RESUMO
Acute skeletal muscle injury initiates a process of necrosis, debris clearance, and ultimately tissue regeneration via myogenesis. While skeletal muscle stem cells (MuSCs) are responsible for populating the proliferative myogenic progenitor pool to fuel muscle repair, recruited and resident immune cells have a central role in the regulation of muscle regeneration via the execution of phagocytosis and release of soluble factors that act directly on MuSCs to regulate myogenic differentiation. Therefore, the timing of MuSC proliferation and differentiation is closely linked to the populations and behaviors of immune cells present within skeletal muscle. This has important implications for aging and muscle repair, as systemic changes in immune system function contribute to a decline in muscle regenerative capacity. Here, we present adapted protocols for the isolation of mononuclear cells from skeletal muscles for the quantification of immune cell populations using flow cytometry. We also describe a cardiotoxin skeletal muscle injury protocol and detail the expected outcomes including immune cell infiltration to the injured sites and formation of new myocytes. As immune cell function is substantially influenced by aging, we extend these approaches and outcomes to aged mice.
Assuntos
Envelhecimento , Modelos Animais de Doenças , Músculo Esquelético , Regeneração , Animais , Camundongos , Músculo Esquelético/lesões , Músculo Esquelético/metabolismo , Envelhecimento/fisiologia , Desenvolvimento Muscular , Citometria de Fluxo/métodos , Diferenciação Celular , Proliferação de CélulasRESUMO
Rationale: MG53's known function in facilitating tissue repair and anti-inflammation has broad applications to regenerative medicine. There is controversy regarding MG53's role in the development of type 2 diabetes mellitus. Objective: This study aims to address this controversy - whether MG53's myokine function contributes to inhibition of insulin signaling in muscle, heart, and liver tissues. Study design: We determined the binding affinity of the recombinant human MG53 (rhMG53) to the insulin receptor extracellular domain (IR-ECD) and found low affinity of interaction with Kd (>480 nM). Using cultured C2C12 myotubes and HepG2 cells, we found no effect of rhMG53 on insulin-stimulated Akt phosphorylation (p-Akt). We performed in vivo assay with C57BL/6J mice subjected to insulin stimulation (1 U/kg, intraperitoneal injection) and observed no effect of rhMG53 on insulin-stimulated p-Akt in muscle, heart and liver tissues. Conclusion: Overall, our data suggest that rhMG53 can bind to the IR-ECD, however has a low likelihood of a physiologic role, as the Kd for binding is ~10,000 higher than the physiologic level of MG53 present in the serum of rodents and humans (~10 pM). Our findings question the notion proposed by Xiao and colleagues - whether targeting circulating MG53 opens a new therapeutic avenue for type 2 diabetes mellitus and its complications.
Assuntos
Insulina , Fígado , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt , Receptor de Insulina , Animais , Humanos , Camundongos , Fosforilação/efeitos dos fármacos , Receptor de Insulina/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fígado/metabolismo , Fígado/efeitos dos fármacos , Insulina/metabolismo , Insulina/farmacologia , Miocárdio/metabolismo , Células Hep G2 , Músculo Esquelético/metabolismo , Músculo Esquelético/efeitos dos fármacos , Masculino , Transdução de Sinais/efeitos dos fármacos , Diabetes Mellitus Tipo 2/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Citocinas/metabolismo , Proteínas de MembranaRESUMO
In persons with a spinal cord injury (SCI), resistance training using neuromuscular electrical stimulation (NMES-RT) increases lean mass in the lower limbs. However, whether protein supplementation in conjunction with NMES-RT further enhances this training effect is unknown. In this randomized controlled pilot trial, 15 individuals with chronic SCI engaged in 3 times/week NMES-RT, with (NMES+PRO, n = 8) or without protein supplementation (NMES, n = 7), for 12 weeks. Before and after the intervention, whole body and regional body composition (DXA) and fasting glucose and insulin concentrations were assessed in plasma. Adherence to the intervention components was ≥96%. Thigh lean mass was increased to a greater extent after NMES+PRO compared to NMES (0.3 (0.2, 0.4) kg; p < 0.001). Furthermore, fasting insulin concentration and Homeostatic Model Assessment for Insulin Resistance (HOMA-IR) were decreased similarly in both groups (fasting insulin: 1 [-9, 11] pmolâL-1; HOMA-IR: 0.1 [-0.3, 0.5] AU; both p ≥ 0.617). Twelve weeks of home-based NMES-RT increased thigh lean mass, an effect that was potentiated by protein supplementation. In combination with the excellent adherence and apparent improvement in cardiometabolic health outcomes, these findings support further investigation through a full-scale randomized controlled trial.
Assuntos
Composição Corporal , Terapia por Estimulação Elétrica , Treinamento Resistido , Traumatismos da Medula Espinal , Humanos , Traumatismos da Medula Espinal/terapia , Traumatismos da Medula Espinal/fisiopatologia , Traumatismos da Medula Espinal/reabilitação , Masculino , Treinamento Resistido/métodos , Feminino , Adulto , Projetos Piloto , Terapia por Estimulação Elétrica/métodos , Pessoa de Meia-Idade , Suplementos Nutricionais , Resistência à Insulina , Insulina/sangue , Proteínas Alimentares/administração & dosagem , Glicemia/metabolismo , Músculo Esquelético/metabolismoRESUMO
AIMS: Previous proteomics studies in dysferlinopathy muscle have been limited in scope, often utilizing 2D-electrophoresis and yielding only a small number of differential expression calls. To address this gap, this study aimed to employ high-resolution proteomics to explore the proteomic landscapes of dysferlinopathy and analyze the correlation between muscle pathological changes and alterations in protein expression in muscle biopsies. METHODS: We conducted a comprehensive approach to investigate the proteomic profile and disease-associated changes in the muscle tissue proteome from 15 patients with dysferlinopathy, exhibiting varying degrees of dystrophic pathology, alongside age-matched controls. Our methodology encompasses tandem mass tag (TMT)-labeled liquid chromatography-mass spectrometry (LC-MS/MS)-based proteomics, protein-protein interaction (PPI) network analysis, weighted gene co-expression network analysis, and differential expression analysis. Subsequently, we examined the correlation between the expression of key proteins and the clinical characteristics of the patients to identify pathogenic targets associated with DYSF mutations in dysferlinopathy. RESULTS: A total of 1600 differentially expressed proteins were identified, with 1321 showing high expression levels and 279 expressed at lower levels. Our investigation yields a molecular profile delineating the altered protein networks in dysferlinopathy-afflicted skeletal muscle, uncovering dysregulation across numerous cellular pathways and molecular processes, including mRNA metabolic processes, regulated exocytosis, immune response, muscle system processes, energy metabolic processes, and calcium transmembrane transport. Moreover, we observe significant associations between the protein expression of ANXA1, ANXA2, ANXA4, ANXA5, LMNA, PYGM, and the extent of histopathologic changes in muscle biopsies from patients with dysferlinopathy, validated through immunoblotting and immunofluorescence assays. CONCLUSIONS: Through the aggregation of expression data from dysferlinopathy-impacted muscles exhibiting a range of pathological alterations, we identified multiple key proteins associated with the dystrophic pathology of patients with dysferlinopathy. These findings provide novel insights into the pathogenesis of dysferlinopathy and propose promising targets for future therapeutic endeavors.
Assuntos
Biomarcadores , Progressão da Doença , Músculo Esquelético , Distrofia Muscular do Cíngulo dos Membros , Proteômica , Humanos , Masculino , Distrofia Muscular do Cíngulo dos Membros/patologia , Distrofia Muscular do Cíngulo dos Membros/genética , Distrofia Muscular do Cíngulo dos Membros/metabolismo , Feminino , Adulto , Adulto Jovem , Músculo Esquelético/patologia , Músculo Esquelético/metabolismo , Adolescente , Biomarcadores/metabolismo , Criança , Disferlina/genética , Disferlina/metabolismo , Pessoa de Meia-Idade , Pré-Escolar , Mapas de Interação de Proteínas , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Espectrometria de Massas em TandemRESUMO
PURPOSE: The effects of coffee ingestion on skeletal muscle microvascular function are not well understood. This study aimed to investigate the acute effects of coffee intake with varying levels of caffeine on skeletal muscle microvascular reactivity at rest and oxygen extraction during maximal incremental exercise in physically active individuals. METHODS: Twenty healthy young male participants were administered coffee with low caffeine (3 mg/kg body weight; LC), high caffeine (6 mg/kg body weight; HC), and placebo (decaf) in different sessions. Skeletal muscle reactivity indexes, including tissue saturation index 10s slope (TSI10) and TSI half time recovery (TSI ½) following 5-minute ischemia were measured at rest and were measured at baseline and post-coffee consumption using near-infrared spectroscopy (NIRS). Post-coffee intake, NIRS was also used to measure microvascular oxygen extraction during exercise via maximal incremental exercise. Peak oxygen consumption and peak power output (Wpeak) were simultaneously evaluated. RESULTS: Post-coffee consumption, TSI10 was significantly higher in the LC condition compared to placebo (p = 0.001) and significantly higher in the HC condition compared to placebo (p < 0.001). However, no difference was detected between LC and HC conditions (p = 0.527). HC condition also showed significant less TSI ½ compared to placebo (p = 0.005). However, no difference was detected for microvascular oxygen extraction during exercise, despite the greater Wpeak found for HC condition (p < 0.001) compared to placebo. CONCLUSION: Coffee ingestion with high caffeine level (6 mg/kg body weight) significantly enhanced skeletal muscle reactivity at rest. However, the improvement of exercise performance with coffee intake is not accompanied by alterations in muscle oxygen extraction.
Assuntos
Cafeína , Café , Estudos Cross-Over , Exercício Físico , Músculo Esquelético , Consumo de Oxigênio , Descanso , Humanos , Masculino , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/metabolismo , Cafeína/administração & dosagem , Cafeína/farmacologia , Exercício Físico/fisiologia , Adulto Jovem , Descanso/fisiologia , Espectroscopia de Luz Próxima ao Infravermelho , Adulto , Microcirculação/efeitos dos fármacos , Método Duplo-Cego , Oxigênio/sangueRESUMO
Titin is a multidomain protein of striated and smooth muscles of vertebrates. The protein consists of repeating immunoglobulin-like (Ig) and fibronectin-like (FnIII) domains, which are ß-sandwiches with a predominant ß-structure, and also contains disordered regions. In this work, the methods of atomic force microscopy (AFM), X-ray diffraction, and Fourier transform infrared spectroscopy were used to study the morphology and structure of aggregates of rabbit skeletal muscle titin obtained in two different solutions: 0.15 M glycine-KOH, pH 7.0 and 200 mM KCl, 10 mM imidazole, pH 7.0. According to AFM data, skeletal muscle titin formed amorphous aggregates of different morphologies in the above two solutions. Amorphous aggregates of titin formed in a solution containing glycine consisted of much larger particles than aggregates of this protein formed in a solution containing KCl. The "KCl-aggregates" according to AFM data had the form of a "sponge"-like structure, while amorphous "glycine-aggregates" of titin formed "branching" structures. Spectrofluorometry revealed the ability of "glycine-aggregates" of titin to bind to the dye thioflavin T (TT), and X-ray diffraction revealed the presence of one of the elements of the amyloid cross ß-structure, a reflection of ~4.6 Å, in these aggregates. These data indicate that "glycine-aggregates" of titin are amyloid or amyloid-like. No similar structural features were found in "KCl-aggregates" of titin; they also did not show the ability to bind to thioflavin T, indicating the non-amyloid nature of these titin aggregates. Fourier transform infrared spectroscopy revealed differences in the secondary structure of the two types of titin aggregates. The data we obtained demonstrate the features of structural changes during the formation of intermolecular bonds between molecules of the giant titin protein during its aggregation. The data expand the understanding of the process of amyloid protein aggregation.
Assuntos
Conectina , Microscopia de Força Atômica , Músculo Esquelético , Agregados Proteicos , Conectina/química , Conectina/metabolismo , Conectina/genética , Coelhos , Animais , Músculo Esquelético/metabolismo , Músculo Esquelético/química , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios X , BenzotiazóisRESUMO
This study investigated the effect of knockout of six Hsp70 genes (orthologues of the mammalian genes Hspa1a, Hspa1b, Hspa2, and Hspa8) on age-related changes in gene expression in the legs of Drosophila melanogaster, which contain predominantly skeletal muscle bundles. For this, the leg transcriptomic profile was examined in males of the w^(1118) control strain and the Hsp70^(-) strain on the 7th, 23rd and 47th days of life. In w^(1118) flies, an age-related decrease in the locomotion (climbing) speed (a marker of functional state and endurance) was accompanied by a pronounced change in the transcriptomic profile of the leg skeletal muscles, which is conservative in nature. In Hsp70^(-) flies, the median lifespan was shorter and the locomotion speed was significantly lower compared to the control; at the same time, complex changes in the age-related dynamics of the skeletal muscle transcriptome were observed. Mass spectrometry-based quantitative proteomics showed that 47-day-old Hsp70^(-) flies, compared with w^(1118) flies, demonstrated multidirectional changes in the contents of key enzymes of glucose metabolism and fat oxidation (glycolysis, pentose phosphate pathway, Krebs cycle, beta-oxidation, and oxidative phosphorylation). Such dysregulation may be associated with a compensatory increase in the expression of other genes encoding chaperones (small Hsp, Hsp40, 60, and 70), which regulate specific sets of target proteins. Taken together, our data show that knockout of six Hsp70 genes slightly reduced the median lifespan of flies, but significantly reduced the locomotion speed, which may be associated with complex changes in the transcriptome of the leg skeletal muscles and with multidirectional changes in the contents of key enzymes of energy metabolism.
Assuntos
Proteínas de Drosophila , Drosophila melanogaster , Proteínas de Choque Térmico HSP70 , Locomoção , Longevidade , Músculo Esquelético , Transcriptoma , Animais , Drosophila melanogaster/genética , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Masculino , Locomoção/fisiologia , Locomoção/genética , Músculo Esquelético/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Longevidade/genética , Envelhecimento/genética , Envelhecimento/metabolismo , Técnicas de Inativação de GenesRESUMO
It is not clear if fat oxidation is attenuated at higher exercise intensities, when exercising with a small muscle mass, and therefore, we studied leg fat oxidation during graded one-leg exercise. Ten males (age: 27 ± 2 years, body mass: 82 ± 3 kg, BMI: 24 ± 1 kg m-2, VÌO2max: 49 ± 2 mL min-1 kg-1) performed one-leg exercise at 25% of maximal workload (Wmax) for 30 min, followed by 120-min exercise at 55% Wmax with the contralateral leg, and finally 30-min exercise at 85% Wmax with the first leg. Blood was sampled from an artery and both femoral veins, and blood flow was determined using Doppler ultrasound. Muscle biopsies were obtained before and after 30 min at each workload. One-way RM ANOVA was applied to determine the impact of exercise intensity. Data are expressed as mean ± SEM. From rest through exercise average blood flow (0.4 ± 0.1, 2.1 ± 0.1, 2.6 ± 0.2, 3.7 ± 0.2 L min-1) and oxygen uptake across the leg (0.03 ± 0.01, 0.23 ± 0.02, 0.35 ± 0.03, 0.53 ± 0.04 L min-1) increased with exercise intensity (p < 0.001). Leg RQ (0.76 ± 0.04, 0.86 ± 0.02,0.87 ± 0.01, 0.92 ± 0.01, p < 0.001), leg plasma FA uptake (2 ± 2, 46 ± 8,83 ± 9, 114 ± 16 µmol min-1; p < 0.001) and rate of leg fat oxidation (0.016 ± 0.005, 0.062 ± 0.012, 0.075 ± 0.011, 0.084 ± 0.018 g min-1, p < 0.007) increased with exercise intensity. Muscle-free carnitine content was unchanged from rest at 25% Wmax and decreased after 30 min exercise at 55% and 85% Wmax (17.4 ± 1.6, 16.6 ± 0.7, 14.5 ± 1.2, 10.5 ± 1.0 mmol/kg dry muscle, respectively; p < 0.006). During incremental one-leg exercise, the rate of leg fat oxidation was not attenuated with increasing exercise intensity, probably due to an insufficient muscle metabolic stress response.
Assuntos
Exercício Físico , Perna (Membro) , Músculo Esquelético , Oxirredução , Consumo de Oxigênio , Fluxo Sanguíneo Regional , Humanos , Masculino , Adulto , Exercício Físico/fisiologia , Consumo de Oxigênio/fisiologia , Perna (Membro)/fisiologia , Músculo Esquelético/fisiologia , Músculo Esquelético/metabolismo , Tecido Adiposo/metabolismo , Adulto Jovem , Joelho/fisiologia , Ultrassonografia Doppler , Metabolismo dos Lipídeos/fisiologiaRESUMO
INTRODUCTION: In patients admitted to the intensive care unit (ICU), muscle mass is inversely associated with mortality. Although muscle mass can be estimated with 24-h urinary creatinine excretion (UCE), its use for risk prediction in individual patients is limited because age-, sex-, weight- and length-specific reference values for UCE are lacking. The ratio between measured creatinine clearance (mCC) and estimated glomerular filtration rate (eGFR) might circumvent this constraint. The main goal was to assess the association of the mCC/eGFR ratio in ICU patients with all-cause hospital and long-term mortality. METHODS: The mCC/eGFR ratio was determined in patients admitted to our ICU between 2005 and 2021 with KDIGO acute kidney injury (AKI) stage 0-2 and an ICU stay ≥ 24 h. mCC was calculated from UCE and plasma creatinine and indexed to 1.73 m2. mCC/eGFR was analyzed by categorizing patients in mCC/eGFR quartiles and as continuous variable. RESULTS: Seven thousand five hundred nine patients (mean age 61 ± 15 years; 38% female) were included. In-hospital mortality was 27% in the lowest mCC/eGFR quartile compared to 11% in the highest quartile (P < 0.001). Five-year post-hospital discharge actuarial mortality was 37% in the lowest mCC/eGFR quartile compared to 19% in the highest quartile (P < 0.001). mCC/eGFR ratio as continuous variable was independently associated with in-hospital mortality in multivariable logistic regression (odds ratio: 0.578 (95% CI: 0.465-0.719); P < 0.001). mCC/eGFR ratio as continuous variable was also significantly associated with 5-year post-hospital discharge mortality in Cox regression (hazard ratio: 0.27 (95% CI: 0.22-0.32); P < 0.001). CONCLUSIONS: The mCC/eGFR ratio is associated with both in-hospital and long-term mortality and may be an easily available index of muscle mass in ICU patients.
Assuntos
Creatinina , Taxa de Filtração Glomerular , Mortalidade Hospitalar , Unidades de Terapia Intensiva , Humanos , Masculino , Feminino , Pessoa de Meia-Idade , Creatinina/sangue , Creatinina/urina , Idoso , Injúria Renal Aguda/mortalidade , Injúria Renal Aguda/sangue , Injúria Renal Aguda/diagnóstico , Estudos Retrospectivos , Músculo Esquelético/metabolismoRESUMO
Low muscle mass is a risk factor for mortality in patients with chronic kidney disease (CKD). However, it is not clear to what extent low muscle mass contributes to this risk, either independently or in combination with metabolic abnormalities and frailty. This study used data from the National Health and Nutrition Examination Survey 1999-2006 and 2011-2018. Low muscle mass was defined as Appendicular Skeletal Mass Index < 7 kg/m2 in men or < 5.5 kg/m2 in women. The follow-up duration was from the first anthropometric and clinical measurements to death or the last follow-up. This study enrolled 2072 patients with CKD. Low muscle mass was associated with a lower risk of metabolic abnormalities, but was associated with an elevated mortality risk. Conversely, central obesity was associated with a higher likelihood of metabolic abnormalities and frailty, yet showed no significant association with mortality risk. Subsequently conducted mediation analysis indicated that the effect of low muscle mass on mortality was direct, not mediated by frailty and metabolic abnormalities. In spite of the inverse relationship between low muscle mass and metabolic abnormalities, low muscle mass are directly associated with an increased risk of all-cause mortality. Low muscle mass may directly contribute to mortality in patients with CKD, independent of metabolic abnormalities and frailty in these patients.
Assuntos
Doenças Metabólicas , Insuficiência Renal Crônica , Humanos , Insuficiência Renal Crônica/mortalidade , Insuficiência Renal Crônica/complicações , Insuficiência Renal Crônica/patologia , Feminino , Masculino , Pessoa de Meia-Idade , Idoso , Doenças Metabólicas/mortalidade , Doenças Metabólicas/complicações , Doenças Metabólicas/patologia , Inquéritos Nutricionais , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Fatores de Risco , Fragilidade/mortalidade , Fragilidade/complicações , Sarcopenia/mortalidade , Sarcopenia/complicações , Sarcopenia/metabolismo , AdultoRESUMO
Current adeno-associated virus (AAV) gene therapy using nature-derived AAVs is limited by non-optimal tissue targeting. In the treatment of muscular diseases (MD), high doses are often required but can lead to severe adverse effects. Here, we rationally design an AAV capsid that specifically targets skeletal muscle to lower treatment doses. We computationally integrate binding motifs of human integrin alphaV beta6, a skeletal muscle receptor, into a liver-detargeting capsid. Designed AAVs show higher productivity and superior muscle transduction compared to their parent. One variant, LICA1, demonstrates comparable muscle transduction to other myotropic AAVs with reduced liver targeting. LICA1's myotropic properties are observed across species, including non-human primate. Consequently, LICA1, but not AAV9, effectively delivers therapeutic transgenes and improved muscle functionality in two mouse MD models (male mice) at a low dose (5E12 vg/kg). These results underline the potential of our design method for AAV engineering and LICA1 variant for MD gene therapy.
Assuntos
Dependovirus , Terapia Genética , Músculo Esquelético , Dependovirus/genética , Animais , Humanos , Músculo Esquelético/metabolismo , Camundongos , Terapia Genética/métodos , Masculino , Vetores Genéticos/genética , Integrinas/metabolismo , Integrinas/genética , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Doenças Musculares/terapia , Doenças Musculares/genética , Transdução Genética , Fígado/metabolismo , Capsídeo/metabolismo , Receptores de Vitronectina/metabolismo , Receptores de Vitronectina/genética , Modelos Animais de Doenças , Células HEK293 , Transgenes , Camundongos Endogâmicos C57BL , Antígenos de NeoplasiasRESUMO
Prolonged strenuous exercise induces oxidative stress, leading to oxidative damage, skeletal muscle fatigue, and reduced exercise performance. The body compensates for oxidative stress through antioxidant actions, while related enzymes alone may not overcome excessive oxidative stress during prolonged strenuous exercise. Phycocyanin is an important antioxidant supplement derived from blue-green algae, which may be helpful in this type of situation. This study determined the effects of phycocyanin on exercise performance from prolonged strenuous exercise. Forty Sprague Dawley male rats were divided into 5 groups (n = 8 /group); Control group (C), Exercise group (E), and Exercise with supplement groups receiving low dose (Phycocyanin = 100 mg/kg BW; ELP) and high dose (Phycocyanin = 200 mg/kg BW; EHP) or vitamin C (Vitamin C = 200 mg/kg BW; VC). Phycocyanin was found to decrease oxidative damage markers, muscle fatigue, and muscle atrophy through the activated AKT/mTOR pathway. This was also found to have greater increases in antioxidants via Nrf2 signaling and increases ATP synthesis, GLUT4 transporters, and insulin signaling due to increased IRS-1/AKT signaling. In conclusion, phycocyanin was found to reduce oxidative damage and muscle atrophy, including an increase in insulin signaling in skeletal muscles leading to increased exercise performance in rats.
Assuntos
Músculo Esquelético , Fator 2 Relacionado a NF-E2 , Estresse Oxidativo , Ficocianina , Condicionamento Físico Animal , Proteínas Proto-Oncogênicas c-akt , Ratos Sprague-Dawley , Transdução de Sinais , Serina-Treonina Quinases TOR , Animais , Estresse Oxidativo/efeitos dos fármacos , Masculino , Fator 2 Relacionado a NF-E2/metabolismo , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Transdução de Sinais/efeitos dos fármacos , Ratos , Ficocianina/farmacologia , Proteínas Substratos do Receptor de Insulina/metabolismo , Fadiga Muscular/efeitos dos fármacos , Antioxidantes/metabolismo , Antioxidantes/farmacologiaRESUMO
Myofibrillar myopathy (MFM) is a group of hereditary myopathies that mainly involves striated muscles. This study aimed to use tandem mass tag (TMT)-based proteomics to investigate the underlying pathomechanisms of two of the most common MFM subtypes, desminopathy and titinopathy. Muscles from 7 patients with desminopathy, 5 with titinopathy and 5 control individuals were included. Samples were labelled with TMT and then underwent high-resolution liquid chromatography-mass spectrometry analysis. Compared with control samples, there were 436 differentially abundant proteins (DAPs) in the desminopathy group and 269 in the titinopathy group. When comparing the desminopathy with the titinopathy group, there were 113 DAPs. In desminopathy, mitochondrial ATP production, muscle contraction, and cytoskeleton organization were significantly suppressed. Activated cellular components and pathways were mostly related to extracellular matrix (ECM). In titinopathy, mitochondrial-related pathways and the cellular component ECM were downregulated, while gluconeogenesis was activated. Direct comparison between desminopathy and titinopathy revealed hub genes that were all involved in glycolytic process. The disparity in glycolysis in the two MFM subtypes is likely due to fiber type switching. This study has revealed disorganization of cytoskeleton and mitochondrial dysfunction as the common pathophysiological processes in MFM, and glycolysis and ECM as the differential pathomechanism between desminopathy and titinopathy. This offers a future direction for targeted therapy for MFM.
Assuntos
Conectina , Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Conectina/genética , Conectina/metabolismo , Proteômica/métodos , Miopatias Congênitas Estruturais/genética , Miopatias Congênitas Estruturais/patologia , Miopatias Congênitas Estruturais/metabolismo , Músculo Esquelético/patologia , Músculo Esquelético/metabolismo , Desmina/genética , Desmina/metabolismo , Glicólise/genética , Mitocôndrias/metabolismo , Mitocôndrias/genética , Mitocôndrias/patologia , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Distrofias Musculares , CardiomiopatiasRESUMO
Estradiol (E2) deficiency arising from menopause is closely related to changes in body composition and declines of muscle mass and strength in elderly women. Whole-body vibration training (WBV) is an emerging approach expected to improve muscle mass and strength of older person, but the underlying mechanisms remain unclear. The balance between protein synthesis and degradation is a determining factor for muscle mass and strength, which is regulated by Akt-mTOR and FoxO1 signal pathway, respectively. In the present study, we firstly determined whether the effects of WBV on muscle mass and strength in ovariectomized female mice was affected by estrogen level, then investigated whether this was associated with Akt-mTOR and FoxO1 signal pathways. We found that (1) WBV, E2 supplementation (E) and WBV combined with E2 supplementation (WBV+E) significantly increased serum estradiol content, quadriceps muscle mass and grip strength in ovariectomized mice, accompanied with alterations of body composition (reducing fat content, increasing lean body mass and lean percent), furthermore, the altered degrees of these indicators by WBV+E were greater than WBV alone; (2) WBV, E and WBV+E remarkably increased the activities of Akt and mTOR and decreased FoxO1 activity, and the changed degrees by WBV+E were greater than WBV alone; (3) Pearson correlation coefficient revealed that serum estradiol content was positively correlated with Akt and mTOR activities, while inversely associated with FoxO1 activity. We concluded that WBV could significantly increase muscle mass and strength in ovariectomized mice, which might achieve through activating Akt-mTOR and suppressing FoxO1 signal pathways, and the improving effect of WBV on muscle mass and strength was better when in the presence of estrogen.
Assuntos
Estradiol , Estrogênios , Proteína Forkhead Box O1 , Força Muscular , Ovariectomia , Serina-Treonina Quinases TOR , Vibração , Animais , Feminino , Vibração/uso terapêutico , Camundongos , Força Muscular/fisiologia , Serina-Treonina Quinases TOR/metabolismo , Estradiol/sangue , Proteína Forkhead Box O1/metabolismo , Estrogênios/sangue , Estrogênios/metabolismo , Transdução de Sinais , Composição Corporal/fisiologia , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiologia , Condicionamento Físico Animal/fisiologia , Condicionamento Físico Animal/métodosRESUMO
Most patients with advanced cancer develop cachexia, a multifactorial syndrome characterized by progressive skeletal muscle wasting. Despite its catastrophic impact on survival, the critical mediators responsible for cancer cachexia development remain poorly defined. Here, we show that a distinct subset of neutrophil-like monocytes, which we term cachexia-inducible monocytes (CiMs), emerges in the advanced cancer milieu and promotes skeletal muscle loss. Unbiased transcriptome analysis reveals that interleukin 36 gamma (IL36G)-producing CD38+ CiMs are induced in chronic monocytic blood cancer characterized by prominent cachexia. Notably, the emergence of CiMs and the activation of CiM-related gene signatures in monocytes are confirmed in various advanced solid cancers. Stimuli of toll-like receptor 4 signaling are responsible for the induction of CiMs. Genetic inhibition of IL36G-mediated signaling attenuates skeletal muscle loss and rescues cachexia phenotypes in advanced cancer models. These findings indicate that the IL36G-producing subset of neutrophil-like monocytes could be a potential therapeutic target in cancer cachexia.
Assuntos
Caquexia , Monócitos , Músculo Esquelético , Neoplasias , Neutrófilos , Caquexia/metabolismo , Caquexia/etiologia , Monócitos/metabolismo , Monócitos/imunologia , Humanos , Neoplasias/complicações , Neoplasias/metabolismo , Neoplasias/imunologia , Neutrófilos/metabolismo , Animais , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Camundongos , Masculino , Transdução de Sinais , Linhagem Celular Tumoral , Receptor 4 Toll-Like/metabolismo , Receptor 4 Toll-Like/genética , Camundongos Endogâmicos C57BL , Interleucinas/metabolismo , Interleucinas/genética , Feminino , Perfilação da Expressão GênicaRESUMO
Ca2+ release from the sarcoplasmic reticulum (SR) plays a central role in excitation-contraction coupling (ECC) in skeletal muscles. However, the mechanism by which activation of the voltage-sensors/dihydropyridine receptors (DHPRs) in the membrane of the transverse tubular system leads to activation of the Ca2+-release channels/ryanodine receptors (RyRs) in the SR is not fully understood. Recent observations showing that a very small Ca2+ leak through RyR1s in mammalian skeletal muscle can markedly raise the background [Ca2+] in the junctional space (JS) above the Ca2+ level in the bulk of the cytosol indicate that there is a diffusional barrier between the JS and the cytosol at large. Here, I use a mathematical model to explore the hypothesis that a sudden rise in Ca2+ leak through DHPR-coupled RyR1s, caused by reduced inhibition at the RyR1 Ca2+/Mg2+ inhibitory I1-sites when the associated DHPRs are activated, is sufficient to enable synchronized responses that trigger a regenerative rise of Ca2+ release that remains under voltage control. In this way, the characteristic response to Ca2+ of RyR channels is key not only for the Ca2+ release mechanism in cardiac muscle and other tissues, but also for the DHPR-dependent Ca2+ release in skeletal muscle.
Assuntos
Canais de Cálcio Tipo L , Cálcio , Músculo Esquelético , Canal de Liberação de Cálcio do Receptor de Rianodina , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Músculo Esquelético/metabolismo , Animais , Cálcio/metabolismo , Canais de Cálcio Tipo L/metabolismo , Retículo Sarcoplasmático/metabolismo , Sinalização do Cálcio/fisiologia , Acoplamento Excitação-Contração , Modelos Biológicos , HumanosRESUMO
Introduction: Introduction: musculoskeletal health has become of increasing interest due to the ageing of the population and the increase in the prevalence of associated diseases. Objective: analyze scientific evidence on the role of nutrition and diet in maintaining muscle and bone health and preventing related diseases. Methods: review of the scientific literature on nutrition and diet in maintaining muscle and bone health. Results and discussion: dietary components such as protein, calcium, magnesium, vitamin D, C, K, B12, among others, have been positively associated with the maintenance of muscle and bone. The Mediterranean diet could slow the onset of sarcopenia and osteoporosis. Conclusion: nutrition is crucial for muscle and bone health.
Introducción: Introducción: la salud musculoesquelética ha adquirido un interés creciente debido al envejecimiento poblacional y al aumento de enfermedades asociadas. Objetivo: analizar la evidencia sobre el papel de la nutrición y la dieta en la salud muscular y ósea y la prevención de enfermedades asociadas. Métodos: revisión de la literatura científica sobre la nutrición y la dieta en el mantenimiento de una adecuada salud muscular y ósea. Resultados y discusión: componentes dietéticos como las proteínas, el calcio, el magnesio y las vitaminas D, C, K, B12, entre otros, se han asociado positivamente con el mantenimiento de la salud muscular y ósea. La dieta mediterránea podría ralentizar la aparición de la sarcopenia y la osteoporosis. Conclusión: la nutrición es crucial para la salud muscular y ósea.
Assuntos
Osso e Ossos , Dieta , Sarcopenia , Humanos , Osso e Ossos/metabolismo , Osso e Ossos/fisiologia , Sarcopenia/prevenção & controle , Sarcopenia/dietoterapia , Dieta Mediterrânea , Osteoporose/prevenção & controle , Osteoporose/dietoterapia , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiologiaRESUMO
Carbohydrates are critical for high-intensity exercise performance. However, the effects of carbohydrate supplementation on muscle metabolism and performance during short-duration high-intensity intermittent exercise remain inadequately explored. Our aim was to address this aspect in a randomized, counterbalanced, double-blinded crossover design. Eleven moderately-to-well-trained males performed high-intensity intermittent cycling receiving carbohydrate (CHO, ~55 g/h) or placebo (PLA) fluid supplementation. Three exercise periods (EX1-EX3) were completed comprising 10 × 45 s at ~105% Wmax interspersed with 135 s rest between bouts and ~20 min between periods. Repeated sprint ability (5 × 6 s sprints with 24 s recovery) was assessed at baseline and after each period. Thigh muscle biopsies were obtained at baseline and before and after EX3 to determine whole-muscle and fiber-type-specific glycogen depletion. No differences were found in muscle glycogen degradation at the whole-muscle (p = 0.683) or fiber-type-specific level (p = 0.763-0.854) with similar post-exercise whole-muscle glycogen concentrations (146 ± 20 and 122 ± 15 mmol·kg-1 dw in CHO and PLA, respectively). Repeated sprint ability declined by ~9% after EX3 with no between-condition differences (p = 0.971) and no overall differences in ratings of perceived exertion (p = 0.550). This was despite distinctions in blood glucose concentrations throughout exercise, reaching post-exercise levels of 5.3 ± 0.2 and 4.1 ± 0.2 mmol·L-1 (p < 0.001) in CHO and PLA, respectively, accompanied by fivefold higher plasma insulin levels in CHO (p < 0.001). In conclusion, we observed no effects of carbohydrate ingestion on net muscle glycogen breakdown or sprint performance during short-duration high-intensity intermittent exercise despite elevated blood glucose and insulin levels. These results therefore question the efficacy of carbohydrate supplementation strategies in high-intensity intermittent sports.
Assuntos
Desempenho Atlético , Estudos Cross-Over , Carboidratos da Dieta , Glicogênio , Músculo Esquelético , Humanos , Masculino , Glicogênio/metabolismo , Carboidratos da Dieta/administração & dosagem , Método Duplo-Cego , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiologia , Desempenho Atlético/fisiologia , Adulto Jovem , Adulto , Treinamento Intervalado de Alta Intensidade , Glicemia/metabolismo , Insulina/sangue , Suplementos Nutricionais , Ciclismo/fisiologiaRESUMO
BACKGROUND: Duchenne muscular dystrophy (DMD) is an incurable neuromuscular disease leading to progressive skeletal muscle weakness and fatigue. Cell transplantation in murine models has shown promise in supplementing the lack of the dystrophin protein in DMD muscles. However, the establishment of novel, long-term, relevant methods is needed to assess its efficiency on the DMD motor function. By applying newly developed methods, this study aimed to evaluate the functional and molecular effects of cell therapy-mediated dystrophin supplementation on DMD muscles. METHODS: Dystrophin was supplemented in the gastrocnemius of a 5-week-old immunodeficient DMD mouse model (Dmd-null/NSG) by intramuscular xenotransplantation of healthy human immortalized myoblasts (Hu5/KD3). A long-term time-course comparative study was conducted between wild-type, untreated DMD, and dystrophin supplemented-DMD mouse muscle functions and histology. A novel GO-ATeam2 transgenic DMD mouse model was also generated to assess in vivo real-time ATP levels in gastrocnemius muscles during repeated contractions. RESULTS: We found that 10.6% dystrophin supplementation in DMD muscles was sufficient to prevent low values of gastrocnemius maximal isometric contraction torque (MCT) at rest, while muscle fatigue tolerance, assessed by MCT decline after treadmill running, was fully ameliorated in 21-week-old transplanted mice. None of the dystrophin-supplemented fibers were positive for muscle damage markers after treadmill running, with 85.4% demonstrating the utilization of oxidative metabolism. Furthermore, ATP levels in response to repeated muscle contractions tended to improve, and mitochondrial activity was significantly enhanced in dystrophin supplemented-fibers. CONCLUSIONS: Cell therapy-mediated dystrophin supplementation efficiently improved DMD muscle functions, as evaluated using newly developed evaluation methods. The enhanced muscle fatigue tolerance in 21-week-old mice was associated with the preferential regeneration of damage-resistant and oxidative fibers, highlighting increased mitochondrial activity, after cell transplantation. These findings significantly contribute to a more in-depth understanding of DMD pathogenesis.
Assuntos
Modelos Animais de Doenças , Distrofina , Fadiga Muscular , Músculo Esquelético , Distrofia Muscular de Duchenne , Animais , Distrofia Muscular de Duchenne/terapia , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patologia , Distrofina/genética , Distrofina/metabolismo , Camundongos , Músculo Esquelético/metabolismo , Humanos , Mioblastos/metabolismo , Camundongos Endogâmicos mdx , Masculino , Contração Muscular , Transplante de Células/métodosRESUMO
Background: Ischemia/reperfusion injury (IRI) is a complex pathological process, triggered by the restoration of blood flow following an interrupted blood supply. While restoring the blood flow is the only option to salvage the ischemic tissue, reperfusion after a prolonged period of ischemia initiates IRI, triggering a cascade of inflammatory responses ultimately leading to neutrophil recruitment to the inflamed tissue, where they release neutrophil extracellular traps (NETs). NETs are web-like structures of decondensed chromatin and neutrophilic proteins, including peptidyl-arginine deiminase 2 and 4 (PAD2, PAD4), that, once outside, can citrullinate plasma proteins, irreversibly changing their conformation and potentially their function. While the involvement of NETs in IRI is known mainly from rodent models, we aimed to determine the effect of NET formation and especially PADs-mediated extracellular protein citrullination in a porcine model of limb IRI. Methods: We conducted our study on amputated pig forelimbs exposed to 1 h or 9 h of ischemia and then reperfused in vivo for 12 h. Limb weight, edema formation, compartmental pressure were measured, and skeletal muscle was analyzed by immunofluorescence (TUNEL assay and dystrophin staining) to evaluate tissue damage. Fibrin tissue deposition, complement deposition and NETs were investigated by immunofluorescence. Citrullinated plasma proteins were immunoprecipitated and citrullinated fibrinogen was identified in the plasma by Western blot and in the tissue by immunofluorescence and Western blot. Results: Our data consolidate the involvement of NETs in a porcine model of limb IRI, correlating their contribution to damage extension with the duration of the ischemic time. We found a massive infiltration of NETs in the group subjected to 9 h ischemia compared to the 1 h and citrullinated fibrinogen levels, in plasma and tissue, were higher in 9 h ischemia group. We propose fibrinogen citrullination as one of the mechanisms contributing to the worsening of IRI. NETs and protein citrullination represent a potential therapeutic target, but approaches are still a matter of debate. Here we introduce the idea of therapeutic approaches against citrullination to specifically inhibit PADs extracellularly, avoiding the downstream effects of hypercitrullination and keeping PADs' and NETs' intracellular regulatory functions.