácidos , Sequência de Bases , Códon , Sequência Conservada , Escherichia coli/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/fisiologia , Fungos/patogenicidade , Genoma Fúngico , Hordeum/genética , Hordeum/metabolismo , Hordeum/microbiologia , Interações Hospedeiro-Patógeno/genética , Dados de Sequência Molecular , Filogenia , Espécies Reativas de Oxigênio/metabolismo , Alinhamento de Sequência , Ustilago/genética , Ustilago/patogenicidade
17.
Alzheimers Dement ; 11(8): 964-74, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25824567

RESUMO

BACKGROUND: Evaluation of brain ß-amyloid by positron emission tomography (PET) imaging can assist in the diagnosis of Alzheimer disease (AD) and other dementias. METHODS: Open-label, nonrandomized, multicenter, phase 3 study to validate the (18)F-labeled ß-amyloid tracer florbetaben by comparing in vivo PET imaging with post-mortem histopathology. RESULTS: Brain images and tissue from 74 deceased subjects (of 216 trial participants) were analyzed. Forty-six of 47 neuritic ß-amyloid-positive cases were read as PET positive, and 24 of 27 neuritic ß-amyloid plaque-negative cases were read as PET negative (sensitivity 97.9% [95% confidence interval or CI 93.8-100%], specificity 88.9% [95% CI 77.0-100%]). In a subgroup, a regional tissue-scan matched analysis was performed. In areas known to strongly accumulate ß-amyloid plaques, sensitivity and specificity were 82% to 90%, and 86% to 95%, respectively. CONCLUSIONS: Florbetaben PET shows high sensitivity and specificity for the detection of histopathology-confirmed neuritic ß-amyloid plaques and may thus be a valuable adjunct to clinical diagnosis, particularly for the exclusion of AD. TRIAL REGISTRATION: ClinicalTrials.govNCT01020838.


Assuntos
Doença de Alzheimer , Peptídeos beta-Amiloides/metabolismo , Compostos de Anilina/farmacocinética , Encéfalo , Placa Amiloide/patologia , Estilbenos/farmacocinética , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/diagnóstico por imagem , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Análise de Variância , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Encéfalo/patologia , Estudos de Coortes , Diagnóstico , Feminino , Humanos , Processamento de Imagem Assistida por Computador , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Tomografia por Emissão de Pósitrons , Escalas de Graduação Psiquiátrica , Curva ROC , Radiografia
18.
Emerg Microbes Infect ; : 2417868, 2024 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-39435487

RESUMO

The complement system is a vital anti-microbial defence mechanism against circulating pathogens. Excessive complement activation can have deleterious outcomes for the host and is consequently tightly modulated by a set of membrane-associated and fluid-phase regulators of complement activation (RCAs). Here, we demonstrate that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) hijacks host cellular RCA members CD55 and CD59 and serum-derived Factor H (FH) to resist antibody-dependent complement-mediated lysis triggered by immunised human sera. Blockage of the biological functions of virion-associated CD55 and CD59 and competition of FH recruitment with functionally inactive recombinant FH-derived short consensus repeats SCR18-20 restore SARS-CoV-2 complement sensitivity in a synergistic manner. Moreover, complement-mediated virolysis is dependent on classical pathway activation and does not occur in the absence of virus-specific antibodies. Altogether, our findings present an intriguing immune escape mechanism that provides novel insights into the immunopathology observed in severe coronavirus disease 2019 (COVID-19).

19.
Plant Cell Physiol ; 53(7): 1232-46, 2012 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22537757

RESUMO

The barley (Hordeum vulgare L.) mutants fch2 and clo-f2 comprise an allelic group of 14 Chl b-deficient lines. The genetic map position of fch2 corresponds to the physical map position of the gene encoding chlorophyllide a oxygenase. This enzyme converts chlorophyllide a to chlorophyllide b and it is essential for Chl b biosynthesis. The fch2 and clo-f2 barley lines were shown to be mutated in the gene for chlorophyllide a oxygenase. A five-base insertion was found in fch2 and base deletions in clo-f2.101, clo-f2.105, clo-f2.2800 and clo-f2.3613. In clo-f2.105 and clo-f2.108, nonsense base exchanges were discovered. All of these mutations led to a premature stop of translation and none of the mutants formed Chl b. The mutant clo-f2.2807 was transcript deficient and formed no Chl b. Missense mutations in clo-f2.102 (leading to the amino acid exchange D495N) and clo-f2.103 (G280D) resulted in a total lack of Chl b, whereas in the missense mutants clo-f2.107 (P419L), clo-f2.109 (A94T), clo-f2.122 (C320Y), clo-f2.123 (A94T), clo-f2.133 (A376V) and clo-f2.181 (L373F) intermediate contents of Chl b were determined. The missense mutations affect conserved residues, and their effect on chlorophyllide a oxygenase is discussed. The mutations in clo-f2.102, clo-f2.103, clo-f2.133 and clo-f2.181 may influence electron transfer as illustrated in the active site of a structural model protein. The changes in clo-f2.107, clo-f2.109, clo-f2.122 and clo-f2.123 may lead to Chlb deficiency by interfering with the regulation of chlorophyllide a oxygenase. The correlation of mutations and phenotypes strongly supports that the barley locus fch2 encodes chlorophyllide a oxygenase.


Assuntos
Hordeum/genética , Mutação de Sentido Incorreto , Oxigenases/genética , Proteínas de Plantas/metabolismo , Alelos , Sequência de Aminoácidos , Domínio Catalítico , Clorofila/genética , Clorofila/metabolismo , Mapeamento Cromossômico , Cromossomos de Plantas/genética , Cromossomos de Plantas/metabolismo , Clonagem Molecular , Códon sem Sentido/genética , Códon sem Sentido/metabolismo , Transporte de Elétrons , Mutação da Fase de Leitura , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Hordeum/enzimologia , Dados de Sequência Molecular , Oxigenases/metabolismo , Fenótipo , Proteínas de Plantas/genética , Estrutura Terciária de Proteína , Alinhamento de Sequência , Sintenia
20.
Environ Microbiol Rep ; 14(1): 60-69, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34797028

RESUMO

Root endospheres house complex and diverse bacterial communities, of which many strains have not been cultivated yet by means of the currently available isolation techniques. The Prospector® (General Automation Lab Technologies, San Carlos, CA, USA), an automated and high-throughput bacterial cultivation system, was applied to analyse the root endomicrobiome of lettuce (Lactuca sativa L.). By using deep sequencing, we compared the results obtained with the Prospector and the traditional solid medium culturing and extinction methods. We found that the species richness did not differ and that the amount of previously uncultured bacteria did not increase, but that the bacterial diversity isolated by the three methods varied. In addition, the tryptic soy broth and King's B media provided a lower, but different, diversity of bacteria than that of Reasoner's 2A (R2A) medium when used within the Prospector system and the number of unique bacterial strains did not weigh up against those isolated with the R2A medium. Thus, to cultivate as broad a variety of bacteria as possible, divergent isolation techniques should be used in parallel. Thanks to its speed and limited manual requirements, the Prospector is a valuable system to enlarge root microbiome culture collections.


Assuntos
Lactuca , Microbiota , Meios de Cultura , Força da Mão , Rios
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