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Localization of mRNAs coding for CMD1, myogenin and the alpha-subunit of the acetylcholine receptor during skeletal muscle development in the chicken.

Myogenin and CMD1, the chicken homologue of MyoD, transactivate the promoter of the alpha-subunit of the acetylcholine receptor (AChR) in chicken fibroblasts. The expression of these three genes was followed by in situ hybridization. In two-day-old embryos the CMD1 gene is expressed shortly before the AChR alpha-subunit and the myogenin genes. At day 19 extrajunctional AChR mRNA clusters have disappeared and myogenin mRNAs are no longer detected in PLD muscle. Moreover, both myogenin and CMD1 mRNA levels increase after muscle denervation in chicks. These data are compatible with a role for myogenic factors in the induction and maintenance of extra-junctional expression of the AChR genes during early muscle development. Using digoxygenin labelled RNA probes, we also show that the mRNAs for the AChR alpha-subunit display a punctated, probably perinuclear distribution, whereas mRNAs for myogenic genes accumulate in the sarcoplasm around subsets of nuclei in the muscle fiber.

Animals↗

Embryonic muscle development in rainbow trout (Oncorhynchus mykiss): a scanning electron microscopy and immunohistological study.

Embryonic muscle development was studied in rainbow trout (Oncorhynchus mykiss) at low and high temperature using scanning electron microscopy (SEM) and immunohistology. Somite development was described starting at stage 16 (Vernier JM. 1969. Ann Embryol Morphogen 4:495-520) for both temperatures, with special interest in their shape and size. Muscle differentiation, associated with somite growth, is characterized by a larger increase in height compared to width and by acquisition of a chevron shape. Thin structures such as striation, sarcomeres, and myofibrils within muscle cells and myotubes were observed starting at the eyed stage (stage 24). Immunohistological analyses showed appearance of embryonic fast myosin at stage 20 in the deep part of the somite. The area where myosin was expressed extended in the somite throughout embryonic development and the presence of myosin was observed in the entire somite at hatching (stage 30). Slow myosin was expressed in a monolayer of superficial cells at the eyed stage and during the entire embryonic development. Then it was expressed in a few layers of cells located in the red muscle area. These results suggest that muscle differentiation, characterized by myosin expression, is engaged at stage 20. Myogenesis starts in the deep part of the somite, near the notochord and progresses laterally to cover the complete somite at hatching when the somite is composed of muscle fibres exhibiting a high degree of maturity. No significant difference was observed in terms of muscular development between low- and high-temperature conditions. J. Exp. Zool. 286:379-389, 2000.

Animals↗

Effects of maternal nutrition and porcine growth hormone (pGH) treatment during gestation on endocrine and metabolic factors in sows, fetuses and pigs, skeletal muscle development, and postnatal growth.

Prenatal growth is very complex and a highly integrated process. Both maternal nutrition and the maternal somatotropic axis play a significant role in coordinating nutrient partitioning and utilization between maternal, placental and fetal tissues. Maternal nutrition may alter the nutrient concentrations and in turn the expression of growth regulating factors such as IGFs and IGFBPs in the blood and tissues, while GH acts in parallel via changing IGFs/IGFBPs and nutrient availability. The similarity in the target components implies that maternal nutrition and the somatotropic axis are closely related to each other and may induce similar effects on placental and fetal growth. Severe restriction of nutrients throughout gestation has a permanent negative effect on fetal and postnatal growth, whereas the effects of both temporary restriction and feeding above requirements during gestation seem to be of transitional character. Advantages in fetal growth gained by maternal growth hormone treatment during early to mid-gestation are not maintained to term, whereas treatment during late or greatest part of gestation increases progeny size at birth, which could be of advantage for postnatal growth. This review summarizes the available knowledge on the effects of different maternal feeding strategies and maternal GH administration during pregnancy and their interactions on metabolic and hormonal (especially IGFs/IGFBPs) status in the feto-maternal unit, skeletal muscle development and growth of the offspring in pigs.

Animal Nutritional Physiological Phenomena↗

Myosin light chain 3 synthesis during chick pectoralis muscle development in ovo.

The level of myosin light chain 3 (LC3) in vertebrate skeletal muscle is developmentally regulated in a tissue-specific manner. We have used the RNA-cDNA hybridization assay to quantitate LC3 mRNA levels at various stages of chick pectoralis muscle development in ovo. The LC3 mRNA was found significantly in breast muscle only on Day 16 in ovo and later, the level of mRNA ranging from about 30 to 32% of that present in adult tissue. These values are in good agreement with the corresponding levels of LC3 in embryonic muscle. These results do not support the earlier reports that the protein and mRNA for LC3 accumulate in a noncoordinate manner in embryonic pectoralis muscle and they suggest that LC3 synthesis in ovo is regulated primarily at the transcriptional level.

Animals↗

Muscle development: molecules of myoblast fusion.

The fusion of myoblasts to make multinucleate muscle fibres is central to muscle development. Recent work on Drosophila has identified two members of the immunoglobulin superfamily that have key roles in controlling the specificity of myoblast fusion.

Animals↗

Heparan sulfates in skeletal muscle development and physiology.

Recent years have seen an emerging interest in the composition of the skeletal muscle extracellular matrix (ECM) and in the developmental and physiological roles of its constituents. Many cell surface-associated and ECM-embedded molecules occur in highly organized spatiotemporal patterns, suggesting important roles in the development and functioning of skeletal muscle. Glycans are historically underrepresented in the study of skeletal muscle ECM, even though studies from up to 30 years ago have demonstrated specific carbohydrates and glycoproteins to be concentrated in neuromuscular junctions (NMJs). Changes in glycan profile and distribution during myogenesis and synaptogenesis hint at an active involvement of glycoconjugates in muscle development. A modest amount of literature involves glycoconjugates in muscle ion housekeeping, but a recent surge of evidence indicates that glycosylation defects are causal for many congenital (neuro)muscular disorders, rendering glycosylation essential for skeletal muscle integrity. In this review, we focus on a single class of ECM-resident glycans and their emerging roles in muscle development, physiology, and pathology: heparan sulfate proteoglycans (HSPGs), notably their heparan sulfate (HS) moiety.

Animals↗

The distribution of alpha-bungarotoxin binding sites of mammalian skeletal muscle developing in vivo.

1. The distribution of alpha-bungarotoxin binding sites on embryonic and neonatal rat skeletal muscle fibres was determined by autoradiography. Most of the bungarotoxin binding could be inhibited by curare. This observation, together with the spatial distribution of toxin-binding sites, indicates that the distribution of bound toxin reflects that of acetylcholine (ACh) receptors on these developing muscle cells.2. At 15 days of embryogenesis, muscle fibres showed an essentially uniform distribution of receptors. By 16 days, many fibres showed an accumulation of receptors in their mid-region. This accumulation was at the same location as histochemically demonstrated cholinesterase activity.3. At 16 days ACh receptors were distributed over the entire length of the fibres, with a gradient of increasing density as the accumulation was appoached. The density of toxin binding sites in the accumulation was greater than the general level on 15 day cells, suggesting that the high junctional density does not develop solely by the loss of extrajunctional receptors.4. The accumulations of ACh receptors became more pronounced and circumscribed with embryonic development, and after birth the extent of the localizations appeared to follow the size of the neuromuscular junction. The extrajunctional receptor density decreased with development, and by 1 week after birth was undetectable by the methods used.5. The results suggest that the high junctional receptor density found on adult, innervated skeletal muscle fibres develops after the formation of the neuromuscular junction.

Age Factors↗

Invertebrate myogenesis: looking back to the future of muscle development.

Recent studies in invertebrates have provided important mechanistic insights into several general aspects of muscle development. Two new genes have been identified that are involved in muscle fusion in Drosophila and a novel maternal component was shown to be responsible for myogenic determination in an ascidian. In addition, genetic analyses of nematode and Drosophila homologues of factors known to be myogenic regulators in other species yielded surprising findings about both the evolutionary conservation and divergence of these functions. Drosophila myogenesis has become a highly informative model for understanding the interplay between the signaling and transcriptional networks that underlie cell-fate specification during embryonic development.

Animals↗

Effect of transforming growth factor-beta on decorin and beta1 integrin expression during muscle development in chickens.

Myoblast-extracellular matrix interactions play a pivotal role in skeletal muscle development. Transforming growth factor-beta (TGF-beta) is a key regulator of muscle cell proliferation and differentiation. The level of TGF-beta expressed will affect the concentration of the extracellular matrix proteoglycan decorin and the cell surface beta1 integrin subunit. The decorin proteoglycan is a regulator of cell growth as well as the organization of the extracellular matrix. The beta1 integrin plays a role in muscle cell attachment, migration, and the formation of multinucleated myotubes. In the current study, chicken myogenic satellite cells isolated from the pectoralis major muscle from the chicken genetic muscle weakness, low score normal (LSN), and normal pectoralis major muscle were used to investigate TGF-beta expression as it relates to decorin and beta1 integrin mRNA expression. The LSN muscle defect is characterized by altered myotube formation and sarcomere structure, and the satellite cells have reduced proliferation and differentiation. The mRNA expression was measured by real-time quantitative reverse transcription PCR. The LSN condition has elevated expression of TGF-beta2 and TGF-beta4 with increased expression of decorin and decreased beta1 integrin during myogenic satellite cell proliferation and differentiation. Normal satellite cell cultures were treated with the addition of exogenous TGF-beta during differentiation to determine if the altered expression of LSN decorin and beta1 integrin was associated with TGF-beta expression. The addition of exogenous TGF-beta decreased decorin expression during differentiation and reduced beta1 integrin expression at 24 and 48 h of differentiation. These results suggested that alteration of decorin expression in the LSN myogenic satellite cells may occur by a mechanism involving factors in addition to TGF-beta, but the addition of exogenous TGF-beta did affect both decorin and beta1 integrin expression. These data, therefore, suggested that TGF-beta might play a pivotal role in chicken skeletal muscle formation through modulation of the expression of both extracellular matrix molecules and cellular receptors important in the control of cell migration and growth regulation.

Animals↗

Muscle development in thyroidectomised chickens (Gallus domesticus).

The metabolic and contractile activity of muscle was determined in immature cockerels made hypothyroid by surgical thyroidectomy at 6 weeks of age. Four weeks after thyroidectomy the activity of Mg2+-activated myofibrillar ATPase and total phosphorylase was reduced in the fast-phasic, posterior latissimus dorsi (PLD) and scapulotriceps (ST) muscles. The activities of these enzymes were unaffected in the slow-tonic, anterior latissimus dorsi (ALD) muscle. Thyroidectomy had no effect on length of the muscles studied but reduced the weight of the ALD and ST muscles. These results suggest that hypothyroidism results in a "slowing down" of fast-phasic muscles, although it does not affect the activity of slow-tonic muscles.

Adenosine Triphosphatases↗

Segregation of the myogenic cell lineage in mouse muscle development.

With increasing interest in the idea of therapeutic implantation of normal muscle precursor cells into muscle lacking the protein product of the dystrophin gene, it has become important to obtain enriched populations of myogenic cells from biopsied muscle sources. Myogenic cells for implantation are highly favoured as they are the only cells that will fuse readily with host muscle fibres into which they are implanted, thus carrying the introduced gene into the target fibre with the maximum of efficiency. Second, myogenic cells appear less immunogenic than those of a non-myogenic nature; and third, the use of mononuclear myogenic cells may permit the introduction of multiple copies of a deficient gene into the patient's own cells. From a mixed population of cells obtained by the enzymic disaggregation of neonatal murine muscle we have selected, utilising a modification of the panning technique, for a cell population rich in myogenic cells. Segregation was accomplished using Mab H28, an antibody to the mouse neuronal cell adhesion molecule (N-CAM), derived from mouse/rat hybridoma cells. Following incubation with Mab H28, disaggregated muscle was applied to the surface of a bacteriological grade dish previously coated with anti-rat immunoglobulin. Cells segregated into two populations; those bearing N-CAM, and hence labelled with Mab H28, were adherent to the dish, whereas those not expressing N-CAM remained in suspension. Use of this technique, which involves minimal cell loss, resulted in the segregation of prefusion myogenic cells together with fibroblasts in the 'non-adherent' fraction, whereas cells in the adherent fraction consisted of a highly enriched population of actively dividing myogenic cells.

Animals↗

Growth and muscle development characteristics of purebred Angus and Brahman bulls.

Thirty-eight purebred bulls (10 to 17 mo of age) were used to determine the effects of breed (Angus or Brahman) and slaughter weight (60, 80, 90 or 100% of the average mature dam's weight for the respective breed) on growth and muscle development characteristics. Angus bulls grazed summer forage after weaning whereas Brahman bulls were fed to simulate gains achieved on forage by Angus. Bulls were then placed in a confinement feedlot for finishing to their appropriate slaughter weight (293, 369, 411, and 469 kg for Angus and 307, 427, 464 and 520 kg for Brahman). No major differences due to breed were found for predicted carcass composition. The LD muscle from Brahman bulls contained more total DNA (2.27 v. 1.19 g), more total protein (768.22 v. 593.59 g) and generally less total lipid (70.56 v. 101.26 g) when expressed on a total muscle (wet tissue) basis. The percentages and areas for all three muscle fiber types were not affected by breed. As carcass weight increased, muscle weights, total protein, lipid, protein:DNA and muscle fiber size for the three fiber types increased. Total DNA content increased only up to the 90% weight group and then leveled off. The percentage of alpha R fibers decreased while the percentage of alpha W fibers increased with increasing carcass weight. These data suggest that slaughtering animals based on a percentage of their dam's mature weight seems to be a practical method for making comparisons of animals on an equivalent compositional basis. Moreover, it appears that histochemical and biochemical evaluations of skeletal muscle can successfully identify what point in the growth cycle an animal is in.

Aging↗

Geographutoxin-sensitive and insensitive sodium currents in mouse skeletal muscle developing in situ.

1. The whole-cell voltage-clamp technique was used to examine developmental changes of Na+ current properties in single fibres of mouse flexor digitorum brevis muscles developing in situ from birth to 20 days post-natal. 2. Geographutoxin II (GTX II), a novel polypeptide toxin from the marine snail Conus geographus, distinguished two different types of voltage-sensitive Na+ currents: GTX II-sensitive and GTX II-insensitive currents, which corresponded respectively to currents with high or low TTX sensitivity. 3. Voltage-dependent activation and inactivation of the GTX II-insensitive currents occurred at membrane potentials 10-20 mV more negative than those for the GTX II-sensitive currents. 4. The GTX II-insensitive current in fibres from mice older than 8 days inactivated more slowly than the GTX II-sensitive current. However, in fibres from younger mice, the two currents decayed with similar speed. 5. The mean specific Na+ conductance (gNa) for the total (GTX II-sensitive plus GTX II-insensitive) Na+ channels was 0.22 mS/muF at a Na+ concentration of 5 mM at birth. The total gNa increased 6-fold to 1.32 mS/muF during the first 20 days after birth. 6. The mean specific gNa for the GTX II-insensitive channels was 0.15 mS/muF at birth, remained at approximately the same level for the first 8 days, and then decreased progressively to become undetectable by day 16. 7. In muscle fibres denervated 12 days after birth, the GTX II-insensitive gNa increased over the next 8 days, whereas the total gNa increased less than normal. 8. By contrast, in fibres denervated on day 4, the total gNa increased more than normal in the following 8 days, and the GTX II-insensitive specific gNa increased above the level seen at birth. 9. Half-maximal activation and inactivation potentials of the total and the GTX II-insensitive currents shifted in the negative direction by 9-17 mV in the first 8 days after birth. 10. We conclude that the regulatory effects of innervation on the total gNa are either suppressive or enhancing depending on the stage of development. On the other hand, denervation elicits an increase in GTX II-insensitive Na+ currents at all ages studied.

Aging↗

The E protein HEB is preferentially expressed in developing muscle.

Myogenesis is regulated by the MyoD class of myogenic regulatory factors (MRFs). These basic helix-loop-helix transcription factors dimerize with E proteins to bind conserved E-box sequences in the promoter regions of muscle-specific genes. Perhaps due to their expression in a wide array of tissues, the specific interactions of E proteins with different MRFs have been largely ignored. Likewise, the expression of E proteins in muscle tissue remains mostly uncharacterized. We investigated the expression of the E proteins HEB, E12, and E47 in rat L6 myoblasts, which express only embryonic and fast (2X) myosin heavy chains (MyHCs) in vitro, C2C12 myosatellite cells, and a number of muscle tissues, to determine whether myosin heavy chain diversity is mirrored by diversity in E protein or MRF expression. Although L6 and C2C12 myotubes demonstrate strong expression of embryonic and 2X (fast) MyHCs, immunofluorescence demonstrated the additional expression of type 1 (slow), 2A, and 2B MyHCs in the C2C12 cell line. Immunofluorescence and western blot analyses show that HEB was expressed in differentiating L6 myoblasts, C2C12 cells, and neonatal rat primary myotubes. In contrast, E12 and E47 expression was not detected in either cell line or in any adult muscle tissue examined. These data strongly implicate HEB in the development of skeletal muscle. However, because HEB is expressed in L6 myoblasts, C2C12 myosatellite cells, and neonatal hindlimb muscles, it is unlikely to be involved in a fiber type-specific manner, and may have a more general role in differentiation of myotubes.

Animals↗

Actin and myosin genes are transcriptionally regulated during mouse skeletal muscle development.

During primary and secondary myotube formation in utero and subsequent maturation of muscle fibers after birth there are complex changes in the pattern of contractile protein gene expression at the RNA and protein levels. In order to determine the degree of transcriptional regulation of actin and myosin genes we have carried out "nuclear run-on" experiments using nuclei prepared from the limb muscle of mice at 14.5, 15.5, 17.5, and 18.5 days in utero and at 10-12 and 12.5 days after birth. We show that transitions in the expression of these genes in vivo are regulated transcriptionally. Transcription of the sarcomeric alpha-actins changes from cardiac to predominantly skeletal actin over this time period; transcription of the beta-actin gene is repressed. The myosin heavy chain and myosin light chain genes also undergo transcriptional transitions during muscle development. Notably, transcription from the MLC3F promoter is activated after that of the MLC1F promoter, which is part of the same gene. These results are discussed in the context of published RNA data.

Actins↗

The expression pattern of contractile and intermediate filament proteins in developing skeletal muscle and rhabdomyosarcoma of childhood: diagnostic and prognostic utility.

In order to investigate whether rhabdomyosarcoma (RMS) can be related to equivalent stages of skeletal muscle development, muscle tissue of 21 human foetuses and 112 primary RMSs were characterized immunohistochemically using antibodies directed against vimentin, desmin, muscle-specific actin (HHF35), sarcomeric actin (sr-actin), smooth muscle actin (sm-actin), and troponin-T. During fetal skeletal muscle development, all myotubes/fibres of the first and second generations expressed desmin, HHF35, and sr-actin. Vimentin was almost exclusively present in immature primary and secondary myotubes/fibres. Troponin-T was expressed in immature myotubes/fibres of the first and second generations as well as mature fibres of the second generation. Sm-actin was never expressed. Vimentin was expressed in 96 per cent of primary and 98 per cent of relapsed RMS; HHF35 in 96 and 98 per cent, respectively; desmin in 95 and 100 per cent; troponin-T in 82 and 75 per cent; sr-actin in 71 and 86 per cent; and sm-actin in 13 and 17 per cent. The proportion of RMS cells reacting with vimentin, HHF35, and desmin was consistently higher than those expressing sr-actin and troponin-T. Neither the shape nor size of neoplastic RMS cells nor the histopathological types were related to the expression pattern of the investigated markers. RMS with aberrant expression of two or more markers predicted a worse prognosis than RMS in which at most one marker was aberrantly expressed (25 per cent and 54 per cent 10-year survival, P = 0.01). These results demonstrate that HHF35, desmin, sr-actin, and troponin-T have the potential to confirm the commitment of the tumours to the myogenic pathway which supports the diagnosis of RMS. However, it was impossible to relate RMS to equivalent stages of skeletal muscle development. Aberrant marker expression by RMS cells correlated significantly with patients' survival.

Actins↗

Three zebrafish MEF2 genes delineate somitic and cardiac muscle development in wild-type and mutant embryos.

The zebrafish is an important experimental system for vertebrate embryology, and is well suited to the molecular analysis of muscle development. Transcription factors, such as the MEF2s, regulate skeletal and cardiac muscle-specific genes during development. We report the identification of three zebrafish MEF2 genes which, like their mammalian counterparts, encode factors that function as DNA-binding transcriptional activators of muscle specific promoters. The pattern of MEF2 expression in zebrafish defines discrete cell populations in the developing somites and heart and has mechanistic implications for developmental regulation of the MEF2 genes, when compared with other species. Alteration of MEF2 expression in two mutants affecting somitogenesis provides insight into the control of muscle formation in the embryo.

Amino Acid Sequence↗