RESUMEN
The extent of muscle pathology in sedentary adult mdx mice is very low and treadmill exercise is often used to increase myofibre necrosis; however, the early events in dystrophic muscle and blood in response to treadmill exercise (leading to myofibre necrosis) are unknown. This study describes in detail two standardised protocols for the treadmill exercise of mdx mice and profiles changes in molecular and cellular events after a single 30 min treadmill session (Protocol A) or after 4 weeks of (twice weekly) treadmill exercise (Protocol B). Both treadmill protocols increased multiple markers of muscle damage. We conclude that a single 30 min treadmill exercise session is a sufficient and conveniently fast screening test and could be used in 'proof-of-concept' studies to evaluate the benefits of pre-clinical drugs in vivo. Myofibre necrosis, blood serum CK and oxidative stress (specifically the ratio of oxidised to reduced protein thiols) are reliable markers of muscle damage after exercise; many parameters demonstrated high biological variation including changes in mRNA levels for key inflammatory cytokines in muscle. The sampling (sacrifice and tissue collection) time after exercise for these parameters is critical. A more precise understanding of the changes in dystrophic muscle after exercise aims to identify biomarkers and new potential therapeutic drug targets for Duchenne Muscular Dystrophy.
Asunto(s)
Modelos Animales de Enfermedad , Prueba de Esfuerzo/veterinaria , Ratones Endogámicos mdx , Músculo Esquelético/patología , Distrofia Muscular Animal/patología , Condicionamiento Físico Animal/fisiología , Animales , Prueba de Esfuerzo/métodos , Inflamación/genética , Inflamación/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Contracción Muscular/fisiología , Músculo Esquelético/metabolismo , Distrofia Muscular Animal/metabolismo , Necrosis , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Reasons for the progressive age-related loss of skeletal muscle mass and function, namely sarcopenia, are complex. Few studies describe sarcopenia in mice, although this species is the mammalian model of choice for genetic intervention and development of pharmaceutical interventions for muscle degeneration. One factor, important to sarcopenia-associated neuromuscular change, is myofibre denervation. Here we describe the morphology of the neuromuscular compartment in young (3 month) compared to geriatric (29 month) old female C57Bl/6J mice. There was no significant difference in the size or number of motoneuron cell bodies at the lumbar level (L1-L5) of the spinal cord at 3 and 29 months. However, in geriatric mice, there was a striking increase (by â¼2.5 fold) in the percentage of fully denervated neuromuscular junctions (NMJs) and associated deterioration of Schwann cells in fast extensor digitorum longus (EDL), but not in slow soleus muscles. There were also distinct changes in myofibre composition of lower limb muscles (tibialis anterior (TA) and soleus) with a shift at 29 months to a faster phenotype in fast TA muscle and to a slower phenotype in slow soleus muscle. Overall, we demonstrate complex changes at the NMJ and muscle levels in geriatric mice that occur despite the maintenance of motoneuron cell bodies in the spinal cord. The challenge is to identify which components of the neuromuscular system are primarily responsible for the marked changes within the NMJ and muscle, in order to selectively target future interventions to reduce sarcopenia.
Asunto(s)
Neuronas Motoras/patología , Unión Neuromuscular/patología , Médula Espinal/patología , Envejecimiento , Animales , Femenino , Vértebras Lumbares/patología , Ratones , Ratones Endogámicos C57BL , Neuronas Motoras/metabolismo , Desnervación Muscular , Fibras Musculares de Contracción Rápida/patología , Fibras Musculares de Contracción Lenta/patología , Músculo Esquelético/patología , Fenotipo , Sarcopenia/patología , Células de Schwann/patología , Factores de TiempoRESUMEN
Three-dimensional optical coherence tomography (3D-OCT) was used to image the structure and pathology of skeletal muscle tissue from the treadmill-exercised mdx mouse model of human Duchenne muscular dystrophy. Optical coherence tomography (OCT) images of excised muscle samples were compared with co-registered hematoxylin and eosin-stained and Evans blue dye fluorescence histology. We show, for the first time, structural 3D-OCT images of skeletal muscle dystropathology well correlated with co-located histology. OCT could identify morphological features of interest and necrotic lesions within the muscle tissue samples based on intrinsic optical contrast. These findings demonstrate the utility of 3D-OCT for the evaluation of small-animal skeletal muscle morphology and pathology, particularly for studies of mouse models of muscular dystrophy.
Asunto(s)
Distrofia Muscular de Duchenne/patología , Tomografía de Coherencia Óptica/métodos , Animales , Colorantes , Modelos Animales de Enfermedad , Azul de Evans , Humanos , Imagenología Tridimensional , Masculino , Ratones , Ratones Endogámicos mdx , Músculo Esquelético/patología , Necrosis , Tomografía de Coherencia Óptica/instrumentaciónRESUMEN
Ceroid and lipofuscin are autofluorescent granules thought to be generated as a consequence of chronic oxidative stress. Because ceroid and lipofuscin are persistent in tissue, their measurement can provide a lifetime history of exposure to chronic oxidative stress. Although ceroid and lipofuscin can be measured by quantification of autofluorescent granules, current methods rely on subjective assessment. Furthermore, there has not been any evaluation of variables affecting quantitative measurements. The article describes a simple statistical approach that can be readily applied to quantitate ceroid and lipofuscin. Furthermore, it is shown that several factors, including magnification tissue thickness and tissue level, can affect precision and sensitivity. After optimizing for these factors, the authors show that ceroid and lipofuscin can be measured reproducibly in the skeletal muscle of dystrophic mice (ceroid) and aged mice (lipofuscin).
Asunto(s)
Ceroide/análisis , Lipofuscina/análisis , Músculo Cuádriceps/química , Envejecimiento/metabolismo , Animales , Interpretación Estadística de Datos , Femenino , Fluorescencia , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos mdx , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patología , Músculo Cuádriceps/patología , Sensibilidad y EspecificidadRESUMEN
We hypothesise that the sarcolemma of an actively growing myofibre has different properties to the sarcolemma of a mature adult myofibre. Such fundamentally different properties have clinical consequences for the onset, and potential therapeutic targets, of various skeletal muscle diseases that first manifest either during childhood (e.g. Duchenne muscular dystrophy, DMD) or after cessation of the main growth phase (e.g. dysferlinopathies). These characteristics are also relevant to the selection of both tissue culture and in vivo models employed to study such myopathies and the molecular regulation of adult myofibres. During growth, multinucleated myofibres increase enormously in size and volume with dramatic increases in length (up to ~600 mm). This is in striking contrast with most mononucleated cells such as fibroblasts, that remain at a relatively small size (~10-20 µm diameter). The consequences of a dynamic, expanding sarcolemma during growth, compared with that of an adult myofibre of a fixed length, are discussed with respect to various aspects of muscle biology.
Asunto(s)
Desarrollo de Músculos , Músculo Esquelético/crecimiento & desarrollo , Enfermedades Musculares/patología , Animales , Desarrollo Óseo , Modelos Animales de Enfermedad , Matriz Extracelular/metabolismo , Humanos , Músculo Esquelético/citología , Músculo Esquelético/metabolismo , Distrofia Muscular de Cinturas/genética , Distrofia Muscular de Cinturas/patología , Distrofia Muscular de Cinturas/fisiopatología , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/patología , Distrofia Muscular de Duchenne/fisiopatología , Miofibrillas/fisiología , Necrosis , Sarcolema/metabolismoAsunto(s)
Factor I del Crecimiento Similar a la Insulina/metabolismo , Mecanotransducción Celular , Contracción Muscular , Músculo Esquelético/patología , Músculo Esquelético/fisiopatología , Esfuerzo Físico , Adulto , Humanos , Hipertrofia/patología , Hipertrofia/fisiopatología , Tamaño de los ÓrganosRESUMEN
Here, we characterise new strains of normal and dystrophic (mdx) mice that overexpress Class 2 IGF-1 Ea in skeletal myofibres. We show that transgenic mice have increased muscle levels of IGF-1 (approximately 13-26 fold) and show striking muscle hypertrophy (approximately 24-56% increase in mass). Adult normal muscles were resistant to elevated IGF-1; they reached adult steady state and maintained the same mass from 3 to 12 months. By contrast, dystrophic muscles from mdx/IGF-1(C2:Ea) mice continued to increase in mass during adulthood. IGF-1 signalling was evident only in muscles that were growing as a result of normal postnatal development (23-day-old mice) or regenerating in response to endogenous necrosis (adult mdx mice). Increased phosphorylation of Akt at Ser473 was not evident in fasted normal adult transgenic muscles, but was 1.9-fold higher in fasted normal young transgenic muscles compared with age-matched wild-type controls and fourfold higher in fasted adult mdx/IGF-1(C2:Ea) compared with mdx muscles. Muscles of adult mdx/IGF-1(C2:Ea) mice showed higher p70(S6K)(Thr421/Ser424) phosphorylation and both young transgenic and adult mdx/IGF-1(C2:Ea) mice had higher phosphorylation of rpS6(Ser235/236). The level of mRNA encoding myogenin was increased in normal young (but not adult) transgenic muscles, indicating enhanced myogenic differentiation. These data demonstrate that elevated IGF-1 has a hypertrophic effect on skeletal muscle only in growth situations.
Asunto(s)
Factor I del Crecimiento Similar a la Insulina/metabolismo , Músculo Esquelético/crecimiento & desarrollo , Músculo Esquelético/patología , Envejecimiento/patología , Animales , Peso Corporal , Regulación de la Expresión Génica , Hipertrofia , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Ratones Endogámicos mdx , Fibras Musculares Esqueléticas/enzimología , Fibras Musculares Esqueléticas/patología , Músculo Esquelético/enzimología , Distrofia Muscular Animal/sangre , Distrofia Muscular Animal/enzimología , Distrofia Muscular Animal/patología , Miogenina/genética , Miogenina/metabolismo , Tamaño de los Órganos , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TORRESUMEN
Tumour Necrosis Factor (TNF) plays a major role in exacerbating necrosis of dystrophic muscle; however, the precise molecular mechanism underlying this effect of TNF is unknown. This study investigates the role that p53 plays in TNF-mediated necrosis of dystrophic myofibres by inhibiting p53 using pifithrin-alpha and three pifithrin-beta analogues. Tissue culture studies using C2C12 myoblasts established that pifithrin-alpha was toxic to differentiating myoblasts at concentrations greater than 10 muM. While non-toxic concentrations of pifithrin-alpha did not prevent the TNF-mediated inhibition of myoblast differentiation, Western blots indicated that nuclear levels of p53 were higher in TNF-treated myoblasts indicating that TNF does elevate p53. In contrast, in vivo studies in adult mdx mice showed that pifithrin-alpha significantly reduced myofibre necrosis that resulted from voluntary wheel running over 48 h. These results support the hypothesis that p53 plays some role in TNF-mediated necrosis of dystrophic muscle and present a potential new target for therapeutic interventions.
Asunto(s)
Benzotiazoles/farmacología , Distrofia Muscular Animal/patología , Tolueno/análogos & derivados , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/fisiología , Animales , Muerte Celular/efectos de los fármacos , Muerte Celular/efectos de la radiación , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/efectos de la radiación , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Células Cultivadas , Femenino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos mdx , Modelos Biológicos , Distrofia Muscular Animal/metabolismo , Mioblastos Esqueléticos/efectos de los fármacos , Mioblastos Esqueléticos/patología , Mioblastos Esqueléticos/fisiología , Mioblastos Esqueléticos/efectos de la radiación , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Tolueno/farmacología , Factor de Necrosis Tumoral alfa/farmacología , Proteína p53 Supresora de Tumor/metabolismo , Rayos UltravioletaRESUMEN
The ability of very old animals to make new muscle after injury remains controversial. This issue has major implications for the regenerative potential of damaged geriatric human muscle, to age-related loss of muscle mass (sarcopenia) and to the proposed need for muscle stem cell therapy for the aged. To further address issues of inherent myogenic capacity and the role of host systemic factors in new muscle formation, whole muscle grafts were transplanted between geriatric (aged 27-29 months) and young (3 months) C57Bl/6J mice and compared with autografts in geriatric and young mice. Grafts were sampled at 5 and 10 days for histological analysis. Inflammation and formation of new myotubes was strikingly impaired at 5 days in the geriatric muscle autografts. However, there was a strong inflammatory response by the geriatric hosts to young muscle grafts and geriatric muscles provoked an inflammatory response by young hosts at 5 days. At 10 days, extensive myotube formation in geriatric muscle autografts (equivalent to that seen in young autografts and both other groups) confirmed excellent intrinsic capacity of myogenic (stem) cells to proliferate and fuse. The key conclusion is that a weaker chemotactic stimulus by damaged geriatric muscle, combined with a reduced inflammatory response of old hosts, results in delayed inflammation in geriatric muscle autografts. This delay is transient. Once inflammation occurs, myogenesis can proceed. The presence of well developed myotubes in old muscle autografts at 10 days confirms a very good inherent myogenic response of geriatric skeletal muscle.
Asunto(s)
Músculo Esquelético/fisiología , Regeneración , Células Madre/citología , Animales , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BLRESUMEN
PURPOSE OF REVIEW: The aim of this overview is to highlight the multiple ways in which oxidative stress could be exacerbating muscle wasting. Understanding these interactions in vivo will assist in identifying opportunities for more targeted therapies to reduce skeletal muscle wasting. RECENT FINDINGS: There are many excellent reviews describing how oxidative stress can damage cellular macromolecules, as well as cause deleterious effects through the modulation of signalling pathways. In this overview, we highlight the potential for complex and possibly paradoxical interactions in vivo. Signalling pathways are discussed, using examples involving nuclear factor-kappa B, apoptosis signal-regulating kinase 1 and Akt. Oxidative stress may also be involved in complex interactions with other factors capable of stimulating the loss of muscle mass, possibly through amplifying feedback cycles. This is discussed using examples related to calcium and tumour necrosis factor. SUMMARY: There is convincing evidence that oxidative stress can increase protein catabolism. The challenge is to demonstrate that oxidative stress is a significant player in the complex interplay that leads to the in-vivo muscle wasting that is caused by a range of conditions and diseases.
Asunto(s)
Caquexia/etiología , Atrofia Muscular/etiología , Estrés Oxidativo , Transducción de Señal/fisiología , Caquexia/prevención & control , Calcio/metabolismo , Calcio/fisiología , Humanos , MAP Quinasa Quinasa Quinasa 5/metabolismo , Atrofia Muscular/prevención & control , FN-kappa B/metabolismo , FN-kappa B/fisiología , Proteínas Proto-Oncogénicas c-akt/fisiología , Especies Reactivas de Oxígeno , Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/fisiologíaRESUMEN
1. Inflammation, particularly the pro-inflammatory cytokine tumour necrosis factor (TNF), increases necrosis of skeletal muscle. Depletion of inflammatory cells, such as neutrophils, cromolyn blockade of mast cell degranulation or pharmacological blockade of TNF reduces necrosis of dystrophic myofibres in the mdx mouse model of the lethal childhood disease Duchenne muscular dystrophy (DMD). 2. Insulin-like growth factor-1 (IGF-1) is a very important cytokine for maintenance of skeletal muscle mass and the transgenic overexpression of IGF-1 within muscle cells reduces necrosis of dystrophic myofibres in mdx mice. Thus, IGF-1 usually has the opposite effect to TNF. 3. Activation of TNF signalling via the c-Jun N-terminal kinase (JNK) can inhibit IGF-1 signalling by phosphorylation and conformational changes in insulin receptor substrate (IRS)-1 downstream of the IGF-1 receptor. Such silencing of IGF-1 signalling in situations where inflammatory cytokines are elevated has many implications for skeletal muscle in vivo. 4. The basis for these interactions between TNF and IGF-1 is discussed with specific reference to clinical consequences for myofibre necrosis in DMD and also for the wasting (atrophy) of skeletal muscles that occurs in very old people and in cachexia associated with inflammatory disorders.
Asunto(s)
Factor I del Crecimiento Similar a la Insulina/fisiología , Músculo Esquelético/fisiología , Receptor Cross-Talk/fisiología , Transducción de Señal/fisiología , Factor de Necrosis Tumoral alfa/fisiología , Animales , Humanos , Factor I del Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/metabolismo , Músculo Esquelético/patología , Necrosis , Transducción de Señal/genética , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Duchenne muscular dystrophy (DMD) is a lethal X-linked disease caused by the absence of functional dystrophin. Abnormal excitation-contraction (E-C) coupling has been reported in dystrophic muscle fibers from mdx mice, and alterations in E-C coupling components may occur as a direct result of dystrophin deficiency. We hypothesized that muscle-specific overexpression of insulin-growth factor-1 (IGF-I) would reduce E-C coupling failure in mdx muscle. Mechanically skinned extensor digitorum longus muscle fibers from mdx mice displayed a faster decline in depolarization-induced force responses (DIFR); however, there were no differences in sarcoplasmic reticulum (SR)-mediated Ca(2+) resequestration or in the properties of the contractile apparatus when compared with nondystrophic controls. The rate of DIFR decline was restored to control levels in fibers from transgenic mdx mice that overexpressed IGF-I in skeletal muscle (mdx/IGF-I mice). Dystrophic muscles have a lower transcript level of a specific dihydropyridine receptor (DHPR) isoform, and IGF-I-mediated changes in E-C coupling were associated with increased transcript levels of specific DHPR isoforms involved in Ca(2+) regulation. Importantly, IGF-I overexpression also increased the sensitivity of the contractile apparatus to Ca(2+). The results demonstrate that IGF-I can ameliorate fundamental aspects of E-C coupling failure in dystrophic muscle fibers and that these effects are important for the improvements in cellular function induced by this growth factor.
Asunto(s)
Factor I del Crecimiento Similar a la Insulina/metabolismo , Contracción Muscular , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/metabolismo , Distrofia Muscular de Duchenne/metabolismo , Animales , Calcio/metabolismo , Canales de Calcio Tipo L/genética , Canales de Calcio Tipo L/metabolismo , Modelos Animales de Enfermedad , Factor I del Crecimiento Similar a la Insulina/genética , Masculino , Ratones , Ratones Endogámicos mdx , Ratones Transgénicos , Fuerza Muscular , Músculo Esquelético/fisiopatología , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/fisiopatología , Isoformas de Proteínas/metabolismo , ARN Mensajero/metabolismo , Retículo Sarcoplasmático/metabolismo , Factores de TiempoRESUMEN
Extreme loss of skeletal muscle mass (atrophy) occurs in human muscles that are not used. In striking contrast, skeletal muscles do not rapidly waste away in hibernating mammals such as bears, or aestivating frogs, subjected to many months of inactivity and starvation. What factors regulate skeletal muscle mass and what mechanisms protect against muscle atrophy in some species? Severe atrophy also occurs with ageing and there is much clinical interest in reducing such loss of muscle mass and strength (sarcopenia). In the meat industry, a key aim is optimizing the control of skeletal muscle growth and meat quality. The impaired response of muscle to insulin resulting in diabetes, that is a consequence of the metabolic impact of increasing obesity and fat deposition in humans, is also of increasing clinical concern. Intensive research in these fields, combined with mouse models, is reviewed with respect to the molecular control of muscle growth (myogenesis) and atrophy/hypertrophy and fat deposition (adipogenesis) in skeletal muscle, with a focus on IGF-1/insulin signaling.
Asunto(s)
Tejido Adiposo/anatomía & histología , Músculo Esquelético/anatomía & histología , Adipogénesis , Tejido Adiposo/crecimiento & desarrollo , Tejido Adiposo/fisiología , Envejecimiento/patología , Animales , Proteínas de Drosophila , Hibernación , Humanos , Hipertrofia , Insulina/fisiología , Factor I del Crecimiento Similar a la Insulina/fisiología , Carne , Ratones , Modelos Biológicos , Desarrollo de Músculos , Músculo Esquelético/crecimiento & desarrollo , Músculo Esquelético/fisiología , Atrofia Muscular/patología , Ranidae , Especificidad de la Especie , UrsidaeRESUMEN
Dramatic clinical success in the treatment of chronic inflammatory diseases has resulted from the use of anti-cytokine therapies including specific blocking antibodies, soluble receptors and traps to silence the actions of inflammatory cytokines such as tumour necrosis factor alpha (TNFalpha) and interleukin-1 (IL-1). Two agents used clinically to block the functional activity of TNFalpha protein are Remicade (an antibody) and Enbrel (a soluble TNF receptor). These tools are now being extended to many other clinical disorders. We have a specific interest in the treatment of muscle diseases. In order to study the effects of novel anti-cytokine drugs on mouse models of human disease, such drugs must be investigated to determine whether they are indeed effective in blocking the inflammatory response in mouse. This has been carried out by means of a simple in vivo bioassay. Histological examination of transverse sections from whole muscle autografts in C57BL/10ScSn mice sampled at 5 days after transplantation provides an excellent assay model and clearly shows that Remicade and Enbrel block the acute inflammatory cell response in vivo. This graft model has also been used to show that a single intraperitoneal injection of Remicade (10 microg/g) is long-lived and effective when administered at 1 week and even 4 weeks prior to the assay. Enbrel is highly effective when injected twice at -3 days and -1 day (2 x 100 microg) before muscle grafting but shows no inhibition of the inflammatory response after a single injection (100 microg) 1 week prior to grafting. This striking ablation of inflammation by pharmacological blockage of TNFalpha is in marked contrast to the lack of any effect in TNFalpha null mice. This simple reproducible in vivo assay model in mice can be used to evaluate the efficacy of many novel anti-cytokine interventions designed to block inflammation.
Asunto(s)
Anticuerpos Monoclonales/farmacología , Inmunoglobulina G/farmacología , Músculo Esquelético/efectos de los fármacos , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Animales , Anticuerpos Monoclonales/administración & dosificación , Relación Dosis-Respuesta a Droga , Etanercept , Femenino , Inmunoglobulina G/administración & dosificación , Inflamación/tratamiento farmacológico , Infliximab , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/inmunología , Músculo Esquelético/trasplante , Receptores del Factor de Necrosis Tumoral/administración & dosificación , Trasplante Autólogo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Transgenic mice that overexpress insulin-like growth factor-1 (IGF-I) specifically in skeletal muscle have generated much information about the role of this factor for muscle growth and remodelling and provide insight for therapeutic applications of IGF-I for different pathological states and ageing. However, difficulties arise when attempting to critically compare the significance of data obtained in vivo by using different genetically engineered mouse lines and various experimental models. Complications arise due to complexity of the IGF-I system, since multiple transcripts of the IGF-I gene encode different isoforms generated by alternate promoter usage, differential splicing and post-translational modification, and how IGF-I gene expression relates to its diverse autocrine, paracrine and endocrine modes of action in vivo has still to be elucidated. In addition, there are problems related to specification of the exact IGF-I isoform used, expression patterns of the promoters, and availability of the transgene product under different experimental conditions. This review discusses the factors that must be considered when reconciling data from cumulative studies on IGF-I in striated muscle growth and differentiation using genetically modified mice. Critical evaluation of the literature focuses specifically on: (1) the importance of detailed information about the IGF-I isoforms and their mode of action (local, systemic or both); (2) expression pattern and strength of the promoters used to drive transgenic IGF-I in skeletal muscle cells (mono and multi-nucleated); (3) local compared with systemic action of the transgene product and possible indirect effects of transgenic IGF-I due to upregulation of other genes within skeletal muscle; (4) re-interpretation of these results in light of the most recent approaches to the dissection of IGF-I function. Full understanding of these complex in vivo issues is essential, not only for skeletal muscle but for many other tissues, in order to effectively extend observations derived from transgenic studies into potential clinical situations.
Asunto(s)
Factor I del Crecimiento Similar a la Insulina/biosíntesis , Factor I del Crecimiento Similar a la Insulina/genética , Ratones Transgénicos , Músculo Esquelético/metabolismo , Envejecimiento , Empalme Alternativo , Animales , Secuencia de Bases , Modelos Animales de Enfermedad , Exones , Humanos , Intrones , Ratones , Datos de Secuencia Molecular , Regiones Promotoras Genéticas , Isoformas de Proteínas , Estructura Terciaria de Proteína , ARN Mensajero/metabolismo , Ratas , TransgenesRESUMEN
Loss of the nerve supply to skeletal muscle results in a relentless loss of muscle mass (atrophy) over time. The ability of insulin-like growth factor-1 to reduce atrophy resulting from denervation was examined after transection of the sciatic nerve in transgenic MLC/mIGF-1 mice that over-express mIGF-1 specifically in differentiated myofibres. The cross sectional area (CSA) of all types of myofibres and specifically type IIB myofibres was measured in tibialis anterior muscles from transgenic and wild-type mice at 28 days after denervation. There was a marked myofibre atrophy ( approximately 60%) in the muscles of wild-type mice over this time with increased numbers of myofibres with small CSA. In the muscles of MLC/mIGF-1 mice, over-expression of mIGF-1 reduced the rate of denervation induced myofibre atrophy by approximately 30% and preserved myofibres with larger CSA, compared to wild-type muscles. It is proposed that the protective effect of mIGF-1 on denervated myofibres might be due to reduced protein breakdown.
Asunto(s)
Factor I del Crecimiento Similar a la Insulina/genética , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Regulación hacia Arriba/fisiología , Animales , Peso Corporal/fisiología , Tamaño de la Célula , Desnervación , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Regulación hacia Abajo/fisiología , Ratones , Ratones Transgénicos , Fibras Musculares Esqueléticas/patología , Proteínas Musculares/metabolismo , Músculo Esquelético/patología , Músculo Esquelético/fisiopatología , Atrofia Muscular/genética , Atrofia Muscular/fisiopatología , Neuropatía Ciática/complicaciones , Neuropatía Ciática/patología , Neuropatía Ciática/fisiopatología , Factores de TiempoRESUMEN
Necrosis of dystrophic myofibers in Duchenne muscular dystrophy and mdx mice results from defects in the subsarcolemmal protein dystrophin that cause membrane fragility and tears in the sarcolemma, and these lead to the destruction of the myofibers. The present study specifically tests whether overexpression of mIGF-1 in mdx/mIGF-1 transgenic mice reduces myofiber breakdown during the acute onset phase of dystrophy (at 21 days). The extent of muscle damage and Evans blue dye (EBD) staining of myofibers was quantitated histologically for mdx/mIGF-1 and their mdx littermates from 15 to 30 days of age. Overexpression of mIGF-1 strikingly reduced the extent of myofiber damage (histology and EBD staining) by up to 97% in tibialis anterior and quadriceps muscles at 21-22 days after birth. In the mdx diaphragm, the onset of muscle breakdown was earlier (by 15 days after birth) but no significant protective effect of IGF-1 was apparent within the first month of age in mdx/IGF-1 mice. These novel observations show that increased mIGF-1 within mdx myofibers specifically reduces the breakdown of dystrophic muscle during the acute onset of muscle degeneration. This mechanism of action can account for the long-term reduced severity of the dystropathology in mdx mice that overexpress mIGF-1 and provides promising opportunities for therapeutic strategies.
Asunto(s)
Terapia Genética/métodos , Factor I del Crecimiento Similar a la Insulina/genética , Fibras Musculares Esqueléticas/patología , Distrofia Muscular de Duchenne/terapia , Animales , Factor I del Crecimiento Similar a la Insulina/análisis , Ratones , Ratones Endogámicos mdx , Ratones Transgénicos , Fibras Musculares de Contracción Rápida/metabolismo , Músculo Esquelético/química , Músculo Esquelético/citología , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patología , Necrosis , Sarcolema/química , Sarcolema/metabolismoRESUMEN
Early myogenic events in regenerating whole muscle grafts were compared between transgenic MLC/mIGF-1 mice with skeletal muscle-specific overexpression of the Exon-1 Ea isoform of insulin-like growth factor-1 (mIGF-1) and control FVB mice, from day 3 to day 21 after transplantation. Immunocytochemistry with antibodies against desmin showed that skeletal muscle-specific overexpression of IGF-1 did not affect the pattern of myoblast activation or proliferation or the onset and number of myotubes formed in regenerating whole muscle grafts. Hypertrophied myotubes were observed in MLC/mIGF grafts at day 7 after transplantation, although such hypertrophy was transient, and the transgenic and control grafts had a similar appearance at later time points (days 10, 14, and 21). Immunostaining with antibodies to platelet endothelial cell adhesion molecule-1, which identifies endothelial cells, demonstrated no difference in the formation of new vascular network in grafts of transgenic and control mice. Skeletal muscle-specific overexpression of mIGF-1 does not appear to stimulate the early events associated with myogenesis during regeneration of whole muscle grafts.