Myostatin and MyoD family expression in skeletal muscle of IGF-1 knockout mice

Insulin-like growth factor-1 (IGF-1) is a positive regulator in proliferation and differentiation of skeletal muscle cells, while myostatin (MSTN) is a member of transforming growth factor beta superfamily that acts as a negative regulator of skeletal muscle mass. The present study was performed to detail whether a correlation exists between MSTN and IGF-1 in skeletal muscle of IGF-1 knockout mice (IGF-1(-/-)) and their wild type (WT; i.e., IGF-1(+/+)) littermates. The body weight of IGF-1(-/-) animals was 32% that of WT littermates. The fiber cross-sectional areas (CSA) and number of fibers in M. rectus femoris of IGF-1(-/-) animals were 49 and 59% those of WT animals, respectively. Thus, muscle hypoplasia of IGF-1(-/-) undoubtedly was confirmed. Myostatin mRNA levels and protein levels were similar between M. gastrocnemius of IGF-1(-/-) and WT animals. Myostatin immunoreactivity was similarly localized in muscle fibers of both IGF-1(-/-) and WT M. rectus femoris. The mRNA levels of MyoD family (Myf5, MyoD, MRF4, myogenin) were differentially expressed in IGF-1(-/-)M. gastrocnemius, in which the mRNA expression of MRF4 and myogenin was significantly lower, whereas there were no changes in the mRNA expression of Myf5 and MyoD. These findings first describe that myostatin expression is not influenced by intrinsic failure of IGF-1, although MRF4 and myogenin are downregulated.     

血清miRNA-93、miRNA-223联合miRNA-324-3p可作为诊断PCOS的生物标记物

本研究拟基于最近的miRNA报道,结合前期关于肌肉生长抑素(myostatin, MSTN)的相关研究,探究多囊卵巢综合症(ploycystic ovary syndrome, PCOS)患者血清miRNA和生长抑素的表达及其相关临床病理特征。本研究选取160例在湘南学院附属医院妇科就诊的女性为研究对象,通过PCR、ELISA等方法比较PCOS患者血清miRNA和生长抑素的表达情况,并且分析其相关临床病理特征。结果显示,通过PCR的方法发现mi RNA-93、mi RNA-223在PCOS患者的血清中显著升高,而mi RNA-4522、mi RNA-6767-5p和mi RNA-324-3p则表达下降。本研究建立了3个miRNA的预测模型,并证实预测模型在筛选组和验证组中有很好的敏感性和特异性,可以有效区分PCOS患者和正常人群,但研究结果也发现,miRNA模型结合MSTN没有更好的诊断价值。     

Mighty is a novel promyogenic factor in skeletal myogenesis

Genetic analysis has revealed an important function in myogenesis for Myostatin, a member of the TGF-beta superfamily. However, the cascade of genes that responds to Myostatin signalling to regulate myogenesis is not well understood. Thus, a suppressive subtraction hybridization to identify such genes was undertaken and here we report the cloning and characterization of a novel gene, Mighty. Mighty is expressed in a variety of different tissues but appears to be specifically regulated by Myostatin in skeletal muscle. Overexpression of Mighty in C2C12 cells results in early withdrawal of myoblasts from the cell cycle, enhanced and accelerated differentiation and hypertrophy of myotubes. Most importantly, Mighty overexpression leads to increased and earlier expression of MyoD and increased secretion of another known differentiation inducing factor, IGF-II. Furthermore, viral expression of Mighty in mdx mice resulted in an increase in the number of larger healthy muscle fibers. Given its role in myogenesis, we propose that Mighty is a critical promyogenic factor which plays a key role in the signalling pathway downstream of Myostatin.     

Complex Microsatellite Dynamics in the Myostatin Gene Within Ruminants

A microsatellite has previously been identified in myostatin in cattle. Sequencing of this region from other artiodactyls coupled with phylogenetic analysis has been used to uncover the potential origins of the microsatellite event, which appears either to have been born twice or to have been gained and lost within ruminants. While caprids and ovids share the ancestral state with pigs and other mammals, microsatellite activity (length polymorphism) is uncovered in both deer and bovids. The dynamic process of microsatellite evolution, including birth, is discussed here in light of several models. Finally, these models are evaluated in the context of patterns of microsatellite conservation between closely related mammalian genomes.     

Overexpression of Myostatin2 in zebrafish reduces the expression of dystrophin associated protein complex (DAPC) which leads to muscle dystrophy

Myostatin, a member of the TGF-β superfamily, is a potent negative regulator of skeletal muscle and growth. Previously, we reported Mstn1 from zebrafish and studied its influence on muscle development. In this study, we identified another form of Myostatin protein which is referred to as Mstn2. The size of Mstn2 cDNA is 1342 bp with 109 and 132 bp of 5′ and 3′-untranslated regions (UTRs), respectively. The coding region is 1101 bp encoding 367 amino acids. The identity between zebrafish Mstn1 and 2 is 66%. The phylogenetic tree revealed that the Mstn2 is an ancestral form of Mstn1. To study the functional aspects, we overexpressed mstn2 and noticed that embryos became less active and the juveniles with bent and curved phenotypes when compared to the control. The RT-PCR and in situ hybridization showed concurrent reduction of dystrophin associated protein complex (DAPC). In cryosection and in situ hybridization, we observed the disintegration of somites, lack of transverse myoseptum and loss of muscle integrity due to the failure of muscle attachment in mstn2 overexpressed embryos. Immunohistochemistry and western blot showed that there was a reduction of dystrophin, dystroglycan and sarcoglycan at translational level in overexpressed embryos. Taken together, these results indicate the suitability of zebrafish as an excellent animal model and our data provide the first in vivo evidence of muscle attachment failure by the overexpression of mstn2 and it leads to muscle loss which results in muscle dystrophy that may contribute to Duchenne syndrome and other muscle related diseases. A. Anusha Amali and Cliff Ji-Fan Lin contributed equally.     

鳜肌肉生长抑制素Myostatin cDNA克隆与组织表达分析

利用RT-PCR和cDNA末端快速扩增法(RACE)克隆了鳜(Siniperca chuatsi)肌肉生长抑制素(myostatin,MSTN)cDNA序列,并分析了该基因的结构特征和亲缘关系。鳜MSTN cDNA序列全长2627bp,包括5′端非翻译区117bp、3′端非翻译区1376bp和开放阅读框(ORF)1134bp,共编码377个氨基酸,含22个氨基酸的信号肽。鳜MSTN具有脊椎动物MSTN的共同序列特征,含有1个蛋白酶水解位点RARR和9个保守的半胱氨酸残基。脊椎动物MSTN氨基酸序列的亲缘关系分析表明,鳜与其他鱼类聚为一支。RT-PCR分析表明,鳜MSTN在成体不同组织中的表达情况不同,其中,卵巢、肾、眼、肌肉、心、脑、皮肤和胃中有表达,肝胰脏未见表达。     

Effect of myostatin on turkey myogenic satellite cells and embryonic myoblasts

Myostatin (GDF-8) inhibits the activation, proliferation, and differentiation of myogenic satellite cells. The relative importance of this growth factor is demonstrated in myostatin-null mice and cattle possessing defective myostatin genes. These defects result in greatly enhanced musculature. In the present study, we examined the effect of myostatin on turkey myogenic satellite cells and embryonic myoblasts. Compared with controls (P<0.05), proliferation of both turkey embryonic myoblasts and satellite cells was inhibited between 26 and 45% in serum-free medium containing 20 ng/mL myostatin. While individual turkey satellite cell clones differed in their responsiveness to myostatin, there were no significant differences in the responsiveness of fast and slow growing cells as groups (P>0.05). A slow growing clone that exhibited the greatest response to myostatin also exhibited the greatest depression of differentiation with this growth factor (P<0.05). All other turkey satellite cell clones exhibited similar responses to the differentiation depressing effects of myostatin (P>0.05). However, myostatin had no effect on differentiation of turkey embryonic myoblasts (P>0.05). When exposed to myostatin, 4 of 6 proliferating clones and all differentiating clones increased their expression of decorin, a growth inhibitor (P<0.05). The present study demonstrates that myostatin inhibits the proliferation and differentiation of satellite cells and suggests a role for decorin in myostatin action in muscle development.     

Discovery of a follistatin-derived myostatin inhibitory peptide

Follistatin is well known as an inhibitor of transforming growth factor (TGF)-β superfamily ligands including myostatin and activin A. Myostatin, a negative regulator of muscle growth, is a promising target with which to treat muscle atrophic diseases. Here, we focused on the N-terminal domain (ND) of follistatin (Fst) that interacts with the type I receptor binding site of myostatin. Through bioassay of synthetic ND-derived fragment peptides, we identified DF-3, a new myostatin inhibitory 14-mer peptide which effectively inhibits myostatin, but fails to inhibit activin A or TGF-β1, in an in vitro luciferase reporter assay. Injected intramuscularly, DF-3 significantly increases skeletal muscle mass in mice and consequently, it can serve as a platform for development of muscle enhancement based on myostatin inhibition.