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1.
Nat Commun ; 11(1): 5074, 2020 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-33033265

RESUMO

Touch and itch sensations are crucial for evoking defensive and emotional responses, and light tactile touch may induce unpleasant itch sensations (mechanical itch or alloknesis). The neural substrate for touch-to-itch conversion in the spinal cord remains elusive. We report that spinal interneurons expressing Tachykinin 2-Cre (Tac2Cre) receive direct Aß low threshold mechanoreceptor (LTMR) input and form monosynaptic connections with GRPR neurons. Ablation or inhibition markedly reduces mechanical but not acute chemical itch nor noxious touch information. Chemogenetic inhibition of Tac2Cre neurons also displays pronounced deficit in chronic dry skin itch, a type of chemical itch in mice. Consistently, ablation of gastrin-releasing peptide receptor (GRPR) neurons, which are essential for transmitting chemical itch, also abolishes mechanical itch. Together, these results suggest that innocuous touch and chemical itch information converge on GRPR neurons and thus map an exquisite spinal circuitry hard-wired for converting innocuous touch to irritating itch.


Assuntos
Rede Nervosa/fisiopatologia , Prurido/fisiopatologia , Tato/fisiologia , Animais , Comportamento Animal , Injeções Espinhais , Luz , Potenciais da Membrana , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Precursores de Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Receptores da Bombesina/metabolismo , Pele/patologia , Medula Espinal/fisiopatologia , Sinapses/metabolismo , Taquicininas/metabolismo
2.
Nat Commun ; 11(1): 4766, 2020 09 21.
Artigo em Inglês | MEDLINE | ID: mdl-32958778

RESUMO

Germline telomere maintenance defects are associated with an increased incidence of inflammatory diseases in humans, yet whether and how telomere dysfunction causes inflammation are not known. Here, we show that telomere dysfunction drives pATM/c-ABL-mediated activation of the YAP1 transcription factor, up-regulating the major pro-inflammatory factor, pro-IL-18. The colonic microbiome stimulates cytosolic receptors activating caspase-1 which cleaves pro-IL-18 into mature IL-18, leading to recruitment of interferon (IFN)-γ-secreting T cells and intestinal inflammation. Correspondingly, patients with germline telomere maintenance defects exhibit DNA damage (γH2AX) signaling together with elevated YAP1 and IL-18 expression. In mice with telomere dysfunction, telomerase reactivation in the intestinal epithelium or pharmacological inhibition of ATM, YAP1, or caspase-1 as well as antibiotic treatment, dramatically reduces IL-18 and intestinal inflammation. Thus, telomere dysfunction-induced activation of the ATM-YAP1-pro-IL-18 pathway in epithelium is a key instigator of tissue inflammation.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Inflamação/patologia , Telômero/patologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Antibacterianos/uso terapêutico , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Caspase 1/metabolismo , Proteínas de Ciclo Celular/antagonistas & inibidores , Proteínas de Ciclo Celular/genética , Criança , Colo/metabolismo , Colo/microbiologia , Colo/patologia , Gastroenteropatias/patologia , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/fisiologia , Humanos , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/microbiologia , Interleucina-18/genética , Interleucina-18/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Camundongos Mutantes , Fosforilação , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Transdução de Sinais , Telomerase/genética , Telomerase/metabolismo
3.
Arch Biochem Biophys ; 689: 108437, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32526201

RESUMO

Skin aging is influenced by several genetic, physiological, and environmental factors. In particular, ultraviolet (UV) exposure is an important factor involved in inducing skin photoaging. Autophagy controlling homeostatic balance between the synthesis, degradation, and recycling of cellular organelles and proteins plays important regulatory roles in several biological processes, including aging. The opioid neuropeptide α-neoendorphin (named NEP) is an endogenous decapeptide (N-YGGFLRKYPK-C) that activates the kappa opioid receptor and exhibits certain anti-aging and anti-wrinkling effects on skin cells; however, its action mechanism has not yet been elucidated. Therefore, the aim of this study was to determine the effects of NEP on anti-skin aging and autophagy activation in human dermal fibroblast cells. Western blot results showed that NEP down-regulates the production of phospho-mammalian target of rapamycin (p-mTOR), whereas increases the expression of key autophagy-related molecules such as Beclin-1, Atg5-Atg12, and LC3-II. The immunocytochemical analysis performed with anti-LC3-II antibody also showed that the autophagic indicators, autophagosomes are formed by NEP. These results suggest that NEP can activate cellular autophagy through mTOR-Beclin-1-mediated signaling pathway. It was also revealed by CM-H2DCF-DA assay and Western blottings that NEP can reduce the production of ultraviolet B (UVB)-induced reactive oxygen species (ROS) like with N-acetylcysteine (NAC), resulting in decreasing the expression levels of skin aging-related proteins, such as phospho-ERK (p-ERK), phospho-p38 (p-p38), and phospho-JNK (p-JNK). Furthermore, NEP could increase the type I procollagen production, while decreasing MMP-1, MMP-2, and MMP-9 activities. Taken together, the results demonstrate that NEP can reduce UVB-induced photoaging by activating autophagy.


Assuntos
Autofagia , Endorfinas/metabolismo , Precursores de Proteínas/metabolismo , Envelhecimento da Pele/efeitos da radiação , Raios Ultravioleta/efeitos adversos , Linhagem Celular , Derme/citologia , Derme/metabolismo , Derme/efeitos da radiação , Fibroblastos/citologia , Fibroblastos/metabolismo , Fibroblastos/efeitos da radiação , Humanos , Pró-Colágeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo
4.
Chem Biol Interact ; 323: 109063, 2020 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-32224134

RESUMO

Exposure to TiO2 NPs induces several cellular alterations after NPs uptake including disruption of cytoskeleton that is crucial for lung physiology but is not considered as a footprint of cell damage. We aimed to investigate cytoskeleton disturbances and the impact on cell migration induced by an acute TiO2 NPs exposure (24 h) and the recovery capability after 6 days of NPs-free treatment, which allowed investigating if cytoskeleton damage was reversible. Exposure to TiO2 NPs (10 µg/cm2) for 24 h induced a decrease 20.2% and 25.1% in tubulin and actin polymerization. Exposure to TiO2 NPs (10 µg/cm2) for 24 h followed by 6 days of NPs-free had a decrease of 26.6% and 21.3% in tubulin and actin polymerization, respectively. The sustained exposure for 7 days to 1 µg/cm2 and 10 µg/cm2 induced a decrease of 22.4% and 30.7% of tubulin polymerization respectively, and 28.7% and 46.2% in actin polymerization. In addition, 24 h followed 6 days of NPs-free exposure of TiO2 NPs (1 µg/cm2 and 10 µg/cm2) decreased cell migration 40.7% and 59.2%, respectively. Cells exposed (10 µg/cm2) for 7 days had a decrease of 65.5% in cell migration. Ki67, protein surfactant B (SFTPB) and matrix metalloprotease 2 (MMP2) were analyzed as genes related to lung epithelial function. The results showed a 20% of Ki67 upregulation in cells exposed for 24 h to 10 µg/cm2 TiO2 NPs while a downregulation of 20% and 25.8% in cells exposed to 1 µg/cm2 and 10 µg/cm2 for 24 h followed by 6 days of NPs-free exposure. Exposure to 1 µg/cm2 and 10 µg/cm2 for 24 h and 7 days upregulates SFTPB expression in 53% and 59% respectively, MMP2 expression remain unchanged. In conclusion, exposure of TiO2 NPs affected cytoskeleton of lung epithelial cells irreversibly but this damage was not cumulative.


Assuntos
Citoesqueleto/patologia , Células Epiteliais/patologia , Pulmão/patologia , Nanopartículas/toxicidade , Titânio/toxicidade , Células A549 , Actinas/metabolismo , Movimento Celular/efeitos dos fármacos , Tamanho Celular , Sobrevivência Celular/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , Endocitose , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/ultraestrutura , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Antígeno Ki-67/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Nanopartículas/ultraestrutura , Polimerização , Precursores de Proteínas/metabolismo , Proteínas Associadas a Surfactantes Pulmonares/metabolismo , Tubulina (Proteína)/metabolismo
5.
Nat Commun ; 11(1): 1995, 2020 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-32332732

RESUMO

Gut microbial dysbiosis is associated with the development of autoimmune disease, but the mechanisms by which microbial dysbiosis affects the transition from asymptomatic autoimmunity to inflammatory disease are incompletely characterized. Here, we identify intestinal barrier integrity as an important checkpoint in translating autoimmunity to inflammation. Zonulin family peptide (zonulin), a potent regulator for intestinal tight junctions, is highly expressed in autoimmune mice and humans and can be used to predict transition from autoimmunity to inflammatory arthritis. Increased serum zonulin levels are accompanied by a leaky intestinal barrier, dysbiosis and inflammation. Restoration of the intestinal barrier in the pre-phase of arthritis using butyrate or a cannabinoid type 1 receptor agonist inhibits the development of arthritis. Moreover, treatment with the zonulin antagonist larazotide acetate, which specifically increases intestinal barrier integrity, effectively reduces arthritis onset. These data identify a preventive approach for the onset of autoimmune disease by specifically targeting impaired intestinal barrier function.


Assuntos
Artrite Reumatoide/prevenção & controle , Permeabilidade da Membrana Celular/efeitos dos fármacos , Disbiose/complicações , Haptoglobinas/antagonistas & inibidores , Mucosa Intestinal/efeitos dos fármacos , Oligopeptídeos/administração & dosagem , Precursores de Proteínas/antagonistas & inibidores , Adulto , Animais , Artrite Experimental/sangue , Artrite Experimental/imunologia , Artrite Experimental/microbiologia , Artrite Experimental/prevenção & controle , Artrite Reumatoide/sangue , Artrite Reumatoide/imunologia , Artrite Reumatoide/microbiologia , Translocação Bacteriana/efeitos dos fármacos , Translocação Bacteriana/imunologia , Células CACO-2 , Permeabilidade da Membrana Celular/imunologia , Estudos de Coortes , Estudos Transversais , Disbiose/imunologia , Disbiose/microbiologia , Feminino , Microbioma Gastrointestinal/imunologia , Haptoglobinas/metabolismo , Voluntários Saudáveis , Humanos , Íleo/citologia , Íleo/efeitos dos fármacos , Íleo/microbiologia , Íleo/patologia , Mucosa Intestinal/citologia , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Precursores de Proteínas/sangue , Precursores de Proteínas/metabolismo , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo
6.
Nat Commun ; 11(1): 1662, 2020 04 03.
Artigo em Inglês | MEDLINE | ID: mdl-32245955

RESUMO

The proteasome is an essential protein-degradation machinery in eukaryotic cells that controls protein turnover and thereby the biogenesis and function of cell organelles. Chloroplasts import thousands of nuclear-encoded precursor proteins from the cytosol, suggesting that the bulk of plastid proteins is transiently exposed to the cytosolic proteasome complex. Therefore, there is a cytosolic equilibrium between chloroplast precursor protein import and proteasomal degradation. We show here that a shift in this equilibrium, induced by mild genetic proteasome impairment, results in elevated precursor protein abundance in the cytosol and significantly increased accumulation of functional photosynthetic complexes in protein import-deficient chloroplasts. Importantly, a proteasome lid mutant shows improved photosynthetic performance, even in the absence of an import defect, signifying that functional precursors are continuously degraded. Hence, turnover of plastid precursors in the cytosol represents a mechanism to constrain thylakoid membrane assembly and photosynthetic electron transport.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiologia , Cloroplastos/metabolismo , Complexo de Proteínas do Centro de Reação Fotossintética/metabolismo , Complexo de Endopeptidases do Proteassoma/genética , Proteínas de Cloroplastos/metabolismo , Citosol/metabolismo , Mutação , Fotossíntese , Plantas Geneticamente Modificadas , Complexo de Endopeptidases do Proteassoma/metabolismo , Precursores de Proteínas/metabolismo , Proteólise , Estresse Fisiológico
7.
Neuron ; 106(6): 1009-1025.e10, 2020 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-32302532

RESUMO

Calorie-rich diets induce hyperphagia and promote obesity, although the underlying mechanisms remain poorly defined. We find that short-term high-fat-diet (HFD) feeding of mice activates prepronociceptin (PNOC)-expressing neurons in the arcuate nucleus of the hypothalamus (ARC). PNOCARC neurons represent a previously unrecognized GABAergic population of ARC neurons distinct from well-defined feeding regulatory AgRP or POMC neurons. PNOCARC neurons arborize densely in the ARC and provide inhibitory synaptic input to nearby anorexigenic POMC neurons. Optogenetic activation of PNOCARC neurons in the ARC and their projections to the bed nucleus of the stria terminalis promotes feeding. Selective ablation of these cells promotes the activation of POMC neurons upon HFD exposure, reduces feeding, and protects from obesity, but it does not affect food intake or body weight under normal chow consumption. We characterize PNOCARC neurons as a novel ARC neuron population activated upon palatable food consumption to promote hyperphagia.


Assuntos
Núcleo Arqueado do Hipotálamo/fisiologia , Dieta Hiperlipídica , Comportamento Alimentar/fisiologia , Neurônios GABAérgicos/fisiologia , Hiperfagia , Obesidade , Ganho de Peso/fisiologia , Animais , Núcleo Arqueado do Hipotálamo/citologia , Núcleo Arqueado do Hipotálamo/metabolismo , Neurônios GABAérgicos/metabolismo , Camundongos , Inibição Neural/fisiologia , Neurônios/metabolismo , Neurônios/fisiologia , Optogenética , Pró-Opiomelanocortina/metabolismo , Precursores de Proteínas/metabolismo , Receptores Opioides/metabolismo , Núcleos Septais/fisiologia
8.
Nature ; 580(7803): 376-380, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32296182

RESUMO

Mechanosensory feedback from the digestive tract to the brain is critical for limiting excessive food and water intake, but the underlying gut-brain communication pathways and mechanisms remain poorly understood1-12. Here we show that, in mice, neurons in the parabrachial nucleus that express the prodynorphin gene (hereafter, PBPdyn neurons) monitor the intake of both fluids and solids, using mechanosensory signals that arise from the upper digestive tract. Most individual PBPdyn neurons are activated by ingestion as well as the stimulation of the mouth and stomach, which indicates the representation of integrated sensory signals across distinct parts of the digestive tract. PBPdyn neurons are anatomically connected to the digestive periphery via cranial and spinal pathways; we show that, among these pathways, the vagus nerve conveys stomach-distension signals to PBPdyn neurons. Upon receipt of these signals, these neurons produce aversive and sustained appetite-suppressing signals, which discourages the initiation of feeding and drinking (fully recapitulating the symptoms of gastric distension) in part via signalling to the paraventricular hypothalamus. By contrast, inhibiting the same population of PBPdyn neurons induces overconsumption only if a drive for ingestion exists, which confirms that these neurons mediate negative feedback signalling. Our findings reveal a neural mechanism that underlies the mechanosensory monitoring of ingestion and negative feedback control of intake behaviours upon distension of the digestive tract.


Assuntos
Ingestão de Alimentos , Retroalimentação , Neurônios/fisiologia , Animais , Encefalinas/genética , Encefalinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Trato Gastrointestinal Superior/fisiologia
9.
J Dairy Sci ; 103(5): 3961-3970, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32171508

RESUMO

Increased intestinal permeability has been shown to be involved in several diseases associated with low-grade chronic inflammation, including obesity and metabolic syndrome. In the last decade, growing evidence shows the beneficial effects of probiotic-containing food supplementation on these conditions. In this crossover intervention study on 28 asymptomatic overweight adults, we tested the effects of a 3-wk kefir supplementation compared with a 3-wk milk supplementation on serum zonulin levels. The effects on serum glucose, triacylglycerols, low-density lipoproteins, high-density lipoproteins, total cholesterol, markers of inflammation (C-reactive protein and adiponectin), anthropometric variables, mood, and appetite were also determined. Kefir supplementation resulted in a greater improvement of serum zonulin levels (F = 6.812, η2 = 0.275), whereas a significant yet similar improvement in lipid profile and serum glucose levels was found in both supplementations. Positive mood was slightly but significantly enhanced with kefir supplementation, and reduced with milk supplementation. The C-reactive protein, adiponectin, and appetite were unaffected. In conclusion, supplementation with both dairy products had health beneficial effects, but only kefir showed an effect on the intestinal barrier dysfunction marker.


Assuntos
Haptoglobinas/metabolismo , Kefir , Leite , Sobrepeso/metabolismo , Precursores de Proteínas/metabolismo , Adulto , Animais , Biomarcadores/sangue , Estudos Cross-Over , Feminino , Humanos , Absorção Intestinal , Mucosa Intestinal/metabolismo , Masculino , Pessoa de Meia-Idade , Leite/metabolismo , Sobrepeso/sangue , Triglicerídeos/metabolismo
10.
J Virol ; 94(11)2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32213612

RESUMO

The structural protein Gag is the only viral component required for retroviral budding from infected cells. Each of the three conserved domains-the matrix (MA), capsid (CA), and nucleocapsid (NC) domains-drives different phases of viral particle assembly and egress. Once virus assembly is complete, retroviruses, like most enveloped viruses, utilize host proteins to catalyze membrane fission and to free progeny virions. These proteins are members of the endosomal sorting complex required for transport (ESCRT), a cellular machinery that coats the inside of budding necks to perform membrane-modeling events necessary for particle abscission. The ESCRT is recruited through interactions with PTAP and LYPXnL, two highly conserved sequences named late (L) domains, which bind TSG101 and Alix, respectively. A TSG101-binding L-domain was identified in the p2 region of the feline immunodeficiency virus (FIV) Gag protein. Here, we show that the human protein Alix stimulates the release of virus from FIV-expressing human cells. Furthermore, we demonstrate that the Alix Bro1 domain rescues FIV mutants lacking a functional TSG101-interacting motif, independently of the entire p2 region and of the canonical Alix-binding L-domain(s) in FIV Gag. However, in contrast to the effect on human immunodeficiency virus type 1 (HIV-1), the C377,409S double mutation, which disrupts both CCHC zinc fingers in the NC domain, does not abrogate Alix-mediated virus rescue. These studies provide insight into conserved and divergent mechanisms of lentivirus-host interactions involved in virus budding.IMPORTANCE FIV is a nonprimate lentivirus that infects domestic cats and causes a syndrome that is reminiscent of AIDS in humans. Based on its similarity to HIV with regard to different molecular and biochemical properties, FIV represents an attractive model for the development of strategies to prevent and/or treat HIV infection. Here, we show that the Bro1 domain of the human cellular protein Alix is sufficient to rescue the budding of FIV mutants devoid of canonical L-domains. Furthermore, we demonstrate that the integrity of the CCHC motifs in the Gag NC domain is dispensable for Alix-mediated rescue of virus budding, suggesting the involvement of other regions of the Gag viral protein. Our research is pertinent to the identification of a conserved yet mechanistically divergent ESCRT-mediated lentivirus budding process in general, and to the role of Alix in particular, which underlies the complex viral-cellular network of interactions that promote late steps of the retroviral life cycle.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Ciclo Celular/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Produtos do Gene gag/metabolismo , HIV-1/fisiologia , Vírus da Imunodeficiência Felina/fisiologia , Precursores de Proteínas/metabolismo , Liberação de Vírus , Animais , Proteínas de Ligação ao Cálcio/genética , Gatos , Proteínas de Ciclo Celular/genética , Complexos Endossomais de Distribuição Requeridos para Transporte/genética , Produtos do Gene gag/genética , Células HEK293 , Humanos , Mutação de Sentido Incorreto , Domínios Proteicos , Precursores de Proteínas/genética , Dedos de Zinco
11.
Biochem Soc Trans ; 48(1): 71-82, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-31922184

RESUMO

Chloroplasts are photosynthetic plant organelles descended from a bacterial ancestor. The vast majority of chloroplast proteins are synthesized in the cytosol and then imported into the chloroplast post-translationally. Translocation complexes exist in the organelle's outer and inner envelope membranes (termed TOC and TIC, respectively) to facilitate protein import. These systems recognize chloroplast precursor proteins and mediate their import in an energy-dependent manner. However, many unanswered questions remain regarding mechanistic details of the import process and the participation and functions of individual components; for example, the cytosolic events that mediate protein delivery to chloroplasts, the composition of the TIC apparatus, and the nature of the protein import motor all require resolution. The flux of proteins through TOC and TIC varies greatly throughout development and in response to specific environmental cues. The import process is, therefore, tightly regulated, and it has emerged that the ubiquitin-proteasome system (UPS) plays a key role in this regard, acting at several different steps in the process. The UPS is involved in: the selective degradation of transcription factors that co-ordinate the expression of chloroplast precursor proteins; the removal of unimported chloroplast precursor proteins in the cytosol; the inhibition of chloroplast biogenesis pre-germination; and the reconfiguration of the TOC apparatus in response to developmental and environmental signals in a process termed chloroplast-associated protein degradation. In this review, we highlight recent advances in our understanding of protein import into chloroplasts and how this process is regulated by the UPS.


Assuntos
Cloroplastos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Transporte Proteico/fisiologia , Ubiquitina/metabolismo , Proteínas de Cloroplastos/metabolismo , Citosol/metabolismo , Organelas/metabolismo , Fotossíntese , Proteínas de Plantas/metabolismo , Plantas/anatomia & histologia , Plantas/metabolismo , Precursores de Proteínas/metabolismo , Proteólise
12.
FASEB J ; 34(2): 2541-2553, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31908023

RESUMO

Brain-derived neurotrophic factor precursor (proBDNF) has been reported to strengthen the dysfunction of monocytes/macrophages in animal studies. However, it is still unknown the roles of proBDNF in the dysfunction of monocytes in the inflammatory diseases in humans. In the present study, we showed that proBDNF and pan neurotrophic receptor p75 were significantly upregulated in monocytes from healthy donors (HD) after lipopolysaccharide treatment. Exogenous proBDNF treatment upregulated CD40 and proinflammatory cytokines expression in monocytes including interleukin (IL)-1ß, IL-6, and tumor necrosis factor (TNF)-α. In Stanford type-A acute aortic dissection (AAD) patients, proBDNF was upregulated in CD14+ CD163+ CX3CR1+ M2- but not CD14+ CD68+ CCR2+ M1-like monocytes. In addition, sera from AAD patients activated gene expression of proinflammatory cytokines in cultured PBMCs from HD, which was attenuated by proBDNF monoclonal antibody (Ab-proB) treatment. These findings suggested that upregulation of proBDNF in M2-like monocytes may contribute to the proinflammatory response in the AAD.


Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Citocinas/metabolismo , Macrófagos/metabolismo , Monócitos/metabolismo , Precursores de Proteínas/metabolismo , Adulto , Aneurisma Dissecante/metabolismo , Células Cultivadas , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima
13.
Diabetes ; 69(4): 670-680, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31896552

RESUMO

The signal peptide of preproinsulin is a major source for HLA class I autoantigen epitopes implicated in CD8 T cell (CTL)-mediated ß-cell destruction in type 1 diabetes (T1D). Among them, the 10-mer epitope located at the C-terminal end of the signal peptide was found to be the most prevalent in patients with recent-onset T1D. While the combined action of signal peptide peptidase and endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) is required for processing of the signal peptide, the mechanisms controlling signal peptide trimming and the contribution of the T1D inflammatory milieu on these mechanisms are unknown. Here, we show in human ß-cells that ER stress regulates ERAP1 gene expression at posttranscriptional level via the IRE1α/miR-17-5p axis and demonstrate that inhibition of the IRE1α activity impairs processing of preproinsulin signal peptide antigen and its recognition by specific autoreactive CTLs during inflammation. These results underscore the impact of ER stress in the increased visibility of ß-cells to the immune system and position the IRE1α/miR-17 pathway as a central component in ß-cell destruction processes and as a potential target for the treatment of autoimmune T1D.


Assuntos
Aminopeptidases/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Estresse do Retículo Endoplasmático/fisiologia , Endorribonucleases/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Antígenos de Histocompatibilidade Menor/metabolismo , Precursores de Proteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Aminopeptidases/genética , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Regulação para Baixo/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Endorribonucleases/genética , Células HEK293 , Humanos , Inflamação/genética , Inflamação/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/imunologia , Interferon gama/farmacologia , Interleucina-1beta/farmacologia , MicroRNAs/genética , MicroRNAs/metabolismo , Antígenos de Histocompatibilidade Menor/genética , Proteínas Serina-Treonina Quinases/genética , Regulação para Cima
14.
Int J Mol Sci ; 21(2)2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31952213

RESUMO

Mutations in the hepatitis B virus (HBV) genome can potentially lead to vaccination failure, diagnostic escape, and disease progression. However, there are no reports on viral gene expression and large hepatitis B surface antigen (HBsAg) antigenicity alterations due to mutations in HBV isolated from a Bangladeshi population. Here, we sequenced the full genome of the HBV isolated from a clinically infected patient in Bangladesh. The open reading frames (ORFs) (P, S, C, and X) of the isolated HBV strain were successfully amplified and cloned into a mammalian expression vector. The HBV isolate was identified as genotype C (sub-genotype C2), serotype adr, and evolutionarily related to strains isolated in Indonesia, Malaysia, and China. Clinically significant mutations, such as preS1 C2964A, reverse transcriptase domain I91L, and small HBsAg N3S, were identified. The viral P, S, C, and X genes were expressed in HEK-293T and HepG2 cells by transient transfection with a native subcellular distribution pattern analyzed by immunofluorescence assay. Western blotting of large HBsAg using preS1 antibody showed no staining, and preS1 ELISA showed a significant reduction in reactivity due to amino acid mutations. This mutated preS1 sequence has been identified in several Asian countries. To our knowledge, this is the first report investigating changes in large HBsAg antigenicity due to preS1 mutations.


Assuntos
Antígenos de Superfície da Hepatite B/imunologia , Vírus da Hepatite B/imunologia , Hepatite B/imunologia , Mutação , Precursores de Proteínas/imunologia , Bangladesh , Sequência de Bases , Genoma Viral/genética , Genótipo , Células HEK293 , Células Hep G2 , Hepatite B/virologia , Antígenos de Superfície da Hepatite B/genética , Antígenos de Superfície da Hepatite B/metabolismo , Vírus da Hepatite B/classificação , Vírus da Hepatite B/genética , Humanos , Masculino , Fases de Leitura Aberta/genética , Filogenia , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Homologia de Sequência do Ácido Nucleico , Sequenciamento Completo do Genoma , Adulto Jovem
15.
Anesth Analg ; 130(1): 248-257, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31166231

RESUMO

BACKGROUND: Persistent use of prescription opioids beyond the period of surgical recovery is a large part of a public health problem linked to the current opioid crisis in the United States. However, few studies have been conducted to examine whether morphine reward is influenced by acute pain and injury. METHODS: In a mouse model of incisional injury and minor trauma, animals underwent conditioning, extinction, and drug-primed reinstatement with morphine to examine the rewarding properties of morphine in the presence of acute incisional injury and drug-induced relapse, respectively. In addition, we sought to determine whether these behaviors were influenced by kappa opioid receptor signaling and measured expression of prodynorphin messenger RNA in the nucleus accumbens and medial prefrontal cortex after conditioning and before reinstatement with morphine and incisional injury. RESULTS: In the presence of incisional injury, we observed enhancement of morphine reward with morphine-conditioned place preference but attenuated morphine-primed reinstatement to reward. This adaptation was not present in animals conditioned 12 days after incisional injury when nociceptive sensitization had resolved; however, they showed enhancement of morphine-primed reinstatement. Prodynorphin expression was greatly enhanced in the nucleus accumbens and medial prefrontal cortex of mice with incisional injury and morphine conditioning and remained elevated up to drug-primed reinstatement. These changes were not observed in mice conditioned 12 days after incisional injury. Further, kappa opioid receptor blockade with norbinaltorphimine before reinstatement reversed the attenuation induced by injury. CONCLUSIONS: These findings suggest enhancement of morphine reward as a result of incisional injury but paradoxically a protective adaptation with incisional injury from drug-induced relapse resulting from kappa opioid receptor activation in the reward circuitry. Remote injury conferred no such protection and appeared to enhance reinstatement.


Assuntos
Dor Aguda/tratamento farmacológico , Comportamento Animal/efeitos dos fármacos , Morfina/farmacologia , Antagonistas de Entorpecentes/farmacologia , Receptores Opioides kappa/agonistas , Recompensa , Ferimentos Penetrantes/tratamento farmacológico , Dor Aguda/metabolismo , Dor Aguda/fisiopatologia , Dor Aguda/psicologia , Animais , Condicionamento Psicológico/efeitos dos fármacos , Modelos Animais de Doenças , Encefalinas/genética , Encefalinas/metabolismo , Extinção Psicológica/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Limiar da Dor/efeitos dos fármacos , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Receptores Opioides kappa/metabolismo , Transdução de Sinais , Ferimentos Penetrantes/metabolismo , Ferimentos Penetrantes/fisiopatologia , Ferimentos Penetrantes/psicologia
16.
Neurobiol Aging ; 87: 49-59, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31882186

RESUMO

Disruption of brain-derived neurotrophic factor (BDNF) biosynthesis and/or signaling has been implicated in the pathogenesis of Alzheimer's disease (AD). We used postmortem brain and fluid samples from 20 patients with variable severity of AD and 11 controls to investigate whether BDNF levels in serum and brain tissue correlated with hippocampal pathology. Total BDNF, precursor BDNF (pro-BDNF), and mature BDNF were measured in cerebrospinal fluid, serum, and 3 postmortem brain regions. Histological markers for AD pathology, the BDNF cognate receptor (TrkB), and glia were measured in the hippocampus (HIP). Lower pro-BDNF levels were observed in the entorhinal and frontal cortices in AD cases compared with controls. AD cases also exhibited significantly lower staining densities of the cognate BDNF receptor TrkB in the HIP compared with controls, and TrkB staining was inversely correlated with both Amylo-Glo and pTau staining in the same region, suggesting a relationship between the density of the cognate BDNF receptor and accumulation of AD pathology. In addition, higher serum pro-BDNF levels correlated with lower HIP pro-BDNF levels and higher pTau staining in the HIP. Total BDNF levels in cortical regions were also negatively correlated with Amylo-Glo staining in the HIP suggesting that reduced BDNF cortical levels might influence hippocampal amyloid accumulation. These results strongly suggest that altered BDNF and TrkB receptors are involved in AD pathology and therefore warrant investigations into therapies involving the BDNF pathway.


Assuntos
Doença de Alzheimer/etiologia , Doença de Alzheimer/metabolismo , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Hipocampo/metabolismo , Precursores de Proteínas/metabolismo , Coloração e Rotulagem/métodos , Proteínas tau/metabolismo , Idoso , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Fator Neurotrófico Derivado do Encéfalo/sangue , Feminino , Hipocampo/patologia , Humanos , Masculino , Glicoproteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Fosforilação , Precursores de Proteínas/sangue , Receptor trkB/metabolismo
17.
Sci Adv ; 5(12): eaax0292, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31840061

RESUMO

The mechanistic basis for the biogenesis of peptide hormones and growth factors is poorly understood. Here, we show that the conserved endoplasmic reticulum membrane translocon-associated protein α (TRAPα), also known as signal sequence receptor 1, plays a critical role in the biosynthesis of insulin. Genetic analysis in the nematode Caenorhabditis elegans and biochemical studies in pancreatic ß cells reveal that TRAPα deletion impairs preproinsulin translocation while unexpectedly disrupting distal steps in insulin biogenesis including proinsulin processing and secretion. The association of common intronic single-nucleotide variants in the human TRAPα gene with susceptibility to type 2 diabetes and pancreatic ß cell dysfunction suggests that impairment of preproinsulin translocation and proinsulin trafficking may contribute to the pathogenesis of type 2 diabetes.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Insulina/biossíntese , Glicoproteínas de Membrana/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Receptores de Peptídeos/metabolismo , Animais , Estresse do Retículo Endoplasmático , Insulina/metabolismo , Secreção de Insulina , Precursores de Proteínas/metabolismo , Proteínas Recombinantes de Fusão/metabolismo
18.
EBioMedicine ; 50: 224-237, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31757777

RESUMO

BACKGROUND: Alcohol intake is a well-known lifestyle risk factor for CRC, and an increasing number of studies have revealed that alcohol intake is also tightly associated with CRC metastasis. However, the effect of alcohol on CRC metastasis and its underlying mechanism remain unclear. METHODS: A retrospective cohort study was performed to investigate the characteristics of patients with alcohol-related CRC. The effects of ethanol on the biological behaviours of CRC cells were assessed through in vivo and in vitro assays using the Lieber-DeCarli ethanol liquid diet and ethanol, respectively. The ethanol-mediated signalling pathway and downstream factors were screened through ELISA, western blot, immunofluorescence and co-immunoprecipitation. FINDINGS: Most patients with alcohol-related CRC, particularly those with tumour metastasis, were characterized by a notably higher circulating ethanol level and a lower systemic acetaldehyde level. Moreover, CRC cells accumulated in ethanol, but not acetaldehyde, to notably higher levels compared with adjacent normal cells. Alcohol intake significantly promoted CRC metastasis via the ethanol-mediated TGF-ß/Smad/Snail axis, and ethanol induced the cytoplasmic mislocalization of RUNX3 and further promoted the aggressiveness of CRC by targeting Snail. Pirfenidone (PFD) significantly eliminated the effects of ethanol on CRC metastasis by specifically blocking TGF-ß signalling. INTERPRETATION: Alcohol intake plays a vital role in CRC metastasis via the ethanol-mediated TGF-ß/RUNX3/Snail axis, and PFD might be a novel therapeutic management strategy for CRC.


Assuntos
Consumo de Bebidas Alcoólicas/efeitos adversos , Neoplasias Colorretais/etiologia , Neoplasias Colorretais/metabolismo , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Transdução de Sinais , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Biomarcadores , Linhagem Celular Tumoral , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Humanos , Incidência , Camundongos , Metástase Neoplásica , Neurofisinas/metabolismo , Ligação Proteica , Precursores de Proteínas/metabolismo , Transporte Proteico , Fator de Crescimento Transformador beta1/metabolismo , Vasopressinas/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
19.
Yakugaku Zasshi ; 139(11): 1403-1415, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31685737

RESUMO

For my Ph.D. research topic, I isolated endogenous morphine-like analgesic dipeptide, kyotorphin, which mediates Met-enkephalin release, and discovered kyotorphin synthetase, a putative receptor and antagonist. Furthermore, I succeeded in purifying µ-opioid receptor and functional reconstitution with purified G proteins. After receiving my full professor position at Nagasaki University in 1996, I worked on two topics of research, molecular mechanisms of chronic pain through lysophosphatidic acid (LPA) and identification and characterization of neuroprotective protein, prothymosin α. In a series of studies, we have shown that LPA signaling defines the molecular mechanisms of neuropathic pain and fibromyalgia in terms of development and maintenance. Above all, the discovery of feed-forward system in LPA production and pain memory may contribute to better understanding of chronic pain and future analgesic drug discovery. Regarding prothymosin α, we first discovered it as neuronal necrosis-inhibitory molecule through two independent mechanisms, such as toll-like receptor and F0/F1 ATPase, both which protect neurons through indirect mechanisms. Prothymosin α is released by non-classical and non-vesicular mechanisms on various stresses, such as ischemia, starvation, and heat-shock. Thus it may be called a new type of neuroprotective damage-associated molecular patterns (DAMPs)/Alarmins. Heterozygotic mice showed a defect in memory-learning and neurogenesis as well as anxiogenic behaviors. Small peptide, P6Q derived from prothymosin α retains neuroprotective actions, which include blockade of cerebral hemorrhage caused by late treatment with tissue plasminogen activator in the stroke model in mice.


Assuntos
Dor Crônica/etiologia , Dor Crônica/genética , Fármacos Neuroprotetores , Precursores de Proteínas , Receptores de Ácidos Lisofosfatídicos/fisiologia , Transdução de Sinais/fisiologia , Timosina/análogos & derivados , Animais , Endorfinas , Humanos , Camundongos , Precursores de Proteínas/metabolismo , ATPases Translocadoras de Prótons , Receptores de Ácidos Lisofosfatídicos/metabolismo , Receptores Opioides , Acidente Vascular Cerebral , Timosina/metabolismo , Receptores Toll-Like
20.
Int J Mol Sci ; 20(23)2019 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-31775361

RESUMO

Metastases in thyroid cancer are associated with aggressive disease and increased patient morbidity, but the factors driving metastatic progression are unclear. The precursor for nerve growth factor (proNGF) is increased in primary thyroid cancers, but its expression or significance in metastases is not known. In this study, we analysed the expression of proNGF in a retrospective cohort of thyroid cancer lymph node metastases (n = 56), linked with corresponding primary tumours, by automated immunohistochemistry and digital quantification. Potential associations of proNGF immunostaining with clinical and pathological parameters were investigated. ProNGF staining intensity (defined by the median h-score) was significantly higher in lymph node metastases (h-score 94, interquartile range (IQR) 50-147) than in corresponding primary tumours (57, IQR 42-84) (p = 0.002). There was a correlation between proNGF expression in primary tumours and corresponding metastases, where there was a 0.68 (95% CI 0 to 1.2) increase in metastatic tumour h-score for each unit increase in the primary tumour h-score. However, larger tumours (both primary and metastatic) had lower proNGF expression. In a multivariate model, proNGF expression in nodal metastases was negatively correlated with lateral neck disease and being male. In conclusion, ProNGF is expressed in locoregional metastases of thyroid cancer and is higher in lymph node metastases than in primary tumours, but is not associated with high-risk clinical features.


Assuntos
Adenocarcinoma Folicular/secundário , Biomarcadores Tumorais/metabolismo , Carcinoma Papilar/secundário , Fator de Crescimento Neural/metabolismo , Precursores de Proteínas/metabolismo , Neoplasias da Glândula Tireoide/patologia , Adenocarcinoma Folicular/metabolismo , Adulto , Idoso , Carcinoma Papilar/metabolismo , Feminino , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Estudos Retrospectivos , Neoplasias da Glândula Tireoide/metabolismo
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