RESUMEN
Skeletal muscle dysfunction and atrophy are characteristic features accompanying chronic conditions. Epigenetic events regulate muscle mass and function maintenance. We hypothesized that the pattern of epigenetic events (muscle-enriched microRNAs and histone acetylation) and acetylation of transcription factors known to signal muscle wasting may differ between early- and late-time points in skeletal muscles of mice exposed to hindlimb immobilization (I) and recovery following I. Body and muscle weights, grip strength, muscle-enriched microRNAs, histone deacetylases (HDACs), acetylation of proteins, histones, and transcription factors (TF), myogenic TF factors, and muscle phenotype were assessed in gastrocnemius of mice exposed to periods (1, 2, 3, 7, 15, and 30 days, I groups) of hindlimb immobilization, and in those exposed to reloading for different periods of time (1, 3, 7, 15, and 30 days, R groups) following 7-day immobilization. Compared to non-immobilized controls, muscle weight, limb strength, microRNAs, especially miR-486, SIRT1 levels, and slow- and fast-twitch cross-sectional areas were decreased in mice of I groups, whereas Pax7 and acetylated FoxO1 and FoxO3 levels were increased. Muscle reloading following splint removal improved muscle mass loss, strength, and fiber atrophy, by increasing microRNAs, particularly miR-486, and SIRT1 content, while decreasing acetylated FoxO1 and FoxO3 levels. In this mouse model of disuse muscle atrophy, muscle-enriched microRNAs, especially miR-486, through Pax7 regulation delayed muscle cell differentiation following unloading of gastrocnemius muscle. Acetylation of FoxO1 and 3 seemed to drive muscle mass loss and atrophy, while deacetylation of these factors through SIRT1 would enable the muscle fibers to regenerate. J. Cell. Physiol. 232: 1415-1427, 2017. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Epigénesis Genética , Suspensión Trasera , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiología , Acetilación , Animales , Peso Corporal , Femenino , Histona Desacetilasas/metabolismo , Histonas/metabolismo , Ratones Endogámicos C57BL , MicroARNs/genética , MicroARNs/metabolismo , Desarrollo de Músculos/genética , Fibras Musculares Esqueléticas/metabolismo , Factores de Tiempo , Factores de Transcripción/metabolismo , Soporte de PesoRESUMEN
Strategies to treat cachexia are still at its infancy. Enhanced muscle protein breakdown and ubiquitin-proteasome system are common features of cachexia associated with chronic conditions including lung cancer (LC). Poly(ADP-ribose) polymerases (PARP), which play a major role in chromatin structure regulation, also underlie maintenance of muscle metabolism and body composition. We hypothesized that protein catabolism, proteolytic markers, muscle fiber phenotype, and muscle anabolism may improve in respiratory and limb muscles of LC-cachectic Parp-1-deficient (Parp-1-/- ) and Parp-2-/- mice. In diaphragm and gastrocnemius of LC (LP07 adenocarcinoma) bearing mice (wild type, Parp-1-/- , and Parp-2-/- ), PARP activity (ADP-ribose polymers, pADPr), redox balance, muscle fiber phenotype, apoptotic nuclei, tyrosine release, protein ubiquitination, muscle-specific E3 ligases, NF-κB signaling pathway, markers of muscle anabolism (Akt, mTOR, p70S6K, and mitochondrial DNA) were evaluated along with body and muscle weights, and limb muscle force. Compared to wild type cachectic animals, in both respiratory and limb muscles of Parp-1-/- and Parp-2-/- cachectic mice: cancer induced-muscle wasting characterized by increased PARP activity, protein oxidation, tyrosine release, and ubiquitin-proteasome system (total protein ubiquitination, atrogin-1, and 20S proteasome C8 subunit) were blunted, the reduction in contractile myosin and atrophy of the fibers was attenuated, while no effects were seen in other structural features (inflammatory cells, internal or apoptotic nuclei), and markers of muscle anabolism partly improved. Activation of either PARP-1 or -2 is likely to play a role in muscle protein catabolism via oxidative stress, NF-κB signaling, and enhanced proteasomal degradation in cancer-induced cachexia. Therapeutic potential of PARP activity inhibition deserves attention.
Asunto(s)
Caquexia/etiología , Neoplasias Pulmonares/complicaciones , Fibras Musculares Esqueléticas/enzimología , Proteínas Musculares/metabolismo , Músculo Esquelético/enzimología , Estrés Oxidativo , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Proteolisis , Animales , Apoptosis , Biomarcadores/metabolismo , Caquexia/enzimología , Caquexia/genética , Caquexia/patología , Línea Celular Tumoral , Diafragma/enzimología , Diafragma/patología , Femenino , Genotipo , Neoplasias Pulmonares/enzimología , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Ratones de la Cepa 129 , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias Musculares/metabolismo , Mitocondrias Musculares/patología , Fibras Musculares Esqueléticas/patología , Músculo Esquelético/patología , FN-kappa B/metabolismo , Tamaño de los Órganos , Fenotipo , Poli(ADP-Ribosa) Polimerasa-1/deficiencia , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasas/deficiencia , Poli(ADP-Ribosa) Polimerasas/genética , Complejo de la Endopetidasa Proteasomal/metabolismo , Transducción de Señal , Factores de Tiempo , UbiquitinaciónRESUMEN
BACKGROUND: Current treatment options for cachexia, which impairs disease prognosis, are limited. Muscle-enriched microRNAs and protein acetylation are involved in muscle wasting including lung cancer (LC) cachexia. Poly(ADP-ribose) polymerases (PARP) are involved in muscle metabolism. We hypothesized that muscle-enriched microRNA, protein hyperacetylation, and expression levels of myogenic transcription factors (MTFs) and downstream targets, muscle loss and function improve in LC cachectic Parp-1(−/−) and Parp-2(−/−) mice. METHODS: Body and muscle weights, grip strength, muscle phenotype, muscle-enriched microRNAs (miR-1, -133, -206, and -486), protein acetylation, acetylated levels of FoxO1, FoxO3, and PGC-1α, histone deacetylases (HDACs) including SIRT1, MTFs, and downstream targets (α-actin, PGC-1α, and creatine kinase) were evaluated in diaphragm and gastrocnemius of LC (LP07 adenocarcinoma) wild type (WT), Parp-1(−/−) and Parp-2−/− mice. RESULTS: Compared to WT cachectic animals, in both respiratory and limb muscles of Parp-1(−/−) and Parp-2(−/−) cachectic mice: downregulation of muscle-specific microRNAs was counterbalanced especially in gastrocnemius of Parp-1(−/−) mice; increased protein acetylation was attenuated (improvement in HDAC3, SIRT-1, and acetylated FoxO3 levels in both muscles, acetylated FoxO1 levels in the diaphragm); reduced MTFs and creatine kinase levels were mitigated; body and muscle weights, strength, and muscle fiber sizes improved, while tumor weight and growth decreased. CONCLUSIONS: These molecular findings may explain the improvements seen in body and muscle weights, limb muscle force and fiber sizes in both Parp-1(−/−) and Parp-2(−/−) cachectic mice. GENERAL SIGNIFICANCE: PARP-1 and -2 play a role in cancer-induced cachexia, thus selective pharmacological inhibition of PARP-1 and -2 may be of interest in clinical settings.
Asunto(s)
Caquexia/metabolismo , Neoplasias Pulmonares/metabolismo , MicroARNs/genética , Poli(ADP-Ribosa) Polimerasas/metabolismo , Acetilación , Animales , Caquexia/genética , Neoplasias Pulmonares/genética , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Poli(ADP-Ribosa) Polimerasa-1 , Poli(ADP-Ribosa) Polimerasas/genéticaRESUMEN
Patients with chronic heart failure (CHF) experience exercise intolerance, fatigue and muscle wasting, which negatively influence their survival. We hypothesized that treatment with either the antioxidant N-acetyl cysteine (NAC) or the proteasome inhibitor bortezomib of rats with monocrotaline-induced CHF may restore inspiratory and limb muscle mass, function, and structure through several molecular mechanisms involved in protein breakdown and metabolism in the diaphragm and gastrocnemius. In these muscles of CHF-cachectic rats with and without treatment with NAC or bortezomib (N = 10/group) and non-cachectic controls, proteolysis (tyrosine release, proteasome activities, ubiquitin-proteasome markers), oxidative stress, inflammation, mitochondrial function, myosin, NF-κB transcriptional activity, muscle structural abnormalities, and fiber morphometry were analyzed together with muscle and cardiac functions. In diaphragm and gastrocnemius of CHF-cachectic rats, tyrosine release, proteasome activity, protein ubiquitination, atrogin-1, MURF-1, NF-κB activity, oxidative stress, inflammation, and structural abnormalities were increased, while muscle and cardiac functions, myosin content, slow- and fast-twitch fiber sizes, and mitochondrial activity were decreased. Concomitant treatment of CHF-cachectic rats with NAC or bortezomib improved protein catabolism, oxidative stress, inflammation, muscle fiber sizes, function and damage, superoxide dismutase and myosin levels, mitochondrial function (complex I, gastrocnemius), cardiac function and decreased NF-κB transcriptional activity in both muscles. Treatment of CHF-cachectic animals with NAC or bortezomib attenuated the functional (heart, muscles), biological, and structural alterations in muscles. Nonetheless, future studies conducted in actual clinical settings are warranted in order to assess the potential beneficial effects and safety concerns of these pharmacological agents on muscle mass loss and wasting in CHF-cachectic patients.
Asunto(s)
Acetilcisteína/administración & dosificación , Bortezomib/administración & dosificación , Insuficiencia Cardíaca/tratamiento farmacológico , Estrés Oxidativo/efectos de los fármacos , Animales , Diafragma/efectos de los fármacos , Diafragma/metabolismo , Diafragma/patología , Insuficiencia Cardíaca/inducido químicamente , Insuficiencia Cardíaca/metabolismo , Humanos , Mitocondrias/metabolismo , Monocrotalina/toxicidad , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , FN-kappa B/metabolismo , Proteolisis/efectos de los fármacos , RatasRESUMEN
Cachexia is a relevant comorbid condition of chronic diseases including cancer. Inflammation, oxidative stress, autophagy, ubiquitin-proteasome system, nuclear factor (NF)-κB, and mitogen-activated protein kinases (MAPK) are involved in the pathophysiology of cancer cachexia. Currently available treatment is limited and data demonstrating effectiveness in in vivo models are lacking. Our objectives were to explore in respiratory and limb muscles of lung cancer (LC) cachectic mice whether proteasome, NF-κB, and MAPK inhibitors improve muscle mass and function loss through several molecular mechanisms. Body and muscle weights, limb muscle force, protein degradation and the ubiquitin-proteasome system, signaling pathways, oxidative stress and inflammation, autophagy, contractile and functional proteins, myostatin and myogenin, and muscle structure were evaluated in the diaphragm and gastrocnemius of LC (LP07 adenocarcinoma) bearing cachectic mice (BALB/c), with and without concomitant treatment with NF-κB (sulfasalazine), MAPK (U0126), and proteasome (bortezomib) inhibitors. Compared to control animals, in both respiratory and limb muscles of LC cachectic mice: muscle proteolysis, ubiquitinated proteins, autophagy, myostatin, protein oxidation, FoxO-1, NF-κB and MAPK signaling pathways, and muscle abnormalities were increased, while myosin, creatine kinase, myogenin, and slow- and fast-twitch muscle fiber size were decreased. Pharmacological inhibition of NF-κB and MAPK, but not the proteasome system, induced in cancer cachectic animals, a substantial restoration of muscle mass and force through a decrease in muscle protein oxidation and catabolism, myostatin, and autophagy, together with a greater content of myogenin, and contractile and functional proteins. Attenuation of MAPK and NF-κB signaling pathway effects on muscles is beneficial in cancer-induced cachexia.
Asunto(s)
Autofagia , Caquexia/tratamiento farmacológico , Caquexia/etiología , Neoplasias Pulmonares/complicaciones , Neoplasias Pulmonares/tratamiento farmacológico , Debilidad Muscular/tratamiento farmacológico , Músculo Esquelético/patología , Proteolisis , Animales , Autofagia/efectos de los fármacos , Ácidos Borónicos/farmacología , Ácidos Borónicos/uso terapéutico , Bortezomib , Butadienos/farmacología , Butadienos/uso terapéutico , Línea Celular Tumoral , Inflamación/complicaciones , Inflamación/patología , Inflamación/fisiopatología , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/fisiopatología , Ratones , Ratones Endogámicos BALB C , Contracción Muscular/efectos de los fármacos , Proteínas Musculares/metabolismo , Debilidad Muscular/etiología , Debilidad Muscular/patología , Debilidad Muscular/fisiopatología , Músculo Esquelético/anomalías , Músculo Esquelético/efectos de los fármacos , Músculo Esquelético/fisiopatología , FN-kappa B/metabolismo , Nitrilos/farmacología , Nitrilos/uso terapéutico , Oxidación-Reducción , Proteolisis/efectos de los fármacos , Pirazinas/farmacología , Pirazinas/uso terapéutico , Transducción de Señal/efectos de los fármacos , Sulfasalazina/farmacología , Sulfasalazina/uso terapéutico , Tirosina/metabolismo , Aumento de Peso/efectos de los fármacosRESUMEN
BACKGROUND: Disuse muscle atrophy is a major comorbidity in patients with chronic diseases including cancer. We sought to explore the kinetics of molecular mechanisms shown to be involved in muscle mass loss throughout time in a mouse model of disuse muscle atrophy and recovery following immobilization. METHODS: Body and muscle weights, grip strength, muscle phenotype (fiber type composition and morphometry and muscle structural alterations), proteolysis, contractile proteins, systemic troponin I, and mitochondrial content were assessed in gastrocnemius of mice exposed to periods (1, 2, 3, 7, 15 and 30 days) of non-invasive hindlimb immobilization (plastic splint, I cohorts) and in those exposed to reloading for different time-points (1, 3, 7, 15, and 30 days, R cohorts) following a seven-day period of immobilization. Groups of control animals were also used. RESULTS: Compared to non-exposed controls, muscle weight, limb strength, slow- and fast-twitch cross-sectional areas, mtDNA/nDNA, and myosin content were decreased in mice of I cohorts, whereas tyrosine release, ubiquitin-proteasome activity, muscle injury and systemic troponin I levels were increased. Gastrocnemius reloading following splint removal improved muscle mass loss, strength, fiber atrophy, injury, myosin content, and mtDNA/nDNA, while reducing ubiquitin-proteasome activity and proteolysis. CONCLUSIONS: A consistent program of molecular and cellular events leading to reduced gastrocnemius muscle mass and mitochondrial content and reduced strength, enhanced proteolysis, and injury, was seen in this non-invasive mouse model of disuse muscle atrophy. Unloading of the muscle following removal of the splint significantly improved the alterations seen during unloading, characterized by a specific kinetic profile of molecular events involved in muscle regeneration. These findings have implications in patients with chronic diseases including cancer in whom physical activity may be severely compromised.
Asunto(s)
Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Animales , Enfermedad Crónica , ADN/análisis , Modelos Animales de Enfermedad , Femenino , Inmovilización/efectos adversos , Ratones , Ratones Endogámicos C57BL , Mitocondrias Musculares/metabolismo , Mitocondrias Musculares/fisiología , Fuerza Muscular/fisiología , Músculo Esquelético/química , Músculo Esquelético/patología , Músculo Esquelético/fisiopatología , Atrofia Muscular/fisiopatología , Proteolisis , Reacción en Cadena en Tiempo Real de la Polimerasa , Factores de Tiempo , Troponina I/análisisRESUMEN
Mechanical ventilation (MV), using high tidal volumes (V(T)), causes lung (ventilator-induced lung injury [VILI]) and distant organ injury. Additionally, sepsis is characterized by increased oxidative stress. We tested whether MV is associated with enhanced oxidative stress in sepsis, the commonest underlying condition in clinical acute lung injury. Protein carbonylation and nitration, antioxidants, and inflammation (immunoblotting) were evaluated in diaphragm, gastrocnemius, soleus, myocardium, and lungs of nonseptic and septic (cecal ligation and puncture 24 hours before MV) rats undergoing MV (n = 7 per group) for 150 minutes using 3 different strategies (low V(T) [V(T) = 9 mL/kg], moderate V(T) [V(T) = 15 mL/kg], and high V(T) [V(T) = 25 mL/kg]) and in nonventilated control animals. Compared with nonventilated control animals, in septic and nonseptic rodents (1) diaphragms, limb muscles, and myocardium of high-V(T) rats exhibited a decrease in protein oxidation and nitration levels, (2) antioxidant levels followed a specific fiber-type distribution in slow- and fast-twitch muscles, (3) tumor necrosis factor α (TNF-α) levels were higher in respiratory and limb muscles, whereas no differences were observed in myocardium, and (4) in lungs, protein oxidation was increased, antioxidants were rather decreased, and TNF-α remained unmodified. In this model of VILI, oxidative stress does not occur in distant organs or skeletal muscles of rodents after several hours of MV with moderate-to-high V(T), whereas protein oxidation levels were increased in the lungs of the animals. Inflammatory events were moderately expressed in skeletal muscles and lungs of the MV rats. Concomitant sepsis did not strongly affect the MV-induced effects on muscles, myocardium, or lungs in the rodents.