Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 38
Filtrar
Mais filtros










Intervalo de ano de publicação
1.
PLoS Pathog ; 18(5): e1010532, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35533195

RESUMO

Ebola virus (EBOV) VP35 is a polyfunctional protein involved in viral genome packaging, viral polymerase function, and host immune antagonism. The mechanisms regulating VP35's engagement in different functions are not well-understood. We previously showed that the host E3 ubiquitin ligase TRIM6 ubiquitinates VP35 at lysine 309 (K309) to facilitate virus replication. However, how K309 ubiquitination regulates the function of VP35 as the viral polymerase co-factor and the precise stage(s) of the EBOV replication cycle that require VP35 ubiquitination are not known. Here, we generated recombinant EBOVs encoding glycine (G) or arginine (R) mutations at VP35/K309 (rEBOV-VP35/K309G/-R) and show that both mutations prohibit VP35/K309 ubiquitination. The K309R mutant retains dsRNA binding and efficient type-I Interferon (IFN-I) antagonism due to the basic residue conservation. The rEBOV-VP35/K309G mutant loses the ability to efficiently antagonize the IFN-I response, while the rEBOV-VP35/K309R mutant's suppression is enhanced. The replication of both mutants was significantly attenuated in both IFN-competent and -deficient cells due to impaired interactions with the viral polymerase. The lack of ubiquitination on VP35/K309 or TRIM6 deficiency disrupts viral transcription with increasing severity along the transcriptional gradient. This disruption of the transcriptional gradient results in unbalanced viral protein production, including reduced synthesis of the viral transcription factor VP30. In addition, lack of ubiquitination on K309 results in enhanced interactions with the viral nucleoprotein and premature nucleocapsid packaging, leading to dysregulation of virus assembly. Overall, we identified a novel role of VP35 ubiquitination in coordinating viral transcription and assembly.


Assuntos
Ebolavirus , Doença pelo Vírus Ebola , Ebolavirus/metabolismo , Humanos , Lisina/genética , Lisina/metabolismo , Proteínas do Nucleocapsídeo/metabolismo , Ubiquitinação , Proteínas Virais Reguladoras e Acessórias/genética , Proteínas Virais Reguladoras e Acessórias/metabolismo , Transcrição Viral
2.
Cell Rep ; 38(10): 110434, 2022 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-35263596

RESUMO

Type I interferons (IFN-I) are essential to establish antiviral innate immunity. Unanchored (or free) polyubiquitin (poly-Ub) has been shown to regulate IFN-I responses. However, few unanchored poly-Ub interactors are known. To identify factors regulated by unanchored poly-Ub in a physiological setting, we developed an approach to isolate unanchored poly-Ub from lung tissue. We identified the RNA helicase DHX16 as a potential pattern recognition receptor (PRR). Silencing of DHX16 in cells and in vivo diminished IFN-I responses against influenza virus. These effects extended to members of other virus families, including Zika and SARS-CoV-2. DHX16-dependent IFN-I production requires RIG-I and unanchored K48-poly-Ub synthesized by the E3-Ub ligase TRIM6. DHX16 recognizes a signal in influenza RNA segments that undergo splicing and requires its RNA helicase motif for direct, high-affinity interactions with specific viral RNAs. Our study establishes DHX16 as a PRR that partners with RIG-I for optimal activation of antiviral immunity requiring unanchored poly-Ub.


Assuntos
Proteína DEAD-box 58 , Interferon Tipo I , RNA Helicases , RNA Viral , Receptores Imunológicos , Infecção por Zika virus , Zika virus , COVID-19 , Proteína DEAD-box 58/imunologia , Humanos , Imunidade Inata , Interferon Tipo I/imunologia , RNA Helicases/imunologia , Receptores Imunológicos/imunologia , SARS-CoV-2 , Proteínas com Motivo Tripartido , Zika virus/genética , Infecção por Zika virus/imunologia
3.
Nature ; 585(7825): 414-419, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32641828

RESUMO

Zika virus (ZIKV) belongs to the family Flaviviridae, and is related to other viruses that cause human diseases. Unlike other flaviviruses, ZIKV infection can cause congenital neurological disorders and replicates efficiently in reproductive tissues1-3. Here we show that the envelope protein (E) of ZIKV is polyubiquitinated by the E3 ubiquitin ligase TRIM7 through Lys63 (K63)-linked polyubiquitination. Accordingly, ZIKV replicates less efficiently in the brain and reproductive tissues of Trim7-/- mice. Ubiquitinated E is present on infectious virions of ZIKV when they are released from specific cell types, and enhances virus attachment and entry into cells. Specifically, K63-linked polyubiquitin chains directly interact with the TIM1 (also known as HAVCR1) receptor of host cells, which enhances virus entry in cells as well as in brain tissue in vivo. Recombinant ZIKV mutants that lack ubiquitination are attenuated in human cells and in wild-type mice, but not in live mosquitoes. Monoclonal antibodies against K63-linked polyubiquitin specifically neutralize ZIKV and reduce viraemia in mice. Our results demonstrate that the ubiquitination of ZIKV E is an important determinant of virus entry, tropism and pathogenesis.


Assuntos
Ubiquitinação , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/metabolismo , Internalização do Vírus , Zika virus/metabolismo , Zika virus/patogenicidade , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Encéfalo/metabolismo , Linhagem Celular , Culicidae/citologia , Culicidae/virologia , Endossomos/metabolismo , Feminino , Receptor Celular 1 do Vírus da Hepatite A/metabolismo , Humanos , Masculino , Fusão de Membrana , Camundongos , Especificidade de Órgãos , Poliubiquitina/imunologia , Poliubiquitina/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Tropismo Viral , Viremia/imunologia , Viremia/prevenção & controle , Viremia/virologia , Replicação Viral , Zika virus/química , Zika virus/genética , Infecção por Zika virus/prevenção & controle , Infecção por Zika virus/virologia
4.
Cell Death Dis ; 11(6): 444, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32518230

RESUMO

The skin is a high turnover organ, and its constant renewal depends on the rapid proliferation of its progenitor cells. The energy requirement for these metabolically active cells is met by mitochondrial respiration, an ATP generating process driven by a series of protein complexes collectively known as the electron transport chain (ETC) that is located on the inner membrane of the mitochondria. However, reactive oxygen species (ROS) like superoxide, singlet oxygen, peroxides are inevitably produced during respiration and disrupt macromolecular and cellular structures if not quenched by the antioxidant system. The oxidative damage caused by mitochondrial ROS production has been established as the molecular basis of multiple pathophysiological conditions, including aging and cancer. Not surprisingly, the mitochondria are the primary organelle affected during chronological and UV-induced skin aging, the phenotypic manifestations of which are the direct consequence of mitochondrial dysfunction. Also, deletions and other aberrations in the mitochondrial DNA (mtDNA) are frequent in photo-aged skin and skin cancer lesions. Recent studies have revealed a more innate role of the mitochondria in maintaining skin homeostasis and pigmentation, which are affected when the essential mitochondrial functions are impaired. Some common and rare skin disorders have a mitochondrial involvement and include dermal manifestations of primary mitochondrial diseases as well as congenital skin diseases caused by damaged mitochondria. With studies increasingly supporting the close association between mitochondria and skin health, its therapeutic targeting in the skin-either via an ATP production boost or free radical scavenging-has gained attention from clinicians and aestheticians alike. Numerous bioactive compounds have been identified that improve mitochondrial functions and have proved effective against aged and diseased skin. In this review, we discuss the essential role of mitochondria in regulating normal and abnormal skin physiology and the possibility of targeting this organelle in various skin disorders.


Assuntos
Mitocôndrias/metabolismo , Envelhecimento da Pele/patologia , Dermatopatias/metabolismo , Dermatopatias/patologia , Pele/metabolismo , Pele/patologia , Animais , Homeostase , Humanos , Regeneração
5.
Clin Exp Allergy ; 48(12): 1676-1687, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30244512

RESUMO

BACKGROUND: Ragweed pollen extract (RWPE) induces TLR4-NFκB-CXCL-dependent recruitment of ROS-generating neutrophils to the airway and OGG1 DNA glycosylase-dependent excision of oxidatively induced 8-OH-Gua DNA base lesions from the airway epithelial cell genome. Administration of free 8-OH-Gua base stimulates RWPE-induced allergic lung inflammation. These studies suggest that stimulation of innate receptors and their adaptor by allergenic extracts initiates excision of a set of DNA base lesions that facilitate innate/allergic lung inflammation. OBJECTIVE: To test the hypothesis that stimulation of a conserved innate receptor/adaptor pathway by allergenic extracts induces excision of a set of pro-inflammatory oxidatively induced DNA base lesions from the lung genome that stimulate allergic airway inflammation. METHODS: Wild-type (WT), Tlr4KO, Tlr2KO, Myd88KO, and TrifKO mice were intranasally challenged once or repeatedly with cat dander extract (CDE), and innate or allergic inflammation and gene expression were quantified. We utilized GC-MS/MS to quantify a set of oxidatively induced DNA base lesions after challenge of naïve mice with CDE. RESULTS: A single CDE challenge stimulated innate neutrophil recruitment that was partially dependent on TLR4 and TLR2, and completely on Myd88, but not TRIF. A single CDE challenge stimulated MyD88-dependent excision of DNA base lesions 5-OH-Cyt, FapyAde, and FapyGua from the lung genome. A single challenge of naïve WT mice with 5-OH-Cyt stimulated neutrophilic lung inflammation. Multiple CDE instillations stimulated MyD88-dependent allergic airway inflammation. Multiple administrations of 5-OH-Cyt with CDE stimulated allergic sensitization and allergic airway inflammation. CONCLUSIONS AND CLINICAL RELEVANCE: We show for the first time that CDE challenge stimulates MyD88-dependent excision of DNA base lesions. Our data suggest that the resultant-free base(s) contribute to CDE-induced innate/allergic lung inflammation. We suggest that blocking the MyD88 pathway in the airways with specific inhibitors may be a novel targeted strategy of inhibiting amplification of innate and adaptive immune inflammation in allergic diseases by oxidatively induced DNA base lesions.


Assuntos
Citosina/análogos & derivados , Dano ao DNA/efeitos dos fármacos , Hipersensibilidade/etiologia , Hipersensibilidade/metabolismo , Pulmão/metabolismo , Estresse Oxidativo , Alérgenos/imunologia , Animais , Biomarcadores , Gatos , Cromatografia Gasosa , Citosina/farmacologia , Citosina/toxicidade , Modelos Animais de Doenças , Hipersensibilidade/patologia , Imunidade Inata , Imunoglobulina E/imunologia , Pulmão/imunologia , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/metabolismo , Pneumonia/etiologia , Pneumonia/metabolismo , Pneumonia/patologia , Espécies Reativas de Oxigênio , Espectrometria de Massas em Tandem
6.
Redox Biol ; 18: 43-53, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29940424

RESUMO

8-Oxoguanine DNA glycosylase 1 (OGG1) initiates the base excision repair pathway by removing one of the most abundant DNA lesions, 8-oxo-7,8-dihydroguanine (8-oxoG). Recent data showed that 8-oxoG not only is a pro-mutagenic genomic base lesion, but also functions as an epigenetic mark and that consequently OGG1 acquire distinct roles in modulation of gene expression. In support, lack of functional OGG1 in Ogg1-/- mice led to an altered expression of genes including those responsible for the aberrant innate and adaptive immune responses and susceptibility to metabolic disorders. Therefore, the present study examined stimulus-driven OGG1-DNA interactions at whole genome level using chromatin immunoprecipitation (ChIP)-coupled sequencing, and the roles of OGG1 enriched on the genome were validated by molecular and system-level approaches. Results showed that signaling levels of cellular ROS generated by TNFα, induced enrichment of OGG1 at specific sites of chromatinized DNA, primarily in the regulatory regions of genes. OGG1-ChIP-ed genes are associated with important cellular and biological processes and OGG1 enrichment was limited to a time scale required for immediate cellular responses. Prevention of OGG1-DNA interactions by siRNA depletion led to modulation of NF-κB's DNA occupancy and differential expression of genes. Taken together these data show TNFα-ROS-driven enrichment of OGG1 at gene regulatory regions in the chromatinized DNA, which is a prerequisite to modulation of gene expression for prompt cellular responses to oxidant stress.


Assuntos
Dano ao DNA , DNA Glicosilases/metabolismo , Redes Reguladoras de Genes , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Cromatina/genética , Cromatina/metabolismo , DNA/genética , DNA/metabolismo , Células HEK293 , Humanos , Fator de Necrose Tumoral alfa/metabolismo
7.
Biol Reprod ; 99(5): 1100-1112, 2018 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-29893818

RESUMO

Term and preterm parturition are associated with oxidative stress (OS)-induced p38 mitogen-activated protein kinase (p38MAPK)-mediated fetal tissue (amniochorion) senescence. p38MAPK activation is a complex cell- and stimulant-dependent process. Two independent pathways of OS-induced p38MAPK activation were investigated in amnion epithelial cells (AECs) in response to cigarette smoke extract (CSE: a validated OS inducer in fetal cells): (1) the OS-mediated oxidation of apoptosis signal-regulating kinase (ASK)-1 bound Thioredoxin (Trx[SH]2) dissociates this complex, creating free and activated ASK1-signalosome and (2) transforming growth factor-mediated activation of (TGF)-beta-activated kinase (TAK)1 and TGF-beta-activated kinase 1-binding protein (TAB)1. AECs isolated from normal term, not-in-labor fetal membranes increased p38MAPK in response to CSE and downregulated it in response to antioxidant N-acetylcysteine. In AECs, both Trx and ASK1 were localized; however, they remained dissociated and not complexed, regardless of conditions. Silencing either ASK1 or its downstream effectors (MKK3/6) did not affect OS-induced p38MAPK activation. Conversely, OS increased TGF-beta's release from AECs and increased phosphorylation of both p38MAPK and TAB1. Silencing of TAB1, but not TAK1, prevented p38MAPK activation, which is indicative of TAB1-mediated autophosphorylation of p38MAPK, an activation mechanism seldom seen. OS-induced p38MAPK activation in AECs is ASK1-Trx signalosome-independent and is mediated by the TGF-beta pathway. This knowledge will help to design strategies to reduce p38MAPK activation-associated pregnancy risks.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Âmnio/citologia , Células Epiteliais/metabolismo , Estresse Oxidativo , Fator de Crescimento Transformador beta/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Adulto , Ativação Enzimática , Células Epiteliais/efeitos dos fármacos , Feminino , Humanos , MAP Quinase Quinase Quinase 5/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Fosforilação , Gravidez , RNA Interferente Pequeno/farmacologia , Fumaça , Nicotiana/química
8.
Am J Physiol Lung Cell Mol Physiol ; 313(6): L1058-L1068, 2017 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-28798252

RESUMO

A mucosal oxidative burst is a hallmark response to pollen exposure that promotes allergic inflammatory responses. Reactive species constituents of oxidative stress signal via the modification of cellular molecules including nucleic acids. One of the most abundant forms of oxidative genomic base damage is 8-oxo-7,8-dihydroguanine (8-oxoG), which is removed from DNA by 8-oxoguanine DNA glycosylase 1 (OGG1). OGG1 in complex with 8-oxoG acts as a GDP-GTP exchange factor and induces acute inflammation; however, the mechanism(s) by which OGG1 signaling regulates allergic airway inflammation is not known. Here, we postulate that the OGG1 signaling pathway differentially altered the levels of small regulatory RNAs and increased the expression of T helper 2 (Th2) cytokines in ragweed pollen extract (RWPE)-challenged lungs. To determine this, the lungs of sensitized mice expressing or lacking OGG1 were challenged with RWPE and/or with OGG1's excision product 8-oxoG. The responses in lungs were assessed by next-generation sequencing, as well as various molecular and histological approaches. The results showed that RWPE challenge induced oxidative burst and damage to DNA and activated OGG1 signaling, resulting in the differential expression of 84 micro-RNAs (miRNAs), which then exacerbated antigen-driven allergic inflammation and histological changes in the lungs. The exogenous administration of the downregulated let-7b-p3 mimetic or inhibitors of upregulated miR-23a or miR-27a decreased eosinophil recruitment and mucus and collagen production via controlling the expression of IL-4, IL-5, and IL-13. Together, these data demonstrate the roles of OGG1 signaling in the regulation of antigen-driven allergic immune responses via differential expression of miRNAs upstream of Th2 cytokines and eosinophils.


Assuntos
Antígenos de Plantas/toxicidade , Dano ao DNA , Hipersensibilidade/imunologia , MicroRNAs/imunologia , Extratos Vegetais/toxicidade , Eosinofilia Pulmonar/imunologia , Células Th2/imunologia , Animais , Linhagem Celular Transformada , Citocinas/genética , Citocinas/imunologia , DNA Glicosilases/genética , DNA Glicosilases/imunologia , Hipersensibilidade/genética , Hipersensibilidade/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , MicroRNAs/genética , Eosinofilia Pulmonar/induzido quimicamente , Eosinofilia Pulmonar/genética , Eosinofilia Pulmonar/patologia , Células Th2/patologia
10.
Braz. j. microbiol ; 48(2): 326-332, April.-June 2017. tab, graf
Artigo em Inglês | LILACS | ID: biblio-839372

RESUMO

Abstract Stress tolerance is a key attribute that must be considered when using yeast cells for industrial applications. High temperature is one factor that can cause stress in yeast. High environmental temperature in particular may exert a natural selection pressure to evolve yeasts into thermotolerant strains. In the present study, three yeasts (Saccharomyces cerevisiae, MC4, and Kluyveromyces marxianus, OFF1 and SLP1) isolated from hot environments were exposed to increased temperatures and were then compared with a laboratory yeast strain. Their resistance to high temperature, oxidative stress, and antioxidant response were evaluated, along with the fatty acid composition of their cell membranes. The SLP1 strain showed a higher specific growth rate, biomass yield, and biomass volumetric productivity while also showing lower duplication time, reactive oxygen species (ROS) production, and lipid peroxidation. In addition, the SLP1 strain demonstrated more catalase activity after temperature was increased, and this strain also showed membranes enriched in saturated fatty acids. It is concluded that the SLP1 yeast strain is a thermotolerant yeast with less oxidative stress and a greater antioxidant response. Therefore, this strain could be used for fermentation at high temperatures.


Assuntos
Saccharomyces cerevisiae/fisiologia , Estresse Fisiológico , Kluyveromyces/fisiologia , Estresse Oxidativo , Antioxidantes/metabolismo , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/efeitos da radiação , Saccharomyces cerevisiae/química , Kluyveromyces/crescimento & desenvolvimento , Kluyveromyces/efeitos da radiação , Kluyveromyces/química , Peroxidação de Lipídeos , Catalase/análise , Membrana Celular/química , Espécies Reativas de Oxigênio/metabolismo , Biomassa , Ácidos Graxos/análise , Temperatura Alta
12.
Braz J Microbiol ; 48(2): 326-332, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28094115

RESUMO

Stress tolerance is a key attribute that must be considered when using yeast cells for industrial applications. High temperature is one factor that can cause stress in yeast. High environmental temperature in particular may exert a natural selection pressure to evolve yeasts into thermotolerant strains. In the present study, three yeasts (Saccharomyces cerevisiae, MC4, and Kluyveromyces marxianus, OFF1 and SLP1) isolated from hot environments were exposed to increased temperatures and were then compared with a laboratory yeast strain. Their resistance to high temperature, oxidative stress, and antioxidant response were evaluated, along with the fatty acid composition of their cell membranes. The SLP1 strain showed a higher specific growth rate, biomass yield, and biomass volumetric productivity while also showing lower duplication time, reactive oxygen species (ROS) production, and lipid peroxidation. In addition, the SLP1 strain demonstrated more catalase activity after temperature was increased, and this strain also showed membranes enriched in saturated fatty acids. It is concluded that the SLP1 yeast strain is a thermotolerant yeast with less oxidative stress and a greater antioxidant response. Therefore, this strain could be used for fermentation at high temperatures.


Assuntos
Antioxidantes/metabolismo , Kluyveromyces/fisiologia , Estresse Oxidativo , Saccharomyces cerevisiae/fisiologia , Estresse Fisiológico , Biomassa , Catalase/análise , Membrana Celular/química , Ácidos Graxos/análise , Temperatura Alta , Kluyveromyces/química , Kluyveromyces/crescimento & desenvolvimento , Kluyveromyces/efeitos da radiação , Peroxidação de Lipídeos , Espécies Reativas de Oxigênio/metabolismo , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/crescimento & desenvolvimento , Saccharomyces cerevisiae/efeitos da radiação
13.
Mech Ageing Dev ; 161(Pt A): 51-65, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-27343030

RESUMO

Age-associated changes in lung structure and function are some of the most important predictors of overall health, cognitive activities and longevity. Common to all aging cells is an increase in oxidatively modified DNA bases, primarily 8-oxo-7,8-dihydroguanine (8-oxoG). It is repaired via DNA base excision repair pathway driven by 8-oxoguanine DNA glycosylase-1 (OGG1-BER), whose role in aging has been the focus of many studies. This study hypothesizes that signaling and consequent gene expression during cellular response to OGG1-BER "wires" senescence/aging processes. To test OGG1-BER was mimicked by repeatedly exposing diploid lung fibroblasts cells and airways of mice to 8-oxoG base. Results showed that repeated exposures led to G1 cell cycle arrest and pre-matured senescence of cultured cells in which over 1000 genes were differentially expressed -86% of them been identical to those in naturally senesced cells. Gene ontology analysis of gene expression displayed biological processes driven by small GTPases, phosphoinositide 3-kinase and mitogen activated kinase cascades both in cultured cells and lungs. These results together, points to a new paradigm about the role of DNA damage and repair by OGG1 in aging and age-associated disease processes.


Assuntos
Envelhecimento/metabolismo , DNA Glicosilases/metabolismo , Reparo do DNA , Fibroblastos/enzimologia , Pulmão/enzimologia , Envelhecimento/patologia , Animais , Linhagem Celular , Fibroblastos/patologia , Pontos de Checagem da Fase G1 do Ciclo Celular , Humanos , Pulmão/patologia , Camundongos
14.
Mediators Inflamm ; 2016: 3762561, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27524866

RESUMO

Asthma is characterized by reversible airway narrowing, shortness of breath, wheezing, coughing, and other symptoms driven by chronic inflammatory processes, commonly triggered by allergens. In 90% of asthmatics, most of these symptoms can also be triggered by intense physical activities and severely exacerbated by environmental factors. This condition is known as exercise-induced asthma (EIA). Current theories explaining EIA pathogenesis involve osmotic and/or thermal alterations in the airways caused by changes in respiratory airflow during exercise. These changes, along with existing airway inflammatory conditions, are associated with increased cellular levels of reactive oxygen species (ROS) affecting important biomolecules including DNA, although the underlying molecular mechanisms have not been completely elucidated. One of the most abundant oxidative DNA lesions is 8-oxoguanine (8-oxoG), which is repaired by 8-oxoguanine DNA glycosylase 1 (OGG1) during the base excision repair (BER) pathway. Whole-genome expression analyses suggest a cellular response to OGG1-BER, involving genes that may have a role in the pathophysiology of EIA leading to mast cell degranulation, airway hyperresponsiveness, and bronchoconstriction. Accordingly, this review discusses a potential new hypothesis in which OGG1-BER-induced gene expression is associated with EIA symptoms.


Assuntos
Asma/metabolismo , DNA Glicosilases/metabolismo , Reparo do DNA , Exercício Físico , Guanina/análogos & derivados , Animais , Broncoconstrição , DNA/análise , Guanina/química , Humanos , Inflamação , Peroxidação de Lipídeos , Mastócitos/citologia , Camundongos , Estresse Oxidativo , Condicionamento Físico Animal , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais
15.
J Allergy Clin Immunol ; 137(5): 1506-1513.e2, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-26586036

RESUMO

BACKGROUND: The National Health and Nutrition Examination Survey identified several pollens and cat dander as among the most common allergens that induce allergic sensitization and allergic diseases. We recently reported that ragweed pollen extract (RWPE) requires Toll-like receptor 4 (TLR4) to stimulate CXCL-mediated innate neutrophilic inflammation, which in turn facilitates allergic sensitization and airway inflammation. Myeloid differentiation protein 2 (MD2) is a TLR4 coreceptor, but its role in pollen- and cat dander-induced innate and allergic inflammation has not been critically evaluated. OBJECTIVE: We sought to elucidate the role of MD2 in inducing pollen- and cat dander-induced innate and allergic airway inflammation. METHODS: TCM(Null) (TLR4(Null), CD14(Null), MD2(Null)), TLR4(Hi), and TCM(Hi) cells and human bronchial epithelial cells with small interfering RNA-induced downregulation of MD2 were stimulated with RWPE, other pollen allergic extracts, or cat dander extract (CDE), and activation of nuclear factor κB (NF-κB), secretion of the NF-κB-dependent CXCL8, or both were quantified. Wild-type mice or mice with small interfering RNA knockdown of lung MD2 were challenged intranasally with RWPE or CDE, and innate and allergic inflammation was quantified. RESULTS: RWPE stimulated MD2-dependent NF-κB activation and CXCL secretion. Likewise, Bermuda, rye, timothy, pigweed, Russian thistle, cottonwood, walnut, and CDE stimulated MD2-dependent CXCL secretion. RWPE and CDE challenge induced MD2-dependent and CD14-independent innate neutrophil recruitment. RWPE induced MD2-dependent allergic sensitization and airway inflammation. CONCLUSIONS: MD2 plays an important role in induction of allergic sensitization to cat dander and common pollens relevant to human allergic diseases.


Assuntos
Alérgenos/imunologia , Alérgenos Animais/imunologia , Antígeno 96 de Linfócito/imunologia , Pólen/imunologia , Hipersensibilidade Respiratória/imunologia , Animais , Antígenos de Plantas/imunologia , Líquido da Lavagem Broncoalveolar/imunologia , Gatos/imunologia , Linhagem Celular , Citocinas/imunologia , Humanos , Imunidade Inata , Pulmão/imunologia , Pulmão/metabolismo , Antígeno 96 de Linfócito/genética , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mucinas/metabolismo , NF-kappa B/imunologia , Extratos Vegetais/imunologia , RNA Mensageiro/metabolismo
16.
Am J Respir Cell Mol Biol ; 54(1): 81-90, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26086549

RESUMO

Neutrophil recruitment is a hallmark of rapid innate immune responses. Exposure of airways of naive mice to pollens rapidly induces neutrophil recruitment. The innate mechanisms that regulate pollen-induced neutrophil recruitment and the contribution of this neutrophilic response to subsequent induction of allergic sensitization and inflammation need to be elucidated. Here we show that ragweed pollen extract (RWPE) challenge in naive mice induces C-X-C motif ligand (CXCL) chemokine synthesis, which stimulates chemokine (C-X-C motif) receptor 2 (CXCR2)-dependent recruitment of neutrophils into the airways. Deletion of Toll-like receptor 4 (TLR4) abolishes CXCL chemokine secretion and neutrophil recruitment induced by a single RWPE challenge and inhibits induction of allergic sensitization and airway inflammation after repeated exposures to RWPE. Forced induction of CXCL chemokine secretion and neutrophil recruitment in mice lacking TLR4 also reconstitutes the ability of multiple challenges of RWPE to induce allergic airway inflammation. Blocking RWPE-induced neutrophil recruitment in wild-type mice by administration of a CXCR2 inhibitor inhibits the ability of repeated exposures to RWPE to stimulate allergic sensitization and airway inflammation. Administration of neutrophils derived from naive donor mice into the airways of Tlr4 knockout recipient mice after each repeated RWPE challenge reconstitutes allergic sensitization and inflammation in these mice. Together these observations indicate that pollen-induced recruitment of neutrophils is TLR4 and CXCR2 dependent and that recruitment of neutrophils is a critical rate-limiting event that stimulates induction of allergic sensitization and airway inflammation. Inhibiting pollen-induced recruitment of neutrophils, such as by administration of CXCR2 antagonists, may be a novel strategy to prevent initiation of pollen-induced allergic airway inflammation.


Assuntos
Antígenos de Plantas/imunologia , Imunidade Inata , Pulmão/imunologia , Infiltração de Neutrófilos , Neutrófilos/imunologia , Extratos Vegetais/imunologia , Pneumonia/imunologia , Hipersensibilidade Respiratória/imunologia , Animais , Antialérgicos/farmacologia , Anti-Inflamatórios/farmacologia , Modelos Animais de Doenças , Humanos , Imunidade Inata/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ativação de Neutrófilo , Infiltração de Neutrófilos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Pneumonia/metabolismo , Pneumonia/prevenção & controle , Espécies Reativas de Oxigênio/imunologia , Espécies Reativas de Oxigênio/metabolismo , Receptores de Interleucina-8B/antagonistas & inibidores , Receptores de Interleucina-8B/imunologia , Receptores de Interleucina-8B/metabolismo , Hipersensibilidade Respiratória/metabolismo , Hipersensibilidade Respiratória/prevenção & controle , Fatores de Tempo , Receptor 4 Toll-Like/deficiência , Receptor 4 Toll-Like/genética
17.
Immunol Lett ; 168(2): 306-12, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26493475

RESUMO

Extraocular muscles (EOM) are preferentially involved in myasthenia gravis (MG) and acetylcholine receptor (AChR) antibody positive MG patients may occasionally present with isolated ocular symptoms. Although experimental autoimmune myasthenia gravis (EAMG) induced by whole AChR immunization closely mimics clinical and immunopathological aspects of MG, EOM are usually not affected. We have previously developed an EAMG model, which imitates EOM symptoms of MG by immunization of human leukocyte antigen (HLA) transgenic mice with α or γ-subunits of human AChR (H-AChR). To investigate the significance of the ϵ-subunit in ocular MG, we immunized HLA-DR3 and HLA-DQ8 transgenic mice with recombinant H-AChR ϵ-subunit expressed in Escherichia coli. HLA-DR3 transgenic mice showed significantly higher clinical ocular and generalized MG severity scores and lower grip strength values than HLA-DQ8 mice. H-AChR ϵ-subunit-immunized HLA-DR3 transgenic mice had higher serum anti-AChR antibody (IgG, IgG1, IgG2b, IgG2c and IgM) levels, neuromuscular junction IgG and complement deposit percentages than ϵ-subunit-immunized HLA-DQ8 transgenic mice. Control mice immunized with E. coli extract or complete Freund adjuvant (CFA) did not show clinical and immunopathological features of ocular and generalized EAMG. Lymph node cells of ϵ-subunit-immunized HLA-DR3 mice showed significantly higher proliferative responses than those of ϵ-subunit-immunized HLA-DQ8 mice, crude E. coli extract-immunized and CFA-immunized transgenic mice. Our results indicate that the human AChR ϵ-subunit is capable of inducing myasthenic muscle weakness. Diversity of the autoimmune responses displayed by mice expressing different HLA class II molecules suggests that the interplay between HLA class II alleles and AChR subunits might have a profound impact on the clinical course of MG.


Assuntos
Antígenos HLA-DQ/imunologia , Antígeno HLA-DR3/imunologia , Miastenia Gravis Autoimune Experimental/imunologia , Músculos Oculomotores/imunologia , Animais , Proliferação de Células , Ensaio de Imunoadsorção Enzimática , Adjuvante de Freund/imunologia , Antígenos HLA-DQ/genética , Antígeno HLA-DR3/genética , Humanos , Imunização/métodos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Linfócitos/imunologia , Linfócitos/patologia , Camundongos Transgênicos , Microscopia de Fluorescência , Miastenia Gravis Autoimune Experimental/induzido quimicamente , Miastenia Gravis Autoimune Experimental/genética , Músculos Oculomotores/metabolismo , Músculos Oculomotores/patologia , Subunidades Proteicas/imunologia , Receptores Colinérgicos/imunologia
18.
J Biol Chem ; 290(41): 24636-48, 2015 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-26245904

RESUMO

Why mammalian cells possess multiple DNA glycosylases (DGs) with overlapping substrate ranges for repairing oxidatively damaged bases via the base excision repair (BER) pathway is a long-standing question. To determine the biological role of these DGs, null animal models have been generated. Here, we report the generation and characterization of mice lacking Neil2 (Nei-like 2). As in mice deficient in each of the other four oxidized base-specific DGs (OGG1, NTH1, NEIL1, and NEIL3), Neil2-null mice show no overt phenotype. However, middle-aged to old Neil2-null mice show the accumulation of oxidative genomic damage, mostly in the transcribed regions. Immuno-pulldown analysis from wild-type (WT) mouse tissue showed the association of NEIL2 with RNA polymerase II, along with Cockayne syndrome group B protein, TFIIH, and other BER proteins. Chromatin immunoprecipitation analysis from mouse tissue showed co-occupancy of NEIL2 and RNA polymerase II only on the transcribed genes, consistent with our earlier in vitro findings on NEIL2's role in transcription-coupled BER. This study provides the first in vivo evidence of genomic region-specific repair in mammals. Furthermore, telomere loss and genomic instability were observed at a higher frequency in embryonic fibroblasts from Neil2-null mice than from the WT. Moreover, Neil2-null mice are much more responsive to inflammatory agents than WT mice. Taken together, our results underscore the importance of NEIL2 in protecting mammals from the development of various pathologies that are linked to genomic instability and/or inflammation. NEIL2 is thus likely to play an important role in long term genomic maintenance, particularly in long-lived mammals such as humans.


Assuntos
DNA Glicosilases/deficiência , DNA Glicosilases/genética , DNA/metabolismo , Genoma/genética , Transcrição Gênica , Envelhecimento/genética , Envelhecimento/metabolismo , Animais , Linhagem Celular , DNA/genética , Dano ao DNA , Técnicas de Inativação de Genes , Instabilidade Genômica , Homeostase , Humanos , Inflamação/genética , Inflamação/metabolismo , Camundongos , Oxirredução , RNA Polimerase II/metabolismo , Telômero/genética
19.
Free Radic Biol Med ; 89: 20-33, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26187872

RESUMO

Reactive oxygen species (ROS) generated by environmental exposures, and endogenously as by-products of respiration, oxidatively modify biomolecules including DNA. Accumulation of ROS-induced DNA damage has been implicated in various diseases that involve inflammatory processes, and efficient DNA repair is considered critical in preventing such diseases. One of the most abundant DNA base lesions is 7,8-dihydro-8-oxoguanine (8-oxoG), which is repaired by the 8-oxoguanine DNA glycosylase 1 (OGG1)-initiated base-excision repair (OGG1-BER) pathway. Recent studies have shown that the OGG1-BER by-product 8-oxoG base forms a complex with cytosolic OGG1, activating small GTPases and downstream cell signaling in cultured cells and lungs. This implies that persistent OGG1-BER could result in signaling leading to histological changes in airways. To test this, we mimicked OGG1-BER by repeatedly challenging airways with its repair product 8-oxoG base. Gene expression was analyzed by RNA sequencing (RNA-Seq) and qRT-PCR, and datasets were evaluated by gene ontology and statistical tools. RNA-Seq analysis identified 3252 differentially expressed transcripts (2435 up- and 817 downregulated, ≥ 3-fold change). Among the upregulated transcripts, 2080 mRNAs were identified whose encoded protein products were involved in modulation of the actin family cytoskeleton, extracellular matrix, cell adhesion, cadherin, and cell junctions, affecting biological processes such as tissue development, cell-to-cell adhesion, cell communication, and the immune system. These data are supported by histological observations showing epithelial alterations, subepithelial fibrosis, and collagen deposits in the lungs. These data imply that continuous challenge by the environment and consequent OGG1-BER-driven signaling trigger gene expression consistent with airway remodeling.


Assuntos
Remodelação das Vias Aéreas/genética , DNA Glicosilases/metabolismo , Reparo do DNA/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Guanina/análogos & derivados , Animais , Biomarcadores/metabolismo , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , DNA Glicosilases/genética , Reparo do DNA/efeitos dos fármacos , Feminino , Ontologia Genética , Guanina/farmacologia , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Pulmão/citologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Estresse Oxidativo
20.
Free Radic Biol Med ; 81: 107-18, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25614460

RESUMO

Reactive oxygen species inflict oxidative modifications on various biological molecules, including DNA. One of the most abundant DNA base lesions, 8-oxo-7,8-dihydroguanine (8-oxoG) is repaired by 8-oxoguanine DNA glycosylase-1 (OGG1) during DNA base excision repair (OGG1-BER). 8-OxoG accumulation in DNA has been associated with various pathological and aging processes, although its role is unclear. The lack of OGG1-BER in Ogg1(-/-) mice resulted in decreased inflammatory responses and increased susceptibility to infections and metabolic disorders. Therefore, we proposed that OGG1 and/or 8-oxoG base may have a role in immune and homeostatic processes. To test our hypothesis, we challenged mouse lungs with OGG1-BER product 8-oxoG base and changes in gene expression were determined by RNA sequencing and data were analyzed by Gene Ontology and statistical tools. RNA-Seq analysis identified 1592 differentially expressed (≥ 3-fold change) transcripts. The upregulated mRNAs were related to biological processes, including homeostatic, immune-system, macrophage activation, regulation of liquid-surface tension, and response to stimulus. These processes were mediated by chemokines, cytokines, gonadotropin-releasing hormone receptor, integrin, and interleukin signaling pathways. Taken together, these findings point to a new paradigm showing that OGG1-BER plays a role in various biological processes that may benefit the host, but when in excess could be implicated in disease and/or aging processes.


Assuntos
DNA Glicosilases/genética , Reparo do DNA , DNA/genética , Pneumonia/genética , Transcriptoma , Administração Intranasal , Animais , Líquido da Lavagem Broncoalveolar/química , DNA/metabolismo , DNA Glicosilases/deficiência , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Guanina/administração & dosagem , Guanina/análogos & derivados , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/patologia , Redes e Vias Metabólicas/genética , Redes e Vias Metabólicas/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Anotação de Sequência Molecular , Estresse Oxidativo , Pneumonia/induzido quimicamente , Pneumonia/imunologia , Pneumonia/patologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...