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OBJECTIVE: The goal of this investigation was to evaluate circulating and skeletal muscle inflammatory biomarkers between maintenance hemodialysis (MHD) and demographic-matched control subjects (CON) before and after ingestion of a protein-rich meal. DESIGN AND METHODS: CON (n = 8; 50 ± 2 years; 31 ± 1 kg/m2) and MHD patients (n = 8; 56 ± 5 years; 32 ± 2 kg/m2) underwent a basal blood draw and muscle biopsy and serial blood draws after the ingestion of a mixed meal on a nondialysis day. Plasma advanced glycation end products (AGEs) and markers of oxidation were assessed via liquid chromatography-tandem mass spectrometry before and after the meal (+240 min). Circulating inflammatory cytokines and soluble receptors for AGE (sRAGE) isoforms (endogenous secretory RAGEs and cleaved RAGEs) were determined before and after the meal (+240 min). Basal muscle was probed for inflammatory cytokines and protein expression of related signaling components (RAGE, Toll-like receptor 4, oligosaccharyltransferase subunit 48, TIR-domain-containing adapter-inducing interferon-ß, total IκBα, and pIκBα). RESULTS: Basal circulating AGEs were 7- to 343-fold higher (P < .001) in MHD than those in CON, but only MG-H1 increased in CON after the meal (P < .001). There was a group effect (MHD > CON) for total sRAGEs (P = .02) and endogenous secretory RAGEs (P < .001) and a trend for cleaved RAGEs (P=.09), with no meal effect. In addition, there was a group effect (MHD < CON; P < .05) for circulating fractalkine, interleukin (IL)10, IL17A, and IL1ß and a trend (P < .10) for IL6 and macrophage inflammatory protein 1 alpha, whereas tumor necrosis factor alpha was higher in MHD (P < .001). In muscle, Toll-like receptor 4 (P = .03), TIR-domain-containing adapter-inducing interferon-ß (P = .002), and oligosaccharyltransferase subunit 48 (P = .02) expression was lower in MHD than that in CON, whereas IL6 was higher (P = .01) and IL8 (P = .08) tended to be higher in MHD. CONCLUSION: Overall, MHD exhibited an exaggerated, circulating, and skeletal muscle inflammatory biomarker environment, and the meal did not appreciably affect the inflammatory status.
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Productos Finales de Glicación Avanzada , Receptor Toll-Like 4 , Humanos , Productos Finales de Glicación Avanzada/metabolismo , Interleucina-6 , Biomarcadores , Interferón beta , Ingestión de AlimentosRESUMEN
Invasive fungal infections are an important cause of morbidity and mortality affecting primarily immunocompromised patients. While fungal identification to the species level is critical to providing appropriate therapy, it can be slow and laborious and often relies on subjective morphological criteria. The use of matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry has the potential to speed up and improve the accuracy of identification. In this multicenter study, we evaluated the accuracy of the Vitek MS v3.0 system in identifying 1,601 clinical mold isolates compared to identification by DNA sequence analysis and supported by morphological and phenotypic testing. Among the 1,519 isolates representing organisms in the v3.0 database, 91% (n = 1,387) were correctly identified to the species level. An additional 27 isolates (2%) were correctly identified to the genus level. Fifteen isolates were incorrectly identified, due to either a single incorrect identification (n = 13) or multiple identifications from different genera (n = 2). In those cases, when a single identification was provided that was not correct, the misidentification was within the same genus. The Vitek MS v3.0 was unable to identify 91 (6%) isolates, despite repeat testing. These isolates were distributed among all the genera. When considering all isolates tested, even those that were not represented in the database, the Vitek MS v3.0 provided a single correct identification 98% of the time. These findings demonstrate that the Vitek MS v3.0 system is highly accurate for the identification of common molds encountered in the clinical mycology laboratory.
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Pruebas Diagnósticas de Rutina/métodos , Hongos/aislamiento & purificación , Infecciones Fúngicas Invasoras/diagnóstico , Técnicas Microbiológicas/métodos , Bases de Datos Factuales , Errores Diagnósticos , Hongos/química , Hongos/clasificación , Humanos , Infecciones Fúngicas Invasoras/microbiología , Análisis de Secuencia de ADN , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
The accuracy and robustness of the Vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) system was evaluated by identifying mycobacteria from automated liquid-medium systems using patient samples. This is the first report to demonstrate that proteins within the liquid medium, its supplements, and decontamination reagents for nonsterile patient samples do not generate misidentification or false-positive results by use of the Vitek MS v3.0 system. Prior to testing with patient samples, a seeded-culture study was conducted to challenge the accuracy of the Vitek MS system at identifying mycobacteria from liquid medium by mimicking a clinical workflow. Seventy-seven Mycobacterium strains representing 21 species, seeded in simulated sputum, were decontaminated, inoculated into BacT/Alert MP liquid culture medium, incubated until positivity, and identified using the Vitek MS system. A total of 383 liquid cultures were tested, of which 379 (99%) were identified correctly to the species/complex/group level, 4 (1%) gave a "no-identification" result, and no misidentifications were observed. Following the simulated-sputum study, a total of 73 smear-positive liquid-medium cultures detected using BD BBL MGIT and VersaTREK Myco liquid media were identified by the Vitek MS system. Sixty-four cultures (87.7%) were correctly identified to the species/complex/group level; 7 (9.6%) resulted in no identification; and 2 (2.7%) were misidentified at the species level. These results indicate that the Vitek MS v3.0 system is an accurate tool for routine diagnostics of Mycobacterium species isolated from liquid cultures.
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Técnicas Bacteriológicas/métodos , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Mycobacterium/clasificación , Mycobacterium/aislamiento & purificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Técnicas Bacteriológicas/instrumentación , Medios de Cultivo , Pruebas Diagnósticas de Rutina , Humanos , Mycobacterium/química , Esputo/microbiologíaRESUMEN
Skeletal muscle insulin resistance is a hallmark of Type 2 diabetes (T2DM) and may be exacerbated by protein modifications by methylglyoxal (MG), known as dicarbonyl stress. The glyoxalase enzyme system composed of glyoxalase 1/2 (GLO1/GLO2) is the natural defense against dicarbonyl stress, yet its protein expression, activity, and regulation remain largely unexplored in skeletal muscle. Therefore, this study investigated dicarbonyl stress and the glyoxalase enzyme system in the skeletal muscle of subjects with T2DM (age: 56 ± 5 yr.; BMI: 32 ± 2 kg/m2) compared with lean healthy control subjects (LHC; age: 27 ± 1 yr.; BMI: 22 ± 1 kg/m2). Skeletal muscle biopsies obtained from the vastus lateralis at basal and insulin-stimulated states of the hyperinsulinemic (40 mU·m-2·min-1)-euglycemic (5 mM) clamp were analyzed for proteins related to dicarbonyl stress and glyoxalase biology. At baseline, T2DM had increased carbonyl stress and lower GLO1 protein expression (-78.8%), which inversely correlated with BMI, percent body fat, and HOMA-IR, while positively correlating with clamp-derived glucose disposal rates. T2DM also had lower NRF2 protein expression (-31.6%), which is a positive regulator of GLO1, while Keap1 protein expression, a negative regulator of GLO1, was elevated (207%). Additionally, insulin stimulation during the clamp had a differential effect on NRF2, Keap1, and MG-modified protein expression. These data suggest that dicarbonyl stress and the glyoxalase enzyme system are dysregulated in T2DM skeletal muscle and may underlie skeletal muscle insulin resistance. Whether these phenotypic differences contribute to the development of T2DM warrants further investigation.
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Diabetes Mellitus Tipo 2/enzimología , Resistencia a la Insulina , Lactoilglutatión Liasa/metabolismo , Carbonilación Proteica , Músculo Cuádriceps/enzimología , Adulto , Aldehído Reductasa/metabolismo , Glucemia/metabolismo , Estudios de Casos y Controles , Femenino , Gliceraldehído-3-Fosfato Deshidrogenasas/metabolismo , Humanos , Insulina/sangre , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Masculino , Persona de Mediana Edad , Factor 2 Relacionado con NF-E2/metabolismo , Triosa-Fosfato Isomerasa/metabolismoRESUMEN
Although there is evidence for metabolic dysfunction and chronic inflammation in Alzheimer's disease (AD), circulating levels of soluble receptor for advanced glycation end products (sRAGE) and the receptor for advanced glycation end products (RAGE) ligand S100B have not been characterized. sRAGE is an important mediator in disease as it can act as a ligand decoy for RAGE and attenuate downstream inflammatory signaling. Cognitively healthy elderly and AD participants with and without type 2 diabetes (n = 135) were stratified according to the clinical dementia rating (CDR; 0 = normal cognition (NC); ≥0.5 = AD). Total serum sRAGE, endogenous secretory RAGE (esRAGE), and S100B were assayed via ELISAs, and cleaved RAGE (cRAGE) and the cRAGE : esRAGE ratio were calculated. cRAGE : esRAGE was lower in AD compared to NC (p < 0.05). Metabolic substratifications were used to investigate the factors that influence sRAGE pathology in AD. Stratification by BMI classification, median fat mass, median HOMA-IR, median insulin, and median amylin were all metabolic or anthropometric factors which significantly interacted with sRAGE profiles within AD subjects. There were no significant differences in serum S100B between groups. These characterizations of sRAGE contribute evidence to the link between impaired metabolism and cognitive decline due to AD.
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Enfermedad de Alzheimer/metabolismo , Isoformas de Proteínas/metabolismo , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Anciano , Femenino , Productos Finales de Glicación Avanzada/metabolismo , Humanos , Insulina/metabolismo , Masculino , Subunidad beta de la Proteína de Unión al Calcio S100/metabolismoRESUMEN
The soluble receptor for advanced glycation end products (sRAGE) may be protective against inflammation associated with obesity and type 2 diabetes (T2DM). The aim of this study was to determine the distribution of sRAGE isoforms and whether sRAGE isoforms are associated with risk of T2DM development in subjects spanning the glucose tolerance continuum. In this retrospective analysis, circulating total sRAGE and endogenous secretory RAGE (esRAGE) were quantified via ELISA, and cleaved RAGE (cRAGE) was calculated in 274 individuals stratified by glucose tolerance status (GTS) and obesity. Group differences were probed by ANOVA, and multivariate ordinal logistic regression was used to test the association between sRAGE isoform concentrations and the proportional odds of developing diabetes, vs. normal glucose tolerance (NGT) or impaired glucose tolerance (IGT). When stratified by GTS, total sRAGE, cRAGE, and esRAGE were all lower with IGT and T2DM, while the ratio of cRAGE to esRAGE (cRAGE:esRAGE) was only lower (P < 0.01) with T2DM compared with NGT. When stratified by GTS and obesity, cRAGE:esRAGE was higher with obesity and lower with IGT (P < 0.0001) compared with lean, NGT. In ordinal logistic regression models, greater total sRAGE (odds ratio, 0.91; P < 0.01) and cRAGE (odds ratio, 0.84; P < 0.01) were associated with lower proportional odds of developing T2DM. Reduced values of sRAGE isoforms observed with both obesity and IGT are independently associated with greater proportional odds of developing T2DM. The mechanisms by which each respective isoform contributes to obesity and insulin resistance may reveal novel treatment strategies for diabetes.
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Diabetes Mellitus Tipo 2/sangre , Intolerancia a la Glucosa/sangre , Obesidad/sangre , Receptor para Productos Finales de Glicación Avanzada/sangre , Adolescente , Adulto , Anciano , Envejecimiento/metabolismo , Glucemia/análisis , Índice de Masa Corporal , Diabetes Mellitus Tipo 2/complicaciones , Progresión de la Enfermedad , Femenino , Intolerancia a la Glucosa/complicaciones , Prueba de Tolerancia a la Glucosa , Hemoglobina Glucada/análisis , Humanos , Isomerismo , Masculino , Persona de Mediana Edad , Obesidad/complicaciones , Sobrepeso/sangre , Sobrepeso/complicaciones , Receptor para Productos Finales de Glicación Avanzada/química , Estudios Retrospectivos , Adulto JovenRESUMEN
Glyoxalase I (GLO1) is the primary enzyme for detoxification of the reactive dicarbonyl methylglyoxal (MG). Loss of GLO1 promotes accumulation of MG resulting in a recapitulation of diabetic phenotypes. We previously demonstrated attenuated GLO1 protein in skeletal muscle from individuals with type 2 diabetes (T2D). However, whether GLO1 attenuation occurs prior to T2D and the mechanisms regulating GLO1 abundance in skeletal muscle are unknown. GLO1 expression and activity were determined in skeletal muscle tissue biopsies from 15 lean healthy individuals (LH, BMI: 22.4 ± 0.7) and 5 individuals with obesity (OB, BMI: 32.4 ± 1.3). GLO1 protein was attenuated by 26 ± 0.3 % in OB compared to LH skeletal muscle (p = 0.019). Similar reductions for GLO1 activity were observed (p = 0.102). NRF2 and Keap1 expression were equivocal between groups despite a 2-fold elevation in GLO1 transcripts in OB skeletal muscle (p = 0.008). GLO1 knock-down (KD) in human immortalized myotubes promoted downregulation of muscle contraction and organization proteins indicating the importance of GLO1 expression for skeletal muscle function. SIRT1 KD had no effect on GLO1 protein or activity whereas, SIRT2 KD attenuated GLO1 protein by 28 ± 0.29 % (p < 0.0001) and GLO1 activity by 42 ± 0.12 % (p = 0.0150). KD of NAMPT also resulted in attenuation of GLO1 protein (28 ± 0.069 %, p = 0.003), activity (67 ± 0.09 %, p = 0.011) and transcripts (50 ± 0.13 %, p = 0.049). Neither the provision of the NAD+ precursors NR nor NMN were able to prevent this attenuation in GLO1 protein. However, NR did augment GLO1 specific activity (p = 0.022 vs NAMPT KD). These perturbations did not alter GLO1 acetylation status. SIRT1, SIRT2 and NAMPT protein levels were all equivocal in skeletal muscle tissue biopsies from individuals with obesity and lean individuals. These data implicate NAD+-dependent regulation of GLO1 in skeletal muscle independent of altered GLO1 acetylation and provide rationale for exploring NR supplementation to rescue attenuated GLO1 abundance and activity in conditions such as obesity.
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Citocinas , Lactoilglutatión Liasa , Músculo Esquelético , Nicotinamida Fosforribosiltransferasa , Obesidad , Sirtuina 2 , Humanos , Músculo Esquelético/metabolismo , Lactoilglutatión Liasa/metabolismo , Lactoilglutatión Liasa/genética , Nicotinamida Fosforribosiltransferasa/metabolismo , Nicotinamida Fosforribosiltransferasa/genética , Sirtuina 2/metabolismo , Sirtuina 2/genética , Citocinas/metabolismo , Masculino , Obesidad/metabolismo , Obesidad/genética , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Femenino , Adulto , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Proteína 1 Asociada A ECH Tipo Kelch/genética , Regulación de la Expresión Génica , Persona de Mediana Edad , Sirtuina 1/metabolismo , Sirtuina 1/genéticaRESUMEN
Breakthrough fungal infections in patients on antimicrobial prophylaxis during allogeneic hematopoietic cell transplantation (allo-HCT) represent a major and often unexplained cause of morbidity and mortality. Candida parapsilosis is a common cause of invasive candidiasis and has been classified as a high-priority fungal pathogen by the World Health Organization. In high-risk allo-HCT recipients on micafungin prophylaxis, we show that heteroresistance (the presence of a phenotypically unstable, low-frequency subpopulation of resistant cells (~1 in 10,000)) underlies breakthrough bloodstream infections by C. parapsilosis. By analyzing 219 clinical isolates from North America, Europe and Asia, we demonstrate widespread micafungin heteroresistance in C. parapsilosis. Standard antimicrobial susceptibility tests, such as broth microdilution or gradient diffusion assays, which guide drug selection for invasive infections, fail to detect micafungin heteroresistance in C. parapsilosis. To facilitate rapid detection of micafungin heteroresistance in C. parapsilosis, we constructed a predictive machine learning framework that classifies isolates as heteroresistant or susceptible using a maximum of ten genomic features. These results connect heteroresistance to unexplained antifungal prophylaxis failure in allo-HCT recipients and demonstrate a proof-of-principle diagnostic approach with the potential to guide clinical decisions and improve patient care.
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Obesity prevalence in the US has nearly tripled since 1975 and a parallel increase in prevalence of type 2 diabetes (T2D). Obesity promotes a myriad of metabolic derangements with insulin resistance (IR) being perhaps the most responsible for the development of T2D and other related diseases such as cardiovascular disease. The precarious nature of IR development is such that it provides a valuable target for the prevention of further disease development. However, the mechanisms driving IR are numerous and complex making the development of viable interventions difficult. The development of metabolic derangement in the context of obesity promotes accumulation of reactive metabolites such as the reactive alpha-dicarbonyl methylglyoxal (MG). MG accumulation has long been appreciated as a marker of disease progression in patients with T2D as well as the development of diabetic complications. However, recent evidence suggests that the accumulation of MG occurs with obesity prior to T2D onset and may be a primary driving factor for the development of IR and T2D. Further, emerging evidence also suggests that this accumulation of MG with obesity may be a result in a loss of MG detoxifying capacity of glyoxalase I. In this review, we will discuss the evidence that posits MG accumulation because of GLO1 attenuation is a novel target mechanism of the development of metabolic derangement. In addition, we will also explore the regulation of GLO1 and the strategies that have been investigated so far to target GLO1 regulation for the prevention and treatment of metabolic derangement.
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Significance: Accumulation of reactive oxygen species (ROS) is known to promote cellular damage in multiple cell types. In skeletal muscle, ROS has been implicated in disuse-induced muscle atrophy. However, the molecular origin and mechanism of how disuse promotes ROS and muscle dysfunction remains unclear. Recent Advances: Recently, we implicated membrane lipids of mitochondria to be a potential source of ROS to promote muscle atrophy. Critical Issues: In this review, we discuss evidence that changes in mitochondrial lipids represent a physiologically relevant process by which disuse promotes mitochondrial electron leak and oxidative stress. Future Directions: We further discuss lipid hydroperoxides as a potential downstream mediator of ROS to induce muscle atrophy. Antioxid. Redox Signal. 38, 338-351.
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Estrés Oxidativo , Fosfolípidos , Humanos , Especies Reactivas de Oxígeno/metabolismo , Fosfolípidos/metabolismo , Estrés Oxidativo/fisiología , Mitocondrias/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismoRESUMEN
BMI-matched normal- (NGT, n = 10, 41 ± 4y, 35.6 ± 3.0 kg/m2 ) and abnormal-glucose-tolerant (AGT, n = 16, 51 ± 3y, 34.3 ± 1.5 kg/m2 ) participants were evaluated for body composition, metabolic health (oral glucose tolerance test [OGTT]), and VO2 max. Participants also completed a treadmill walking test at 65% VO2 max for 30 min. Total sRAGE, esRAGE, sTLR2, and sTLR4 were assessed via ELISA, and cRAGE was calculated. AGT exhibited greater (p < 0.05) body fat % (+24%), fasting plasma glucose (+37%), OGTT AUC (+59%), and HOMA-IR (+55%) and lower (p < 0.05) VO2 max (-24%). sTLR2 was 33% lower in AGT than NGT (main effect, p = 0.034). However, sTLR2 did not change (p > 0.05) following AE. sTLR4 tended to be 36% lower in AGT than NGT (main effect, p = 0.096) and did not change following AE (p > 0.05). Total sRAGE and isoforms were similar (p > 0.05) between groups and did not change following AE (p > 0.05). sTLR2 was correlated with (p < 0.05) basal BG (r = -0.505) and OGTT AUC (r = -0.687). sTLR4 was correlated with basal BG (p < 0.10, r = -0.374) and OGTT AUC (p < 0.05, r = -0.402). Linear regressions were predictive of sTLRs in the basal state (sTLR2: R2 = 0.641, p = 0.01; sTLR4: R2 = 0.566, p = 0.037) and after acute exercise state (sTLR2: R2 = 0.681, p = 0.004, sTLR4: R2 = 0.568, p = 0.036).These findings show circulating sTLR profiles are disrupted in AGT and acute AE minimally modulates their levels.
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Tejido Adiposo , Composición Corporal , Humanos , Prueba de Tolerancia a la Glucosa , Tejido Adiposo/metabolismo , Glucosa/metabolismo , Ejercicio Físico , Glucemia/metabolismoRESUMEN
Nearly 40% of Americans have obesity and are at increased risk for developing type 2 diabetes. Skeletal muscle is responsible for >80% of insulin-stimulated glucose uptake that is attenuated by the inflammatory milieu of obesity and augmented by aerobic exercise. The receptor for advanced glycation endproducts (RAGE) is an inflammatory receptor directly linking metabolic dysfunction with inflammation. Circulating soluble isoforms of RAGE (sRAGE) formed either by proteolytic cleavage (cRAGE) or alternative splicing (esRAGE) act as decoys for RAGE ligands, thereby counteracting RAGE-mediated inflammation. We aimed to determine if RAGE expression or alternative splicing of RAGE is altered by obesity in muscle, and whether acute aerobic exercise (AE) modifies RAGE and sRAGE. Young (20-34 yr) participants without [n = 17; body mass index (BMI): 22.6 ± 2.6 kg/m2] and with obesity (n = 7; BMI: 32.8 ± 2.9 kg/m2) performed acute aerobic exercise (AE) at 40%, 65%, or 80% of maximal aerobic capacity (VÌo2max; mL/kg/min) on separate visits. Blood was taken before and 30 min after each AE bout. Muscle biopsy samples were taken before, 30 min, and 3 h after the 80% VÌo2max AE bout. Individuals with obesity had higher total RAGE and esRAGE mRNA and RAGE protein (P < 0.0001). In addition, RAGE and esRAGE transcripts correlated to transcripts of the NF-κB subunit P65 (P < 0.05). There was no effect of AE on total RAGE or esRAGE transcripts, or RAGE protein (P > 0.05), and AE tended to decrease circulating sRAGE in particular at lower intensities of exercise. RAGE expression is exacerbated in skeletal muscle with obesity, which may contribute to muscle inflammation via NF-κB. Future work should investigate the consequences of increased skeletal muscle RAGE on the development of obesity-related metabolic dysfunction and potential mitigating strategies.NEW & NOTEWORTHY This study is the first to investigate the effects of aerobic exercise intensity on circulating sRAGE isoforms, muscle RAGE protein, and muscle RAGE splicing. sRAGE isoforms tended to diminish with exercise, although this effect was attenuated with increasing exercise intensity. Muscle RAGE protein and gene expression were unaffected by exercise. However, individuals with obesity displayed nearly twofold higher muscle RAGE protein and gene expression, which positively correlated with expression of the P65 subunit of NF-κB.
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Diabetes Mellitus Tipo 2 , Humanos , Adulto Joven , Ejercicio Físico , Inflamación , Músculo Esquelético , FN-kappa B , Receptor para Productos Finales de Glicación AvanzadaRESUMEN
The benefits of exercise involve skeletal muscle redox state alterations of nicotinamide adenine dinucleotide (NAD) and flavin adenine dinucleotide (FAD). We determined the fiber-specific effects of acute exercise on the skeletal muscle redox state in healthy adults. Muscle biopsies were obtained from 19 participants (11 M, 8 F; 26 ± 4 yr) at baseline (fasted) and 30 min and 3 h after treadmill exercise at 80% maximal oxygen consumption (VÌo2max). Muscle samples were probed for autofluorescence of NADH (excitation at 340-360 nm) and oxidized flavoproteins (Fp; excitation at 440-470 nm) and subsequently, fiber typed to quantify the redox signatures of individual muscle fibers. Redox state was calculated as the oxidation-to-reduction redox ratio: Fp/(Fp + NADH). At baseline, pair-wise comparisons revealed that the redox ratio of myosin heavy chain (MHC) I fibers was 7.2% higher than MHC IIa (P = 0.023, 95% CI: 5.2, 9.2%) and the redox ratio of MHC IIa was 8.0% higher than MHC IIx (P = 0.035, 95% CI: 6.8, 9.2%). MHC I fibers also displayed greater NADH intensity than MHC IIx (P = 0.007) and greater Fp intensity than both MHC IIa (P = 0.019) and MHC IIx (P < 0.0001). Fp intensities increased in all fiber types (main effect, P = 0.039) but redox ratios did not change (main effect, P = 0.483) 30 min after exercise. The change in redox ratio was positively correlated with capillary density in MHC I (rho = 0.762, P = 0.037), MHC IIa fibers (rho = 0.881, P = 0.007), and modestly in MHC IIx fibers (rho = 0. 771, P = 0.103). These findings support the use of redox autofluorescence to interrogate skeletal muscle metabolism.NEW & NOTEWORTHY This study is the first to use autofluorescent imaging to describe differential redox states within human skeletal muscle fiber types with exercise. Our findings highlight an easy and efficacious technique for assessing skeletal muscle redox in humans.
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Músculo Esquelético , NAD , Adulto , Humanos , NAD/metabolismo , Músculo Esquelético/fisiología , Fibras Musculares Esqueléticas/metabolismo , Ejercicio Físico/fisiología , Cadenas Pesadas de Miosina/metabolismo , Oxidación-ReducciónRESUMEN
Reactive oxygen species (ROS) accumulation is a cardinal feature of skeletal muscle atrophy. ROS refers to a collection of radical molecules whose cellular signals are vast, and it is unclear which downstream consequences of ROS are responsible for the loss of muscle mass and strength. Here, we show that lipid hydroperoxides (LOOH) are increased with age and disuse, and the accumulation of LOOH by deletion of glutathione peroxidase 4 (GPx4) is sufficient to augment muscle atrophy. LOOH promoted atrophy in a lysosomal-dependent, proteasomal-independent manner. In young and old mice, genetic and pharmacological neutralization of LOOH or their secondary reactive lipid aldehydes robustly prevented muscle atrophy and weakness, indicating that LOOH-derived carbonyl stress mediates age- and disuse-induced muscle dysfunction. Our findings provide novel insights for the role of LOOH in sarcopenia including a therapeutic implication by pharmacological suppression.
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Sarcopenia , Ratones , Animales , Sarcopenia/patología , Peróxidos Lipídicos/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Atrofia Muscular/metabolismo , Atrofia Muscular/patología , Músculo Esquelético/metabolismo , Estrés OxidativoRESUMEN
Background: Cathepsin B (CTSB) and brain derived neurotrophic factor (BDNF) are increased with aerobic exercise (AE) and skeletal muscle has been identified as a potential source of secretion. However, the intensity of AE and the potential for skeletal muscle contributions to circulating CTSB and BDNF have not been fully studied in humans. Objective: Determine the effects of AE intensity on circulating and skeletal muscle CTSB and BDNF expression profiles. Methods: Young healthy subjects (nâ=â16) completed treadmill-based AE consisting of VO2max and calorie-matched acute AE sessions at 40%, 65% and 80% VO2max. Fasting serum was obtained before and 30-minutes after each bout of exercise. Skeletal muscle biopsies (vastus lateralis) were taken before, 30-minutes and 3-hours after the 80% bout. Circulating CTSB and BDNF were assayed in serum. CTSB protein, BDNF protein and mRNA expression were measured in skeletal muscle tissue. Results: Serum CTSB increased by 20±7% (pâ=â0.02) and 30±18% (pâ=â0.04) after 80% and VO2max AE bouts, respectively. Serum BDNF showed a small non-significant increase (6±3%; pâ=â0.09) after VO2max. In skeletal muscle tissue, proCTSB increased 3âh-post AE (87±26%; pâ<â0.01) with no change in CTSB gene expression. Mature BDNF protein decreased (31±35%; pâ=â0.03) while mRNA expression increased (131±41%; pâ<â0.01) 3âh-post AE. Skeletal muscle fiber typing revealed that type IIa and IIx fibers display greater BDNF expression compared to type I (pâ=â0.02 and pâ<â0.01, respectively). Conclusions: High intensity AE elicits greater increases in circulating CTSB compared with lower intensities. Skeletal muscle protein and gene expression corroborate the potential role of skeletal muscle in generating and releasing neuroprotective exerkines into the circulation.NEW AND NOTEWORTHY: 1) CTSB is enriched in the circulation in an aerobic exercise intensity dependent manner. 2) Skeletal muscle tissue expresses both message and protein of CTSB and BDNF. 3) BDNF is highly expressed in glycolytic skeletal muscle fibers.
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Thioredoxin-interacting protein (TXNIP) negatively effects the redox state and growth signaling via its interactions with thioredoxin (TRX) and regulated in development and DNA damage response 1 (REDD1), respectively. TXNIP expression is downregulated by pathways activated during aerobic exercise (AE), via posttranslational modifications (PTMs; serine phosphorylation and ubiquitination). The purpose of this investigation was to determine the effects of acute AE on TXNIP expression, posttranslational modifications, and its interacting partners, REDD1 and TRX. Fifteen healthy adults performed 30 min of aerobic exercise (80% VÌo2max) with muscle biopsies taken before, immediately following, and 3 h following the exercise bout. To explore potential mechanisms underlying our in vivo findings, primary human myotubes were exposed to two models of exercise, electrical pulse stimulation (EPS) and palmitate-forskolin-ionomycin (PFI). Immediately following exercise, TXNIP protein decreased, but returned to preexercise levels 3 h after exercise. These results were replicated in our PFI exercise model only. Although not statistically significant, there was a trending main effect in serine-phosphorylation status of TXNIP (P = 0.07) immediately following exercise. REDD1 protein decreased 3 h after exercise. AE had no effect on TRX protein expression, gene expression, or the activity of its reducing enzyme, thioredoxin reductase. Consequently, AE had no effect on the TRX: TXNIP interaction. Our results indicate that AE leads to acute reductions in TXNIP and REDD1 protein expression. However, these changes did not result in alterations in the TRX: TXNIP interaction and could not be entirely explained by alterations in TXNIP PTMs or changes in TRX expression or activity.NEW & NOTEWORTHY Aerobic exercise is an effective tool in the prevention and treatment of several chronic metabolic diseases. However, the mechanisms through which these benefits are conferred have yet to be fully elucidated. Our data reveal a novel effect of aerobic exercise on reducing the protein expression of molecular targets that negatively impact redox and insulin/growth signaling in skeletal muscle. These findings contribute to the expanding repository of molecular signatures provoked by aerobic exercise.
Asunto(s)
Proteínas Portadoras , Ejercicio Físico , Músculo Esquelético , Factores de Transcripción/metabolismo , Proteínas Portadoras/metabolismo , Humanos , Insulina/metabolismo , Músculo Esquelético/metabolismo , Oxidación-Reducción , Transducción de SeñalRESUMEN
Candida metapsilosis is a member of the Candida parapsilosis species complex, a group of opportunistic human pathogens. Of all the members of this complex, C. metapsilosis is the least virulent, and accounts for a small proportion of invasive Candida infections. Previous studies established that all C. metapsilosis isolates are hybrids, originating from a single hybridization event between two lineages, parent A and parent B. Here, we use MinION and Illumina sequencing to characterize a C. metapsilosis isolate that originated from a separate hybridization. One of the parents of the new isolate is very closely related to parent A. However, the other parent (parent C) is not the same as parent B. Unlike C. metapsilosis AB isolates, the C. metapsilosis AC isolate has not undergone introgression at the mating type-like locus. In addition, the A and C haplotypes are not fully collinear. The C. metapsilosis AC isolate has undergone loss of heterozygosity with a preference for haplotype A, indicating that this isolate is in the early stages of genome stabilization.
Asunto(s)
Candida parapsilosis , Candidiasis , Antifúngicos , Candida/genética , Candida parapsilosis/genética , Candidiasis/genética , Genoma , Humanos , Hibridación GenéticaRESUMEN
Invasive fungal infections (IFI) remain a serious threat to immunocompromised hosts. Current diagnostic methods, including fungal culture and antigen detection, are slow and often lack specificity. Rapid diagnostic tools with increased sensitivity and specificity could improve the care of patients with IFI. Recently, Luminex Molecular Diagnostics (Toronto, Canada) developed 23 analyte-specific reagents (ASRs) for the detection of the most common clinically relevant fungi. This study's objective was to evaluate the sensitivity and specificity of a subset of these ASRs for fungal isolates and clinical specimens. Previously characterized fungal and bacterial isolates (n = 110), blood culture specimens (n = 34), and respiratory specimens (n = 44) were tested using either a Candida 7-plex panel (Candida albicans, Candida glabrata, Candida tropicalis, Candida parapsilosis, Candida lusitaniae, Candida guilliermondii, and Candida krusei) or a mold 11-plex panel (Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger, Aspergillus terreus, Scedosporium prolificans, Scedosporium apiospermum, Fusarium oxysporum/Fusarium solani, Rhizopus arrhizus, Rhizopus microsporus, Mucor indicus, and Cunninghamella bertholletiae). The Candida 7-plex panel correctly identified all Candida isolates as confirmed by fungal culture and biochemical tests, for a sensitivity and specificity of 100%. The mold 11-plex panel correctly identified all mold isolates tested except for A. niger. Fungal isolates of Rhizopus and Mucor species were not detected, either, although they could represent species other than those targeted by the ASRs. Further evaluation will be necessary to confirm the sensitivities of some of the mold ASRs. Implementation of these ASRs will allow same-day detection of fungal DNA in clinical specimens.
Asunto(s)
Química Analítica , Técnicas de Laboratorio Clínico/métodos , Hongos/química , Hongos/clasificación , Micología/métodos , Micosis/diagnóstico , Micosis/microbiología , Hongos/aislamiento & purificación , Humanos , Sensibilidad y EspecificidadRESUMEN
The intestinal microbiota is a complex community of bacteria, archaea, viruses, protists and fungi1,2. Although the composition of bacterial constituents has been linked to immune homeostasis and infectious susceptibility3-7, the role of non-bacterial constituents and cross-kingdom microbial interactions in these processes is poorly understood2,8. Fungi represent a major cause of infectious morbidity and mortality in immunocompromised individuals, although the relationship of intestinal fungi (that is, the mycobiota) with fungal bloodstream infections remains undefined9. We integrated an optimized bioinformatics pipeline with high-resolution mycobiota sequencing and comparative genomic analyses of fecal and blood specimens from recipients of allogeneic hematopoietic cell transplant. Patients with Candida bloodstream infection experienced a prior marked intestinal expansion of pathogenic Candida species; this expansion consisted of a complex dynamic between multiple species and subspecies with a stochastic translocation pattern into the bloodstream. The intestinal expansion of pathogenic Candida spp. was associated with a substantial loss in bacterial burden and diversity, particularly in the anaerobes. Thus, simultaneous analysis of intestinal fungi and bacteria identifies dysbiosis states across kingdoms that may promote fungal translocation and facilitate invasive disease. These findings support microbiota-driven approaches to identify patients at risk of fungal bloodstream infections for pre-emptive therapeutic intervention.