Lineage Tracing Reveals a Subset of Reserve Muscle Stem Cells Capable of Clonal Expansion under Stress

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Single-Cell Analysis of the Muscle Stem Cell Hierarchy Identifies Heterotypic Communication Signals Involved in Skeletal Muscle Regeneration

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The JAK2/STAT3 Signal Pathway Regulates the Expression of Genes Related to Skeletal Muscle Development and Energy Metabolism in Mice and Mouse Skeletal Muscle Cells

The gene encoding a β-galactosidase from Entevobacter cloacae GAO was cloned and expressed in Escherichia coli. The nucleotide sequence of the insert of a positive clone had an open reading frame of 3084 bp that encoded a polypeptide of 1028 amino acid residues with a calculated molecular mass of 116,677 daltons. The amino acid sequence of β-galactosidase deduced from the nucleotide sequence, especially the sequence around the putative active site and of the fourteen regions, showed significant homology to β-galactosidases of other microorganisms, E. coli, Klebsiella pneumoniae, Lactobacillus bulgaricus, and Clostridium acetobutylicum.     

Aging Disrupts Muscle Stem Cell Function by Impairing Matricellular WISP1 Secretion from Fibro-Adipogenic Progenitors

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Metabolic Maturation during Muscle Stem Cell Differentiation Is Achieved by miR-1/133a-Mediated Inhibition of the Dlk1-Dio3 Mega Gene Cluster

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Muscle Stem Cells Give Rise to Rhabdomyosarcomas in a Severe Mouse Model of Duchenne Muscular Dystrophy

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骨髓间充质干细胞成肌和成脂分化的调控

骨髓间充质干细胞(mesenchymal stem cells,MSCs)是来源于骨髓基质的一类具有高度自我更新能力和多向分化潜能的成体干细胞.因其具有容易获取、体外扩增方便迅速、移植排斥反应较弱等优点而成为临床应用的理想细胞模型.骨髓间充质干细胞向成肌和成脂的分化对动物机体内肌肉和脂肪的组成具有直接影响,因而与肉品质及人类健康息息相关.本文综述了骨髓间充质干细胞定向分化为骨骼肌细胞和脂肪细胞的过程及其调控机制,并重点分析了关键调控因子PRDM16(PR domain-containing16)和骨形态发生蛋白(bone morphogenetic proteins,BMPs)在骨髓间充质干细胞成肌和成脂分化中的作用.     

环境胁迫下植物MAPK多叠级联响应(英文)

Plant mitogen-activated protein kinases(MAPKs) are involved in growth,evelopment and responses to endogenous and environmental cues．which link stimuli that areactivated by external sensors to cellular responses．In Arabidopsis,as amodel,all of MAP kinase genes have been listed and classified．Based on the Arabidopsis MAPK families．a number of MAPk inase genes in other plant species have been recently isolated and classified．Most of the cloned MAPk inase genes can be activated by avariety of stresss timuli including pathogen infection．wounding．temperature,drought．salinity．osmolarity．UV irradiation．ozone and reactive oxygen species．Some tools and strategies are used to investigate their functions and signal pathways under different environmental stresses．indicating complexity and crosstalk of plant MAPk inase signaling pathways．It is still necessary to explore more novel tools and strategies to clarify MAPK signaling pathways,and how to apply the MAPK cascade to improve the resistance of crop to abiotic and biotic stress     

环境胁迫下植物MAPK多叠级联响应

植物MAP(mitogen-activated protein)激酶涉及植物的生长发育、对内源和外界环境刺激的反应.MAP激酶能将胞外感受器引起的刺激传递到胞内引起细胞的反应.拟南芥(Arabidopsis thaliana)作为模式植物,其全部的MAP激酶已经列出并进行了分类.根据已分类的拟南芥MAP激酶家族,已经分离出大量的MAP激酶基因,并将它们进行分类,发现它们大多能被包括病原、创伤、温度、干旱、盐、渗透、紫外线辐射、臭氧和活性氧等胁迫刺激激活.通过研究在不同环境胁迫下的功能和信号路径,发现植物MAP激酶信号传递系统是复杂且相互交错的.需要开发一些新的工具和策略去阐明MAPK信号传递路径,以及如何利用MAPK系统去改善农作物对生物和非生物胁迫的抗性.     

Lactate Contributes to Glyceroneogenesis and Glyconeogenesis in Skeletal Muscle by Reversal of Pyruvate Kinase

Phosphoenolpyruvate (PEP) generated from pyruvate is required for de novo synthesis of glycerol and glycogen in skeletal muscle. One possible pathway involves synthesis of PEP from the citric acid cycle intermediates via PEP carboxykinase, whereas another could involve reversal of pyruvate kinase (PK). Earlier studies have reported that reverse flux through PK can contribute carbon precursors for glycogen synthesis in muscle, but the physiological importance of this pathway remains uncertain especially in the setting of high plasma glucose. In addition, although PEP is a common intermediate for both glyconeogenesis and glyceroneogenesis, the importance of reverse PK in de novo glycerol synthesis has not been examined. Here we studied the contribution of reverse PK to synthesis of glycogen and the glycerol moiety of acylglycerols in skeletal muscle of animals with high plasma glucose. Rats received a single intraperitoneal bolus of glucose, glycerol, and lactate under a fed or fasted state. Only one of the three substrates was ¹³C-labeled in each experiment. After 3 h of normal awake activity, the animals were sacrificed, and the contribution from each substrate to glycogen and the glycerol moiety of acylglycerols was evaluated. The fraction of ¹³C labeling in glycogen and the glycerol moiety exceeded the possible contribution from either plasma glucose or muscle oxaloacetate. The reverse PK served as a common route for both glyconeogenesis and glyceroneogenesis in the skeletal muscle of rats with high plasma glucose. The activity of pyruvate carboxylase was low in muscle, and no PEP carboxykinase activity was detected.     

Muscle Stem Cells Exhibit Distinct Clonal Dynamics in Response to Tissue Repair and Homeostatic Aging

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Wnt3a诱导小鼠胚胎干细胞向心肌细胞分化的研究

目的：研究Wnt3a在诱导小鼠胚胎干细胞心肌细胞分化中的作用和原理。方法：设计不同浓度,不同成分的Wnt3a条件培养基对小鼠胚胎干细胞诱导分化,对分化细胞进行形态学鉴定,通过免疫细胞化学检测心肌肌钙蛋白-T（cTnT）的表达,通过RT．PCR检测肌球蛋白重链（ot．MHC）和肌球蛋白轻链（MLC．2v）的表达。结果：Wnt3a诱导小鼠胚胎干细胞分化为心肌样细胞,分化细胞具有自动收缩性,免疫细胞化学检测心肌肌钙蛋白．T（cTllT）表达阳性,RT．PCR检测肌球蛋白重链（d—MHC）和肌球蛋白轻链（MLC-2v）表达阳性。经典Wnt信号途径的抑制剂Frizzled一8／Fc,能够抑制Wnt3a的诱导分化作用。结论：Wnt3a通过经典Wnt信号途径诱导小鼠胚胎干细胞向心肌细胞分化。     

Notch3在血管平滑肌细胞中对Erk1/2的调控作用研究

Erk1/2活性在血管许多细胞功能中具有重要影响,而Notch3主要表达在动脉平滑肌细胞中,并且是发育过程中动脉成熟所必需的.为了探讨Notch3在血管平滑肌细胞中对Erk1/2信号通路的调控作用,采用siRNA基因敲除Notch3,γ-分泌酶抑制剂DAPT抑制Notch信号通路,质粒转染过表达Notch3活性区等方法,用Western印迹检测Notch3对血管平滑肌细胞中Erk1/2磷酸化水平,即Erk1/2活性的影响.同时,利用活性氧自由基（ROS）诱导激活Erk1/2;siRNA敲除Notch3表达致使血管平滑肌细胞中Erk1/2的磷酸化水平显著降低,并且抑制了ROS诱导的Erk1/2激活;同样,Notch通路抑制剂DAPT也抑制了ROS诱导的Erk1/2激活;而Notch3活性区NICD的过表达并没有改变血管平滑肌细胞中Erk1/2的磷酸化水平,但其延缓了ROS激活后Erk1/2活性的衰减.上述结果表明,Notch3可在血管平滑肌细胞中调控Erk1/2活性以及ROS诱导的Erk1/2信号激活.     

Visualization of G3BP Stress Granules Dynamics in Live Primary Cells

SGs can be visualized in cells by immunostaining of specific protein components or polyA+ mRNAs. SGs are highly dynamic and the study of their assembly and fate is important to understand the cellular response to stress. The deficiency in key factors of SGs like G3BP (RasGAP SH3 domain Binding Protein) leads to developmental defects in mice and alterations of the Central Nervous System. To study the dynamics of SGs in cells from an organism, one can culture primary cells and follow the localization of a transfected tagged component of SGs. We describe time-lapse experiment to observe G3BP1-containing SGs in Mouse Embryonic Fibroblasts (MEFs). This technique can also be used to study G3BP-containing SGs in live neurons, which is crucial as it was recently shown that these SGs are formed at the onset of neurodegenerative diseases like Alzheimer''s disease. This approach can be adapted to any other cellular body and granule protein component, and performed with transgenic animals, allowing the live study of granules dynamics for example in the absence of a specific factor of these granules.     

Transplantation of Induced Pluripotent Stem Cell-derived Mesoangioblast-like Myogenic Progenitors in Mouse Models of Muscle Regeneration

Patient-derived iPSCs could be an invaluable source of cells for future autologous cell therapy protocols. iPSC-derived myogenic stem/progenitor cells similar to pericyte-derived mesoangioblasts (iPSC-derived mesoangioblast-like stem/progenitor cells: IDEMs) can be established from iPSCs generated from patients affected by different forms of muscular dystrophy. Patient-specific IDEMs can be genetically corrected with different strategies (e.g. lentiviral vectors, human artificial chromosomes) and enhanced in their myogenic differentiation potential upon overexpression of the myogenesis regulator MyoD. This myogenic potential is then assessed in vitro with specific differentiation assays and analyzed by immunofluorescence. The regenerative potential of IDEMs is further evaluated in vivo, upon intramuscular and intra-arterial transplantation in two representative mouse models displaying acute and chronic muscle regeneration. The contribution of IDEMs to the host skeletal muscle is then confirmed by different functional tests in transplanted mice. In particular, the amelioration of the motor capacity of the animals is studied with treadmill tests. Cell engraftment and differentiation are then assessed by a number of histological and immunofluorescence assays on transplanted muscles. Overall, this paper describes the assays and tools currently utilized to evaluate the differentiation capacity of IDEMs, focusing on the transplantation methods and subsequent outcome measures to analyze the efficacy of cell transplantation.     

Differential Regulation of NOTCH2 and NOTCH3 Contribute to Their Unique Functions in Vascular Smooth Muscle Cells

Notch signaling is a key regulator of vascular smooth muscle cell (VSMC) phenotypes, including differentiation, proliferation, and cell survival. However, the exact contribution of the individual Notch receptors has not been thoroughly delineated. In this study, we identify unique roles for NOTCH2 and NOTCH3 in regulating proliferation and cell survival in cultured VSMCs. Our results indicate that NOTCH2 inhibits PDGF-B-dependent proliferation and its expression is decreased by PDGF-B. In contrast, NOTCH3 promotes proliferation and receptor expression is increased by PDGF-B. Additionally, data show that NOTCH3, but not NOTCH2 protects VSMCs from apoptosis and apoptosis mediators degrade NOTCH3 protein. We identified three pro-survival genes specifically regulated by NOTCH3 in cultured VSMCs and in mouse aortas. This regulation is mediated through MAP kinase signaling, which we demonstrate can be activated by NOTCH3, but not NOTCH2. Overall, this study highlights discrete roles for NOTCH2 and NOTCH3 in VSMCs and connects these roles to specific upstream regulators that control their expression.     

Sustained co-cultivation with human placenta-derived MSCs enhances ALK5/Smad3 signaling in human breast epithelial cells,leading to EMT and differentiation

The interaction between mammary epithelial cells and their surrounding microenvironment are important in the development of the mammary gland. Thus, mesenchymal stem cells (MSCs), which retain pluripotency for various mesenchymal lineages, may provide a permissive environment for the morphologic alteration and differentiation of mammary epithelial cells. To this end, we investigated whether the interactions between mammary epithelial cells and human placenta-derived MSCs (hPMSC) affect the morphology, proliferation, and differentiation of epithelial cells in a co-culture system. We show that after co-culture with hPMSCs, human mammary epithelial cell lines (MCF-10F and HEMC) underwent significant morphologic alterations and a dramatic increase in ductal–alveolar branching, which was accompanied by a decrease or loss of the epithelial marker E-cadherin and a gain of the mesenchymal markers, α-SMA and vimentin. MCF-10F and HEMC proliferation was also inhibited in the presence of hPMSCs, and this retardation in growth was due to cell cycle arrest. Furthermore, in MCF-10F and HMEC cells, hPMSCs induced the production of lipid droplets, milk fat globule protein, and milk protein lactoferrin, which are markers of functional mammary differentiation. We also noticed an elevation in ALK5 and phosphorylated Smad3 protein levels upon hPMSC co-culture. Strikingly, the changes in morphology, proliferation, and differentiation were reversed by treatment with ALK5 or Smad3 knockdown in MCF-10F/hPMSC co-cultures. Collectively, our findings suggest that co-cultivation with hPMSCs leads to epithelial to mesenchymal transition (EMT) and differentiation of human breast epithelial cells through the ALK5/Smad3 signaling pathway.