RESUMO
The current study aimed to investigate the hypothesis that purinergic receptors P2Y1 and P2Y2 play a regulatory role in gene expression in unloaded muscle. ATP is released from cells through pannexin channels, and it interacts with P2Y1 and P2Y2 receptors, leading to the activation of markers of protein catabolism and a reduction in protein synthesis. To test this hypothesis thirty-two rats were randomly divided into four groups (8 per group): a non-treated control group (C), a group subjected to three days of hindlimb unloading with a placebo (HS), a group subjected to three days of hindlimb unloading treated with a P2Y1 receptor inhibitor, MRS2179 (HSM), and a group subjected to three days of hindlimb unloading treated with a P2Y2 receptor inhibitor, AR-C 118925XX (HSA). This study revealed several key findings following three days of soleus muscle unloading: 1: Inhibition of P2Y1 or P2Y2 receptors prevented the accumulation of ATP, the increase in IP3 receptor content, and the decrease in the phosphorylation of GSK-3beta. This inhibition also mitigated the reduction in the rate of protein synthesis. However, it had no significant effect on the markers of mTORC1-dependent signaling. 2: Blocking P2Y1 receptors prevented the unloading-induced upregulation of phosphorylated p38MAPK and partially reduced the increase in MuRF1mRNA expression. 3: Blocking P2Y2 receptors prevented muscle atrophy during unloading, partially maintained the levels of phosphorylated ERK1/2, reduced the increase in mRNA expression of MAFbx, ubiquitin, and IL-6 receptor, prevented the decrease in phosphorylated AMPK, and attenuated the increase in phosphorylated p70S6K. Taken together, these results suggest that the prevention of muscle atrophy during unloading, as achieved by the P2Y2 receptor inhibitor, is likely mediated through a reduction in catabolic processes and maintenance of energy homeostasis. In contrast, the P2Y1 receptor appears to play a relatively minor role in muscle atrophy during unloading.
Assuntos
Músculo Esquelético , Transdução de Sinais , Animais , Ratos , Trifosfato de Adenosina/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Receptores Purinérgicos P2Y1/metabolismo , Receptores Purinérgicos P2Y2/genética , Receptores Purinérgicos P2Y2/metabolismoRESUMO
Skeletal muscle unloading leads to the decreased electrical activity and decline of muscle tone. AIMS: Current study evaluated the effect of muscle tone preservation achieved by tetanus toxin (TeNT) treatment on signaling pathways regulating atrophic processes during unloading. MAIN METHODS: Four groups of rats were used: non-treated control (C), control rats with TeNT administration (CT), 7 days of unloading/hindlimb suspension with placebo (HS), and 7 days of unloading with TeNT administration (HST). KEY FINDINGS: Absolute and relative force of tetanic contractions was decreased by 65% in soleus muscle of HS rats when compared with C. Treatment with TeNT significantly lessened force decline in soleus muscle of HST rats when compared with HS. TeNT administration increased myosin heavy chain I beta (MyHC Iß) expression in CT rats and prevented MyHC Iß loss in HST group when compared with C rats. Desmin content was lower by 31.4% (p < 0.05) in HS group when compared with HST. Calpain-1 expression was increased in HS group when compared with C, CT and HST. There was a decrease in p-p70S6K content (41%, p < 0,05) and an increase in p-eEF2 content (77%, p < 0,05) in HS group when compared with C, while there were no significant differences in the content of these proteins between HST, CT and C groups. SIGNIFICANCE: Treatment with TeNT significantly diminished unloading-induced decline of soleus muscle mass and mechanical properties and affected the regulation of MyHC Iß expression. These effects are mediated by signaling pathways regulating protein synthesis and degradation.
Assuntos
Proteínas do Citoesqueleto , Tono Muscular , Animais , Proteínas do Citoesqueleto/metabolismo , Elevação dos Membros Posteriores/fisiologia , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Cadeias Pesadas de Miosina/metabolismo , Ratos , Ratos WistarRESUMO
This Special Issue presents some of the most recent studies on the skeletal muscle denervation [...].
Assuntos
Denervação Muscular , Músculo Esquelético , Humanos , Músculo Esquelético/patologia , Atrofia Muscular/patologiaRESUMO
The ubiquitously expressed adaptor protein Shc exists in three isoforms p46Shc, p52Shc, and p66Shc, which execute distinctly different actions in cells. The role of p46Shc is insufficiently studied, and the purpose of this study was to further investigate its functional significance. We developed unique rat mutants lacking p52Shc and p46Shc isoforms (p52Shc/46Shc-KO) and carried out histological analysis of skeletal and cardiac muscle of parental and genetically modified rats with impaired gait. p52Shc/46Shc-KO rats demonstrate severe functional abnormalities associated with impaired gait. Our analysis of p52Shc/46Shc-KO rat axons and myelin sheets in cross-sections of the sciatic nerve revealed the presence of significant anomalies. Based on the lack of skeletal muscle fiber atrophy and the presence of sciatic nerve abnormalities, we suggest that the impaired gait in p52Shc/46Shc-KO rats might be due to the sensory feedback from active muscle to the brain locomotor centers. The lack of dystrophin in some heart muscle fibers reflects damage due to dilated cardiomyopathy. Since rats with only p52Shc knockout do not display the phenotype of p52Shc/p46Shc-KO, abnormal locomotion is likely to be caused by p46Shc deletion. Our data suggest a previously unknown role of 46Shc actions and signaling in regulation of gait.
Assuntos
Cardiomiopatia Dilatada/genética , Marcha/genética , Proteína 1 de Transformação que Contém Domínio 2 de Homologia de Src/genética , Animais , Cardiomiopatia Dilatada/patologia , Técnicas de Inativação de Genes , Músculo Esquelético/patologia , Músculo Esquelético/fisiopatologia , Isoformas de Proteínas/genética , Ratos Transgênicos , Nervo Isquiático/patologiaRESUMO
Skeletal muscle unloading results in atrophy. We hypothesized that pannexin 1 ATP-permeable channel (PANX1) is involved in the response of muscle to unloading. We tested this hypothesis by blocking PANX1, which regulates efflux of ATP from the cytoplasm. Rats were divided into six groups (eight rats each): non-treated control for 1 and 3 days of the experiments (1C and 3C, respectively), 1 and 3 days of hindlimb suspension (HS) with placebo (1H and 3H, respectively), and 1 and 3 days of HS with PANX1 inhibitor probenecid (PRB; 1HP and 3HP, respectively). When compared with 3C group there was a significant increase in ATP in soleus muscle of 3H and 3HP groups (32 and 51%, respectively, p < 0.05). When compared with 3H group, 3HP group had: (1) lower mRNA expression of E3 ligases MuRF1 and MAFbx (by 50 and 38% respectively, p < 0.05) and MYOG (by 34%, p < 0.05); (2) higher phosphorylation of p70S6k and p90RSK (by 51 and 35% respectively, p < 0.05); (3) lower levels of phosphorylated eEF2 (by 157%, p < 0.05); (4) higher level of phosphorylated GSK3ß (by 189%, p < 0.05). In conclusion, PANX1 ATP-permeable channels are involved in the regulation of muscle atrophic processes by modulating expression of E3 ligases, and protein translation and elongation processes during unloading.
Assuntos
Conexinas/metabolismo , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Transdução de Sinais , Animais , Elevação dos Membros Posteriores , Masculino , Músculo Esquelético/patologia , Atrofia Muscular/patologia , Ratos , Ratos WistarRESUMO
Unloading leads to skeletal muscle atrophy via the upregulation of MuRF-1 and MAFbx E3-ligases expression. Reportedly, histone deacetylases (HDACs) 4 and 5 may regulate the expression of MuRF1 and MAFbx. To examine the HDAC-dependent mechanisms involved in the control of E3-ubiquitin ligases expression at the early stages of muscle unloading we used HDACs 4 and 5 inhibitor LMK-235 and HDAC 4 inhibitor Tasqinimod (Tq). Male Wistar rats were divided into four groups (eight rats per group): nontreated control (C), three days of unloading/hindlimb suspension (HS) and three days HS with HDACs inhibitor LMK-235 (HSLMK) or Tq (HSTq). Treatment with LMK-235 diminished unloading-induced of MAFbx, myogenin (MYOG), ubiquitin and calpain-1 mRNA expression (p < 0.05). Tq administration had no effect on the expression of E3-ligases. The mRNA expression of MuRF1 and MAFbx was significantly increased in both HS and HSTq groups (1.5 and 4.0 folds, respectively; p < 0.05) when compared with the C group. It is concluded that during three days of muscle unloading: (1) the HDACs 4 and 5 participate in the regulation of MAFbx expression as well as the expression of MYOG, ubiquitin and calpain-1; (2) the inhibition of HDAC 4 has no effect on MAFbx expression. Therefore, HDAC 5 is perhaps more important for the regulation of MAFbx expression than HDAC 4.
Assuntos
Histona Desacetilases/metabolismo , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Proteínas Ligases SKP Culina F-Box/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Calpaína/metabolismo , Elevação dos Membros Posteriores/fisiologia , Masculino , Atrofia Muscular/metabolismo , Miogenina/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Ubiquitina/metabolismoRESUMO
To test the hypothesis that p38α-MAPK plays a critical role in the regulation of E3 ligase expression and skeletal muscle atrophy during unloading, we used VX-745, a selective p38α inhibitor. Three groups of rats were used: non-treated control (C), 3 days of unloading/hindlimb suspension (HS), and 3 days HS with VX-745 inhibitor (HSVX; 10 mg/kg/day). Total weight of soleus muscle in HS group was reduced compared to C (72.3 ± 2.5 vs 83.0 ± 3 mg, respectively), whereas muscle weight in the HSVX group was maintained (84.2 ± 5 mg). The expression of muscle RING-finger protein-1 (MuRF1) mRNA was significantly increased in the HS group (165%), but not in the HSVX group (127%), when compared with the C group. The expression of muscle-specific E3 ubiquitin ligases muscle atrophy F-box (MAFbx) mRNA was increased in both HS and HSVX groups (294% and 271%, respectively) when compared with C group. The expression of ubiquitin mRNA was significantly higher in the HS (423%) than in the C and HSVX (200%) groups. VX-745 treatment blocked unloading-induced upregulation of calpain-1 mRNA expression (HS: 120%; HSVX: 107%). These results indicate that p38α-MAPK signaling regulates MuRF1 but not MAFbx E3 ligase expression and inhibits skeletal muscle atrophy during early stages of unloading.
Assuntos
Proteína Quinase 14 Ativada por Mitógeno/metabolismo , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/metabolismo , Inibidores de Proteínas Quinases/administração & dosagem , Piridazinas/administração & dosagem , Pirimidinas/administração & dosagem , Animais , Calpaína/genética , Calpaína/metabolismo , Elevação dos Membros Posteriores , Interleucina-6/metabolismo , Masculino , Proteína Quinase 14 Ativada por Mitógeno/antagonistas & inibidores , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Atrofia Muscular/tratamento farmacológico , Biossíntese de Proteínas/efeitos dos fármacos , Biossíntese de Proteínas/genética , Proteólise/efeitos dos fármacos , Ratos , Ratos Wistar , Proteínas Ligases SKP Culina F-Box/genética , Proteínas Ligases SKP Culina F-Box/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina/genética , Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
BACKGROUND: Animal studies showed that alcoholic myopathy is characterized by the reduction in myofiber cross-sectional area (CSA) and by impaired anabolic signaling. The goal of this study was to compare changes in CSA and fiber type composition with modifications in anabolic and catabolic signaling pathways at the early stages of alcohol misuse in humans. METHODS: Skeletal muscle samples from 7 male patients with chronic alcohol abuse (AL; 47.7 ± 2.0 years old; alcohol misuse duration 7.7 ± 0.6 years) were compared with muscle from a control group of 7 healthy men (C; 39.7 ± 5.0 years old). Biopsies from vastus lateralis muscles were taken and analyzed for the changes in fiber type composition, fiber CSA, and for the alterations in anabolic and catabolic signaling pathways. RESULTS: AL patients did not have detectable clinical myopathy symptoms or muscle fiber atrophy, but the relative proportion of fast fibers was increased. There was a significant decrease in IGF-1 in plasma and IRS-1 protein content in muscle of AL group. Levels of total and phosphorylated p70S6K1, GSK3ß, and p90RSK1 were not different between AL and C groups. Muscle of AL patients had increased mRNA expression of HSP70 and HSP90. A marker of anabolic pathway p-4E-BP1 was decreased, while catabolic markers (MuRF-1, MAFbx, ubiquitinated proteins) were increased in AL patients when compared with C group. CONCLUSIONS: At the early stages of alcohol misuse in humans, changes in the regulation of anabolic and catabolic signaling pathways precede the development of skeletal muscle atrophy and manifestation of clinical symptoms of alcoholic myopathy.
Assuntos
Alcoolismo/metabolismo , Alcoolismo/patologia , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Transdução de Sinais/fisiologia , Adulto , Alcoolismo/complicações , Humanos , Masculino , Metabolismo/efeitos dos fármacos , Metabolismo/fisiologia , Pessoa de Meia-Idade , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/induzido quimicamente , Atrofia Muscular/metabolismo , Atrofia Muscular/patologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Unloading causes rapid skeletal muscle atrophy due to increased protein degradation via activation of calpains and decreased protein synthesis. Our study elucidated role of calpain-1 in the regulation of ubiquitin proteasome pathway (UPP) and anabolic processes mediated by Akt-mTOR-p70S6K and MAPK-Erk (p90RSK) signaling. We hypothesized that blocking calpain will inhibit activation of UPP and decrease protein degradation resulting in reduction of unloading-induced skeletal muscle atrophy. Rats were divided into three groups: non-treated control (C), three day hindlimb suspension with (HSPD) or without (HS) treatment with calpain inhibitor PD150606. When compared with control PD150606 treatment during unloading: 1) attenuated loss of muscle mass, 2) prevented accumulation of calpain-1 (1.8-fold in HS vs 1.3-fold in HSPD) and ubiquitin (2.3-fold in HS vs 0.7-fold in HSPD) mRNA and ubiquitinated proteins (1.6-fold in HS vs 0.8-fold in HSPD), 3) prevented decrease in the pAkt (0.4-fold in HS vs 1-fold in HSPD) and pFOXO3 (0.2-fold in HS vs 1.2-fold in HSPD) levels, 4) prevented increase in MAFbx (3.8-fold in HS vs 1.3-fold in HSPD) and eEF2k (1.8-fold in HS vs 0.6-fold in HSPD) mRNA. Our study indicates that blocking of calpain during unloading decreases skeletal muscle atrophy by inhibiting UPP activation and preserving anabolic signaling.
Assuntos
Acrilatos/farmacologia , Calpaína/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Proteínas Musculares/metabolismo , Atrofia Muscular/metabolismo , Proteólise/efeitos dos fármacos , Animais , Calpaína/antagonistas & inibidores , Imobilização , Masculino , Proteínas Musculares/antagonistas & inibidores , Atrofia Muscular/tratamento farmacológico , Atrofia Muscular/patologia , Ratos , Ratos WistarRESUMO
Skeletal muscle abnormalities and atrophy during unloading are accompanied by the accumulation of excess calcium in the sarcoplasm. We hypothesized that calcium accumulation may occur, among other mechanisms, due to the inhibition of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) activity. Consequently, the use of the SERCA activator will reduce the level of calcium in the sarcoplasm and prevent the negative consequences of muscle unloading. Wistar rats were randomly assigned into one of three groups (eight rats per group): control rats with placebo (C), 7 days of unloading/hindlimb suspension with placebo (7HS), and 7 days of unloading treated with SERCA activator CDN1163 (7HSC). After seven days of unloading the soleus muscle, the 7HS group displayed increased fatigue in the ex vivo test, a significant increase in the level of calcium-dependent CaMK II phosphorylation and the level of tropomyosin oxidation, as well as a decrease in the content of mitochondrial DNA and protein, slow-type myosin mRNA, and the percentage of slow-type muscle fibers. All of these changes were prevented in the 7HSC group. Moreover, treatment with CDN1163 blocked a decrease in the phosphorylation of p70S6k, an increase in eEF2 phosphorylation, and an increase in MuRF-1 mRNA expression. Nevertheless, there were no differences in the degree of fast and slow muscle fiber atrophy between the 7HS and 7HSC groups. Conclusion: SERCA activation during 7 days of unloading prevented an increase in soleus fatigue, the decrease of slow-type myosin, mitochondrial markers, and markers of calcium homeostasis but had no effect on muscle atrophy.
Assuntos
Cálcio , Músculo Esquelético , Ratos , Animais , Ratos Wistar , Atrofia Muscular/tratamento farmacológico , Retículo EndoplasmáticoRESUMO
Current study tested a hypothesis that during skeletal muscle unloading, calcium-dependent signaling pathways, markers of protein synthesis, and expression of E3 ubiquitin ligases can be regulated by metformin. Thirty-two male Wistar rats were randomly assigned into one of four groups: nontreated control (3C), control rats treated with metformin (3CM), 3 days of unloading/hindlimb suspension with placebo (3HS), and 3 days of unloading treated with metformin (3HSM). In soleus muscle of HS group level of phospho-AMP-activated protein kinase (p-AMPK) was decreased by 46% while ATP content was increased by 49% when compared with the control group. There was an increase of the level of phospho-CaMK II (483%) and an upregulation of Calcineurin (CaN), SERCA2a, and Calpain-1 mRNA expression (87%, 41%, and 62%, respectively, P < 0.05) in the HS group relative to the control. HS group also had increased mRNA expression of MuRF1, MAFbx, and ubiquitin (167%, 146%, and 191%, respectively, P < 0.05) when compared with the control soleus muscle. Metformin treatment impeded unloading-induced changes in soleus muscle. In conclusion, metformin treatment during 3 days of soleus muscle unloading: 1) prevented the decrease of p-AMPK and increase of ATP content; 2) affected regulation of calcium-dependent signaling pathways via level of CaMK II phosphorylation or CaMK II, CaN, SERCA2a, and Calpain-1 mRNA expression; 3) attenuated an increase in the expression of critical markers of ubiquitin-proteasome pathways MuRF1, MAFbx, and ubiquitin while not affecting the unloading-induced increase of ULK-1 marker of autophagic/lysosomal pathway.NEW & NOTEWORTHY Current study for the first time tested the hypothesis that during 3 days of soleus muscle unloading, calcium-dependent signaling pathways, markers of protein synthesis, and the expression of E3 ubiquitin ligases can be regulated by metformin. Treatment with metformin during unloading: prevented the decrease of p-AMPK and increase of ATP content, affected regulation of calcium-dependent signaling pathways, and attenuated an increase of critical markers of ubiquitin-proteasome pathways. Nevertheless, metformin treatment has not prevented soleus muscle atrophy.
Assuntos
Metformina , Ubiquitina , Masculino , Ratos , Animais , Ubiquitina/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Cálcio/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Metformina/farmacologia , Metformina/uso terapêutico , Calpaína/metabolismo , Ratos Wistar , Elevação dos Membros Posteriores/fisiologia , Atrofia Muscular/metabolismo , Músculo Esquelético/fisiologia , Calcineurina/metabolismo , RNA Mensageiro/metabolismo , Trifosfato de Adenosina/metabolismoRESUMO
The dystrophinglycoprotein complex (DGC) provides an essential link from the muscle fibre cytoskeleton to the extracellular matrix. In dystrophic humans and mdx mice, mutations in the dystrophin gene disrupt the structure of the DGC causing severe damage to muscle fibres. In frog muscles, transmission of force laterally from an activated fibre to the muscle surface occurs without attenuation, but lateral transmission of force has not been demonstrated in mammalian muscles. A unique 'yoke' apparatus was developed that attached to the epimysium of muscles midway between the tendons and enabled the measurement of lateral force. We now report that in muscles of young wild-type (WT) mice and rats, compared over a wide range of longitudinal forces, forces transmitted laterally showed little or no decrement. In contrast, for muscles of mdx mice and very old rats, forces transmitted laterally were impaired severely. Muscles of both mdx mice and very old rats showed major reductions in the expression of dystrophin. We conclude that during contractions, forces developed by skeletal muscles of young WT mice and rats are transmitted laterally from fibre to fibre through the DGC without decrement. In contrast, in muscles of dystrophic or very old animals, disruptions in DGC structure and function impair lateral transmission of force causing instability and increased susceptibility of fibres to contraction-induced injury.
Assuntos
Envelhecimento/fisiologia , Contração Muscular/fisiologia , Força Muscular/fisiologia , Músculo Esquelético/fisiologia , Distrofia Muscular Animal/fisiopatologia , Animais , Western Blotting , Distrofina/genética , Distrofina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos mdx , Distrofia Muscular Animal/metabolismo , RatosRESUMO
Ossabaw swine fed excess kilocalorie diet develop metabolic syndrome (MS) characterized by obesity, hypertension, insulin resistance, and glucose intolerance with/without dyslipidemia. The purpose of this study was to test the hypothesis that MS would have a detrimental effect on skeletal muscle structure and cause changes in the expression of myosin heavy chains (MHCs). Adult male Ossabaw swine were fed for 24 wk high-fructose or high-fat/cholesterol/fructose diets to induce normolipidemic MS (MetS) or dyslipidemic MS (DMetS), respectively, and were compared with the lean swine on control diet. MetS swine showed mild MS, lacking increases in total and low density lipoprotein (LDL) cholesterol, both of which were highly upregulated in DMetS swine. There was an â¼1.2-fold increase in the cross-sectional areas of muscle fibers in MetS and DMetS groups compared with control for biceps femoris and plantaris muscles. In plantaris muscles, DMetS diet caused an â¼2-fold decrease in slow MHC mRNA and protein expression and an â¼1.2- to 1.8-fold increase in the number of intramyocellular lipid (IMCL) droplets without large changes in the size of the droplets. There was a trend to the decrease in slow MHC expression in muscles of swine on MetS diet. The number of IMCL droplets in muscle fibers of the MetS group was comparable to controls. These data correlate well with the data on total plasma cholesterol (control = 60, MetS = 70, and DMetS = 298 mg/dl) and LDL (control = 29, MetS = 30, and DMetS = 232 mg/dl). We conclude that structural changes observed in skeletal muscle of obese Ossabaw swine correlate with those previously reported for obese humans.
Assuntos
Dieta , Síndrome Metabólica/metabolismo , Músculo Esquelético/metabolismo , Obesidade/metabolismo , Anatomia Transversal , Animais , Western Blotting , Colesterol/sangue , Imuno-Histoquímica , Metabolismo dos Lipídeos/fisiologia , Masculino , Síndrome Metabólica/genética , Síndrome Metabólica/patologia , Fibras Musculares de Contração Rápida/patologia , Fibras Musculares de Contração Rápida/fisiologia , Fibras Musculares Esqueléticas/patologia , Fibras Musculares Esqueléticas/fisiologia , Fibras Musculares de Contração Lenta/patologia , Fibras Musculares de Contração Lenta/fisiologia , Músculo Esquelético/citologia , Músculo Esquelético/patologia , Cadeias Pesadas de Miosina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Suínos , Porco Miniatura , Triglicerídeos/sangueRESUMO
An association between oxidative stress and muscle atrophy and weakness in vivo is supported by elevated oxidative damage and accelerated loss of muscle mass and force with aging in CuZn-superoxide dismutase-deficient (Sod1(-/-)) mice. The purpose was to determine the basis for low specific force (N/cm(2)) of gastrocnemius muscles in Sod1(-/-) mice and establish the extent to which structural and functional changes in muscles of Sod1(-/-) mice resemble those associated with normal aging. We tested the hypothesis that muscle weakness in Sod1(-/-) mice is due to functionally denervated fibers by comparing forces during nerve and direct muscle stimulation. No differences were observed for wild-type mice at any age in the forces generated in response to nerve and muscle stimulation. Nerve- and muscle-stimulated forces were also not different for 4-wk-old Sod1(-/-) mice, whereas, for 8- and 20-mo-old mice, forces during muscle stimulation were 16 and 30% greater, respectively, than those obtained using nerve stimulation. In addition to functional evidence of denervation with aging, fiber number was not different for Sod1(-/-) and wild-type mice at 4 wk, but 50% lower for Sod1(-/-) mice by 20 mo, and denervated motor end plates were prevalent in Sod1(-/-) mice at both 8 and 20 mo and in WT mice by 28 mo. The data suggest ongoing denervation in muscles of Sod1(-/-) mice that results in fiber loss and muscle atrophy. Moreover, the findings support using Sod1(-/-) mice to explore mechanistic links between oxidative stress and the progression of deficits in muscle structure and function.
Assuntos
Contração Muscular , Fibras Musculares Esqueléticas/enzimologia , Força Muscular , Debilidade Muscular , Músculo Esquelético/enzimologia , Músculo Esquelético/inervação , Atrofia Muscular/enzimologia , Superóxido Dismutase/deficiência , Fatores Etários , Envelhecimento/metabolismo , Animais , Estimulação Elétrica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fibras Musculares Esqueléticas/patologia , Músculo Esquelético/patologia , Atrofia Muscular/genética , Atrofia Muscular/patologia , Atrofia Muscular/fisiopatologia , Estresse Oxidativo , Superóxido Dismutase/genética , Superóxido Dismutase-1RESUMO
Daptomycin is a lipopeptide antibiotic with strong bactericidal effects against Gram-positive bacteria and minor side effects on skeletal muscles. The type and magnitude of the early effect of daptomycin on skeletal muscles of rats was quantified by histopathology, examination of contractile properties, Evans Blue Dye uptake, and effect on the patch repair process. A single dose of daptomycin of up to 200 mg/kg had no effect on muscle fibers. A dose of 150 mg/kg of daptomycin, twice per day for 3 days, produced a small number of myofibers (Assuntos
Antibacterianos/toxicidade
, Daptomicina/toxicidade
, Músculo Esquelético/efeitos dos fármacos
, Animais
, Antibacterianos/administração & dosagem
, Peso Corporal/efeitos dos fármacos
, Membrana Celular/efeitos dos fármacos
, Membrana Celular/fisiologia
, Daptomicina/administração & dosagem
, Azul Evans
, Imuno-Histoquímica
, Injeções Intravenosas
, Masculino
, Microscopia de Fluorescência
, Contração Muscular/efeitos dos fármacos
, Contração Muscular/fisiologia
, Fibras Musculares Esqueléticas/efeitos dos fármacos
, Músculo Esquelético/patologia
, Músculo Esquelético/fisiologia
, Tamanho do Órgão/efeitos dos fármacos
, Ratos
, Ratos Sprague-Dawley
, Sarcolema/efeitos dos fármacos
RESUMO
Skeletal muscle is composed of muscle fibers and an extracellular matrix (ECM). The collagen fiber network of the ECM is a major contributor to the passive force of skeletal muscles at high strain. We investigated the effect of aging on the biomechanical and structural properties of epimysium of the tibialis anterior muscles (TBA) of rats to understand the mechanisms responsible for the age-related changes. The biomechanical properties were tested directly in vitro by uniaxial extension of epimysium. The presence of age-related changes in the arrangement and size of the collagen fibrils in the epimysium was examined by scanning electron microscopy (SEM). A mathematical model was subsequently developed based on the structure-function relationships that predicted the compliance of the epimysium. Biomechanically, the epimysium from old rats was much stiffer than that of the young rats. No differences were found in the ultrastructure and thickness of the epimysium or size of the collagen fibrils between young and old rats. The changes in the arrangement and size of the collagen fibrils do not appear to be the principal cause of the increased stiffness of the epimysium from the old rats. Other changes in the structural composition of the epimysium from old rats likely has a strong effect on the increased stiffness. The age-related increase in the stiffness of the epimysium could play an important role in the impaired lateral force transmission in the muscles of the elderly.
Assuntos
Envelhecimento/fisiologia , Matriz Extracelular/fisiologia , Modelos Biológicos , Músculo Esquelético/fisiologia , Animais , Anisotropia , Fenômenos Biomecânicos/métodos , Simulação por Computador , Elasticidade , Ratos , Ratos Endogâmicos Lew , Estresse MecânicoRESUMO
A micromechanical model has been developed to investigate the mechanical properties of the epimysium. In the present model, the collagen fibers in the epimysium are embedded randomly in the ground substance. Two parallel wavy collagen fibers and the surrounding ground substance are used as the repeat unit (unit cell), and the epimysium is considered as an aggregate of unit cells. Each unit cell is distributed in the epimysium with some different angle to the muscle fiber direction. The model allows the progressive straightening of the collagen fiber as well as the effects of fiber reorientation. The predictions of the model compare favorably against experiment. The effects of the collagen fiber volume fraction, collagen fiber waviness at the rest length and the mechanical properties of the collagen fibers and the ground substance are analyzed. This model allows the analysis of mechanical behavior of most soft tissues if appropriate experimental data are available.
Assuntos
Colágeno/fisiologia , Matriz Extracelular/fisiologia , Modelos Biológicos , Músculo Esquelético/fisiologia , Algoritmos , Animais , Fenômenos Biomecânicos , Elasticidade , Ratos , Estresse MecânicoRESUMO
We tested whether NF-κB pathway is indispensable for the increase in expression of E3-ligases and unloading-induced muscle atrophy using IKKß inhibitor IMD-0354. Three groups of rats were used: nontreated control (C), 3 days of unloading/hindlimb suspension with (HS+IMD) or without (HS) IMD-0354. Levels of IκBα were higher in HS+IMD (1.16-fold) and lower in HS (0.82-fold) when compared with C group. IMD-0354 treatment during unloading: had no effect on loss of muscle mass; increased mRNA levels of MuRF1 and MAFbx; increased levels of pFoxO3; and had no effect on levels of Bcl-3, p105, and p50 proteins. Our study for the first time showed that inhibiting IKKß in vivo during 3-day unloading failed to diminish expression of ubiquitin ligases and prevent muscle atrophy.
Assuntos
Quinase I-kappa B/antagonistas & inibidores , Proteínas Musculares/genética , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Proteínas com Motivo Tripartido/genética , Ubiquitina-Proteína Ligases/genética , Animais , Benzamidas/farmacologia , Inibidores Enzimáticos/farmacologia , Proteína Forkhead Box O3/metabolismo , Elevação dos Membros Posteriores/efeitos adversos , Quinase I-kappa B/metabolismo , Masculino , Proteínas Musculares/metabolismo , Músculo Esquelético/efeitos dos fármacos , Atrofia Muscular/etiologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Proteínas Ligases SKP Culina F-Box/genética , Proteínas Ligases SKP Culina F-Box/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Damage to peripheral nerve tissue may cause loss of function in both the nerve and the targeted muscles it innervates. This study compared the repair capability of engineered nerve conduit (ENC), engineered fibroblast conduit (EFC), and autograft in a 10-mm tibial nerve gap. ENCs were fabricated utilizing primary fibroblasts and the nerve cells of rats on embryonic day 15 (E15). EFCs were fabricated utilizing primary fibroblasts only. Following a 12-week recovery, nerve repair was assessed by measuring contractile properties in the medial gastrocnemius muscle, distal motor nerve conduction velocity in the lateral gastrocnemius, and histology of muscle and nerve. The autografts, ENCs and EFCs reestablished 96%, 87% and 84% of native distal motor nerve conduction velocity in the lateral gastrocnemius, 100%, 44% and 44% of native specific force of medical gastrocnemius, and 63%, 61% and 67% of native medial gastrocnemius mass, respectively. Histology of the repaired nerve revealed large axons in the autograft, larger but fewer axons in the ENC repair, and many smaller axons in the EFC repair. Muscle histology revealed similar muscle fiber cross-sectional areas among autograft, ENC and EFC repairs. In conclusion, both ENCs and EFCs promoted nerve regeneration in a 10-mm tibial nerve gap repair, suggesting that the E15 rat nerve cells may not be necessary for nerve regeneration, and EFC alone can suffice for peripheral nerve injury repair.
RESUMO
During muscle contraction, the integrity of the myotendinous junction (MTJ) is important for the transmission of force from muscle to tendon. We evaluated the contractile and structural characteristics of 3-dimensional (3-D) skeletal muscle constructs co-cultured with engineered self-organized tendon constructs (n = 4), or segments of adult (n = 4) or fetal (n = 5) rat-tail tendon. We hypothesized that the co-culture of tendon and muscle would produce constructs with viable muscle-tendon interfaces that remain intact during generation of force. Construct diameter (lm) and maximum isometric force (microN) were measured, and specific force (kPa) was determined. After measure of force, constructs were loaded at a constant strain rate until failure and surface strains were recorded optically across the tendon, the muscle and the interface and used to determine the tangent modulus (passive stiffness) of the construct. Frozen samples were used for Trichrome Masson staining and immunofluorescent analysis of the MTJ-specific protein paxillin. No differences were observed between the groups with respect to diameter, maximum force, or specific force. The MTJ was robust and withstood tensile loading beyond the physiological strain range. The majority of the constructs failed in the muscle region. At the MTJ, there is an increase in the expression and localization of paxillin. In conclusion, using 3 sources of tendon tissue, we successfully engineered 3-D muscle-tendon constructs with functionally viable MTJ, characterized by structural features and protein expression patterns resembling neonatal MTJs in vivo.