Transgenic mice that express Cre recombinase under control of a skeletal muscle-specific promoter from mef2c

Genes expressed in skeletal muscle are often required in other tissues. This is particularly the case for cardiac and smooth muscle, both contractile tissues that share numerous characteristics with skeletal muscle, such that targeted inactivation can lead to embryonic lethality prior to a requirement for gene function in skeletal muscle. Thus, it is essential that conditional inactivation approaches are developed to disrupt genes specifically in skeletal muscle. In this report, we describe a transgenic mouse that expresses Cre recombinase under the control of a skeletal muscle-specific promoter from the mef2c gene. Cre expression in this transgenic line is completely restricted to skeletal muscle from early in development and is present in all skeletal muscles, including those of epaxial and hypaxial origins and in fast and slow fibers. This early skeletal muscle-specific Cre line will be a useful tool to define the function of genes specifically in skeletal muscle.     

骨髓间充质干细胞成肌分化在骨骼肌再生中的应用

骨骼肌良好的再生能力是由于肌卫星细胞的存在,然而肌卫星细胞的数量仅占骨骼肌细胞数量的1%~ 5%,当肌肉损伤时,仅依靠这些卫星细胞还不足以促进骨骼肌修复与再生,并且这种再生能力会随着年龄的增大而衰减,并不能修复损伤严重的骨骼肌。骨髓间充质干细胞（BMSC）因其多向分化潜能,旁分泌潜能,免疫调节能力及容易获取等特点广泛用于损伤骨骼肌的修复与再生。但在某种程度上,仅仅采用BMSC治疗损伤的骨骼肌仍不能达到满意的效果。因此,大量研究采用药物、生物材料、细胞及细胞因子对BMSC进行预处理不仅可改善它的移植率,还可显著促进其向骨骼肌分化,从而最大限度的发掘骨骼肌间充质干细胞的成肌分化潜能以促进骨骼肌的修复。因此,本篇综述旨在概括BMSC成肌分化在骨骼肌再生中的应用。     

Polymorphic forms of troponin T and troponin C and their localization in striated muscle cell types

1. 1. Immunochemical studies have shown that the major forms of troponin T present in fast skeletal, slow skeletal and cardiac muscles are different proteins.

2. 2. Similar studies indicate that the major form of troponin C present in fast skeletal muscles differs from troponin C present in slow skeletal and cardiac muscle cells. The forms of troponin C present in slow skeletal and cardiac muscles are immunochemically very similar.

3. 3. The antibodies to the polymorphic forms of troponin T and troponin C are specific for the muscle type, except in the case of the slow skeletal and cardiac muscle forms of troponin C.

4. 4. By the immunoperoxidase technique, it has been shown that the fast skeletal muscle troponin T is localized in type II cells and slow skeletal muscle troponin T in type I cells.

5. 5. Fast skeletal muscle troponin C is present in type II cells and a different troponin C, identified by its reaction with the antibody against cardiac troponin C, is present in type I cells.

6. 6. It is concluded that in normal adult skeletal muscle, fast muscle forms of troponin I, troponin T and troponin C are present together as a homocomplex in type II cells and the slow muscle forms exist as an analagous homocomplex in type I cells.

     

Fatness and skeletal maturity of Belgian boys 12 through 17 years of age

Relationships between fatness and skeletal maturity are considered in a nationwide sample of 14,259 Belgian boys 12 through 17 years of age (The Leuven Growth Study of Belgian Boys). Absolute fatness was estimated from four skinfolds using the Drinkwater and Ross technique and from the sum of four skinfolds, and was related to skeletal maturity assessed by the Tanner-Whitehouse method (I and II). In addition, comparisons were made between the fattest 5% and leanest 5% of the boys at each age level. Correlations between the indices of fatness and skeletal age and relative skeletal age (the difference between skeletal and chronological ages) are positive and generally low, ranging from 0.12 to 0.39. They tend to decrease with age from 12 to 17 years. Comparisons between the extreme groups indicate that the leanest boys are more delayed in skeletal maturity, by about 0.8 years, than the fattest boys are advanced, by about 0.5 years. Stature data for the same boys are consistent with the skeletal maturity data and thus suggest that the size differences between the extreme groups are due in part to maturity differences. Over the age span 12 through 20 years, the leanest boys are reduced in stature by about – 1.2 standard deviations, while the fattest boys are larger in stature by about +0.6 standard deviation units. The size differences, however, persist after skeletal maturity is attained so that there may be a specific role for fatness in influencing statural growth.     

大鼠骨骼肌快肌肌钙蛋白T的同工型

骨骼肌快肌的收缩主要是由钙离子通过肌钙蛋白所调节控制。这些肌钙蛋白位于肌纤维之中。肌蛋白包括肌钙蛋白T、肌钙蛋白C、肌钙蛋白I。采用双向聚丙烯酰胺凝胶电泳和免疫学技术,对大鼠胚胎、新生大鼠和成年大鼠的骨能肌快肌肌钙蛋白T的同工型进行了研究。在成年大鼠的骨能肌快肌中,发现了10种肌钙蛋白T同工型。在大鼠胚胎和新生大鼠的骨能肌中,发现了7种肌钙蛋白T同工型。作为不同动物、不同发育阶段和不同组织发育的特殊标记,这些肌钙蛋白T同工型具有重要意义[动物学报49(3)：362—369,2003]。     

Bone regeneration via skeletal cell lineage plasticity: All hands mobilized for emergencies

An emerging concept is that quiescent mature skeletal cells provide an important cellular source for bone regeneration. It has long been considered that a small number of resident skeletal stem cells are solely responsible for the remarkable regenerative capacity of adult bones. However, recent in vivo lineage‐tracing studies suggest that all stages of skeletal lineage cells, including dormant pre‐adipocyte‐like stromal cells in the marrow, osteoblast precursor cells on the bone surface and other stem and progenitor cells, are concomitantly recruited to the injury site and collectively participate in regeneration of the damaged skeletal structure. Lineage plasticity appears to play an important role in this process, by which mature skeletal cells can transform their identities into skeletal stem cell‐like cells in response to injury. These highly malleable, long‐living mature skeletal cells, readily available throughout postnatal life, might represent an ideal cellular resource that can be exploited for regenerative medicine.     

Differentially expressed fibroblast growth factors regulate skeletal muscle development through autocrine and paracrine mechanisms

Several FGF family members are expressed in skeletal muscle; however, the roles of these factors in skeletal muscle development are unclear. We examined the RNA expression, protein levels, and biological activities of the FGF family in the MM14 mouse skeletal muscle cell line. Proliferating skeletal muscle cells express FGF-1, FGF-2, FGF-6, and FGF-7 mRNA. Differentiated myofibers express FGF-5, FGF-7, and reduced levels of FGF-6 mRNA. FGF-3, FGF-4, and FGF-8 were not detectable by RT-PCR in either proliferating or differentiated skeletal muscle cells. FGF-I and FGF-2 proteins were present in proliferating skeletal muscle cells, but undetectable after terminal differentiation. We show that transfection of expression constructs encoding FGF-1 or FGF-2 mimics the effects of exogenously applied FGFs, inhibiting skeletal muscle cell differentiation and stimulating DNA synthesis. These effects require activation of an FGF tyrosine kinase receptor as they are blocked by transfection of a dominant negative mutant FGF receptor. Transient transfection of cells with FGF-1 or FGF-2 expression constructs exerted a global effect on myoblast DNA synthesis, as greater than 50% of the nontransfected cells responded by initiating DNA synthesis. The global effect of cultures transfected with FGF-2 expression vectors was blocked by an anti-FGF-2 monoclonal antibody, suggesting that FGF-2 was exported from the transfected cells. Despite the fact that both FGF-l and FGF-2 lack secretory signal sequences, when expressed intracellularly, they regulate skeletal muscle development. Thus, production of FGF-1 and FGF-2 by skeletal muscle cells may act as a paracrine and autocrine regulator of skeletal muscle development in vivo.     

The abundance of calmodulin mRNAs is regulated in phosphorylase kinase-deficient skeletal muscle

In the I/Lyn mouse strain a mutation on the X chromosome results in a deficiency of the major calmodulin-regulated enzyme in skeletal muscle, phosphorylase kinase. Calmodulin has been identified as the delta-subunit of phosphorylase kinase, and it is estimated that approximately 40% of the total calmodulin in rabbit skeletal muscle is associated with the phosphorylase kinase hexadecamer (alpha, beta, gamma, delta)4. The absence of phosphorylase kinase in I/Lyn skeletal muscle results in a reduction in the total amount of calmodulin. The mechanisms affecting this reduction were investigated by comparing the abundance and heterogeneities in calmodulin mRNAs between normal and phosphorylase kinase-deficient skeletal muscles. The results demonstrate that in normal tissue there are four species of calmodulin mRNA distinguished by their molecular weight. All four of these species are present in the deficient tissue, and none of them are preferentially reduced. However, there is a 54% reduction in all four mRNAs as well as in calmodulin in the deficient skeletal muscle relative to normal skeletal muscle. These results indicate that the expression of calmodulin mRNAs is coordinated with the expression of its major enzyme target in skeletal muscle.     

Comparability in skeletal maturation research

Comparability is a fundamental issue in skeletal maturation research. Since the introduction of the first edition of the Greulich-Pyle Atlas and the Tanner-Whitehouse method, a number of methodologic reports have appeared regarding potential sources of error, reliability and replicability in the assessment of skeletal maturity from hand-wrist radiographs. Some of these reports are mentioned and two recent examples of methodologic studies are cited. Maximum reliability of skeletal assessments can be expected only when there is strict adherence to carefully standardized investigative procedures. Technical as well as human factors must be taken into account to insure minimal variation in findings within and between laboratories over time. Single or serial skeletal radiographs uniformly taken on properly identified subjects constitute a valuable permanent record of biologic maturation. while the film image can be considered as objective evidence of skeletal maturity, a subjective element is introduced in observing and reporting the presence or absence of particular ossification centers, or rating an ossification pattern against a standard. Intra- and interobserver skeletal maturity assessment replicability relates to such factors as motivation, training, assessment method, and quality control procedures. Suggestions are presented to facilitate comparability in skeletal maturation research, including the possibility of preparation and distribution of sets of standardized skeletal radiographs for periodic determination and improvement of assessor reliability.     

A fetal skeletal muscle actin mRNA in the mouse and its identity with cardiac actin mRNA

We compare a recombinant cDNA plasmid (pAF81) complementary to a fetal skeletal muscle actin mRNA with a plasmid (pAM91) complementary to the actin mRNA expressed in adult skeletal muscle. The two mRNAs are significantly diverged in silent nucleotide positions; they are coexpressed in fetal skeletal muscle, and in differentiating muscle cell cultures their accumulation begins coordinately. The sequence of pAF81 shows that the amino acid sequence of mouse fetal skeletal muscle actin is almost identical to that of adult bovine cardiac actin. Hybridization of pAF81 to RNA from different mouse tissues shows that fetal skeletal muscle actin mRNA is very homologous or identical to fetal and adult cardiac actin mRNA. Only one gene homologous to pAF81 is detected on blots of restricted mouse DNA. We conclude that this gene must be expressed both in fetal skeletal muscle and in fetal heart. Whereas mRNA transcribed from this gene is the major actin mRNA species in adult heart, it is present in low amounts, if at all, in adult skeletal muscle.     

Current topics for teaching skeletal muscle physiology

Contractions of skeletal muscles provide the stability and power for all body movements. Consequently, any impairment in skeletal muscle function results in some degree of instability or immobility. Factors that influence skeletal muscle structure and function are therefore of great interest both scientifically and clinically. Injury, disease, and old age are among the factors that commonly contribute to impairment in skeletal muscle function. The goal of this article is to update current concepts of skeletal muscle physiology. Particular emphasis is placed on mechanisms of injury, repair, and adaptation in skeletal muscle as well as mechanisms underlying the declining skeletal muscle structure and function associated with aging. For additional materials please refer to the "Skeletal Muscle Physiology" presentation located on the American Physiological Society Archive of Teaching Resources Web site (http://www.apsarchive.org).     

New insights into the role of sphingosine 1-phosphate and lysophosphatidic acid in the regulation of skeletal muscle cell biology

Lysophospholipids are bioactive molecules that are implicated in the control of fundamental biological processes such as proliferation, differentiation, survival and motility in different cell types. Here we review the role of sphingosine 1-phosphate (S1P) and lysophosphatidic acid (LPA) in the regulation of skeletal muscle biology. Indeed, a wealth of experimental data indicate that these molecules are crucial players in the skeletal muscle regeneration process, acting by controllers of activation, proliferation and differentiation not only of muscle-resident satellite cells but also of mesenchymal progenitors that originate outside the skeletal muscle. Moreover, S1P and LPA are clearly involved in the regulation of skeletal muscle metabolism, muscle adaptation to different physiological needs and resistance to muscle fatigue. Notably, studies accomplished so far, have highlighted the complexity of S1P and LPA signaling in skeletal muscle cells that appears to be further complicated by their close dependence on functional cross-talks with growth factors, hormones and cytokines. Our increasing understanding of bioactive lipid signaling can individuate novel molecular targets aimed at enhancing skeletal muscle regeneration and reducing the fibrotic process that impairs full functional recovery of the tissue during aging, after a trauma or skeletal muscle diseases. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.     

Phosphorylation of the cardiac isoform of calsequestrin in cultured rat myotubes and rat skeletal muscle.

Calsequestrin is a high-capacity Ca(2+)-binding protein and a major constituent of the sarcoplasmic reticulum (SR) of both skeletal and cardiac muscle. Two isoforms of calsequestrin, cardiac and skeletal muscle forms, have been described which are products of separate genes. Purified forms of the two prototypical calsequestrin isoforms, dog cardiac and rabbit fast-twitch skeletal muscle calsequestrins, serve as excellent substrates for casein kinase II and are phosphorylated on distinct sites (Cala, S.E. and Jones, L.R. (1991) J. Biol. Chem 266, 391-398). Dog cardiac calsequestrin is phosphorylated at a 50 to 100-fold greater rate than is rabbit skeletal muscle calsequestrin, and only the dog cardiac isoform contains endogenous Pi on casein kinase II phosphorylation sites. In this study, we identified and examined both calsequestrin isoforms in rat muscle cultures and homogenates to demonstrate that the cardiac isoform of calsequestrin in rat skeletal muscle was phosphorylated in vivo on sites which are phosphorylated by casein kinase II in vitro. Phosphorylation of rat skeletal muscle calsequestrin was not detected. In tissue homogenates, cardiac and skeletal muscle calsequestrin isoforms were both found to be prominent substrates for endogenous casein kinase II activity with cardiac calsequestrin the preferred substrate. In addition, these studies revealed that the cardiac isoform of calsequestrin was the predominant form expressed in skeletal muscle of fetal rats and cultured myotubes.     

Peptide and non-peptide G-protein coupled receptors (GPCRs) in skeletal muscle

G-protein coupled receptors (GPCRs) represent a large class of cell surface receptors that mediate a multitude of functions. Over the years, a number of GPCRs and ancillary proteins have been shown to be expressed in skeletal muscle. Unlike the case with other muscle tissues like cardiac and vascular smooth muscle cells, there has been little attempt at systematically analyzing GPCRs in skeletal muscle. Here we have compiled all the GPCRs that are expressed in skeletal muscle. In addition, we review the known function of these receptors in both skeletal muscle tissue and in cultured skeletal muscle cells.