RESUMEN
Idiopathic inflammatory myopathy (IIM) is an autoimmune disease that invades skeletal muscle; however, the etiology of IIM is still poorly understood. Toll-like receptor (TLR) 4 has been widely reported to take part in the autoimmune inflammation of IIMs. The mammalian target of rapamycin, mTOR, is a key central substance which mediates immune responses and metabolic changes, and also has been confirmed to be involved in the pathogenesis of IIMs. However, the interconnectedness between TLR4 and mTOR in IIM inflammation has not been fully elucidated. We hypothesized that TLR4 may play an important role in IIM inflammatory muscle injury by regulating mTOR. Mice were divided into four groups: a normal control group, IIM animal model (experimental autoimmune myositis, EAM) group, TAK242 intervention group and rapamycin (RAPA) intervention group. The results of EAM mice showed that TLR4, mTOR, nuclear factor-kappa B (NF-κB) and inflammatory factors interleukin-17A (IL-17A) and interferon γ (IFN-γ) mRNA levels were significantly upregulated. These factors were positively correlated with the degree of muscle inflammatory injury. When EAM mice were given the antagonist TAK242 to inhibit the TLR4 pathway, the results demonstrated that both mTOR and NF-κB were downregulated in the muscle of the mice. Muscle staining showed that the inflammatory injury was alleviated and the EAM mouse muscle strength was improved. Then, RAPA was used to inhibit the mTOR pathway, and the inflammatory factors IL-17A and IFN-γ were downregulated in EAM mouse muscle and serum. Consistently, muscle inflammatory injury was significantly reduced, and muscle strength was significantly improved. Our results suggest that TLR4 may regulate inflammatory muscle injury in EAM by activating the mTOR and NF-κB pathways, which provides both an experimental complement for the pathological mechanism of IIM and an encouraging target for the selection of effective treatments.
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Miositis , Enfermedad Autoinmune Experimental del Sistema Nervioso , Serina-Treonina Quinasas TOR , Receptor Toll-Like 4 , Animales , Interferón gamma/metabolismo , Interleucina-17/metabolismo , Mamíferos/metabolismo , Ratones , Músculo Esquelético/metabolismo , Miositis/metabolismo , FN-kappa B/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Receptor Toll-Like 4/metabolismoRESUMEN
BACKGROUND AND OBJECTIVES: To investigate the prevalence of vitamin A and vitamin D deficiency and the associated factors in hospitalized neonates in Xi'an, China. METHODS AND STUDY DESIGN: A total of 524 hospitalized neonates were collected in this study. Serum vitamin A and D concentrations were detected in neonates within two weeks of birth. RESULTS: Serum vitamin A and D concentrations of hospitalized neonates were 0.55±0.21 µmol/L and 42.0±20.6 nmol/L, respectively. They were greater in full-term neonates than in preterm neonates, greater in rural neonates than in urban, and greater in single than in twin (all p<0.001). The prevalence of vitamin A and D deficiency were 14.9% and 33.0%, the prevalence of marginal vitamin A deficiency was 64.7%, and vitamin D insufficiency was 35.1%. Neonatal serum vitamin A and D concentrations were all positively correlated with birth weight and gestational age. Neonatal serum vitamin D concentration was also positively correlated with maternal serum vitamin D concentration. Additionally, neonatal vitamin A concentration was positively correlated with neonatal serum vitamin D concentration. CONCLUSIONS: Vitamin A and vitamin D statuses are compromised in hospitalized neonates in Xi'an, especially in premature neonates, low birth weight neonates, twins, and those born in urban areas. Individualized supplementation with vitamin A and vitamin D in neonates should be a clinical consideration.
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Deficiencia de Vitamina A , Deficiencia de Vitamina D , Femenino , Humanos , Recién Nacido , Prevalencia , Vitamina A , Deficiencia de Vitamina A/epidemiología , Vitamina D , Deficiencia de Vitamina D/epidemiología , VitaminasRESUMEN
OBJECTIVES: To investigate the levels of fat-soluble vitamins A, D, and E in children with obesity and their influencing factors. METHODS: A total of 273 children with obesity who attended the Department of Clinical Nutrition, Xi'an Children's Hospital, from January 2019 to April 2021 were enrolled as the obesity group. A total of 226 children with normal body weight who underwent physical examination during the same period were enrolled as the control group. Anthropometric parameters and body composition were measured for both groups, and the serum concentrations of vitamins A, D, and E were also measured. RESULTS: Compared with the control group, the obesity group had significantly higher serum levels of vitamin A [(1.32±0.21) µmol/L vs (1.16±0.21) µmol/L, P<0.001] and vitamin E [(9.3±1.4) mg/L vs (8.3±1.2) mg/L, P<0.001] and a significant reduction in the level of 25-hydroxyvitamin D [(49±22) nmol/L vs (62±24) nmol/L, P<0.001]. In the obesity group, the prevalence rates of marginal vitamin A deficiency, vitamin D deficiency/insufficiency, and vitamin E insufficiency were 5.5% (15/273), 56.8% (155/273), and 4.0% (11/273), respectively. After adjustment for body mass index Z-score and waist-to-height ratio, serum vitamin A level was positively correlated with age (P<0.001), while vitamins E and 25-hydroxyvitamin D levels were negatively correlated with age in children with obesity (P<0.001). After adjustment for age, the serum levels of vitamin A, vitamin E and 25-hydroxyvitamin D were not correlated with degree of obesity, percentage of body fat, and duration of obesity in children with obesity, while the serum levels of vitamins A and E were positively correlated with waist-to-height ratio (P<0.001). CONCLUSIONS: There are higher serum levels of vitamins A and E in children with obesity, especially in those with abdominal obesity, while serum vitamin D nutritional status is poor and worsens with age. Therefore, vitamin D nutritional status should be taken seriously for children with obesity, and vitamin D supplementation should be performed when necessary.
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Obesidad Infantil , Calcifediol , Niño , Humanos , Vitamina A , Vitamina D , Vitamina E , VitaminasRESUMEN
BACKGROUND AND OBJECTIVES: To investigate the vitamin A, D, E concentrations and status, and analyse the associated risk factors for vitamin A, D, E deficiency in children in Shaanxi Province, Northwest China. METHODS AND STUDY DESIGN: The study included a total of 25,806 children admitted to hospitals in Shaanxi province from January 2019 to December 2019. Fasting venous blood samples were collected to measure serum vitamin A, D, E concentrations. A logistic regression model was performed to estimate the association between risk factors and vitamin A, D, E deficiency and insufficiency. RESULTS: The mean serum vitamin A, D, E concentrations were 0.87±0.33 ïmol/L, 63.7±29.7 nmol/L and 20.8±6.98 ïmol/L, respectively. The prevalence of vitamin A, D, E deficiency was high in neonates (15.1%, 81.5% and 44.9%, respectively). Children living in rural areas were at higher risk for vitamin A, D, E deficiency and insufficiency. Logistic regression analyses revealed that the risk of vitamin D deficiency and insufficiency in children was 7.68 times (95% CI: 6.97-8.47) higher in winter than in summer. With adjustment for gender, season, and living regions, age correlated positively with serum vitamin A concentration (r=0.110, p<0.001), and negatively with vitamin D and E concentrations (r=-0.370 and r=-0.250 both p<0.001). CONCLUSIONS: This study showed that the prevalence of vitamin A, D and E deficiency was extremely high in neonates in Shaanxi province, northwest China. Children living in rural areas and winter had a high risk of vitamin A, D, E deficiency and insufficiency.
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Vitamina A , Deficiencia de Vitamina D , Niño , China/epidemiología , Estudios Transversales , Humanos , Recién Nacido , Prevalencia , Factores de Riesgo , Estaciones del Año , Vitamina D , Deficiencia de Vitamina D/epidemiologíaRESUMEN
Bacteria degrade nicotine mainly using pyridine and pyrrolidine pathways. Previously, we discovered a hybrid of the pyridine and pyrrolidine pathways (the VPP pathway) in Pseudomonas geniculata N1 and characterized its key enzyme, 6-hydroxypseudooxynicotine amine oxidase (HisD). It catalyzes oxidative deamination of 6-hydroxypseudooxynicotine to 6-hydroxy-3-succinoylsemialdehyde-pyridine, which is the crucial step connecting upstream and downstream portions of the VPP pathway. We determined the crystal structure of wild-type HisD to 2.6 Å. HisD is a monomer that contains a flavin mononucleotide, an iron-sulfur cluster, and ADP. On the basis of sequence alignment and structure comparison, a difference has been found among HisD, closely related trimethylamine dehydrogenase (TMADH), and histamine dehydrogenase (HADH). The flavin mononucleotide (FMN) cofactor is not covalently bound to any residue, and the FMN isoalloxazine ring is planar in HisD compared to TMADH or HADH, which forms a 6-S-cysteinyl flavin mononucleotide cofactor and has an FMN isoalloxazine ring in a "butterfly bend" conformation. Based on the structure, docking study, and site-directed mutagenesis, the residues Glu60, Tyr170, Asp262, and Trp263 may be involved in substrate binding. The expanded understanding of the substrate binding mode from this study may guide rational engineering of such enzymes for biodegradation of potential pollutants or for bioconversion to generate desired products.IMPORTANCE Nicotine is a major tobacco alkaloid in tobacco waste. Pyridine and pyrrolidine pathways are the two best-elucidated nicotine metabolic pathways; Pseudomonas geniculata N1 catabolizes nicotine via a hybrid between the pyridine and pyrrolidine pathways. The crucial enzyme, 6-hydroxypseudooxynicotine amine oxidase (HisD), links the upstream and downstream portions of the VPP pathway; however, there is little structural information about this important enzyme. In this study, we determined the crystal structure of HisD from Pseudomonas geniculata N1. Its basic insights about the structure may help us to guide the engineering of such enzymes for bioremediation and bioconversion applications.
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Proteínas Bacterianas/química , Redes y Vías Metabólicas , Nicotina/metabolismo , Pseudomonas/genética , Secuencia de Aminoácidos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Mononucleótido de Flavina/metabolismo , Pseudomonas/enzimología , Piridinas/metabolismo , Pirrolidinas/metabolismo , Alineación de SecuenciaRESUMEN
ATP-binding cassette (ABC) transporters, composed of importers and exporters, form one of the biggest protein superfamilies that transport a variety of substrates across the membrane, powered by ATP hydrolysis. Most ABC transporters are composed of two transmembrane domains and two cytoplasmic nucleotide-binding domains. Also, importers from prokaryotes usually have extra solute-binding proteins in the periplasm that are responsible for the binding of substrates. Structures of importers have been reported that suggested a two-state model for the transport mechanism. Energy-coupling factor (ECF) transporters belong to a new class of ATP-binding cassette importers. Each ECF transporter comprises an energy-coupling module consisting of a transmembrane T protein (EcfT), two nucleotide-binding proteins (EcfA and EcfA'), and another transmembrane substrate-specific binding S protein (EcfS). Despite the similarities with ABC transporters, ECF transporters have different organizational and functional properties. The lack of solute-binding proteins in ECF transporters differentiates them clearly from the canonical ABC importers. Previously reported structures of the EcfS proteins RibU and ThiT clearly demonstrated the binding site of substrate riboflavin and thiamine, respectively. However, the organization of the four different components and the transport mechanism of ECF transporters remain unknown. Here we present the structure of an intact folate ECF transporter from Lactobacillus brevis at a resolution of 3 Å. This structure was captured in an inward-facing, nucleotide-free conformation with no bound substrate. The folate-binding protein FolT is nearly parallel to the membrane and is bound almost entirely by EcfT, which adopts an L shape and connects to EcfA and EcfA' through two coupling helices. Two conserved XRX motifs from the coupling helices of EcfT have a vital role in energy coupling by docking into EcfA-EcfA'. We propose a transport model that involves a substantial conformational change of FolT.
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Proteínas Bacterianas/química , Transportadores de Ácido Fólico/química , Levilactobacillus brevis/química , Transportadoras de Casetes de Unión a ATP/química , Adenosina Trifosfatasas/metabolismo , Adenosina Trifosfato/metabolismo , Secuencias de Aminoácidos , Proteínas Bacterianas/metabolismo , Secuencia Conservada , Cristalografía por Rayos X , Transportadores de Ácido Fólico/metabolismo , Modelos Moleculares , Conformación Proteica , Subunidades de Proteína/química , Subunidades de Proteína/metabolismo , Proteolípidos/metabolismoRESUMEN
Ups1 forms a complex with Mdm35 and is critical for the transport of phosphatidic acid (PA) from the mitochondrial outer membrane to the inner membrane. We report the crystal structure of the Ups1-Mdm35-PA complex and the functional characterization of Ups1-Mdm35 in PA binding and transfer. Ups1 features a barrel-like structure consisting of an antiparallel ß-sheet and three α-helices. Mdm35 adopts a three-helical clamp-like structure to wrap around Ups1 to form a stable complex. The ß-sheet and α-helices of Ups1 form a long tunnel-like pocket to accommodate the substrate PA, and a short helix α2 acts as a lid to cover the pocket. The hydrophobic residues lining the pocket and helix α2 are critical for PA binding and transfer. In addition, a hydrophilic patch on the surface of Ups1 near the PA phosphate-binding site also plays an important role in the function of Ups1-Mdm35. Our study reveals the molecular basis of the function of Ups1-Mdm35 and sheds new light on the mechanism of intramitochondrial phospholipid transport by the MSF1/PRELI family proteins.
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Mitocondrias/metabolismo , Proteínas Mitocondriales/química , Proteínas Mitocondriales/metabolismo , Ácidos Fosfatidicos/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Transporte Biológico , Cardiolipinas/biosíntesis , Cristalografía por Rayos X , Modelos Moleculares , Ácidos Fosfatidicos/química , Fosfolípidos/metabolismo , Conformación Proteica , Estructura Secundaria de ProteínaRESUMEN
MYC2, a basic helix-loop-helix (bHLH) transcription factor, is a key regulator in the activation of jasmonate (JA) response. However, the molecular details of MYC2 involving in methyl jasmonate (MeJA)-induced chilling tolerance of fruit remain largely unclear. In the present work, two MYC2 genes, MaMYC2a and MaMYC2b, and one homolog of the inducer of the C-repeat-binding factor (CBF) gene, MaICE1 were isolated and characterized from banana fruit. MaMYC2s and MaICE1 were found to be all localized in the nucleus. In addition, the proline-rich domain (PRD) and the acidic domain (AD) in the N-terminus were important for the transcriptional activation of MaMYC2 in yeast cells. Unlike MaICE1's constitutive expression, MaMYC2a and MaMYC2b were induced rapidly following MeJA treatment during cold storage. Moreover, protein-protein interaction analysis confirmed that MaMYC2s interacted with MaICE1. The expression of ICE-CBF cold-responsive pathway genes including MaCBF1, MaCBF2, MaCOR1, MaKIN2, MaRD2 and MaRD5 was also significantly induced by MeJA. Taken together, our work provides strong evidence that MaMYC2 is involved in MeJA-induced chilling tolerance in banana fruit through physically interacting and likely functionally coordinating with MaICE1, revealing a novel mechanism for ICE1 in response to cold stress as well as during development of induced chilling tolerance.
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Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/metabolismo , Frío , Ciclopentanos/metabolismo , Frutas/metabolismo , Musa/metabolismo , Oxilipinas/metabolismo , Proteínas de Plantas/metabolismo , Acetatos , Secuencia de Aminoácidos , Regulación de la Expresión Génica de las Plantas , Datos de Secuencia Molecular , Musa/genética , Estructura Terciaria de Proteína , Reacción en Cadena en Tiempo Real de la Polimerasa , Análisis de Secuencia de ADNRESUMEN
Hydroquinone (HQ) and catechol (CC) are important chemical raw materials in the modern industry, unfortunately, which are also high toxic phenolic pollutants. So how to achieve highly sensitive and selective determination HQ and CC is the challenge we face. In the present work, we report a facile strategy to obtain nitrogen and phosphorous co-doped glucose-derived carbon coated CoP nanowires (G-CoP/N,P-C NWs), in which nitrilotriacetic acid (NTA) was as the chelating reagent, glucose was as carbon source, and the precursors were subsequently experienced carbonization and phosphorization process. G-CoP/N,P-C NWs can shorten the distance of the electron transport and expand the reaction area, showing the intriguing electronic conductivity and electrocatalytic abilities. An electrochemical phenolic sensor based on G-CoP/N,P-C NWs is fabricated. The as-prepared sensor showcases the good sensing performance for HQ and CC with comparative linearity ranges of 0.8-900 µM (HQ) and 0.6-800 µM (CC), low limits of detections (LODs) of 0.18 µM (S/N = 3) and 0.12 µM (S/N = 3) for HQ and CC, respectively. Notably, it also displays excellent practical application for the recognition of HQ and CC in the rain water, the tap water, the domestic wastewater and the lake water, which may be a promising candidate in environmental water monitoring and drinking water safety.
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Hidroquinonas , Nanocables , Carbono , Catecoles/análisis , Electrodos , Glucosa , Hidroquinonas/análisis , Fenoles , Aguas Residuales , AguaRESUMEN
Antarctic krill oil is an emerging marine lipid and expected to be a potential functional food due to its diverse nutrients, such as eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), phospholipids, astaxanthin and tocopherols. Although krill oil has been previously proved to have anti-inflammatory activity, there is little information about the relationship between its chemical compositions and anti-inflammatory activity. In this study, the RAW264.7 macrophages model was used to elucidate and compare the anti-inflammatory potential of different krill lipid fractions: KLF-A, KLF-H and KLF-E, which have increasing phospholipids, EPA and DHA contents but decreasing astaxanthin and tocopherols levels. Results showed that all the krill lipid fractions alleviated the inflammatory reaction by inhibition of production of nitric oxide (NO), release of tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß and IL-6 and gene expression of proinflammatory mediators including TNF-α, IL-1ß, IL-6, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS). In addition, KLF-E with the highest phospholipids, EPA and DHA contents showed the strongest inhibition effect on the LPS-induced proinflammatory mediator release and their gene expressions. The results would be helpful to provide powerful insights into the underlying anti-inflammatory mechanism of krill lipid and guiding the production of krill oil products with tailor-made anti-inflammatory activity.
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Biodegradation of aromatic and heterocyclic compounds requires an oxidative ring cleavage enzymatic step. Extensive biochemical research has yielded mechanistic insights about catabolism of aromatic substrates; yet much less is known about the reaction mechanisms underlying the cleavage of heterocyclic compounds such as pyridine-ring-containing ones like 2,5-hydroxy-pyridine (DHP). 2,5-Dihydroxypyridine dioxygenase (NicX) from Pseudomonas putida KT2440 uses a mononuclear nonheme Fe(II) to catalyze the oxidative pyridine ring cleavage reaction by transforming DHP into N-formylmaleamic acid (NFM). Herein, we report a crystal structure for the resting form of NicX, as well as a complex structure wherein DHP and NFM are trapped in different subunits. The resting state structure displays an octahedral coordination for Fe(II) with two histidine residues (His265 and His318), a serine residue (Ser302), a carboxylate ligand (Asp320), and two water molecules. DHP does not bind as a ligand to Fe(II), yet its interactions with Leu104 and His105 function to guide and stabilize the substrate to the appropriate position to initiate the reaction. Additionally, combined structural and computational analyses lend support to an apical dioxygen catalytic mechanism. Our study thus deepens understanding of non-heme Fe(II) dioxygenases.
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Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Dioxigenasas/química , Dioxigenasas/metabolismo , Compuestos Heterocíclicos/metabolismo , Pseudomonas putida/enzimología , Secuencia de Aminoácidos , Catálisis , Cristalografía por Rayos X , Dioxigenasas/clasificación , Dioxigenasas/genética , Hierro , Ligandos , Modelos Moleculares , Oxígeno/metabolismo , Filogenia , Conformación ProteicaRESUMEN
Phospholipase D (PLD) hydrolyzes the phosphodiester bond of glycerophospholipids and produces phosphatidic acid (PA), which acts as a second messenger in many living organisms. A large number of PLDs have been identified in eukaryotes, and are viewed as promising targets for drug design because these enzymes are known to be tightly regulated and to function in the pathophysiology of many human diseases. However, the underlying molecular mechanisms of catalysis and regulation of eukaryotic PLD remain elusive. Here, we determined the crystal structure of full-length plant PLDα1 in the apo state and in complex with PA. The structure shows that the N-terminal C2 domain hydrophobically interacts with the C-terminal catalytic domain that features two HKD motifs. Our analysis reveals the catalytic site, substrate-binding mechanism, and a new Ca2+-binding site that is required for the activation of PLD. In addition, we tested several efficient small-molecule inhibitors against PLDα1, and suggested a possible competitive inhibition mechanism according to structure-based docking analysis. This study explains many long-standing questions about PLDs and provides structural insights into PLD-targeted inhibitor/drug design.
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Proteínas de Arabidopsis/química , Fosfolipasa D/química , Proteínas de Arabidopsis/antagonistas & inhibidores , Proteínas de Arabidopsis/metabolismo , Dominio Catalítico , Cristalografía por Rayos X , Activación Enzimática , Inhibidores Enzimáticos/química , Enzimas , Modelos Moleculares , Fosfolipasa D/antagonistas & inhibidores , Fosfolipasa D/metabolismo , Conformación Proteica , Dominios ProteicosRESUMEN
Isovaleryl-CoA dehydrogenase (IVD), a member of the acyl-CoA dehydrogenase (ACAD) family, is a key enzyme catalyzing the conversion of isovaleryl-CoA to ß-methylcrotonyl-CoA in the third reaction of the leucine catabolism pathway and simultaneously transfers electrons to the electron-transferring flavoprotein (ETF) for ATP synthesis. We previously identified the ETF ortholog in rice blast fungus Magnaporthe oryzae (MoETF) and showed that MoETF was essential for fungal growth, conidiation and pathogenicity. To further investigate the biological function of electron-transferring proteins and clarify the role of leucine catabolism in growth and pathogenesis, we characterized MoIVD (M. oryzae isovaleryl-CoA dehydrogenase). MoIvd is highly conserved in fungi and its expression was highly induced by leucine. The Δmoivd mutants showed reduced growth, decreased conidiation and compromised pathogenicity, while the conidial germination and appressorial formation appeared normal. Consistent with a block in leucine degradation, the Δmoivd mutants accumulated isovaleric acid, grew more slowly, fully lacked pigmentation and completely failed to produce conidia on leucine-rich medium. These defects were largely rescued by raising the extracellular pH, suggesting that the accumulation of isovaleric acid contributes to the growth and conidiation defects. However, the reduced virulence of the mutants was probably due to their inability to overcome oxidative stress, since a large amount of ROS (reactive oxygen species) accumulated in infected host cell. In addition, MoIvd is localized to mitochondria and interacted with its electron receptor MoEtfb, the ß subunit of MoEtf. Taken together, our results suggest that MoIVD functions in leucine catabolism and is required for the vegetative growth, conidiation and full virulence of M. oryzae, providing the first evidence for IVD-mediated leucine catabolism in the development and pathogenesis of plant fungal pathogens.
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Objective: To measure the expression levels of interleukin-17( IL-17) and IL-23 mRNAs in the muscle tissue of the mice with experimental autoimmune myositis( EAM),and investigate the impact of tacrolimus( TAC) treatment on the mRNA levels of IL-17 and IL-23 and its therapeutic effect in EAM mice. Methods: Fifteen female BALB / c mice were divided randomly into three groups,a normal control group,an EAM model group and a TAC-treated group. HE staining was used to observe the pathological changes for evaluating muscle inflammation of EAM mice. The expression levels of IL-17 and IL-23 mRNAs in the muscle tissues were measured by real-time fluorescence quantitative PCR. Then the correlations between the pathological changes and the expressions of IL-17 and IL-23 mRNAs were analyzed by Pearson methods. Results: Compared with the normal controls,the mRNA levels of IL-17 and IL-23 were significantly up-regulated in the EAM model group,and they were down-regulated obviously after TAC treatment. Moreover,the mRNA levels of IL-17 and IL-23 had a significantly positive correlation with the pathological score of the muscle tissues. Compared with the EAM model group,the pathological score of the muscle tissues decreased in the TAC-treated group. Conclusion: TAC can down-regulate mRNA levels of IL-17 and IL-23 in the EAM mice.
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Inmunosupresores/farmacología , Interleucina-17/metabolismo , Interleucina-23/metabolismo , Músculo Esquelético/metabolismo , Enfermedad Autoinmune Experimental del Sistema Nervioso/metabolismo , ARN Mensajero/metabolismo , Tacrolimus/farmacología , Animales , Regulación hacia Abajo , Femenino , Interleucina-17/genética , Interleucina-23/genética , Ratones , Ratones Endogámicos BALB C , Distribución AleatoriaRESUMEN
OBJECTIVE: Toll-like receptor 4 (TLR4) is one of the key players in the development of many autoimmune diseases. To determine the possible role of TLR4 in polymyositis (PM) development, we collected muscle samples from PM patients and mice subjected to an experimental autoimmune myositis (EAM) model. METHODS: We measured TLR4-MyD88 pathway-related factors, interferon-γ (IFN-γ), and interleukin-17A (IL-17A) in EAM mice and PM patients. Then, we observed the changes of above factors and the inflammatory development of EAM mice with TLR4 antagonist TAK-242, IFN-γ, or IL-17A antibody treatment. RESULTS: The expression of TLR4, MyD88, and NF-κB was significantly upregulated in the muscle tissues both in 22 patients with PM and in the EAM model. As expected, increased levels of various cytokines, such as IL-1ß, IL-6, IL-10, IL-12, tumor necrosis factor-α, TGF-ß, IFN-γ, and IL-17A, were evident in the serum of EAM mice. Moreover, mRNA expression levels of IFN-γ and IL-17A were significantly increased in both PM patients and EAM mice. Consistently, the levels of these factors were positively correlated with the degree of muscle inflammation in EAM mice. However, when EAM mice were treated with TLR4 antagonist TAK-242, the expression of IFN-γ and IL-17A was decreased. When the cytokines were neutralized by anti-IFN-γ or anti-IL-17A antibody, the inflammatory development of EAM exacerbated or mitigated. CONCLUSION: The present study provided the important evidence that the TLR4-MyD88 pathway may be involved in the immune mechanisms of PM by mediating IFN-γ and IL-17A.
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Energy-coupling factor (ECF) transporters are a large family of ATP-binding cassette transporters recently identified in microorganisms. Responsible for micronutrient uptake from the environment, ECF transporters are modular transporters composed of a membrane substrate-binding component EcfS and an ECF module consisting of an integral membrane scaffold component EcfT and two cytoplasmic ATP binding/hydrolysis components EcfA/A'. ECF transporters are classified into groups I and II. Currently, the molecular understanding of group-I ECF transporters is very limited, partly due to a lack of transporter complex structural information. Here, we present structures and structure-based analyses of the group-I cobalt ECF transporter CbiMNQO, whose constituting subunits CbiM/CbiN, CbiQ, and CbiO correspond to the EcfS, EcfT, and EcfA components of group-II ECF transporters, respectively. Through reconstitution of different CbiMNQO subunits and determination of related ATPase and transporter activities, the substrate-binding subunit CbiM was found to stimulate CbiQO's basal ATPase activity. The structure of CbiMQO complex was determined in its inward-open conformation and that of CbiO in ß, γ-methyleneadenosine 5'-triphosphate-bound closed conformation. Structure-based analyses revealed interactions between different components, substrate-gating function of the L1 loop of CbiM, and conformational changes of CbiO induced by ATP binding and product release within the CbiMNQO transporter complex. These findings enabled us to propose a working model of the CbiMNQO transporter, in which the transport process requires the rotation or toppling of both CbiQ and CbiM, and CbiN might function in coupling conformational changes between CbiQ and CbiM.