RESUMO
Rapid Diagnostic Tests (RDT) are useful to identify syphilis cases, particularly for hard-to-reach populations and if laboratory services are scarce. However, RDT performance may be suboptimal. We aimed to assess the sensitivity and specificity of a syphilis RDT using well-characterized blood donors' samples. We categorized samples from 811 blood donors into five groups: 1 - Samples with reactive Chemiluminescence (QML), FTA-Abs, and VDRL; 2 - Samples with reactive QML and FTA-Abs, and nonreactive VDRL; 3 - Samples with reactive QML, and nonreactive for other markers (false-positives); 4 - Controls with nonreactive QML; and 5 - Samples reactive for HIV, with nonreactive QML. Sensitivity was tested in groups 1 (overall and according to VDRL titers) and 2; specificity was tested in groups 3â5. The RDT had high specificity, even in samples reactive for HIV. The sensitivity was high (91.9%) in samples with reactive VDRL but varied between 75.0%â100% according to VDRL titers. The overall sensitivity was lower (81.3%) in samples with reactive FTA-Abs and nonreactive VDRL. The RDT is a useful tool to detect active syphilis but may be more limited for cases with very early or remote infection, or those with prior treatment. When higher sensitivity is needed, additional strategies including recurrent testing or laboratory-based tests may be required.
Assuntos
Infecções por HIV , Sífilis , Humanos , Sífilis/diagnóstico , Doadores de Sangue , Testes de Diagnóstico Rápido , Sorodiagnóstico da Sífilis , Sensibilidade e Especificidade , Infecções por HIV/diagnósticoRESUMO
Abstract Rapid Diagnostic Tests (RDT) are useful to identify syphilis cases, particularly for hard-to-reach populations and if laboratory services are scarce. However, RDT performance may be suboptimal. We aimed to assess the sensitivity and specificity of a syphilis RDT using well-characterized blood donors' samples. We categorized samples from 811 blood donors into five groups: 1 - Samples with reactive Chemiluminescence (QML), FTA-Abs, and VDRL; 2 - Samples with reactive QML and FTA-Abs, and nonreactive VDRL; 3 - Samples with reactive QML, and nonreactive for other markers (false-positives); 4 - Controls with nonreactive QML; and 5 - Samples reactive for HIV, with nonreactive QML. Sensitivity was tested in groups 1 (overall and according to VDRL titers) and 2; specificity was tested in groups 3‒5. The RDT had high specificity, even in samples reactive for HIV. The sensitivity was high (91.9%) in samples with reactive VDRL but varied between 75.0%‒100% according to VDRL titers. The overall sensitivity was lower (81.3%) in samples with reactive FTA-Abs and nonreactive VDRL. The RDT is a useful tool to detect active syphilis but may be more limited for cases with very early or remote infection, or those with prior treatment. When higher sensitivity is needed, additional strategies including recurrent testing or laboratory-based tests may be required.
RESUMO
Calcitonin Gene-Related Peptide (CGRP) is a potent vasodilator peptide widely distributed in the central nervous system and various peripheral tissues, including cardiac muscle. However, its role in heart protein metabolism remains unknown. We examined the acute effects of CGRP on autophagy and the related signaling pathways in the heart mice and cultured neonatal cardiomyocytes. CGRP (100 µg kg-1; s.c.) or 0.9 % saline was injected in awake male C57B16 mice, and the metabolic profile was determined up to 60 min. In fed mice, CGRP drastically increased glycemia and reduced insulinemia, an effect that was accompanied by reduced cardiac phosphorylation levels of Akt at Ser473 without affecting FoxO. Despite these catabolic effects, CGRP acutely inhibited autophagy as estimated by the decrease in LC3II:LC3I and autophagic flux. In addition, the fasting-induced autophagic flux in mice hearts was entirely abrogated by one single injection of CGRP. In parallel, CGRP stimulated PKA/CREB and mTORC1 signaling and increased the phosphorylation of Unc51-like kinase-1 (ULK1), an essential protein in autophagy initiation. Similar effects were observed in cardiomyocytes, in which CGRP also inhibited autophagic flux and stimulated Akt and FoxO phosphorylation. These ï¬ndings suggest that CGRP in vivo acutely suppresses autophagy in the heart of fed and fasted mice, most likely through the activation of PKA/mTORC1 signaling but independent of Akt.
Assuntos
Autofagia/efeitos dos fármacos , Peptídeo Relacionado com Gene de Calcitonina/fisiologia , Coração/efeitos dos fármacos , Animais , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Camundongos , Miócitos Cardíacos/efeitos dos fármacos , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacosRESUMO
Skeletal muscle has the intrinsic ability to self-repair through a multifactorial process, but many aspects of its cellular and molecular mechanisms are not fully understood. There is increasing evidence that some members of the mammalian ß-galactoside-binding protein family (galectins) are involved in the muscular repair process (MRP), including galectin-3 (Gal-3). However, there are many questions about the role of this protein on muscle self-repair. Here, we demonstrate that endogenous Gal-3 is required for: (i) muscle repair in vivo by using a chloride-barium myolesion mouse model and (ii) mouse primary myoblasts myogenic programming. Injured muscle from Gal-3 knockout mice (GAL3KO) showed persistent inflammation associated with compromised muscle repair and the formation of fibrotic tissue on the lesion site. In GAL3KO mice, osteopontin expression remained high even after 7 and 14 d of the myolesion, while Myoblast differentiation transcription factor (MyoD) and myogenin had decreased their expression. In GAL3KO mouse primary myoblast cell culture, Paired Box 7 (Pax7) detection seems to sustain even when cells are stimulated to differentiation and MyoD expression is drastically reduced. The detection and temporal expression levels of these transcriptional factors appear to be altered in Gal-3-deficient myoblast. Gal-3 expression in wild-type mice for GAL3KO states, both in vivo and in vitro, in sarcoplasm/cytoplasm and myonuclei; as differentiation proceeds, Gal-3 expression is drastically reduced, and its location is confined to the sarcolemma/plasma cell membrane. We also observed a change in the temporal-spatial profile of Gal-3 expression and muscle transcription factors levels during the myolesion. Overall, these results demonstrate that endogenous Gal-3 is required for the skeletal muscle repair process.
Assuntos
Galectina 3/metabolismo , Músculo Esquelético/metabolismo , Animais , Compostos de Bário/administração & dosagem , Compostos de Bário/farmacologia , Cloretos/administração & dosagem , Cloretos/farmacologia , Galectina 3/deficiência , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/patologiaRESUMO
Although we have shown that catecholamines suppress the activity of the Ubiquitin-Proteasome System (UPS) and atrophy-related genes expression through a cAMP-dependent manner in skeletal muscle from rodents, the underlying mechanisms remain unclear. Here, we report that a single injection of norepinephrine (NE; 1 mg kg-1 ; s.c) attenuated the fasting-induced up-regulation of FoxO-target genes in tibialis anterior (TA) muscles by the stimulation of PKA/CREB and Akt/FoxO1 signaling pathways. In addition, muscle-specific activation of PKA by the overexpression of PKA catalytic subunit (PKAcat) suppressed FoxO reporter activity induced by (1) a wild-type; (2) a non-phosphorylatable; (3) a non-phosphorylatable and non-acetylatable forms of FoxO1 and FoxO3; (4) downregulation of FoxO protein content, and probably by (5) PGC-1α up-regulation. Consistently, the overexpression of the PKAcat inhibitor (PKI) up-regulated FoxO activity and the content of Atrogin-1 and MuRF1, as well as induced muscle fiber atrophy, the latter effect being prevented by the overexpression of a dominant negative (d. n.) form of FoxO (d.n.FoxO). The sustained overexpression of PKAcat induced fiber-type transition toward a smaller, slower, and more oxidative phenotype and improved muscle resistance to fatigue. Taken together, our data provide the first evidence that endogenous PKA activity is required to restrain the basal activity of FoxO and physiologically important to maintain skeletal muscle mass.
Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Proteína Forkhead Box O1/metabolismo , Músculo Esquelético/enzimologia , Atrofia Muscular/metabolismo , Animais , Linhagem Celular , Proteína Forkhead Box O3/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/patologia , Mioblastos Esqueléticos/enzimologia , Transdução de SinaisRESUMO
Surface Electromyography (sEMG) signal processing has a disruptive technology potential to enable a natural human interface with artificial limbs and assistive devices. However, this biosignal real-time control interface still presents several restrictions such as control limitations due to a lack of reliable signal prediction and standards for signal processing among research groups. Our paper aims to present and validate our sEMG database through the signal classification performed by the reliable forms of our Extreme Learning Machines (ELM) classifiers, used to maintain a more consistent signal classification. To perform the signal processing, we explore the use of a stochastic filter based on the Antonyan Vardan Transform (AVT) in combination with two variations of our Reliable classifiers (denoted R-ELM and R-Regularized ELM (RELM), respectively), to derive a reliability metric from the system, which autonomously selects the most reliable samples for the signal classification. To validate and compare our database and classifiers with related papers, we performed the classification of the whole of Databases 1, 2, and 6 (DB1, DB2, and DB6) of the NINAProdatabase. Our database presented consistent results, while the reliable forms of ELM classifiers matched or outperformed related papers, reaching average accuracies higher than 99 % for the IEEdatabase, while average accuracies of 75 . 1 % , 79 . 77 % , and 69 . 83 % were achieved for NINAPro DB1, DB2, and DB6, respectively.
Assuntos
Membros Artificiais , Bases de Dados Factuais , Eletromiografia/tendências , Movimento/fisiologia , Adulto , Algoritmos , Amputados , Feminino , Humanos , Aprendizado de Máquina , Masculino , Redes Neurais de Computação , Reconhecimento Automatizado de Padrão , Processamento de Sinais Assistido por Computador , Máquina de Vetores de Suporte , Extremidade Superior/fisiopatologiaRESUMO
BACKGROUND: Stimulation of ß2 -adrenoceptors can promote muscle hypertrophy and fibre type shift, and it can counteract atrophy and weakness. The underlying mechanisms remain elusive. METHODS: Fed wild type (WT), 2-day fasted WT, muscle-specific insulin (INS) receptor (IR) knockout (M-IR-/- ), and MKR mice were studied with regard to acute effects of the ß2 -agonist formoterol (FOR) on protein metabolism and signalling events. MKR mice express a dominant negative IGF1 receptor, which blocks both INS/IGF1 signalling. All received one injection of FOR (300 µg kg-1 subcutaneously) or saline. Skeletal muscles and serum samples were analysed from 30 to 240 min. For the study of chronic effects of FOR on muscle plasticity and function as well as intracellular signalling pathways, fed WT and MKR mice were treated with formoterol (300 µg kg-1 day-1 ) for 30 days. RESULTS: In fed and fasted mice, one injection of FOR inhibited autophagosome formation (LC3-II content, 65%, P ≤ 0.05) that was paralleled by an increase in serum INS levels (4-fold to 25-fold, P ≤ 0.05) and the phosphorylation of Akt (4.4-fold to 6.5-fold, P ≤ 0.05) and ERK1/2 (50% to two-fold, P ≤ 0.05). This led to the suppression (40-70%, P ≤ 0.05) of the master regulators of atrophy, FoxOs, and the mRNA levels of their target genes. FOR enhanced (41%, P ≤ 0.05) protein synthesis only in fed condition and stimulated (4.4-fold to 35-fold, P ≤ 0.05) the prosynthetic Akt/mTOR/p70S6K pathway in both fed and fasted states. FOR effects on Akt signalling during fasting were blunted in both M-IR-/- and MKR mice. Inhibition of proteolysis markers by FOR was prevented only in MKR mice. Blockade of PI3K/Akt axis and mTORC1, but not ERK1/2, in fasted mice also suppressed the acute FOR effects on proteolysis and autophagy. Chronic stimulation of ß2 -adrenoceptors in fed WT mice increased body (11%, P ≤ 0.05) and muscle (15%, P ≤ 0.05) growth and downregulated atrophy-related genes (30-40%, P ≤ 0.05), but these effects were abolished in MKR mice. Increases in muscle force caused by FOR (WT, 24%, P ≤ 0.05) were only partially impaired in MKR mice (12%, P ≤ 0.05), and FOR-induced slow-to-fast fibre type shift was not blocked at all in these animals. In MKR mice, FOR also restored the lower levels of muscle SDH activity to basal WT values and caused a marked reduction (57%, P ≤ 0.05) in the number of centrally nucleated fibers. CONCLUSIONS: NS/IGF1 signalling is necessary for the anti-proteolytic and hypertrophic effects of in vivo ß2 -adrenergic stimulation and appears to mediate FOR-induced enhancement of protein synthesis. INS/IGF1 signalling only partially contributes to gain in strength and does not mediate fibre type transition induced by FOR.
Assuntos
Agonistas de Receptores Adrenérgicos beta 2/farmacologia , Fator de Crescimento Insulin-Like I/metabolismo , Insulina/metabolismo , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Proteostase/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Autofagia/efeitos dos fármacos , Lisossomos/metabolismo , Masculino , Camundongos , Camundongos Knockout , Proteínas Musculares/metabolismo , Força Muscular , Músculo Esquelético/fisiopatologia , Fosfatidilinositol 3-Quinases , Proteólise , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Hypertension causes cardiac hypertrophy, one of the most important risk factors for heart failure (HF). Despite the importance of cardiac hypertrophy as a risk factor for the development of HF, not all hypertrophied hearts will ultimately fail. Alterations of cytoskeletal and sarcolemma-associated proteins are considered markers cardiac remodeling during HF. Dystrophin provides mechanical stability to the plasma membrane through its interactions with the actin cytoskeleton and, indirectly, to extracellular matrix proteins. This study was undertaken to evaluate dystrophin and calpain-1 in the transition from compensated cardiac hypertrophy to HF. Wistar rats were subjected to abdominal aorta constriction and killed at 30, 60 and 90 days post surgery (dps). Cardiac function and blood pressure were evaluated. The hearts were collected and Western blotting and immunofluorescence performed for dystrophin, calpain-1, alpha-fodrin and calpastatin. Statistical analyses were performed and considered significant when p<0.05. After 90 dps, 70% of the animals showed hypertrophic hearts (HH) and 30% hypertrophic+dilated hearts (HD). Systolic and diastolic functions were preserved at 30 and 60 dps, however, decreased in the HD group. Blood pressure, cardiomyocyte diameter and collagen content were increased at all time points. Dystrophin expression was lightly increased at 30 and 60 dps and HH group. HD group showed decreased expression of dystrophin and calpastatin and increased expression of calpain-1 and alpha-fodrin fragments. The first signals of dystrophin reduction were observed as early as 60 dps. In conclusion, some hearts present a distinct molecular pattern at an early stage of the disease; this pattern could provide an opportunity to identify these failure-prone hearts during the development of the cardiac disease. We showed that decreased expression of dystrophin and increased expression of calpains are coincident and could work as possible therapeutic targets to prevent heart failure as a consequence of cardiac hypertrophy.
Assuntos
Biomarcadores/metabolismo , Cardiomegalia/metabolismo , Distrofina/metabolismo , Animais , Pressão Sanguínea , Western Blotting , Cardiomegalia/diagnóstico por imagem , Cardiomegalia/fisiopatologia , Ecocardiografia , Imunofluorescência , Masculino , Ratos , Ratos WistarRESUMO
Although it is well known that chronic hypoxia induces muscle wasting, the effects of intermittent hypoxia on skeletal muscle protein metabolism remain unclear. We hypothesized that acute intermittent hypoxia (AIH), a challenge that activates the hypothalamic-pituitary-adrenal axis, would alter muscle protein homeostasis through a glucocorticoid-dependent mechanism. Three-week-old rats were submitted to adrenalectomy (ADX) and exposed to 8 h of AIH (6% O2 for 40 s at 9-min intervals). Animals were euthanized, and the soleus and extensor digitorum longus (EDL) muscles were harvested and incubated in vitro for measurements of protein turnover. AIH increased plasma levels of corticosterone and induced insulin resistance as estimated by the insulin tolerance test and lower rates of muscle glucose oxidation and the HOMA index. In both soleus and EDL muscles, rates of overall proteolysis increased after AIH. This rise was accompanied by an increased proteolytic activities of the ubiquitin(Ub)-proteasome system (UPS) and lysosomal and Ca2+-dependent pathways. Furthermore, AIH increased Ub-protein conjugates and gene expression of atrogin-1 and MuRF-1, two key Ub-protein ligases involved in muscle atrophy. In parallel, AIH increased the mRNA expression of the autophagy-related genes LC3b and GABARAPl1. In vitro rates of protein synthesis in skeletal muscles did not differ between AIH and control rats. ADX completely blocked the insulin resistance in hypoxic rats and the AIH-induced activation of proteolytic pathways and atrogene expression in both soleus and EDL muscles. These results demonstrate that AIH induces insulin resistance in association with activation of the UPS, the autophagic-lysosomal process, and Ca2+-dependent proteolysis through a glucocorticoid-dependent mechanism.NEW & NOTEWORTHY Since hypoxia is a condition in which the body is deprived of adequate oxygen supply and muscle wasting is induced, the present work provides evidence linking hypoxia to proteolysis through a glucocorticoid-dependent mechanism. We show that the activation of proteolytic pathways, atrophy-related genes, and insulin resistance in rats exposed to acute intermittent hypoxia was abolished by surgical removal of adrenal gland. This finding will be helpful for understanding of the muscle wasting in hypoxemic conditions.
Assuntos
Glucocorticoides/metabolismo , Hipóxia/fisiopatologia , Músculo Esquelético/fisiopatologia , Animais , Cálcio/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Sistema Hipotálamo-Hipofisário/fisiopatologia , Hipóxia/metabolismo , Resistência à Insulina/fisiologia , Lisossomos/metabolismo , Lisossomos/fisiologia , Masculino , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Atrofia Muscular/fisiopatologia , Sistema Hipófise-Suprarrenal/metabolismo , Sistema Hipófise-Suprarrenal/fisiopatologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Complexo de Endopeptidases do Proteassoma/fisiologia , Proteólise , Ratos , Ratos Wistar , Ubiquitina/metabolismoRESUMO
Calcitonin gene-related peptide (CGRP) is a neuropeptide released by motor neuron in skeletal muscle and modulates the neuromuscular transmission by induction of synthesis and insertion of acetylcholine receptor on postsynaptic muscle membrane; however, its role in skeletal muscle protein metabolism remains unclear. We examined the in vitro and in vivo effects of CGRP on protein breakdown and signaling pathways in control skeletal muscles and muscles following denervation (DEN) in rats. In isolated muscles, CGRP (10(-10) to 10(-6)M) reduced basal and DEN-induced activation of overall proteolysis in a concentration-dependent manner. The in vitro anti-proteolytic effect of CGRP was completely abolished by CGRP8-37, a CGRP receptor antagonist. CGRP down-regulated the lysosomal proteolysis, the mRNA levels of LC3b, Gabarapl1 and cathepsin L and the protein content of LC3-II in control and denervated muscles. In parallel, CGRP elevated cAMP levels, stimulated PKA/CREB signaling and increased Foxo1 phosphorylation in both conditions. In denervated muscles and starved C2C12 cells, Rp-8-Br-cAMPs or PKI, two PKA inhibitors, completely abolished the inhibitory effect of CGRP on Foxo1, 3 and 4 and LC3 lipidation. A single injection of CGRP (100 µg kg(-1)) in denervated rats increased the phosphorylation levels of CREB and Akt, inhibited Foxo transcriptional activity, the LC3 lipidation as well as the mRNA levels of LC3b and cathepsin L, two bona fide targets of Foxo. This study shows for the first time that CGRP exerts a direct inhibitory action on autophagic-lysosomal proteolysis in control and denervated skeletal muscle by recruiting cAMP/PKA signaling, effects that are related to inhibition of Foxo activity and LC3 lipidation.