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Many soft and biological materials display so-called 'soft glassy' dynamics; their constituents undergo anomalous random motions and complex cooperative rearrangements. A recent simulation model of one soft glassy material, a coarsening foam, suggested that the random motions of its bubbles are due to the system configuration moving over a fractal energy landscape in high-dimensional space. Here we show that the salient geometrical features of such high-dimensional fractal landscapes can be explored and reliably quantified, using empirical trajectory data from many degrees of freedom, in a model-free manner. For a mayonnaise-like dense emulsion, analysis of the observed trajectories of oil droplets quantitatively reproduces the high-dimensional fractal geometry of the configuration path and its associated local energy minima generated using a computational model. That geometry in turn drives the droplets' complex random motion observed in real space. Our results indicate that experimental studies can elucidate whether the similar dynamics in different soft and biological materials may also be due to fractal landscape dynamics.
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The changes of physicochemical and biochemical parameters of a silty loam (S1) and sandy loam (S2) vineyard soils added with spent mushroom substrate (SMS) or SMS composted with ophite (OF) as rock dust (SMS + OF) were studied. Two doses of SMS or SMS + OF (25 and 100 Mg ha-1) were applied for two consecutive years (2020-2021) and changes of soil physicochemical parameters, and dehydrogenase activity (DHA), respiration (RES), microbial biomass (BIO), and the phospholipid fatty acids (PLFAs) profile were assayed on a temporal basis. The results showed an increase in soil organic carbon (OC) content, total and mineralised N, P, and K, especially when the highest SMS dose was applied to soils. Repeated application caused OC content over time up to 2.3 times higher than initial content in the silty loam soil. This increase was not observed in sandy soil, possibly due to a higher bioavailability of OC, as indicated by the evolution of extractable humic acid/fulvic acid pools. In both soils, all biochemical parameters increased after amendment, being favoured both by the OC and by the presence of OF. Significant positive correlations were found between DHA, RES and BIO, and OC content especially in the first part and then levelled off after the second dose application. Total bacterial or fungal PLFAs patterns reflected the variation of BIO by SMS application. The higher growth of fungi vs. bacterial community in amended soils was recorded after the first SMS application, although the opposite effect occurred after the second application, with similar results in both soils. The findings indicate that the application of SMS or SMS + OF in vineyard soils could be an appropriate agronomic management practice for maintaining soil sustainability, although doses and application times of these amendments should first be evaluated depending on soil texture.
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Agaricales , Poluentes do Solo , Solo/química , Fazendas , Agaricales/química , Carbono , Poluentes do Solo/análise , AreiaRESUMO
Candidatus Liberibacter spp is the most prevalent microorganism in the citrus plant, associated with Citrus Huanglongbing (HLB), which is transmitted by the psyllid vector. In Colombia, the vector Diaphorina citri Kugayama has been reported in different regions, but "Ca. Liberibacter asiaticus" (CLas) has only been detected in insect vectors, not in citrus host plants. To identify the presence and quantify the pathogen in citrus tissues, we employed a combined strategy that involved three techniques based on polymerase chain reaction (PCR). First, we used endpoint PCR with specific primers for CLas (OI1-OI2c) to confirm the infection. Second, we used qPCR with specific primers CIT295a - CIT298 designed on 16S rDNA gene regions to quantify the pathogen load. Finally, we employed droplet digital PCR (ddPCR) to determine the copy number of the pathogen in citrus tissues using the ß-subunit of ribonucleotide reductase (RNR) gene (nrdB) that is specific to CLas. We identified the presence of CLas in citrus plants for the first time in Colombia and quantified its titer in the plant tissue. We employed ddPCR and qPCR to provide crucial information for the country's disease management, control strategies, and general crop health.
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The solute carrier family 16 (SLC16) is comprised of 14 members of the monocarboxylate transporter (MCT) family that play an essential role in the transport of important cell nutrients and for cellular metabolism and pH regulation. MCTs 1-4 have been extensively studied and are involved in the proton-dependent transport of L-lactate, pyruvate, short-chain fatty acids, and monocarboxylate drugs in a wide variety of tissues. MCTs 1 and 4 are overexpressed in a number of cancers, and current investigations have focused on transporter inhibition as a novel therapeutic strategy in cancers. MCT1 has also been used in strategies aimed at enhancing drug absorption due to its high expression in the intestine. Other MCT isoforms are less well characterized, but ongoing studies indicate that MCT6 transports xenobiotics such as bumetanide, nateglinide, and probenecid, whereas MCT7 has been characterized as a transporter of ketone bodies. MCT8 and MCT10 transport thyroid hormones, and recently, MCT9 has been characterized as a carnitine efflux transporter and MCT12 as a creatine transporter. Expressed at the blood brain barrier, MCT8 mutations have been associated with an X-linked intellectual disability, known as Allan-Herndon-Dudley syndrome. Many MCT isoforms are associated with hormone, lipid, and glucose homeostasis, and recent research has focused on their potential roles in disease, with MCTs representing promising novel therapeutic targets. This review will provide a summary of the current literature focusing on the characterization, function, and regulation of the MCT family isoforms and on their roles in drug disposition and in health and disease. SIGNIFICANCE STATEMENT: The 14-member solute carrier family 16 of monocarboxylate transporters (MCTs) plays a fundamental role in maintaining intracellular concentrations of a broad range of important endogenous molecules in health and disease. MCTs 1, 2, and 4 (L-lactate transporters) are overexpressed in cancers and represent a novel therapeutic target in cancer. Recent studies have highlighted the importance of MCTs in glucose, lipid, and hormone homeostasis, including MCT8 in thyroid hormone brain uptake, MCT12 in carnitine transport, and MCT11 in type 2 diabetes.
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Transportadores de Ácidos Monocarboxílicos/metabolismo , Animais , Humanos , Doenças Metabólicas/metabolismo , Transportadores de Ácidos Monocarboxílicos/antagonistas & inibidores , Transportadores de Ácidos Monocarboxílicos/química , Transportadores de Ácidos Monocarboxílicos/genética , Relação Estrutura-Atividade , Distribuição Tecidual , Transcrição GênicaRESUMO
The use of nanoparticles as drug delivery systems has increased in importance in the last decades. Despite the disadvantages of difficulty swallowing, gastric irritation, low solubility, and poor bioavailability, oral administration stands out as the most widely used route for therapeutic treatments, though it may not always be the most effective route. The effect of the first hepatic pass is one of the primary challenges that drugs must overcome to carry out their therapeutic effect. For these reasons, controlled-release systems based on nanoparticles synthesized from biodegradable natural polymers have been reported to be very efficient in enhancing oral delivery in multiple studies. Chitosan has been shown to have an extensive variability of properties and roles in the pharmaceutical and health fields; of its most important properties are the ability to encapsulate and transport drugs within the body and enhance the drug interaction with the target cells, which improves the efficacy of the encapsulated drugs. The physicochemical properties of chitosan give it the ability to form nanoparticles through multiple mechanisms, which will be addressed in this article. The present review article focuses on highlighting the applications of chitosan nanoparticles for oral drug delivery.
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Quitosana , Nanopartículas , Portadores de Fármacos/química , Quitosana/química , Sistemas de Liberação de Medicamentos , Administração Oral , Polímeros/química , Nanopartículas/químicaRESUMO
Pregestational Diabetes Mellitus (PDM) during pregnancy constitutes an unfavorable embryonic and fetal development environment, with a high incidence of congenital malformations (CM). Neural tube defects are the second most common type of CM in children of diabetic mothers (CDM), who also have an elevated risk of developing neurodevelopmental disorders. The mechanisms that lead to these neuronal disorders in CDM are not yet fully understood. The present study aimed to know the effect of hyperglycemia on proliferation, neuronal differentiation percentage, and expression of neuronal differentiation mRNA markers in human umbilical cord Wharton's jelly mesenchymal stem cells (hUCWJMSC) of children from normoglycemic pregnancies (NGP) and PDM. We isolated and characterized hUCWJMSC by flow cytometry, immunofluorescence, RT-PCR and were induced to differentiate into adipocytes, osteocytes, and neurons. Proliferation assays were performed to determine the doubling time, and Nestin, TUBB3, FOXO1, KCNK2, LMO3, and MAP2 mRNA gene expression was assessed by semiquantitative RT-PCR. Hyperglycemia significantly decreased proliferation and neuronal differentiation percentage in NGP and PDM cells treated with 40 mM d-glucose. Nestin mRNA expression decreased under control glycemic conditions, while FOXO1, KCNK2, LMO3, and MAP2 mRNA expression increased during neuronal differentiation in both NGP and PDM cells. On the other hand, under hyperglycemic conditions, Nestin was significantly decreased in cells from NGP but not in cells from PDM, while mRNA expression of FOXO1 and LMO3 was significantly increased in cells from NGP, but not in cells from PDM. We found evidence that maternal PDM, with hyperglycemia in culture, affects the biological properties of fetal cells. All these results could be part of fetal programming.
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Diabetes Mellitus , Hiperglicemia , Células-Tronco Mesenquimais , Efeitos Tardios da Exposição Pré-Natal , Geleia de Wharton , Criança , Feminino , Humanos , Gravidez , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteína Forkhead Box O1/genética , Hiperglicemia/complicações , Fatores Imunológicos , Proteínas com Domínio LIM/genética , Nestina/genéticaRESUMO
BACKGROUND: Omega-3 long chain polyunsaturated fatty acids (LCPUFA) reduce circulating cytokines produced by monocytes. Nevertheless, whether the omega-3 LCPUFA regulate the monocytes and their cytokines in Duchenne muscular dystrophy (DMD) is unknown. The aim of this study was to evaluate whether circulating pro-inflammatory monocytes are increased and whether omega-3 LCPUFA selectively suppress these monocytes and their cytokines in patients with DMD. METHODS: This was a double-blind, randomized, placebo-controlled pilot study carried out in patients with DMD supplemented with omega-3 LCPUFA (n = 6) or sunflower oils (placebo, n = 6) for 6 months. Monocytes and their cytokines were measured at baseline and after 1, 2, 3, and 6 months of supplementation. RESULTS: The anti-inflammatory monocytes (median, [95% CI]) are increased at month 3 (-0.46 [-13.5-9.5] vs. 8.4 [5.5-12.5], p = 0.05) in the omega-3 LCPUFA group compared with the placebo group. The pro-inflammatory monocytes (-5.7 [-63.8-114.1] vs. -51.9 [-91.2 to -25.4], p = 0.026 and -16.4 [-50.8-50.6] vs. -57.9 [-86.9 to -18.5], p = 0.045 at months 3 and 6, respectively) and their cytokine interleukin 6 (-11.9 [-93.5-148.9] vs. -64.7 [-77.8 to -42.6], p = 0.019 at month 6) decreased in the omega-3 LCPUFA group compared with the placebo group. Pro-inflammatory monocytes decreased and anti-inflammatory monocytes were augmented (p < 0.05) during the 6 months of supplementation with omega-3 LCPUFA. CONCLUSIONS: This pilot study suggests that supplementation with omega-3 LCPUFA could have a selective reductive effect on pro-inflammatory monocytes and their cytokines in patients with DMD. These findings also support the performance of studies in a significant population to explore the role of omega-3 LCPUFA on monocyte populations and their cytokines in patients with DMD. This research was registered at clinicaltrials.gov (NCT018264229).
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Ácidos Graxos Ômega-3 , Distrofia Muscular de Duchenne , Suplementos Nutricionais , Método Duplo-Cego , Ácidos Graxos Ômega-3/farmacologia , Ácidos Graxos Ômega-3/uso terapêutico , Humanos , Monócitos , Distrofia Muscular de Duchenne/tratamento farmacológico , Projetos PilotoRESUMO
OBJECTIVES: This study aimed to evaluate whether the expression of circulating dystromiRs and a group of oxidative stress-related (OS-R) miRNAs is associated with muscle injury and circulating metabolic parameters in Duchenne muscular dystrophy (DMD) patients. METHODS: Twenty-four DMD patients were included in this cross-sectional study. Clinical scales to evaluate muscle injury (Vignos, GMFCS, Brooke, and Medical Research Council), enzymatic muscle injury parameters (CPK, ALT, and AST), anthropometry, metabolic indicators, physical activity, serum dystromiRs (miR-1-3p, miR-133a-3p, and miR-206), and OS-R miRNAs (miR-21-5p, miR-31-5p, miR-128-3p, and miR-144-3p) levels were measured in ambulatory and non-ambulatory DMD patients. RESULTS: DystromiRs (except miR-1-3p) and miRNAs OS-R levels were lower (p-value <.05) in the non-ambulatory group than the ambulatory group. The expression of those miRNAs correlated with Vignos scale score (For instance, rho = -0.567, p-value <0.05 for miR-21-5p) and with other scales scores of muscle function and strength. CPK, AST, and ALT concentration correlated with expression of all miRNAs (For instance, rho = 0.741, p-value <.05 between miR-206 level and AST concentration). MiR-21-5p level correlated with glucose concentration (rho = -0.369, p-value = .038), and the miR-1-3p level correlated with insulin concentration (rho = 0.343, p-value = .05). CONCLUSIONS: Non-ambulatory DMD patients have lower circulating dystromiRs and OS-R miRNAs levels than ambulatory DMD patients. The progressive muscle injury is associated with a decrease in the expression of those miRNAs, evidencing DMD progress. These findings add new information about the natural history of DMD.
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MicroRNA Circulante , Insulinas , MicroRNAs , Distrofia Muscular de Duchenne , Biomarcadores , Estudos Transversais , Glucose , Humanos , Músculos/metabolismo , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismoRESUMO
Homogeneous extremely low-frequency electromagnetic fields (ELF-EMFs) alter biological phenomena, including the cell phenotype and proliferation rate. Heterogenous vortex magnetic fields (VMFs), a new approach of exposure to magnetic fields, induce systematic movements on charged biomolecules from target cells; however, the effect of VMFs on living systems remains uncertain. Here, we designed, constructed, and characterized an ELF-VMF-modified Rodin's coil to expose SH-SY5Y cells. Samples were analyzed by performing 2D-differential-gel electrophoresis, identified by MALDI-TOF/TOF, validated by western blotting, and characterized by confocal microscopy. A total of 106 protein spots were differentially expressed; 40 spots were downregulated and 66 were upregulated in the exposed cell proteome, compared to the control cell proteome. The identified spots are associated with cytoskeleton and cell viability proteins, and according to the protein-protein interaction network, a significant interaction among them was found. Our data revealed a decrease in cell survival associated with apoptotic cells without effects on the cell cycle, as well as evident changes in the cytoskeleton. We demonstrated that ELF-VMFs, at a specific frequency and exposure time, alter the cell proteome and structurally affect the target cells. This is the first report showing that VMF application might be a versatile system for testing different hypotheses in living systems, using appropriate exposure parameters.© 2022 Bioelectromagnetics Society.
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Neuroblastoma , Proteoma , Apoptose , Linhagem Celular , Citoesqueleto , Campos Eletromagnéticos , Humanos , Campos MagnéticosRESUMO
Charge selectivity forms the basis of cellular excitation or inhibition by Cys-loop ligand-gated ion channels (LGICs), and is essential for physiological receptor function. There are no reports of naturally occurring mutations in LGICs associated with the conversion of charge selectivity. Here, we report on a CHRNA1 mutation (α1Leu251Arg) in a patient with congenital myasthenic syndrome associated with transformation of the muscle acetylcholine receptor (AChR) into an inhibitory channel. Performing patch-clamp experiments, the AChR was found to be converted into chloride conductance at positive potentials, whereas whole-cell currents at negative potentials, although markedly reduced, were still carried by sodium. Umbrella sampling molecular dynamics simulations revealed constriction of the channel pore radius to 2.4 Å as a result of the mutation, which required partial desolvation of the ions in order to permeate the pore. Ion desolvation was associated with an energetic penalty that was compensated for by the favorable electrostatic interaction of the positively charged arginines with chloride. These findings reveal a mechanism for the transformation of the muscle AChR into an inhibitory channel in a clinical context.
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Acetilcolina/metabolismo , Cloretos/metabolismo , Músculos/metabolismo , Mutação/genética , Receptores Colinérgicos/metabolismo , Linhagem Celular , Células HEK293 , Humanos , Ativação do Canal Iônico/fisiologia , Potenciais da Membrana/fisiologia , Síndromes Miastênicas Congênitas/metabolismo , Técnicas de Patch-Clamp/métodos , Receptores Nicotínicos/metabolismo , Sódio/metabolismoRESUMO
The drug of abuse, γ-hydroxybutyric acid (GHB), is commonly co-ingested with ethanol, resulting in a high incidence of toxicity and death. Our laboratory has previously reported that GHB is a substrate for the monocarboxylate transporters (MCTs), necessary for its absorption, renal clearance, and tissue distribution, including across the blood-brain barrier. Our goal was to investigate the drug-drug interaction (DDI) between GHB and ethanol and to evaluate MCT1 inhibition as a strategy to reverse toxicity. The toxicokinetics of this DDI were investigated, including brain-to-plasma concentration ratios, in the presence and absence of ethanol. The toxicodynamic parameters examined were respiratory depression (breathing frequency, tidal volume) and sedation (time of return-of-righting reflex). Ethanol was administered (2 g/kg i.v.) 5 minutes before the intravenous or oral administration of GHB, and MCT1 inhibitors AZD-3965 and AR-C155858 (5 mg/kg i.v.) were administered 60 minutes after GHB administration. Ethanol administration did not alter the toxicokinetics or respiratory depression caused by GHB after intravenous or oral administration; however, it significantly increased the sedation effect, measured by return-to-righting time. AZD-3965 or AR-C155858 significantly decreased the effects of the co-administration of GHB and ethanol on respiratory depression and sedation of this DDI and decreased brain concentrations and the brain-to-plasma concentration ratio of GHB. The results indicate that ethanol co-administered with GHB increases toxicity and that MCT1 inhibition is effective in reversing toxicity by inhibiting GHB brain uptake when given after GHB-ethanol administration. SIGNIFICANCE STATEMENT: These studies investigated the enhanced toxicity observed clinically when γ-hydroxybutyric acid (GHB) is co-ingested with alcohol and evaluated strategies to reverse this toxicity. The effects of the novel monocarboxylate transporter 1 (MCT1) inhibitors AR-C155858 and AZD-3965 on this drug-drug interaction have not been studied before, and these preclinical studies indicate that MCT1 inhibitors can decrease brain concentrations of GHB by inhibiting brain uptake, even when administered at times after GHB-ethanol. AZD-3965 represents a potential treatment strategy for GHB-ethanol overdoses.
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Etanol/toxicidade , Hidroxibutiratos/toxicidade , Transportadores de Ácidos Monocarboxílicos/antagonistas & inibidores , Pirimidinonas/farmacologia , Simportadores/antagonistas & inibidores , Tiofenos/farmacologia , Uracila/análogos & derivados , Animais , Interações Medicamentosas/fisiologia , Etanol/metabolismo , Hidroxibutiratos/metabolismo , Masculino , Transportadores de Ácidos Monocarboxílicos/metabolismo , Pirimidinonas/uso terapêutico , Ratos , Ratos Sprague-Dawley , Insuficiência Respiratória/induzido quimicamente , Insuficiência Respiratória/tratamento farmacológico , Insuficiência Respiratória/metabolismo , Simportadores/metabolismo , Tiofenos/uso terapêutico , Uracila/farmacologia , Uracila/uso terapêuticoRESUMO
S-metolachlor is a widely used herbicide that may contaminate groundwater when applied to irrigated crops, especially when the soil has a low organic carbon (OC) content. The objective here was to assess the capacity of two organic wastes, namely, green compost (GC) and pelletised organo-mineral manure fertilizer (PM), applied to two soils (S) with different textures at a rate of 10% dry weight to modify the fate of S-metolachlor. The herbicide's Freundlich adsorption coefficient (Kf) increased within a range of 3.2-8.2 times in S + GC and 3.8-6.8 times in S + PM. A positive correlation between adsorption and OC and the coefficient of variation of the OC normalised adsorption coefficients (Kfoc) higher than 20% indicated the evident influence on this process of soil OC content and its nature. The increase in adsorption did not prevent the dissipation of S-metolachlor in the amended soils, although the degradation rate decreased up to ~2 times or was not significantly modified across the different soil types. The S-metolachlor metabolites, metolachlor ethane sulfonic acid and metolachlor oxanilic acid, were detected in the herbicide's dissipation in the unamended soils, but they were not detected in the amended soils. The mobility experiments indicated leached amounts of S-metolachlor higher than 50% in unamended soil. The amounts decreased 1.1-1.7 times and 1.7-1.8 times in the S + GC and S + PM when a saturated flow was applied. Moreover, breakthrough curves indicated a slow leaching kinetics of herbicide in amended soils, with low concentrations continuously detected in the leachates together with a decrease in the maximum peak concentration. The results show the effect of the application of organic wastes especially in sandy soils to promote the immobilisation and/or degradation of S-metolachlor, avoiding its transfer to other environmental compartments.
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Herbicidas , Poluentes do Solo , Acetamidas , Adsorção , Herbicidas/análise , Solo , Poluentes do Solo/análiseRESUMO
Gamma hydroxybutyric acid (GHB) has been approved clinically to treat excessive daytime sleepiness and cataplexy in patients with narcolepsy, alcohol and opioid withdrawal, and as an anesthetic. The use of GHB clinically is limited due to its high abuse potential. The absorption, clearance and tissue uptake of GHB is mediated by proton-dependent and sodium-coupled monocarboxylate transporters (MCTs and SMCTs) and inhibition of these transporters may result in a change in GHB pharmacokinetics and pharmacodynamics. Previous studies have reported that non-steroidal anti-inflammatory drugs (NSAIDs) may inhibit these monocarboxylate transporters. Therefore, the purpose of this work was to analyze the interaction between GHB (at a dose of 600 mg/kg i. v.) and the NSAID, diclofenac, by examining the effects of this drug on the in vivo pharmacokinetics and pharmacodynamics in rat studies. The pharmacodynamic effect evaluated was respiratory depression, a measure of toxicity observed by GHB at this dose. There was an improvement in the respiratory rate with diclofenac administration suggesting an effect of diclofenac on GHB toxicity. In vitro studies with rat blood brain endothelial cells (RBE4) that express MCT1 indicated that diclofenac can inhibit GHB transport with an IC50 of 10.6 µM at pH 7.4. In vivo studies found a decrease in brain GHB concentrations and a decrease in the brain-to-plasma concentration ratio following diclofenac treatment. With this study we can conclude that diclofenac and potentially other NSAIDs can inhibit the transport of GHB into the brain, therefore decreasing GHB's pharmacodynamic effects and toxicity.
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Encéfalo , Diclofenaco/farmacocinética , Interações Medicamentosas , Hidroxibutiratos/farmacocinética , Transportadores de Ácidos Monocarboxílicos , Insuficiência Respiratória , Simportadores , Anestésicos/farmacocinética , Anestésicos/toxicidade , Animais , Anti-Inflamatórios não Esteroides/farmacocinética , Transporte Biológico Ativo/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Células Cultivadas , Relação Dose-Resposta a Droga , Células Endoteliais/metabolismo , Hidroxibutiratos/toxicidade , Transportadores de Ácidos Monocarboxílicos/antagonistas & inibidores , Transportadores de Ácidos Monocarboxílicos/metabolismo , Ratos , Ratos Sprague-Dawley , Insuficiência Respiratória/induzido quimicamente , Insuficiência Respiratória/tratamento farmacológico , Oxibato de Sódio/farmacocinética , Simportadores/antagonistas & inibidores , Simportadores/metabolismoRESUMO
MUSK encodes the muscle-specific receptor tyrosine kinase (MuSK), a key component of the agrin-LRP4-MuSK-DOK7 signaling pathway, which is essential for the formation and maintenance of highly specialized synapses between motor neurons and muscle fibers. We report a patient with severe early-onset congenital myasthenic syndrome and two novel missense mutations in MUSK (p.C317R and p.A617V). Functional studies show that MUSK p.C317R, located at the frizzled-like cysteine-rich domain of MuSK, disrupts an integral part of MuSK architecture resulting in ablated MuSK phosphorylation and acetylcholine receptor (AChR) cluster formation. MUSK p.A617V, located at the kinase domain of MuSK, enhances MuSK phosphorylation resulting in anomalous AChR cluster formation. The identification and evidence for pathogenicity of MUSK mutations supported the initiation of treatment with ß2-adrenergic agonists with a dramatic improvement of muscle strength in the patient. This work suggests uncharacterized mechanisms in which control of the precise level of MuSK phosphorylation is crucial in governing synaptic structure.
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Mutação , Síndromes Miastênicas Congênitas/diagnóstico , Síndromes Miastênicas Congênitas/genética , Receptores Proteína Tirosina Quinases/genética , Receptores Colinérgicos/genética , Sinapses/genética , Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Agonistas de Receptores Adrenérgicos beta 2/uso terapêutico , Alelos , Substituição de Aminoácidos , Animais , Sistemas CRISPR-Cas , Linhagem Celular , Análise Mutacional de DNA , Feminino , Marcação de Genes , Humanos , Camundongos , Modelos Moleculares , Conformação Molecular , Proteínas Musculares/metabolismo , Síndromes Miastênicas Congênitas/tratamento farmacológico , Síndromes Miastênicas Congênitas/metabolismo , Linhagem , Fosforilação , Receptores Proteína Tirosina Quinases/química , Receptores Proteína Tirosina Quinases/metabolismo , Receptores Colinérgicos/química , Receptores Colinérgicos/metabolismo , Relação Estrutura-Atividade , Sinapses/metabolismoRESUMO
Acetylcholine receptor deficiency is the most common form of the congenital myasthenic syndromes, a heterogeneous collection of genetic disorders of neuromuscular transmission characterized by fatiguable muscle weakness. Most patients with acetylcholine receptor deficiency respond well to acetylcholinesterase inhibitors; however, in some cases the efficacy of acetylcholinesterase inhibitors diminishes over time. Patients with acetylcholine receptor deficiency can also benefit from the addition of a ß2-adrenergic receptor agonist to their medication. The working mechanism of ß2-adrenergic agonists in myasthenic patients is not fully understood. Here, we report the long-term follow-up for the addition of ß2-adrenergic agonists for a cohort of patients with acetylcholine receptor deficiency on anticholinesterase medication that demonstrates a sustained quantitative improvement. Coincidently we used a disease model to mirror the treatment of acetylcholine receptor deficiency, and demonstrate improved muscle fatigue, improved neuromuscular transmission and improved synaptic structure resulting from the addition of the ß2-adrenergic agonist salbutamol to the anticholinesterase medication pyridostigmine. Following an initial improvement in muscle fatiguability, a gradual decline in the effect of pyridostigmine was observed in mice treated with pyridostigmine alone (P < 0.001). Combination therapy with pyridostigmine and salbutamol counteracted this decline (P < 0.001). Studies of compound muscle action potential decrement at high nerve stimulation frequencies (P < 0.05) and miniature end-plate potential amplitude analysis (P < 0.01) showed an improvement in mice following combination therapy, compared to pyridostigmine monotherapy. Pyridostigmine alone reduced postsynaptic areas (P < 0.001) and postsynaptic folding (P < 0.01). Combination therapy increased postsynaptic area (P < 0.001) and promoted the formation of postsynaptic junctional folds (P < 0.001), in particular in fast-twitch muscles. In conclusion, we demonstrate for the first time how the improvement seen in patients from adding salbutamol to their medication can be explained in an experimental model of acetylcholine receptor deficiency, the most common form of congenital myasthenic syndrome. Salbutamol enhances neuromuscular junction synaptic structure by counteracting the detrimental effects of long-term acetylcholinesterase inhibitors on the postsynaptic neuromuscular junction. The results have implications for both autoimmune and genetic myasthenias where anticholinesterase medication is a standard treatment.
Assuntos
Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Albuterol/farmacologia , Inibidores da Colinesterase/uso terapêutico , Síndromes Miastênicas Congênitas/tratamento farmacológico , Junção Neuromuscular/efeitos dos fármacos , Brometo de Piridostigmina/uso terapêutico , Potenciais de Ação/fisiologia , Agonistas de Receptores Adrenérgicos beta 2/uso terapêutico , Albuterol/uso terapêutico , Animais , Inibidores da Colinesterase/farmacologia , Modelos Animais de Doenças , Humanos , Camundongos , Brometo de Piridostigmina/farmacologia , Transmissão Sináptica/efeitos dos fármacosRESUMO
Next generation sequencing techniques were recently used to show mutations in COL13A1 cause synaptic basal lamina-associated congenital myasthenic syndrome type 19. Animal studies showed COL13A1, a synaptic extracellular-matrix protein, is involved in the formation and maintenance of the neuromuscular synapse that appears independent of the Agrin-LRP4-MuSK-DOK7 acetylcholine receptor clustering pathway. Here, we report the phenotypic spectrum of 16 patients from 11 kinships harbouring homozygous or heteroallelic mutations in COL13A1. Clinical presentation was mostly at birth with hypotonia and breathing and feeding difficulties often requiring ventilation and artificial feeding. Respiratory crisis related to recurrent apnoeas, sometimes triggered by chest infections, were common early in life but resolved over time. The predominant pattern of muscle weakness included bilateral ptosis (non-fatigable in adulthood), myopathic facies and marked axial weakness, especially of neck flexion, while limb muscles were less involved. Other features included facial dysmorphism, skeletal abnormalities and mild learning difficulties. All patients tested had results consistent with abnormal neuromuscular transmission. Muscle biopsies were within normal limits or showed non-specific changes. Muscle MRI and serum creatine kinase levels were normal. In keeping with COL13A1 mutations affecting both synaptic structure and presynaptic function, treatment with 3,4-diaminopyridine and salbutamol resulted in motor and respiratory function improvement. In non-treated cases, disease severity and muscle strength improved gradually over time and several adults recovered normal muscle strength in the limbs. In summary, patients with COL13A1 mutations present mostly with severe early-onset myasthenic syndrome with feeding and breathing difficulties. Axial weakness is greater than limb weakness. Disease course improves gradually over time, which could be consistent with the less prominent role of COL13A1 once the neuromuscular junction is mature. This report emphasizes the role of collagens at the human muscle endplate and should facilitate the recognition of this disorder, which can benefit from pharmacological treatment.
Assuntos
Colágeno Tipo XIII/genética , Proteínas Musculares/genética , Síndromes Miastênicas Congênitas/genética , Junção Neuromuscular/metabolismo , Transmissão Sináptica/genética , Adolescente , Adulto , Criança , Feminino , Homozigoto , Humanos , Masculino , Músculo Esquelético/patologia , Mutação/genética , Síndromes Miastênicas Congênitas/diagnóstico , Junção Neuromuscular/genética , Sinapses/genética , Adulto JovemRESUMO
This paper reports the mobility and total balance of chlorotoluron (CTL), flufenacet (FNC) and bromide ion (Br-) throughout a sandy soil profile after the application of spent mushroom substrate (SMS) and green compost (GC). Obtaining mobility dataset is crucial to simulate the herbicides' fate under amended soil scenarios by application pesticide leaching models with regulatory application (FOCUS models). The application of organic residues is nowadays increased to improve the crop yields and there is a gap in the simulations of this kind of amended scenarios. A two-year field experiment involving unamended soil (S) and SMS- or GC-amended soil plots was conducted. CTL, FNC, and Br- were annually applied and their residual concentrations were determined in soil profiles (0-100 cm) regularly sampled. In all the treatments the order of mobility is followed as FNC < CTL < Br-. SMS and GC increased herbicide retention in the top 10 cm by the higher organic carbon (OC) content than the unamended soil, and their ability to increase the soil's water-holding capacity and to decrease water percolation. Simultaneously dissolved organic carbon (DOC) content facilitated herbicide transport being it favoured by the initial soil moisture content and the rainfall shortly after the chemicals' initial application. Over the first year, residual amounts (<2.6%) of Br-, CTL and FNC were leached down to 90-100 cm depth in the three treatments. However, over the second year low CTL and FNC amounts (<1.0%) reached the bottom layer only in S + SMS although high Br- concentrations did so in the three treatments (<20%). According to the total balance of Br-, CTL, and FNC in the soil profiles other processes (degradation, mineralisation, bound residues formation, and/or crop uptake) different from leaching below 1 m depth might play a key role in their dissipation especially in the amended soil profiles. SMS and GC are likely to be used as organic amendments to preserve the soil and water quality but in the case of SMS, its higher DOC content could imply a higher potential risk for groundwater contamination than GC.
Assuntos
Agaricales , Compostagem , Herbicidas , Poluentes do Solo , SoloRESUMO
INTRODUCTION: In Duchenne muscular dystrophy (DMD) muscle is replaced by adipose tissue. The role of dietary intake (DI) in DMD has not been evaluated. In this study we examined body composition, body mass index (BMI), and adequacy of DI in patients with DMD and evaluated the influence of DI on body composition. METHODS: Patients (n = 101; age 3-18 years; BMI 11.8-29.5 kg/m2 ) completed a dietary recall to determine DI and then underwent dual-energy X-ray absorptiometry to determine body composition. RESULTS: Preschool-age and school-age boys with DMD had high total energy intake. Protein intake per kilogram exceeded recommendations. As age increased, the percentage of boys with abnormal BMI and fat mass increased, while lean mass decreased. Dietary intake did not predict body fat or lean mass. DISCUSSION: Age-dependent changes in BD in boys with DMD may be due to endogenous metabolic factors related to the underlying disease process and to disease-related mobility impairments. Muscle Nerve 59:295-302, 2019.
Assuntos
Composição Corporal , Índice de Massa Corporal , Dieta , Distrofia Muscular de Duchenne/patologia , Absorciometria de Fóton , Adolescente , Criança , Pré-Escolar , Estudos Transversais , Proteínas Alimentares , Ingestão de Alimentos , Ingestão de Energia , Feminino , Humanos , Masculino , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/reabilitação , Estado NutricionalRESUMO
INTRODUCTION: Patients with Duchenne muscular dystrophy (DMD) demonstrate decreased bone mineral density (BD). It is not clear which factors exert the greatest impact on BD loss in these patients. METHODS: In 63 patients with DMD, serum cytokines (interleukin [IL]-1, IL-6, and tumor necrosis factor-beta [TNF-ß]), C-reactive protein (CRP), creatine kinase (CK), muscle function (by Vignos scale), body composition, and total BD (the latter 2 measured by dual-energy X-ray absorptiometry, or DEXA) were determined. RESULTS: The main factors associated with BD loss were muscle function (34.0%; ß = -0.139; P < 0.023) and age (36.7%; ß = -0.151; P = 0.004). Cytokines, CRP, body fat mass, and CK did not contribute to BD loss. DISCUSSION: Muscle function and age contribute to BD loss in DMD. We propose that a cut-off of at least 6 points for the Vignos scale and at least 10.5 years of age predict a Z-score of less than or equal to -2.0. Muscle Nerve 59:417-421, 2019.
Assuntos
Envelhecimento/patologia , Densidade Óssea , Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/patologia , Osteoporose/patologia , Absorciometria de Fóton , Tecido Adiposo/patologia , Adiposidade , Adolescente , Composição Corporal , Proteína C-Reativa/análise , Criança , Pré-Escolar , Estudos de Coortes , Citocinas/sangue , Feminino , Humanos , Inflamação/patologia , Masculino , Debilidade Muscular/fisiopatologia , Estudos ProspectivosRESUMO
Trichomonas vaginalis induces cellular damage to the host cells (cytotoxicity) through the proteolytic activity of multiple proteinases of the cysteine type (CPs). Some CPs are modulated by environmental factors such as iron, zinc, polyamines, etc. Thus, the goal of this study was to assess the effect of glucose on T. vaginalis cytotoxicity, proteolytic activity and the particular role of TvCP2 (TVAG_057000) during cellular damage. Cytotoxicity assays showed that glucose-restriction (GR) promotes the highest HeLa cell monolayers destruction (~95%) by trichomonads compared to those grown under high glucose (~44%) condition. Zymography and Western blot using different primary antibodies showed that GR increased the proteolytic activity, amount and secretion of certain CPs, including TvCP2. We further characterized the effect of glucose on TvCP2. TvCP2 increases in GR, localized in vesicles close to the plasma membrane and on the surface of T. vaginalis. Furthermore, pretreatment of GR-trichomonads with an anti-TvCP2r polyclonal antibody specifically reduced the levels of cytotoxicity and apoptosis induction to HeLa cells in a concentration-dependent manner. In conclusion, our data show that GR, as a nutritional stress condition, promotes trichomonal cytotoxicity to the host cells, increases trichomonad proteolytic activity and amount of CPs, such as TvCP2 involved in cellular damage.