RESUMO
Background and Objectives: Nandrolone decanoate (ND) is the most widely used among the anabolic androgenic steroids (AAS), synthetic substances derived from testosterone, to improve muscular and health gains associated with exercises. The AAS leads to physical performance enhancement and presents anti-aging properties, but its abuse is associated with several adverse effects. Supraphysiological doses of AAS with or without physical exercise can cause morphological and functional alterations in neuromuscular interactions. This study aims to investigate the effects of ND supraphysiological doses in neuromuscular interactions, focusing on the soleus muscle and its neuromuscular junctions (NMJs) in rats, associated or not with physical exercise. Materials and Methods: Forty male Sprague Dawley rats were divided into four groups: sedentary and exercised groups, with or without ND at the dose of 10 mg/kg/week. The animals were treated for eight weeks, with intramuscular injections, and the soleus muscle was collected for morphological analyses. Results: The supraphysiological doses of ND in the sedentary group caused muscle degeneration, evidenced by splitting fibers, clusters of small fibers, irregular myofibrils, altered sarcomeres, an increase in collagen deposition and in the number of type I muscle fibers (slow-twitch) and central nuclei, as well as a decrease in fibers with peripheral nuclei. On the other hand, in the ND exercise group, there was an increase in the NMJs diameter with scattering of its acetylcholine receptors, although no major morphological changes were found in the skeletal muscle. Thus, the alterations caused by ND in sedentary rats were partially reversed by physical exercise. Conclusions: The supraphysiological ND exposure in the sedentary rats promoted an increase in muscle oxidative pattern and adverse morphological alterations in skeletal muscle, resulting from damage or post-injury regeneration. In the ND-exercised rats, no major morphological changes were found. Thus, the physical exercise partially reversed the alterations caused by ND in sedentary rats.
Assuntos
Anabolizantes , Nandrolona , Ratos , Masculino , Animais , Decanoato de Nandrolona/farmacologia , Nandrolona/efeitos adversos , Anabolizantes/efeitos adversos , Ratos Wistar , Ratos Sprague-Dawley , Músculo Esquelético/fisiologia , Junção NeuromuscularRESUMO
BACKGROUND: Duchenne muscular dystrophy (DMD) is a severe form of muscular dystrophy without an effective treatment, caused by mutations in the DMD gene, leading to the absence of dystrophin. DMD results in muscle weakness, loss of ambulation, and death at an early age. Metabolomics studies in mdx mice, the most used model for DMD, reveal changes in metabolites associated with muscle degeneration and aging. In DMD, the tongue muscles exhibit unique behavior, initially showing partial protection against inflammation but later experiencing fibrosis and loss of muscle fibers. Certain metabolites and proteins, like TNF-α and TGF-ß, are potential biomarkers for dystrophic muscle characterization. METHODS: To investigate disease progression and aging, we utilized young (1 month old) and old (21-25 months old) mdx and wild-type tongue muscles. Metabolite changes were analyzed using 1H nuclear magnetic resonance, while TNF-α and TGF-ß were assessed using Western blotting to examine inflammation and fibrosis. Morphometric analysis was conducted to assess the extent of myofiber damage between groups. RESULTS: The histological analysis of the mid-belly tongue showed no differences between groups. No differences were found between the concentrations of metabolites from wild-type or mdx whole tongues of the same age. The metabolites alanine, methionine, and 3-methylhistidine were higher, and taurine and glycerol were lower in young tongues in both wild type and mdx (p < 0.001). The metabolites glycine (p < 0.001) and glutamic acid (p = 0.0018) were different only in the mdx groups, being higher in young mdx mice. Acetic acid, phosphocreatine, isoleucine, succinic acid, creatine, and the proteins TNF-α and TGF-ß had no difference in the analysis between groups (p > 0.05). CONCLUSIONS: Surprisingly, histological, metabolite, and protein analysis reveal that the tongue of old mdx remains partially spared from the severe myonecrosis observed in other muscles. The metabolites alanine, methionine, 3-methylhistidine, taurine, and glycerol may be effective for specific assessments, although their use for disease progression monitoring should be cautious due to age-related changes in the tongue muscle. Acetic acid, phosphocreatine, isoleucine, succinate, creatine, TNF-α, and TGF-ß do not vary with aging and remain constant in spared muscles, suggesting their potential as specific biomarkers for DMD progression independent of aging.
Assuntos
Distrofia Muscular de Duchenne , Camundongos , Animais , Distrofia Muscular de Duchenne/genética , Fator de Necrose Tumoral alfa/genética , Creatina , Camundongos Endogâmicos mdx , Fosfocreatina , Glicerol , Isoleucina , Fibras Musculares Esqueléticas , Metionina , Racemetionina , Ácido Acético , Alanina , Progressão da DoençaRESUMO
Background: Duchenne muscular dystrophy (DMD) is a severe form of muscular dystrophy without an effective treatment, caused by mutations in the DMD gene, leading to the absence of dystrophin. DMD results in muscle weakness, loss of ambulation and death at an early age. Metabolomics studies in mdx mice, the most used model for DMD, reveal changes in metabolites associated with muscle degeneration and aging. In DMD, the tongue muscles exhibit unique behavior, initially showing partial protection against inflammation but later experiencing fibrosis and loss of muscle fibers. Certain metabolites and proteins, like TNF-α and TGF-ß, are potential biomarkers for dystrophic muscle characterization. Methods: To investigate disease progression and aging, we utilized young (1-month old) and old (21-25 months old) mdx and wild-type mice. Metabolite changes were analyzed using 1-H Nuclear Magnetic Resonance, while TNF-α and TGF-ß were assessed using Western blotting to examine inflammation, and fibrosis. Morphometric analysis was conducted to assess the extent of myofiber damage between groups. Results: The histological analysis of the tongue showed no differences between groups. No differences were found between the concentrations of metabolites from wild type or mdx animals of the same age. The metabolites alanine, methionine, 3-methylhistidine were higher, and taurine and glycerol were lower in young animals in both wild type and mdx (p < 0.001). The metabolites glycine (p < 0.001) and glutamic acid (p = 0.0018) were different only in the mdx groups, being higher in young mdx mice. Acetic acid, phosphocreatine, isoleucine, succinic acid, creatine and the proteins TNF-α and TGF-ß had no difference in the analysis between groups (p > 0.05). Conclusions: Surprisingly, histological and protein analysis reveals that the tongue of young and old mdx animals is protected from severe myonecrosis observed in other muscles. The metabolites alanine, methionine, 3-methylhistidine, taurine, and glycerol may be effective for specific assessments, although their use for disease progression monitoring should be cautious due to age-related changes. Acetic acid, phosphocreatine, isoleucine, succinate, creatine, TNF-α, and TGF-ß do not vary with aging and remain constant in spared muscles, suggesting their potential as specific biomarkers for DMD progression independent of aging.
RESUMO
INTRODUCTION/AIMS: The mechanisms that underlie the pathogenesis of statin-associated muscle symptoms (SAMS) remain unclear. Pregnancy is associated with increased cholesterol levels. Statins may be useful during pregnancy, but their safety is uncertain. Hence, we investigated the postpartum effects of exposure to rosuvastatin and simvastatin during pregnancy in Wistar rats, targeting the neuromuscular structures. METHODS: Twenty-one pregnant Wistar rats were divided into three groups: control (C) treated with vehicle (dimethylsulfoxide + dH20), simvastatin (S) 62.5 mg/kg/day, and rosuvastatin (R) 10 mg/kg/day. Gavage was performed daily from the gestational days 8 to 20. At weaning, the postpartum mother tissues were collected and subjected to morphological and morphometric analysis of the soleus muscle, associated neuromuscular junctions (NMJs), and the sciatic nerve; protein quantification; quantification of the cholesterol and creatine kinase in the serum; and intramuscular collagen analysis. RESULTS: An increase in morphometric parameters (area, maximum and minimum diameters, Feret diameter, and minimum Feret) was observed in NMJs from the S and R groups in comparison with the C group, and there was also a loss of common NMJ circularity. The number of myofibers with central nuclei was higher in S (17 ± 3.9, P = .0083) and R (18.86 ± 14.42, P = .0498) than in C (6.8 ± 2.6). DISCUSSION: Gestational exposure to statins induced postpartum NMJ morphology alterations in soleus muscle, which may be caused by the remodeling of clusters of nicotinic acetylcholine receptors. This may be associated with the development and progression of SAMS observed in clinical practice.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases , Ratos , Gravidez , Humanos , Feminino , Animais , Ratos Wistar , Inibidores de Hidroximetilglutaril-CoA Redutases/toxicidade , Rosuvastatina Cálcica , Junção Neuromuscular/metabolismo , Músculo Esquelético/metabolismo , Sinvastatina/efeitos adversos , Período Pós-PartoRESUMO
Peripheral nerve injuries (PNI) lead to alterations in the Agrin-LRP4-MuSK pathway. This results in disaggregation of AChRs and change from epsilon (mature, innervated) to gamma (immature, denervated) subunit. Tubulization technique has been shown to be effective for PNI repair and it also allows the use of adjuvants, such as fibrin biopolymer (FB). This study evaluated the effect of the association of tubulization with FB after PNI on AChRs and associated proteins. Fifty-two adults male Wistar rats were used, distributed in 4 experimental groups: Sham Control (S), Denervated Control (D); Tubulization (TB) and Tubulization + Fibrin Biopolymer (TB+FB). Catwalk was performed every 15 days. Ninety days after surgery the right soleus muscles and ischiatic nerves were submitted to the following analyses: (a) morphological and morphometric analysis of AChRs by confocal microscopy; (b) morphological and morphometric analysis of the ischiatic nerve; (c) protein quantification of AChRs: alpha, gama, and epsilon, of Schwann cells, agrin, LRP4, MuSK, rapsyn, MMP3, MyoD, myogenin, MURF1 and atrogin-1. The main results were about the NMJs that in the TB+FB group presented morphological and morphometric approximation (compactness index; area of the AChRs and motor plate) to the S group. In addition, there were also an increase of S100 and AChRε protein expression and a decrease of MyoD. These positive association resulted in AChRs stabilization that potentiate the neuromuscular regeneration, which strengthens the use of TB for severe injuries repair and the beneficial effect of FB, along with tubulization technique.
Assuntos
Traumatismos dos Nervos Periféricos , Ratos , Animais , Masculino , Agrina/farmacologia , Agrina/metabolismo , Fibrina/metabolismo , Distribuição Normal , Ratos Wistar , Junção Neuromuscular/metabolismo , Receptores Colinérgicos/metabolismoRESUMO
High-fat (HF) diets in combination with sedentary lifestyle represent one of the major public health concerns predisposing to obesity and diabetes leading to skeletal muscle atrophy, decreased fiber diameter and muscle mass with accumulation of fat tissue resulting in loss of muscle strength. One strategy to overcome the maleficent effects of HF diet is resistance training, a strategy used to improve muscle mass, reverting the negative effects on obesity-related changes in skeletal muscle. Together with resistance training, supplementation with creatine monohydrate (CrM) in the diet has been used to improve muscle mass and strength. Creatine is a non-essential amino acid that is directly involved in the cross-bridge cycle providing a phosphate group to ADP during the initiation of muscle contraction. Besides its antioxidant and anti-inflammatory effects CrM also upregulates IGF-1 resulting in hyperthophy with an increase in muscle function. However, it is unknown whether CrM supplementation during resistance training would revert the negative effects of high-fat diet on the muscle performance. During 8 weeks we measured muscle performance to climb a 1.1m and 80° ladder with increasing load on trained rats that had received standard diet or high-fat diet, supplemented or not with CrM. We observed that the CrM supplementation up-regulated IGF-1 and phospho-AKT protein levels, suggesting an activation of the IGF1-PI3K-Akt/PKB-mTOR pathway. Moreover, despite the CrM supplementation, HF diet down-regulated several proteins of the IGF1-PI3K-Akt/PKB-mTOR pathway, suggesting that diet lipid content is crucial to maintain or improve muscle function during resistance training.
Assuntos
Creatina/farmacologia , Dieta Hiperlipídica/efeitos adversos , Músculo Esquelético/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Difosfato de Adenosina/química , Animais , Antioxidantes/metabolismo , Suplementos Nutricionais , Inflamação , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Músculo Esquelético/fisiopatologia , Atrofia Muscular/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos , Ratos Wistar , Serina-Treonina Quinases TOR/metabolismo , TemperaturaRESUMO
RESUMO A terapia por laser de baixa intensidade (Low-Level Laser Therapy - LLLT) é utilizada com frequência nas lesões musculares, mas precisa ser investigada em modelo de desnutrição. O objetivo desse estudo foi analisar os efeitos da LLLT na regeneração muscular de ratos submetidos à desnutrição e recuperação proteica. Foram utilizados 40 ratos Wistar, recém-desmamados, divididos em grupo controle (C), que consumiu ração normoproteica (14% caseína), e grupo desnutrido (D), que consumiu ração hipoproteica (6% caseína) por 45 dias e ração normoproteica até o final do experimento. Posteriormente, o músculo tibial anterior direito foi criolesado e tratado com LLLT (AsGaAl 830nm, 30mW, 20J/cm²), três vezes por semana, por 7 e 21 dias. Houve redução da área de inflamação/regeneração no grupo C21 comparado ao D21 (p<0,05), sendo mais evidente com a LLLT (C21L e D21L). O conteúdo de TNF-α foi reduzido após 21 dias da lesão. A área de densidade de tecido conjuntivo (ADTC) foi menor nos grupos C21 e C21L comparados aos respectivos grupos desnutridos (p<0,05). A LLLT reduziu a ADTC no grupo D21L quando comparado do D21 (p<0,05), porém o conteúdo de TGF-β1 não foi influenciado. A área de secção transversa (AST) da fibra muscular aumentou nos grupos 21 dias. A m-TOR apresentou maior conteúdo no grupo C21L quando comparado ao D21L (p<0,05). Concluiu-se que a LLLT favoreceu a regeneração muscular na fase tardia no modelo experimental de desnutrição pós-natal e posterior recuperação proteica.
RESUMEN La terapia por láser de baja intensidad (Low-Level Laser Therapy - LLLT) es utilizada con frecuencia en las lesiones musculares, sin embargo, precisa ser investigada en modelo de desnutrición. El objetivo de ese estudio fue analizar los efectos de la LLLT en la regeneración muscular de ratones sometidos a la desnutrición y a la recuperación proteica. Fueron utilizados 40 ratones Wistar, recién-destetados, divididos en grupo control (C), que consumió ración normoproteica (el 14% caseína), y grupo desnutrido (D), que consumió ración hipoproteica (el 6% caseína) por 45 días y ración normoproteica hasta el final del experimento. Posteriormente, el músculo tibial anterior derecho que tuvo criolesión y fue tratado con LLLT (AsGaAl 830nm, 30mW, 20J/cm²), tres veces a la semana, por 7 y 21 días. Hubo reducción del área de inflamación/regeneración en el grupo C21 comparado al D21 (p<0,05), siendo más evidente con la LLLT (C21L y D21L). El contenido de TNF-α fue reducido después de 21 días de la lesión. El área de densidad de tejido conjuntivo (ADTC) fue más pequeña en los grupos C21 y C21L comparados a los respectivos grupos desnutridos (p<0,05). La LLLT redujo la ADTC en el grupo D21L cuando comparado del D21 (p<0,05), sin embargo, el contenido de TGF-β1 no fue influenciado. El área de sección transversa (AST) de la fibra muscular incrementó en los grupos 21 días. La m-TOR presentó contenido más grande en el grupo C21L cuando comparado al D21L (p<0,05). Se concluyó que la LLLT favoreció la regeneración muscular en la etapa tardía en el modelo experimental de desnutrición posnatal y posterior recuperación proteica.
ABSTRACT Low-Level Laser Therapy - LLLT is used frequently on muscle lesions, but needs to be investigated in a malnutrition model. The aim of this study was to analyze the effects of LLLT on muscle regeneration of rats subjected to malnutrition and protein recovery. Forty recently weaned Wistar rats were used, divided into control group (C), subjected to a normal-protein diet (14% casein), and the malnourished group (D), subjected to a low-protein diet (6% casein) for 45 days and to a normal-protein diet until the end of the experiment. Subsequently, the right tibialis anterior muscle was subjected to cryogenic cooling and treated with LLLT (830 nm AsGaAl, 30 mW, 20 J/cm²), three times a week, for 7 and 21 days. There was a reduction of the inflammation/regeneration area in the C21 group compared to D21 (p<0.05), which became more evident with the LLLT (C21L and D21L). The TNF-α contents were reduced after 21 days of the injury. The connective tissue density area (CTDA) was lower in the C21 and C21L groups compared to the respective malnourished groups (p<0.05). LLLT reduced the CTDA in group D21L in comparison to D21 (p<0.05), but the TGF-β1 contents were not influenced. The cross-sectional area (CSA) of the muscle fiber increased in the 21-day groups. Higher levels of m-TOR were found in the C21L group when compared to D21L (p<0.05). It was concluded that LLLT favored muscle regeneration in the late stage of the experimental model of postnatal malnutrition and subsequent protein recovery.
RESUMO
Duchenne muscular dystrophy (DMD) is a progressive and fatal disease, characterized by the absence of dystrophin, muscle degeneration and cardiorespiratory failure. Creatine kinase is the classic marker to screen for DMD. However, other markers are needed to follow disease progression and to evaluate the response to therapy over longer periods. In the present study, we aim to identify interleukins in the plasma of the mdx mice model of DMD that could serve as biomarkers to monitor dystrophy progression, at distinct stages of the disease (1, 3 and 8â¯months of age). We used deflazacort and omega-3 therapies to validate the biomarkers studied. Plasma levels of TNF-α and TGF-ß were increased in mdx mice in relation to control, at all times studied. Differences in IFN-γ and IL-10 contents, comparing mdx x CTRL, were detected only at the early stage (1 month). IL-6 decreased at 3 and 8â¯months and IL-13 increased at 8â¯months in the mdx compared to control. Deflazacort and omega-3 reduced the plasma levels of the pro-inflammatory (TNF-α, INF-γ, IL-6) and pro-fibrotic (IL-13 and TGF-ß) interleukins and increased the plasma levels of IL-10. It is suggested that TNF-α and TGF-ß in plasma would be the best markers to follow disease progression. IL-6, INF-γ and IL-10 would be suitable markers to the earlier stages of dystrophy and IL-13 a suitable marker to the later stages of dystrophy.
Assuntos
Anti-Inflamatórios/uso terapêutico , Biomarcadores/sangue , Ácidos Graxos Ômega-3/uso terapêutico , Distrofia Muscular de Duchenne/sangue , Distrofia Muscular de Duchenne/tratamento farmacológico , Pregnenodionas/uso terapêutico , Animais , Progressão da Doença , Interferon gama/sangue , Interleucinas/sangue , Camundongos , Fator de Crescimento Transformador beta/sangue , Fator de Necrose Tumoral alfa/sangueRESUMO
Congenital muscular dystrophy with laminin α2 chain deficiency (MDC1A) is one of the most severe forms of muscular disease and is characterized by severe muscle weakness and delayed motor milestones. The genetic basis of MDC1A is well known, yet the secondary mechanisms ultimately leading to muscle degeneration and subsequent connective tissue infiltration are not fully understood. In order to obtain new insights into the molecular mechanisms underlying MDC1A, we performed a comparative proteomic analysis of affected muscles (diaphragm and gastrocnemius) from laminin α2 chain-deficient dy(3K)/dy(3K) mice, using multidimensional protein identification technology combined with tandem mass tags. Out of the approximately 700 identified proteins, 113 and 101 proteins, respectively, were differentially expressed in the diseased gastrocnemius and diaphragm muscles compared with normal muscles. A large portion of these proteins are involved in different metabolic processes, bind calcium, or are expressed in the extracellular matrix. Our findings suggest that metabolic alterations and calcium dysregulation could be novel mechanisms that underlie MDC1A and might be targets that should be explored for therapy. Also, detailed knowledge of the composition of fibrotic tissue, rich in extracellular matrix proteins, in laminin α2 chain-deficient muscle might help in the design of future anti-fibrotic treatments. All MS data have been deposited in the ProteomeXchange with identifier PXD000978 (http://proteomecentral.proteomexchange.org/dataset/PXD000978).
Assuntos
Cálcio/metabolismo , Diafragma/metabolismo , Proteínas da Matriz Extracelular/biossíntese , Laminina/deficiência , Músculo Esquelético/metabolismo , Animais , Diafragma/patologia , Modelos Animais de Doenças , Fibrose/genética , Fibrose/patologia , Expressão Gênica/genética , Perfilação da Expressão Gênica , Laminina/genética , Camundongos , Camundongos Knockout , Músculo Esquelético/patologia , Distrofia Muscular Animal , Espectrometria de Massas em TandemRESUMO
INTRODUCTION: The purpose of this study was to better understand the beneficial effects of doxycycline on the dystrophic muscles of the mdx mouse. METHODS: Doxycycline (DOX) was administered for 36 days, starting on postnatal day 0, via drinking water. Untreated mdx mice received plain water for the same period and served as a control group. RESULTS: DOX decreased the levels of metalloproteinase-9 and tumor necrosis factor-alpha in the biceps brachii and diaphragm of the mdx mice. It also reduced the total amount of calcium in the muscles studied, concomitant with an increase in the levels of calsequestrin 1. CONCLUSIONS: The results show that DOX can affect factors that are important in dystrophic pathogenesis and highlight its potential as a readily accessible therapy in clinical trials for treatment of Duchenne muscular dystrophy.
Assuntos
Antibacterianos/uso terapêutico , Doxiciclina/uso terapêutico , Distrofia Muscular Animal/tratamento farmacológico , Aldeídos/metabolismo , Animais , Antibacterianos/farmacologia , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Calsequestrina , Doxiciclina/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Fatores de Tempo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Duchenne muscular dystrophy (DMD) is the most common childhood myopathy, characterized by muscle loss and cardiorespiratory failure. While the genetic basis of DMD is well established, secondary mechanisms associated with dystrophic pathophysiology are not fully clarified yet. In order to obtain new insights into the molecular mechanisms of muscle dystrophy during earlier stages of the disease, we performed a comparative proteomic profile of the spared extraocular muscles (EOM) vs. affected diaphragm from the mdx mice, using a label based shotgun proteomic approach. Out of the 857 identified proteins, 42 to 62 proteins had differential abundance of peptide ions. The calcium-handling proteins sarcalumenin and calsequestrin-1 were increased in control EOM compared with control DIA, reinforcing the view that constitutional properties of EOM are important for their protection against myonecrosis. The finding that galectin-1 (muscle regeneration), annexin A1 (anti-inflammatory) and HSP 47 (fibrosis) were increased in dystrophic diaphragm provides novel insights into the mechanisms through which mdx affected muscles are able to counteract dystrophy, during the early stage of the disease. Overall, the shotgun technique proved to be suitable to perform quantitative comparisons between distinct dystrophic muscles and allowed the suggestion of new potential biomarkers and drug targets for dystrophinopaties.
Assuntos
Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/patologia , Proteômica , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Músculo Esquelético/metabolismo , Distrofia Muscular de Duchenne/metabolismoRESUMO
INTRODUCTION: The purpose of this study was to determine the effects of suramin, an antifibrotic agent, on cardiac function and remodeling in mdx mice. METHODS: mdx mice (8 months old) received intraperitoneal injections of suramin twice a week for 3 months. Control mdx mice (8 months old) were injected with saline. RESULTS: Suramin improved the electrocardiography profile with the main corrections seen in S- to R-wave ratio, PR interval, and Q amplitude, and a significant decrease in the cardiomyopathy index. Suramin decreased myocardial fibrosis, inflammation, and myonecrosis. CONCLUSIONS: These findings suggest that suramin may be a new adjunctive therapy to help improve cardiomyopathy in DMD.
Assuntos
Antineoplásicos/uso terapêutico , Cardiomiopatias/tratamento farmacológico , Cardiomiopatias/etiologia , Distrofina/deficiência , Distrofia Muscular de Duchenne/complicações , Suramina/uso terapêutico , Fatores Etários , Análise de Variância , Animais , Cardiomiopatias/sangue , Creatina Quinase/sangue , Modelos Animais de Doenças , Eletrocardiografia , Eletroencefalografia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Fibras Musculares Esqueléticas/patologia , Distrofia Muscular de Duchenne/sangue , Distrofia Muscular de Duchenne/genética , Fator de Crescimento Transformador beta/metabolismoRESUMO
INTRODUCTION: In Duchenne muscular dystrophy and in the mdx mouse, muscle fiber degeneration and subsequent fibrosis lead to cardiorespiratory failure. Previously, we demonstrated that the anti-fibrotic agent suramin was effective in decreasing fibrosis in mdx muscles. In this study, we were interested to see whether suramin could affect metalloproteinases (MMP) and improve the functional activity of the mdx diaphragm muscle. METHODS: Zymography was performed to evaluate MMP-2 and MMP-9 activity. Western blotting was used to analyze the levels of beta-dystroglycan. Muscle function was assessed in hemidiaphragm in vitro preparations. RESULTS: We found that suramin affects metalloproteinase-9 activity and increases beta-dystroglycan. Furthermore, suramin also protects against diaphragm muscle fatigue over time. CONCLUSIONS: These results show the potential benefits of suramin in maintaining the structure of the dystrophin-glycoprotein complex.
Assuntos
Diafragma/metabolismo , Distroglicanas/metabolismo , Distrofina/deficiência , Metaloproteinase 9 da Matriz/metabolismo , Suramina/farmacologia , Animais , Diafragma/efeitos dos fármacos , Distroglicanas/biossíntese , Distrofina/genética , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Feminino , Fibrose , Masculino , Inibidores de Metaloproteinases de Matriz/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Fadiga Muscular/efeitos dos fármacos , Fadiga Muscular/fisiologia , Distrofia Muscular de Duchenne/enzimologia , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patologia , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologiaRESUMO
INTRODUCTION: We examined whether doxycycline, an antibiotic member of the tetracycline family, improves the histopathology and muscle function in mdx mice, the experimental model of DMD. METHODS: Doxycycline was administered for 36 days (starting on postnatal day 0) and for 9 months (starting at 8 months of age) in drinking water. Histopathological, biochemical (creatine kinase), and functional (forelimb muscle grip strength) parameters were evaluated in limb, diaphragm, and cardiac muscle. RESULTS: Doxycycline significantly minimized the dystrophic phenotype of skeletal and cardiac muscles and improved forelimb muscle strength. The drug protected muscle fibers against myonecrosis and reduced inflammation. Furthermore, it slowed the progression of myocardial fibrosis. CONCLUSIONS: This study provides evidence that doxycycline may be a potential therapeutic agent for DMD.
Assuntos
Antibacterianos/uso terapêutico , Doxiciclina/uso terapêutico , Coração/efeitos dos fármacos , Força Muscular/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Distrofia Muscular Animal/tratamento farmacológico , Miocárdio/patologia , Animais , Antibacterianos/farmacologia , Doxiciclina/farmacologia , Fibrose , Coração/fisiopatologia , Camundongos , Camundongos Endogâmicos mdx , Força Muscular/fisiologia , Músculo Esquelético/patologia , Músculo Esquelético/fisiopatologia , Distrofia Muscular Animal/patologia , Distrofia Muscular Animal/fisiopatologia , FenótipoRESUMO
In Duchenne muscular dystrophy (DMD) and in the mdx mouse model of DMD, the lack of dystrophin is related to enhanced calcium influx and muscle degeneration. Stretch-activated channels (SACs) might be directly involved in the pathology of DMD, and transient receptor potential cation channels have been proposed as likely candidates of SACs. We investigated the levels of transient receptor potential canonical channel 1 (TRPC1) and the effects of streptomycin, a SAC blocker, in muscles showing different degrees of the dystrophic phenotype. Mdx mice (18 days old, n = 16) received daily intraperitoneal injections of streptomycin (182 mg/kg body wt) for 18 days, followed by removal of the diaphragm, sternomastoid (STN), biceps brachii, and tibialis anterior muscles. Control mdx mice (n = 37) were injected with saline. Western blot analysis showed higher levels of TRPC1 in diaphragm muscle compared with STN and limb muscles. Streptomycin reduced creatine kinase and prevented exercise-induced increases of total calcium and Evans blue dye uptake in diaphragm and in STN muscles. It is suggested that different levels of the stretch-activated calcium channel protein TRPC1 may contribute to the different degrees of the dystrophic phenotype seen in mdx mice. Early treatment designed to regulate the activity of these channels may ameliorate the progression of dystrophy in the most affected muscle, the diaphragm.
Assuntos
Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patologia , Canais de Cátion TRPC/metabolismo , Animais , Western Blotting , Imunofluorescência , Canais Iônicos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos mdx , Fenótipo , Inibidores da Síntese de Proteínas/farmacologia , Estreptomicina/farmacologiaRESUMO
Congenital muscular dystrophy caused by laminin α2 chain deficiency (also known as MDC1A) is a severe and incapacitating disease, characterized by massive muscle wasting. The ubiquitin-proteasome system plays a major role in muscle wasting and we recently demonstrated that increased proteasomal activity is a feature of MDC1A. The autophagy-lysosome pathway is the other major system involved in degradation of proteins and organelles within the muscle cell. However, it remains to be determined if the autophagy-lysosome pathway is dysregulated in muscular dystrophies, including MDC1A. Using the dy(3K)/dy(3K) mouse model of laminin α2 chain deficiency and MDC1A patient muscle, we show here that expression of autophagy-related genes is upregulated in laminin α2 chain-deficient muscle. Moreover, we found that autophagy inhibition significantly improves the dystrophic dy(3K)/dy(3K) phenotype. In particular, we show that systemic injection of 3-methyladenine (3-MA) reduces muscle fibrosis, atrophy, apoptosis and increases muscle regeneration and muscle mass. Importantly, lifespan and locomotive behavior were also greatly improved. These findings indicate that enhanced autophagic activity is pathogenic and that autophagy inhibition holds a promising therapeutic potential in the treatment of MDC1A.
Assuntos
Autofagia , Laminina/antagonistas & inibidores , Laminina/deficiência , Músculos/patologia , Distrofias Musculares/patologia , Adenina/administração & dosagem , Adenina/análogos & derivados , Adenina/farmacologia , Adenina/uso terapêutico , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Autofagia/genética , Comportamento Animal/efeitos dos fármacos , Modelos Animais de Doenças , Quimioterapia Combinada , Fibrose , Regulação da Expressão Gênica , Injeções , Laminina/metabolismo , Leupeptinas/farmacologia , Leupeptinas/uso terapêutico , Camundongos , Atividade Motora/efeitos dos fármacos , Músculos/metabolismo , Músculos/fisiopatologia , Atrofia Muscular/complicações , Atrofia Muscular/tratamento farmacológico , Atrofia Muscular/patologia , Atrofia Muscular/fisiopatologia , Distrofias Musculares/complicações , Distrofias Musculares/tratamento farmacológico , Distrofias Musculares/fisiopatologia , Doenças do Sistema Nervoso Periférico/complicações , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Doenças do Sistema Nervoso Periférico/patologia , Doenças do Sistema Nervoso Periférico/fisiopatologia , Fenótipo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Regeneração , Análise de SobrevidaRESUMO
Fibrosis is a pathological feature observed in patients with Duchenne muscular dystrophy (DMD) and in mdx mice, the experimental model of DMD. We evaluated the effect of suramin, a transforming growth factor-beta 1 (TGF-ß1) blocker, on fibrosis in mdx mice. mdx mice (6 months old) received suramin for 7 weeks. Suramin- and saline-treated (control) mdx mice performed exercise on a treadmill to worsen disease progression. Immunoblotting showed an increase of TGF-ß1 in mdx diaphragm, limb, and cardiac muscles. Suramin decreased creatine kinase in mdx mice and attenuated fibrosis in all muscles studied, except for cardiac muscle. Suramin protected limb muscles against damage and reduced the exercise-induced loss of strength over time. These findings support a role for TGF-ß1 in fibrinogenesis and myonecrosis during the later stages of disease in mdx mice. Suramin might be a useful therapeutic alternative for the treatment of dystrophinopathies.
Assuntos
Fibras Musculares Esqueléticas/patologia , Músculo Esquelético/patologia , Condicionamento Físico Animal , Suramina/farmacologia , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Animais , Creatina Quinase/metabolismo , Modelos Animais de Doenças , Feminino , Fibrose , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Necrose , Condicionamento Físico Animal/efeitos adversos , Suramina/uso terapêutico , Fator de Crescimento Transformador beta1/fisiologiaRESUMO
The bone tissue abnormalities observed in patients with Duchenne muscular dystrophy are frequently attributed to muscle weakness. In this condition, bones receive fewer mechanical stimuli, compromising the process of bone modeling. In the present study we hypothesize that other factors inherent to the disease might be associated with bone tissue impairment, irrespective of the presence of muscle impairment. Mdx mice lack dystrophin and present cycles of muscle degeneration/regeneration that become more intense in the third week of life. As observed in humans with muscular dystrophy, bone tissue abnormalities were found in mdx mice during more intense muscle degeneration due to age. Under these circumstances, muscle deficit is probably one of the factors promoting these changes. To test our hypothesis, we investigated the changes that occur in the femur of mdx mice at 21 days of age when muscle damage is still not significant. The mechanical (structural and material) and biochemical properties and morphometric characteristics of the femur of mdx and control animals were evaluated. The results demonstrated a lower strength, stiffness and energy absorption capacity in mdx femurs. Higher values for structural (load and stiffness) and material (stress, elastic modulus and toughness) properties were observed in the control group. Mdx femurs were shorter and were characterized by a smaller cortical area and thickness and a smaller area of epiphyseal trabecular bone. The hydroxyproline content was similar in the two groups, but there was a significant difference in the Ca/P ratios. Thermogravimetry showed a higher mineral matrix content in cortical bone of control animals. In conclusion, femurs of mdx mice presented impaired mechanical and biochemical properties as well as changes in collagen organization in the extracellular matrix. Thus, mdx mice developed femoral osteopenia even in the absence of significant muscle fiber degeneration. This weakness of the mdx femur is probably due to genetic factors that are directly or indirectly related to dystrophin deficiency.
Assuntos
Fêmur/anatomia & histologia , Fêmur/fisiopatologia , Animais , Fenômenos Biomecânicos/fisiologia , Fêmur/metabolismo , Hidroxiprolina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos mdx , Microscopia , Temperatura , TermogravimetriaRESUMO
Duchenne muscular dystrophy is one of the most common hereditary diseases. Abnormal ion handling renders dystrophic muscle fibers more susceptible to necrosis and a rise in intracellular calcium is an important initiating event in dystrophic muscle pathogenesis. In the mdx mice, muscles are affected with different intensities and some muscles are spared. We investigated the levels of the calcium-binding proteins calsequestrin and calmodulin in the non-spared axial (sternomastoid and diaphragm), limb (tibialis anterior and soleus), cardiac and in the spared extraocular muscles (EOM) of control and mdx mice. Immunoblotting analysis showed a significant increase of the proteins in the spared mdx EOM and a significant decrease in the most affected diaphragm. Both proteins were comparable to the cardiac muscle controls. In limb and sternomastoid muscles, calmodulin and calsequestrin were affected differently. These results suggest that differential levels of the calcium-handling proteins may be involved in the pathogenesis of myonecrosis in mdx muscles. Understanding the signaling mechanisms involving Ca(2+)-calmodulin activation and calsequestrin expression may be a valuable way to develop new therapeutic approaches to the dystrophinopaties.
Assuntos
Calmodulina/metabolismo , Calsequestrina/metabolismo , Músculo Esquelético/metabolismo , Distrofia Muscular de Duchenne/metabolismo , Animais , Western Blotting , Diafragma/metabolismo , Modelos Animais de Doenças , Imunofluorescência , Masculino , Camundongos , Camundongos Endogâmicos mdx , Músculo Esquelético/patologia , Distrofia Muscular de Duchenne/patologia , Miocárdio/metabolismo , Necrose , FenótipoRESUMO
The lack of dystrophin in mdx mice and in Duchenne muscular dystrophy causes sarcolemmal breakdown and increased calcium influx followed by myonecrosis. We examined whether the calcium channel blockers diltiazem and verapamil protect dystrophic muscles from degeneration. Mdx mice received daily intraperitoneal injections of diltiazem or verapamil for 18 days, followed by removal of the sternomastoid, diaphragm, tibialis anterior, and cardiac muscles. Control mdx mice were injected with saline. Both drugs significantly decreased blood creatine kinase levels. Total calcium content was significantly higher in mdx muscles than in control C57Bl/10. Verapamil and diltiazem reduced total calcium content only in diaphragm and cardiac muscle. Histological analysis showed that diltiazem significantly attenuated myonecrosis in diaphragm. Immunoblots showed a significant increase of calsequestrin and beta-dystroglycan levels in some diltiazem- and verapamil-treated muscles. Possible interactions of these drugs with the sarcoplasmic reticulum and sarcolemma may also contribute to the improvement of the dystrophic phenotype.