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Sample records for adult muscle regeneration

  1. Fgf regulates dedifferentiation during skeletal muscle regeneration in adult zebrafish.

    PubMed

    Saera-Vila, Alfonso; Kish, Phillip E; Kahana, Alon

    2016-09-01

    Fibroblast growth factors (Fgfs) regulate critical biological processes such as embryonic development, tissue homeostasis, wound healing, and tissue regeneration. In zebrafish, Fgf signaling plays an important role in the regeneration of the spinal cord, liver, heart, fin, and photoreceptors, although its exact mechanism of action is not fully understood. Utilizing an adult zebrafish extraocular muscle (EOM) regeneration model, we demonstrate that blocking Fgf receptor function using either a chemical inhibitor (SU5402) or a dominant-negative transgenic construct (dnFGFR1a:EGFP) impairs muscle regeneration. Adult zebrafish EOMs regenerate through a myocyte dedifferentiation process, which involves a muscle-to-mesenchyme transition and cell cycle reentry by differentiated myocytes. Blocking Fgf signaling reduced cell proliferation and active caspase 3 levels in the regenerating muscle with no detectable levels of apoptosis, supporting the hypothesis that Fgf signaling is involved in the early steps of dedifferentiation. Fgf signaling in regenerating myocytes involves the MAPK/ERK pathway: inhibition of MEK activity with U0126 mimicked the phenotype of the Fgf receptor inhibition on both muscle regeneration and cell proliferation, and activated ERK (p-ERK) was detected in injured muscles by immunofluorescence and western blot. Interestingly, following injury, ERK2 expression is specifically induced and activated by phosphorylation, suggesting a key role in muscle regeneration. We conclude that the critical early steps of myocyte dedifferentiation in EOM regeneration are dependent on Fgf signaling.

  2. Myocyte Dedifferentiation Drives Extraocular Muscle Regeneration in Adult Zebrafish

    PubMed Central

    Saera-Vila, Alfonso; Kasprick, Daniel S.; Junttila, Tyler L.; Grzegorski, Steven J.; Louie, Ke'ale W.; Chiari, Estelle F.; Kish, Phillip E.; Kahana, Alon

    2015-01-01

    Purpose The purpose of this study was to characterize the injury response of extraocular muscles (EOMs) in adult zebrafish. Methods Adult zebrafish underwent lateral rectus (LR) muscle myectomy surgery to remove 50% of the muscle, followed by molecular and cellular characterization of the tissue response to the injury. Results Following myectomy, the LR muscle regenerated an anatomically correct and functional muscle within 7 to 10 days post injury (DPI). Following injury, the residual muscle stump was replaced by a mesenchymal cell population that lost cell polarity and expressed mesenchymal markers. Next, a robust proliferative burst repopulated the area of the regenerating muscle. Regenerating cells expressed myod, identifying them as myoblasts. However, both immunofluorescence and electron microscopy failed to identify classic Pax7-positive satellite cells in control or injured EOMs. Instead, some proliferating nuclei were noted to express mef2c at the very earliest point in the proliferative burst, suggesting myonuclear reprogramming and dedifferentiation. Bromodeoxyuridine (BrdU) labeling of regenerating cells followed by a second myectomy without repeat labeling resulted in a twice-regenerated muscle broadly populated by BrdU-labeled nuclei with minimal apparent dilution of the BrdU signal. A double-pulse experiment using BrdU and 5-ethynyl-2′-deoxyuridine (EdU) identified double-labeled nuclei, confirming the shared progenitor lineage. Rapid regeneration occurred despite a cell cycle length of 19.1 hours, whereas 72% of the regenerating muscle nuclei entered the cell cycle by 48 hours post injury (HPI). Dextran lineage tracing revealed that residual myocytes were responsible for muscle regeneration. Conclusions EOM regeneration in adult zebrafish occurs by dedifferentiation of residual myocytes involving a muscle-to-mesenchyme transition. A mechanistic understanding of myocyte reprogramming may facilitate novel approaches to the development of molecular

  3. Myogenic regulatory factors during regeneration of skeletal muscle in young, adult, and old rats

    NASA Technical Reports Server (NTRS)

    Marsh, D. R.; Criswell, D. S.; Carson, J. A.; Booth, F. W.

    1997-01-01

    Myogenic factor mRNA expression was examined during muscle regeneration after bupivacaine injection in Fischer 344/Brown Norway F1 rats aged 3, 18, and 31 mo of age (young, adult, and old, respectively). Mass of the tibialis anterior muscle in the young rats had recovered to control values by 21 days postbupivacaine injection but in adult and old rats remained 40% less than that of contralateral controls at 21 and 28 days of recovery. During muscle regeneration, myogenin mRNA was significantly increased in muscles of young, adult, and old rats 5 days after bupivacaine injection. Subsequently, myogenin mRNA levels in young rat muscle decreased to postinjection control values by day 21 but did not return to control values in 28-day regenerating muscles of adult and old rats. The expression of MyoD mRNA was also increased in muscles at day 5 of regeneration in young, adult, and old rats, decreased to control levels by day 14 in young and adult rats, and remained elevated in the old rats for 28 days. In summary, either a diminished ability to downregulate myogenin and MyoD mRNAs in regenerating muscle occurs in old rat muscles, or the continuing myogenic effort includes elevated expression of these mRNAs.

  4. Muscle regeneration by adipose tissue-derived adult stem cells attached to injectable PLGA spheres.

    PubMed

    Kim, MiJung; Choi, Yu Suk; Yang, Seung Hye; Hong, Hea-Nam; Cho, Sung-Woo; Cha, Sang Myun; Pak, Jhang Ho; Kim, Chan Wha; Kwon, Seog Woon; Park, Chan Jeoung

    2006-09-22

    The [corrected] use of adult stem cells for cell-based tissue engineering and regeneration strategies represents a promising approach for skeletal muscle repair. We have evaluated the combination of adipose tissue-derived adult stem cells (ADSCs) obtained from autologous liposuction and injectable poly(lactic-co-glycolic acid) (PLGA) spheres for muscle regeneration. ADSCs attached to PLGA spheres and PLGA spheres alone were cultured in myogenic medium for 21 days and injected subcutaneously into the necks of nude mice. After 30 and 60 days, the mice were sacrificed, and newly formed tissues were analyzed by immunostaining, H and E staining, and RT-PCR. We found that ADSCs attached to PLGA spheres, but not PLGA spheres alone, were able to generate muscle tissue. These findings suggest that ADSCs and PLGA spheres are useful materials for muscle tissue engineering and that their combination can be used in clinical settings for muscle regeneration.

  5. Pax7-expressing satellite cells are indispensable for adult skeletal muscle regeneration.

    PubMed

    Sambasivan, Ramkumar; Yao, Roseline; Kissenpfennig, Adrien; Van Wittenberghe, Laetitia; Paldi, Andràs; Gayraud-Morel, Barbara; Guenou, Hind; Malissen, Bernard; Tajbakhsh, Shahragim; Galy, Anne

    2011-09-01

    Distinct cell populations with regenerative capacity have been reported to contribute to myofibres after skeletal muscle injury, including non-satellite cells as well as myogenic satellite cells. However, the relative contribution of these distinct cell types to skeletal muscle repair and homeostasis and the identity of adult muscle stem cells remain unknown. We generated a model for the conditional depletion of satellite cells by expressing a human diphtheria toxin receptor under control of the murine Pax7 locus. Intramuscular injection of diphtheria toxin during muscle homeostasis, or combined with muscle injury caused by myotoxins or exercise, led to a marked loss of muscle tissue and failure to regenerate skeletal muscle. Moreover, the muscle tissue became infiltrated by inflammatory cells and adipocytes. This localised loss of satellite cells was not compensated for endogenously by other cell types, but muscle regeneration was rescued after transplantation of adult Pax7(+) satellite cells alone. These findings indicate that other cell types with regenerative potential depend on the presence of the satellite cell population, and these observations have important implications for myopathic conditions and stem cell-based therapeutic approaches.

  6. Myomaker is essential for muscle regeneration.

    PubMed

    Millay, Douglas P; Sutherland, Lillian B; Bassel-Duby, Rhonda; Olson, Eric N

    2014-08-01

    Regeneration of injured adult skeletal muscle involves fusion of activated satellite cells to form new myofibers. Myomaker is a muscle-specific membrane protein required for fusion of embryonic myoblasts, but its potential involvement in adult muscle regeneration has not been explored. We show that myogenic basic helix-loop-helix (bHLH) transcription factors induce myomaker expression in satellite cells during acute and chronic muscle regeneration. Moreover, genetic deletion of myomaker in adult satellite cells completely abolishes muscle regeneration, resulting in severe muscle destruction after injury. Myomaker is the only muscle-specific protein known to be absolutely essential for fusion of embryonic and adult myoblasts.

  7. Myomaker is essential for muscle regeneration

    PubMed Central

    Millay, Douglas P.; Sutherland, Lillian B.; Bassel-Duby, Rhonda

    2014-01-01

    Regeneration of injured adult skeletal muscle involves fusion of activated satellite cells to form new myofibers. Myomaker is a muscle-specific membrane protein required for fusion of embryonic myoblasts, but its potential involvement in adult muscle regeneration has not been explored. We show that myogenic basic helix–loop–helix (bHLH) transcription factors induce myomaker expression in satellite cells during acute and chronic muscle regeneration. Moreover, genetic deletion of myomaker in adult satellite cells completely abolishes muscle regeneration, resulting in severe muscle destruction after injury. Myomaker is the only muscle-specific protein known to be absolutely essential for fusion of embryonic and adult myoblasts. PMID:25085416

  8. Mature adult dystrophic mouse muscle environment does not impede efficient engrafted satellite cell regeneration and self-renewal.

    PubMed

    Boldrin, Luisa; Zammit, Peter Steven; Muntoni, Francesco; Morgan, Jennifer Elizabeth

    2009-10-01

    Changes that occur in the skeletal muscle environment with the progress of muscular dystrophies may affect stem cell function and result in impaired muscle regeneration. It has previously been suggested that the success of stem cell transplantation could therefore be dependent both on the properties of the cell itself and on the host muscle environment. Here we engrafted young and mature adult mdx-nude mice, which are the genetic homolog of Duchenne muscular dystrophy, with a small number of satellite cells freshly isolated from young, normal donor mice. We found that the donor satellite cells contributed to muscle regeneration and self-renewal as efficiently within mature adult, as in young, dystrophic host muscle. Donor-derived satellite cells also contributed to robust regeneration after further injury, showing that they were functional despite the more advanced dystrophic muscle environment. These findings provide evidence that muscle tissue in a later stage of dystrophy may be effectively treated by stem cells.

  9. A 3-D cardiac muscle construct for exploring adult marrow stem cell based myocardial regeneration.

    PubMed

    Valarmathi, Mani T; Goodwin, Richard L; Fuseler, John W; Davis, Jeffrey M; Yost, Michael J; Potts, Jay D

    2010-04-01

    Adult bone marrow stromal cells (BMSCs) are capable of differentiating into cardiomyocyte-like cells in vitro and contribute to myocardial regeneration in vivo. Consequently, BMSCs may potentially play a vital role in cardiac repair and regeneration. However, this concept has been limited by inadequate and inconsistent differentiation of BMSCs into cardiomyocytes along with poor survival and integration of neo-cardiomyocytes after implantation into ischemic myocardium. In order to overcome these barriers and to explore adult stem cell based myocardial regeneration, we have developed an in vitro model of three-dimensional (3-D) cardiac muscle using rat ventricular embryonic cardiomyocytes (ECMs) and BMSCs. When ECMs and BMSCs were seeded sequentially onto a 3-D tubular scaffold engineered from topographically aligned type I collagen-fibers and cultured in basal medium for 7, 14, 21, or 28 days, the maturation and co-differentiation into a cardiomyocyte lineage was observed. Phenotypic induction was characterized at morphological, immunological, biochemical and molecular levels. The observed expression of transcripts coding for cardiomyocyte phenotypic markers and the immunolocalization of cardiomyogenic lineage-associated proteins revealed typical expression patterns of neo-cardiomyogenesis. At the biochemical level differentiating cells exhibited appropriate metabolic activity and at the ultrastructural level myofibrillar and sarcomeric organization were indicative of an immature phenotype. Our 3-D co-culture system sustains the ECMs in vitro continuum of differentiation process and simultaneously induces the maturation and differentiation of BMSCs into cardiomyocyte-like cells. Thus, this novel 3-D co-culture system provides a useful in vitro model to investigate the functional role and interplay of developing ECMs and BMSCs during cardiomyogenic differentiation.

  10. Active muscle regeneration following eccentric contraction-induced injury is similar between healthy young and older adults

    PubMed Central

    MacNeil, R. Gavin; Clough, Launa G.; Dirain, Marvin; Sandesara, Bhanuprasad; Pahor, Marco; Manini, Todd M.; Leeuwenburgh, Christiaan

    2013-01-01

    Repair of skeletal muscle after injury is a key aspect of maintaining proper musculoskeletal function. Studies have suggested that regenerative processes, including myogenesis and angiogenesis, are impaired during advanced age, but evidence from humans is limited. This study aimed to compare active muscle regeneration between healthy young and older adults. We evaluated changes in clinical, biochemical, and immunohistochemical indices of muscle regeneration at precisely 2 (T2) and 7 (T3) days following acute muscle injury. Men and women, aged 18-30 and ≥70 years, matched for gender and body mass index, performed 150 unilateral, eccentric contractions of the plantar flexors at 110% of one repetition maximum. Data were analyzed using analysis of covariance, adjusted for gender, habitual physical activity, and baseline level of the outcome. A total of 30 young (n = 15; 22.5 ± 3.7 yr) and older (n = 15; 75.8 ± 5.0 yr) adults completed the study. Following muscle injury, force production declined 16% and 14% in young and older adults, respectively, by T2 and in each group, returned to 93% of baseline strength by T3. Despite modest differences in the pattern of response, postinjury changes in intramuscular concentrations of myogenic growth factors and number of myonuclear (4′,6-diamidino-2-phenylindole+ and paired box 7+) cells were largely similar between groups. Likewise, postinjury changes in serum and intramuscular indices of inflammation (e.g., TNF-α and monocyte chemoattractant protein-1) and angiogenesis (e.g., VEGF and kinase insert domain receptor) did not differ significantly between groups. These findings suggest that declines in physical activity and increased co-morbidity may contribute to age-related impairments in active muscle regeneration rather than aging per se. PMID:23493365

  11. Active muscle regeneration following eccentric contraction-induced injury is similar between healthy young and older adults.

    PubMed

    Buford, Thomas W; MacNeil, R Gavin; Clough, Launa G; Dirain, Marvin; Sandesara, Bhanuprasad; Pahor, Marco; Manini, Todd M; Leeuwenburgh, Christiaan

    2014-06-01

    Repair of skeletal muscle after injury is a key aspect of maintaining proper musculoskeletal function. Studies have suggested that regenerative processes, including myogenesis and angiogenesis, are impaired during advanced age, but evidence from humans is limited. This study aimed to compare active muscle regeneration between healthy young and older adults. We evaluated changes in clinical, biochemical, and immunohistochemical indices of muscle regeneration at precisely 2 (T2) and 7 (T3) days following acute muscle injury. Men and women, aged 18-30 and ≥70 years, matched for gender and body mass index, performed 150 unilateral, eccentric contractions of the plantar flexors at 110% of one repetition maximum. Data were analyzed using analysis of covariance, adjusted for gender, habitual physical activity, and baseline level of the outcome. A total of 30 young (n = 15; 22.5 ± 3.7 yr) and older (n = 15; 75.8 ± 5.0 yr) adults completed the study. Following muscle injury, force production declined 16% and 14% in young and older adults, respectively, by T2 and in each group, returned to 93% of baseline strength by T3. Despite modest differences in the pattern of response, postinjury changes in intramuscular concentrations of myogenic growth factors and number of myonuclear (4',6-diamidino-2-phenylindole+ and paired box 7+) cells were largely similar between groups. Likewise, postinjury changes in serum and intramuscular indices of inflammation (e.g., TNF-α and monocyte chemoattractant protein-1) and angiogenesis (e.g., VEGF and kinase insert domain receptor) did not differ significantly between groups. These findings suggest that declines in physical activity and increased co-morbidity may contribute to age-related impairments in active muscle regeneration rather than aging per se.

  12. MicroRNA-29a in Adult Muscle Stem Cells Controls Skeletal Muscle Regeneration During Injury and Exercise Downstream of Fibroblast Growth Factor-2.

    PubMed

    Galimov, Artur; Merry, Troy L; Luca, Edlira; Rushing, Elisabeth J; Mizbani, Amir; Turcekova, Katarina; Hartung, Angelika; Croce, Carlo M; Ristow, Michael; Krützfeldt, Jan

    2016-03-01

    The expansion of myogenic progenitors (MPs) in the adult muscle stem cell niche is critical for the regeneration of skeletal muscle. Activation of quiescent MPs depends on the dismantling of the basement membrane and increased access to growth factors such as fibroblast growth factor-2 (FGF2). Here, we demonstrate using microRNA (miRNA) profiling in mouse and human myoblasts that the capacity of FGF2 to stimulate myoblast proliferation is mediated by miR-29a. FGF2 induces miR-29a expression and inhibition of miR-29a using pharmacological or genetic deletion decreases myoblast proliferation. Next generation RNA sequencing from miR-29a knockout myoblasts (Pax7(CE/+) ; miR-29a(flox/flox) ) identified members of the basement membrane as the most abundant miR-29a targets. Using gain- and loss-of-function experiments, we confirm that miR-29a coordinately regulates Fbn1, Lamc1, Nid2, Col4a1, Hspg2 and Sparc in myoblasts in vitro and in MPs in vivo. Induction of FGF2 and miR-29a and downregulation of its target genes precedes muscle regeneration during cardiotoxin (CTX)-induced muscle injury. Importantly, MP-specific tamoxifen-induced deletion of miR-29a in adult skeletal muscle decreased the proliferation and formation of newly formed myofibers during both CTX-induced muscle injury and after a single bout of eccentric exercise. Our results identify a novel miRNA-based checkpoint of the basement membrane in the adult muscle stem cell niche. Strategies targeting miR-29a might provide useful clinical approaches to maintain muscle mass in disease states such as ageing that involve aberrant FGF2 signaling.

  13. Redox Control of Skeletal Muscle Regeneration.

    PubMed

    Le Moal, Emmeran; Pialoux, Vincent; Juban, Gaëtan; Groussard, Carole; Zouhal, Hassane; Chazaud, Bénédicte; Mounier, Rémi

    2017-02-06

    Skeletal muscle shows high plasticity in response to external demand. Moreover, adult skeletal muscle is capable of complete regeneration after injury, due to the properties of muscle stem cells (MuSCs), the satellite cells, which follow a tightly regulated myogenic program to generate both new myofibers and new MuSCs for further needs. Although reactive oxygen species (ROS) and reactive nitrogen species (RNS) have long been associated with skeletal muscle physiology, their implication in the cell and molecular processes at work during muscle regeneration is more recent. This review focuses on redox regulation during skeletal muscle regeneration. An overview of the basics of ROS/RNS and antioxidant chemistry and biology occurring in skeletal muscle is first provided. Then, the comprehensive knowledge on redox regulation of MuSCs and their surrounding cell partners (macrophages, endothelial cells) during skeletal muscle regeneration is presented in normal muscle and in specific physiological (exercise-induced muscle damage, aging) and pathological (muscular dystrophies) contexts. Recent advances in the comprehension of these processes has led to the development of therapeutic assays using antioxidant supplementation, which result in inconsistent efficiency, underlying the need for new tools that are aimed at precisely deciphering and targeting ROS networks. This review should provide an overall insight of the redox regulation of skeletal muscle regeneration while highlighting the limits of the use of nonspecific antioxidants to improve muscle function. Antioxid. Redox Signal. 00, 000-000.

  14. MicroRNA deep sequencing in two adult stem cell populations identifies miR-501 as a novel regulator of myosin heavy chain during muscle regeneration

    PubMed Central

    Mizbani, Amir; Luca, Edlira; Rushing, Elisabeth J.

    2016-01-01

    MicroRNAs (miRNAs) are important regulators of skeletal muscle regeneration, but the underlying mechanisms are still incompletely understood. Here, comparative miRNA sequencing analysis of myogenic progenitor cells (MPs) and non-myogenic fibroblast-adipocyte progenitors (FAPs) during cardiotoxin (CTX)-induced muscle injury uncovered miR-501 as a novel muscle-specific miRNA. miR-501 is an intronic miRNA and its expression levels in MPs correlated with its host gene, chloride channel, voltage-sensitive 5 (Clcn5). Pharmacological inhibition of miR-501 dramatically blunted the induction of embryonic myosin heavy chain (MYH3) and, to a lesser extent, adult myosin isoforms during muscle regeneration, and promoted small-diameter neofibers. An unbiased target identification approach in primary myoblasts validated gigaxonin as a target of miR-501 that mimicked the effect of miR-501 inhibition on MYH3 expression. In the mdx mouse model, which models a pathological disease state, not only was miR-501 induced in regenerating skeletal muscle, but also its serum levels were increased, which correlated with the disease state of the animals. Our results suggest that miR-501 plays a key role in adult muscle regeneration and might serve as a novel serum biomarker for the activation of adult muscle stem cells. PMID:27707793

  15. Hindlimb suspension reduces muscle regeneration

    NASA Technical Reports Server (NTRS)

    Mozdziak, P. E.; Truong, Q.; Macius, A.; Schultz, E.

    1998-01-01

    Exposure of juvenile skeletal muscle to a weightless environment reduces growth and satellite cell mitotic activity. However, the effect of a weightless environment on the satellite cell population during muscle repair remains unknown. Muscle injury was induced in rat soleus muscles using the myotoxic snake venom, notexin. Rats were placed into hindlimb-suspended or weightbearing groups for 10 days following injury. Cellular proliferation during regeneration was evaluated using 5-bromo-2'-deoxyuridine (BrdU) immunohistochemistry and image analysis. Hindlimb suspension reduced (P < 0.05) regenerated muscle mass, regenerated myofiber diameter, uninjured muscle mass, and uninjured myofiber diameter compared to weightbearing rats. Hindlimb suspension reduced (P < 0.05) BrdU labeling in uninjured soleus muscles compared to weight-bearing muscles. However, hindlimb suspension did not abolish muscle regeneration because myofibers formed in the injured soleus muscles of hindlimb-suspended rats, and BrdU labeling was equivalent (P > 0.10) on myofiber segments isolated from the soleus muscles of hindlimb-suspended and weightbearing rats following injury. Thus, hindlimb suspension (weightlessness) does not suppress satellite cell mitotic activity in regenerating muscles before myofiber formation, but reduces growth of the newly formed myofibers.

  16. Adult rat motor neurons do not re-establish electrical coupling during axonal regeneration and muscle reinnervation.

    PubMed

    Favero, Morgana; Cangiano, Alberto; Busetto, Giuseppe

    2015-01-01

    Gap junctions (GJs) between neurons are present in both the newborn and the adult nervous system, and although important roles have been suggested or demonstrated in a number of instances, in many other cases a full understanding of their physiological role is still missing. GJs are expressed in the rodent lumbar cord at birth and mediate both dye and electrical coupling between motor neurons. This expression has been proposed to mediate: (i) fast synchronization of motoneuronal spike activity, in turn linked to the process of refinement of neuromuscular connections, and (ii) slow synchronization of locomotor-like oscillatory activity. Soon after birth this coupling disappears. Since in the adult rat regeneration of motor fibers after peripheral nerve injury leads to a recapitulation of synaptic refinement at the target muscles, we tested whether GJs between motor neurons are transiently re-expressed. We found that in conditions of maximal responsiveness of lumbar motor neurons (such as no depression by anesthetics, decerebrate release of activity of subsets of motor neurons, use of temporal and spatial summation by antidromic and orthodromic stimulations, testing of large ensembles of motor neurons) no firing is observed in ventral root axons in response to antidromic spike invasion of nearby counterparts. We conclude that junctional coupling between motor neurons is not required for the refinement of neuromuscular innervation in the adult.

  17. Spontaneous calcium transients manifest in the regenerating muscle and are necessary for skeletal muscle replenishment.

    PubMed

    Tu, Michelle Kim; Borodinsky, Laura Noemi

    2014-07-01

    Tissue regeneration entails replenishing of damaged cells, appropriate cell differentiation and inclusion of regenerated cells into functioning tissues. In adult humans, the capacity of the injured spinal cord and muscle to self-repair is limited. In contrast, the amphibian larva can regenerate its tail after amputation with complete recovery of muscle, notochord and spinal cord. The cellular and molecular mechanisms underlying this phenomenon are still unclear. Here we show that upon injury muscle cell precursors exhibit Ca(2+) transients that depend on Ca(2+) release from ryanodine receptor-operated stores. Blockade of these transients impairs muscle regeneration. Furthermore, inhibiting Ca(2+) transients in the regenerating tail prevents the activation and proliferation of muscle satellite cells, which results in deficient muscle replenishment. These findings suggest that Ca(2+)-mediated activity is critical for the early stages of muscle regeneration, which may lead to developing effective therapies for tissue repair.

  18. Skeletal muscle degeneration and regeneration in mice and flies.

    PubMed

    Rai, Mamta; Nongthomba, Upendra; Grounds, Miranda D

    2014-01-01

    Many aspects of skeletal muscle biology are remarkably similar between mammals and tiny insects, and experimental models of mice and flies (Drosophila) provide powerful tools to understand factors controlling the growth, maintenance, degeneration (atrophy and necrosis), and regeneration of normal and diseased muscles, with potential applications to the human condition. This review compares the limb muscles of mice and the indirect flight muscles of flies, with respect to the mechanisms of adult myofiber formation, homeostasis, atrophy, hypertrophy, and the response to muscle degeneration, with some comment on myogenic precursor cells and common gene regulatory pathways. There is a striking similarity between the species for events related to muscle atrophy and hypertrophy, without contribution of any myoblast fusion. Since the flight muscles of adult flies lack a population of reserve myogenic cells (equivalent to satellite cells), this indicates that such cells are not required for maintenance of normal muscle function. However, since satellite cells are essential in postnatal mammals for myogenesis and regeneration in response to myofiber necrosis, the extent to which such regeneration might be possible in flight muscles of adult flies remains unclear. Common cellular and molecular pathways for both species are outlined related to neuromuscular disorders and to age-related loss of skeletal muscle mass and function (sarcopenia). The commonality of events related to skeletal muscles in these disparate species (with vast differences in size, growth duration, longevity, and muscle activities) emphasizes the combined value and power of these experimental animal models.

  19. Induction of Acute Skeletal Muscle Regeneration by Cardiotoxin Injection.

    PubMed

    Guardiola, Ombretta; Andolfi, Gennaro; Tirone, Mario; Iavarone, Francescopaolo; Brunelli, Silvia; Minchiotti, Gabriella

    2017-01-01

    Skeletal muscle regeneration is a physiological process that occurs in adult skeletal muscles in response to injury or disease. Acute injury-induced skeletal muscle regeneration is a widely used, powerful model system to study the events involved in muscle regeneration as well as the mechanisms and different players. Indeed, a detailed knowledge of this process is essential for a better understanding of the pathological conditions that lead to skeletal muscle degeneration, and it aids in identifying new targeted therapeutic strategies. The present work describes a detailed and reproducible protocol to induce acute skeletal muscle regeneration in mice through a single intramuscular injection of cardiotoxin (CTX). CTX belongs to the family of snake venom toxins and causes myolysis of myofibers, which eventually triggers the regeneration events. The dynamics of skeletal muscle regeneration is evaluated by histological analysis of muscle sections. The protocol also illustrates the experimental procedures for dissecting, freezing, and cutting the Tibialis Anterior muscle, as well as the routine Hematoxylin & Eosin staining that is widely used for subsequent morphological and morphometric analysis.

  20. Muscle repair and regeneration: stem cells, scaffolds, and the contributions of skeletal muscle to amphibian limb regeneration.

    PubMed

    Milner, Derek J; Cameron, Jo Ann

    2013-01-01

    Skeletal muscle possesses a robust innate capability for repair of tissue damage. Natural repair of muscle damage is a stepwise process that requires the coordinated activity of a number of cell types, including infiltrating macrophages, resident myogenic and non-myogenic stem cells, and connective tissue fibroblasts. Despite the proficiency of this intrinsic repair capability, severe injuries that result in significant loss of muscle tissue overwhelm the innate repair process and require intervention if muscle function is to be restored. Recent advances in stem cell biology, regenerative medicine, and materials science have led to attempts at developing tissue engineering-based methods for repairing severe muscle defects. Muscle tissue also plays a role in the ability of tailed amphibians to regenerate amputated limbs through epimorphic regeneration. Muscle contributes adult stem cells to the amphibian regeneration blastema, but it can also contribute blastemal cells through the dedifferentiation of multinucleate myofibers into mononuclear precursors. This fascinating plasticity and its contributions to limb regeneration have prompted researchers to investigate the potential for mammalian muscle to undergo dedifferentiation. Several works have shown that mammalian myotubes can be fragmented into mononuclear cells and induced to re-enter the cell cycle, but mature myofibers are resistant to fragmentation. However, recent works suggest that there may be a path to inducing fragmentation of mature myofibers into proliferative multipotent cells with the potential for use in muscle tissue engineering and regenerative therapies.

  1. Comparative study of muscle regeneration following cardiotoxin and glycerol injury.

    PubMed

    Mahdy, Mohamed A A; Lei, Hsiao Yin; Wakamatsu, Jun-Ichi; Hosaka, Yoshinao Z; Nishimura, Takanori

    2015-11-01

    In the present study, we examined muscle regeneration following two types of chemical injuries, cardiotoxin (CTX) and glycerol, in order to compare their effect on the morphological characteristics during muscle regeneration, in addition we studied the structural changes of the intramuscular connective tissue (IMCT) during the regeneration process, by scanning electron microscopy (SEM) after digestion of the cellular elements of the muscle with sodium hydroxide. Tibialis anterior (TA) muscles of adult male mice were injected either with CTX or glycerol. Muscle degeneration was greater in the CTX-injured model than in the glycerol-injured model at day 4 post injection. Muscle regeneration started at day 7 in both the CTX and glycerol models. However, the CTX-injured model showed a higher myotube density and larger myotube diameter than the glycerol-injured model at days 10 and 14 post injection. On other hand, adipocyte infiltration was detected in the glycerol-injured model. In contrast, no adipocytes could be detected in the CTX-injured model. Furthermore, ultrastructural analysis showed a significant difference in myofiber damage and regeneration between the two models. SEM of the IMCT showed a transient increase in endomysial collagen deposition at early stages of regeneration in the CTX-injured model. In contrast, glycerol-injured model showed slight endomysial collagen deposition. Our results suggest that changes in IMCT affect the efficiency of muscle regeneration. Studying the three dimensional structure of IMCT may help clinical therapies to reduce skeletal muscle fibrosis. To our knowledge this is the first time the changes in IMCT following CTX and glycerol injury using SEM-cell maceration technique have been compared.

  2. Muscle cells provide instructions for planarian regeneration.

    PubMed

    Witchley, Jessica N; Mayer, Mirjam; Wagner, Daniel E; Owen, Jared H; Reddien, Peter W

    2013-08-29

    Regeneration requires both potential and instructions for tissue replacement. In planarians, pluripotent stem cells have the potential to produce all new tissue. The identities of the cells that provide regeneration instructions are unknown. Here, we report that position control genes (PCGs) that control regeneration and tissue turnover are expressed in a subepidermal layer of nonneoblast cells. These subepidermal cells coexpress many PCGs. We propose that these subepidermal cells provide a system of body coordinates and positional information for regeneration, and identify them to be muscle cells of the planarian body wall. Almost all planarian muscle cells express PCGs, suggesting a dual function: contraction and control of patterning. PCG expression is dynamic in muscle cells after injury, even in the absence of neoblasts, suggesting that muscle is instructive for regeneration. We conclude that planarian regeneration involves two highly flexible systems: pluripotent neoblasts that can generate any new cell type and muscle cells that provide positional instructions for the regeneration of any body region.

  3. Macrophage depletion impairs skeletal muscle regeneration: The roles of regulatory factors for muscle regeneration.

    PubMed

    Liu, Xiaoguang; Liu, Yu; Zhao, Linlin; Zeng, Zhigang; Xiao, Weihua; Chen, Peijie

    2017-03-01

    Though macrophages are essential for skeletal muscle regeneration, which is a complex process, the roles and mechanisms of the macrophages in the process of muscle regeneration are still not fully understood. The objective of this study is to explore the roles of macrophages and the mechanisms involved in the regeneration of injured skeletal muscle. One hundred and twelve C57BL/6 mice were randomly divided into muscle contusion and macrophages depleted groups. Their gastrocnemius muscles were harvested at the time points of 12 h, 1, 3, 5, 7, 14 d post-injury. The changes in skeletal muscle morphology were assessed by hematoxylin and eosin (HE) stain. The gene expression was analyzed by real-time polymerase chain reaction. The data showed that CL-liposomes treatment did affect the expression of myogenic regulatory factors (MyoD, myogenin) after injury. In addition, CL-liposomes treatment decreased the expression of regulatory factors of muscle regeneration (HGF, uPA, COX-2, IGF-1, MGF, FGF6) and increased the expression of inflammatory cytokines (TGF-β1, TNF-α, IL-1β, RANTES) in the late stage of regeneration. Moreover, there were significant correlations between macrophages and some regulatory factors (such as HGF, uPA) for muscle regeneration. These results suggested that macrophages depletion impairs skeletal muscle regeneration and that the regulatory factors for muscle regeneration may play important roles in this process.

  4. Transplantation and regeneration of striated muscle.

    PubMed Central

    Allbrook, D.

    1975-01-01

    This lecture explores the factors controlling regeneration and reconstitution of skeletal muscle following vascular and neural injury by giving an account of some experimental work in this field, which is then linked to the problem of the use of whole-muscle transplants in clinical surgery. Images Fig. 2 Fig. 4 Fig. 7 Fig. 8 Fig. 9 PMID:1147539

  5. Muscle regeneration in amphibians and mammals: passing the torch.

    PubMed

    Carlson, Bruce M

    2003-02-01

    Skeletal muscle in both amphibians and mammals possesses a high regenerative capacity. In amphibians, a muscle can regenerate in two distinct ways: as a tissue component of an entire regenerating limb (epimorphic regeneration) or as an isolated entity (tissue regeneration). In the absence of epimorphic regenerative ability, mammals can regenerate muscles only by the tissue mode. This review focuses principally on the regeneration of entire muscles and covers what is known and what remains to be elucidated about fundamental mechanisms underlying muscle regeneration at this level.

  6. Brain and muscle Arnt-like 1 promotes skeletal muscle regeneration through satellite cell expansion

    SciTech Connect

    Chatterjee, Somik; Yin, Hongshan; Nam, Deokhwa; Li, Yong; Ma, Ke

    2015-02-01

    Circadian clock is an evolutionarily conserved timing mechanism governing diverse biological processes and the skeletal muscle possesses intrinsic functional clocks. Interestingly, although the essential clock transcription activator, Brain and muscle Arnt-like 1 (Bmal1), participates in maintenance of muscle mass, little is known regarding its role in muscle growth and repair. In this report, we investigate the in vivo function of Bmal1 in skeletal muscle regeneration using two muscle injury models. Bmal1 is highly up-regulated by cardiotoxin injury, and its genetic ablation significantly impairs regeneration with markedly suppressed new myofiber formation and attenuated myogenic induction. A similarly defective regenerative response is observed in Bmal1-null mice as compared to wild-type controls upon freeze injury. Lack of satellite cell expansion accounts for the regeneration defect, as Bmal1{sup −/−} mice display significantly lower satellite cell number with nearly abolished induction of the satellite cell marker, Pax7. Furthermore, satellite cell-derived primary myoblasts devoid of Bmal1 display reduced growth and proliferation ex vivo. Collectively, our results demonstrate, for the first time, that Bmal1 is an integral component of the pro-myogenic response that is required for muscle repair. This mechanism may underlie its role in preserving adult muscle mass and could be targeted therapeutically to prevent muscle-wasting diseases. - Highlights: • Bmal1 is highly inducible by muscle injury and myogenic stimuli. • Genetic ablation of Bmal1 significantly impairs muscle regeneration. • Bmal1 promotes satellite cell expansion during muscle regeneration. • Bmal1-deficient primary myoblasts display attenuated growth and proliferation.

  7. A Rat Model for Muscle Regeneration in the Soft Palate

    PubMed Central

    Carvajal Monroy, Paola L.; Grefte, Sander; Kuijpers-Jagtman, Anne M.; Helmich, Maria P. A. C.; Ulrich, Dietmar J. O.; Von den Hoff, Johannes W.; Wagener, Frank A. D. T. G.

    2013-01-01

    Background Children with a cleft in the soft palate have difficulties with speech, swallowing, and sucking. Despite successful surgical repositioning of the muscles, optimal function is often not achieved. Scar formation and defective regeneration may hamper the functional recovery of the muscles after cleft palate repair. Therefore, the aim of this study is to investigate the anatomy and histology of the soft palate in rats, and to establish an in vivo model for muscle regeneration after surgical injury. Methods Fourteen adult male Sprague Dawley rats were divided into four groups. Groups 1 (n = 4) and 2 (n = 2) were used to investigate the anatomy and histology of the soft palate, respectively. Group 3 (n = 6) was used for surgical wounding of the soft palate, and group 4 (n = 2) was used as unwounded control group. The wounds (1 mm) were evaluated by (immuno)histochemistry (AZAN staining, Pax7, MyoD, MyoG, MyHC, and ASMA) after 7 days. Results The present study shows that the anatomy and histology of the soft palate muscles of the rat is largely comparable with that in humans. All wounds showed clinical evidence of healing after 7 days. AZAN staining demonstrated extensive collagen deposition in the wound area, and initial regeneration of muscle fibers and salivary glands. Proliferating and differentiating satellite cells were identified in the wound area by antibody staining. Conclusions This model is the first, suitable for studying muscle regeneration in the rat soft palate, and allows the development of novel adjuvant strategies to promote muscle regeneration after cleft palate surgery. PMID:23554995

  8. Regulatory T cells and skeletal muscle regeneration.

    PubMed

    Schiaffino, Stefano; Pereira, Marcelo G; Ciciliot, Stefano; Rovere-Querini, Patrizia

    2017-02-01

    Skeletal muscle regeneration results from the activation and differentiation of myogenic stem cells, called satellite cells, located beneath the basal lamina of the muscle fibers. Inflammatory and immune cells have a crucial role in the regeneration process. Acute muscle injury causes an immediate transient wave of neutrophils followed by a more persistent infiltration of M1 (proinflammatory) and M2 (anti-inflammatory/proregenerative) macrophages. New studies show that injured muscle is also infiltrated by a specialized population of regulatory T (Treg) cells, which control both the inflammatory response, by promoting the M1-to-M2 switch, and the activation of satellite cells. Treg cells accumulate in injured muscle in response to specific cytokines, such as IL-33, and promote muscle growth by releasing growth factors, such as amphiregulin. Muscle repair during aging is impaired due to reduced number of Treg cells and can be enhanced by IL-33 supplementation. Migration of Treg cells could also contribute to explain the effect of heterochronic parabiosis, whereby muscle regeneration of aged mice can be improved by a parabiotically linked young partners. In mdx dystrophin-deficient mice, a model of human Duchenne muscular dystrophy, muscle injury, and inflammation is mitigated by expansion of the Treg-cell population but exacerbated by Treg-cell depletion. These findings support the notion that immunological mechanisms are not only essential in the response to pathogenic microbes and tumor cells but also have a wider homeostatic role in tissue repair, and open new perspectives for boosting muscle growth in chronic muscle disease and during aging.

  9. Biologic-free mechanically induced muscle regeneration

    PubMed Central

    Cezar, Christine A.; Roche, Ellen T.; Vandenburgh, Herman H.; Duda, Georg N.; Walsh, Conor J.; Mooney, David J.

    2016-01-01

    Severe skeletal muscle injuries are common and can lead to extensive fibrosis, scarring, and loss of function. Clinically, no therapeutic intervention exists that allows for a full functional restoration. As a result, both drug and cellular therapies are being widely investigated for treatment of muscle injury. Because muscle is known to respond to mechanical loading, we investigated instead whether a material system capable of massage-like compressions could promote regeneration. Magnetic actuation of biphasic ferrogel scaffolds implanted at the site of muscle injury resulted in uniform cyclic compressions that led to reduced fibrous capsule formation around the implant, as well as reduced fibrosis and inflammation in the injured muscle. In contrast, no significant effect of ferrogel actuation on muscle vascularization or perfusion was found. Strikingly, ferrogel-driven mechanical compressions led to enhanced muscle regeneration and a ∼threefold increase in maximum contractile force of the treated muscle at 2 wk compared with no-treatment controls. Although this study focuses on the repair of severely injured skeletal muscle, magnetically stimulated bioagent-free ferrogels may find broad utility in the field of regenerative medicine. PMID:26811474

  10. Palmdelphin promotes myoblast differentiation and muscle regeneration

    PubMed Central

    Nie, Yaping; Chen, Hu; Guo, Cilin; Yuan, Zhuning; Zhou, Xingyu; Zhang, Ying; Zhang, Xumeng; Mo, Delin; Chen, Yaosheng

    2017-01-01

    Differentiation of myoblasts is essential in the development and regeneration of skeletal muscles to form multinucleated, contractile muscle fibers. However, the process of myoblast differentiation in mammals is complicated and requires to be further investigated. In this study, we found Palmdelphin (Palmd), a cytosolic protein, promotes myoblast differentiation. Palmd is predominantly expressed in the cytosol of myoblasts and is gradually up-regulated after differentiation. Knockdown of Palmd by small interfering RNA (siRNA) in C2C12 markedly inhibits myogenic differentiation, suggesting a specific role of Palmd in the morphological changes of myoblast differentiation program. Overexpression of Palmd in C2C12 enhances myogenic differentiation. Remarkably, inhibition of Palmd results in impaired myotube formation during muscle regeneration after injury. These findings reveal a new cytosolic protein that promotes mammalian myoblast differentiation and provide new insights into the molecular regulation of muscle formation. PMID:28148961

  11. Cripto regulates skeletal muscle regeneration and modulates satellite cell determination by antagonizing myostatin.

    PubMed

    Guardiola, Ombretta; Lafuste, Peggy; Brunelli, Silvia; Iaconis, Salvatore; Touvier, Thierry; Mourikis, Philippos; De Bock, Katrien; Lonardo, Enza; Andolfi, Gennaro; Bouché, Ann; Liguori, Giovanna L; Shen, Michael M; Tajbakhsh, Shahragim; Cossu, Giulio; Carmeliet, Peter; Minchiotti, Gabriella

    2012-11-20

    Skeletal muscle regeneration mainly depends on satellite cells, a population of resident muscle stem cells. However, our understanding of the molecular mechanisms underlying satellite cell activation is still largely undefined. Here, we show that Cripto, a regulator of early embryogenesis, is a novel regulator of muscle regeneration and satellite cell progression toward the myogenic lineage. Conditional inactivation of cripto in adult satellite cells compromises skeletal muscle regeneration, whereas gain of function of Cripto accelerates regeneration, leading to muscle hypertrophy. Moreover, we provide evidence that Cripto modulates myogenic cell determination and promotes proliferation by antagonizing the TGF-β ligand myostatin. Our data provide unique insights into the molecular and cellular basis of Cripto activity in skeletal muscle regeneration and raise previously undescribed implications for stem cell biology and regenerative medicine.

  12. How sex hormones promote skeletal muscle regeneration.

    PubMed

    Velders, Martina; Diel, Patrick

    2013-11-01

    Skeletal muscle regeneration efficiency declines with age for both men and women. This decline impacts on functional capabilities in the elderly and limits their ability to engage in regular physical activity and to maintain independence. Aging is associated with a decline in sex hormone production. Therefore, elucidating the effects of sex hormone substitution on skeletal muscle homeostasis and regeneration after injury or disuse is highly relevant for the aging population, where sarcopenia affects more than 30 % of individuals over 60 years of age. While the anabolic effects of androgens are well known, the effects of estrogens on skeletal muscle anabolism have only been uncovered in recent times. Hence, the purpose of this review is to provide a mechanistic insight into the regulation of skeletal muscle regenerative processes by both androgens and estrogens. Animal studies using estrogen receptor (ER) antagonists and receptor subtype selective agonists have revealed that estrogens act through both genomic and non-genomic pathways to reduce leukocyte invasion and increase satellite cell numbers in regenerating skeletal muscle tissue. Although animal studies have been more conclusive than human studies in establishing a role for sex hormones in the attenuation of muscle damage, data from a number of recent well controlled human studies is presented to support the notion that hormonal therapies and exercise induce added positive effects on functional measures and lean tissue mass. Based on the fact that aging human skeletal muscle retains the ability to adapt to exercise with enhanced satellite cell activation, combining sex hormone therapies with exercise may induce additive effects on satellite cell accretion. There is evidence to suggest that there is a 'window of opportunity' after the onset of a hypogonadal state such as menopause, to initiate a hormonal therapy in order to achieve maximal benefits for skeletal muscle health. Novel receptor subtype selective

  13. Expression of nestin, desmin and vimentin in intact and regenerating muscle spindles of rat hind limb skeletal muscles.

    PubMed

    Cízková, Dana; Soukup, Tomás; Mokrý, Jaroslav

    2009-02-01

    We describe the expression and distribution patterns of nestin, desmin and vimentin in intact and regenerating muscle spindles of the rat hind limb skeletal muscles. Regeneration was induced by intramuscular isotransplantation of extensor digitorum longus (EDL) or soleus muscles from 15-day-old rats into the EDL muscle of adult female inbred Lewis rats. The host muscles with grafts were excised after 7-, 16-, 21- and 29-day survival and immunohistochemically stained. Nestin expression in intact spindles in host muscles was restricted to Schwann cells of sensory and motor nerves. In transplanted muscles, however, nestin expression was also found in regenerating "spindle fibers", 7 and 16 days after grafting. From the 21st day onwards, the regenerated spindle fibers were devoid of nestin immunoreactivity. Desmin was detected in spindle fibers at all developmental stages in regenerating as well as in intact spindles. Vimentin was expressed in cells of the outer and inner capsules of all muscle spindles and in newly formed myoblasts and myotubes of regenerating spindles 7 days after grafting. Our results show that the expression pattern of these intermediate filaments in regenerating spindle fibers corresponds to that found in regenerating extrafusal fibers, which supports our earlier suggestion that they resemble small-diameter extrafusal fibers.

  14. Skeletal muscle dedifferentiation during salamander limb regeneration.

    PubMed

    Wang, Heng; Simon, András

    2016-10-01

    Salamanders can regenerate entire limbs throughout their life. A critical step during limb regeneration is formation of a blastema, which gives rise to the new extremity. Salamander limb regeneration has historically been tightly linked to the term dedifferentiation, however, with refined research tools it is important to revisit the definition of dedifferentiation in the context. To what extent do differentiated cells revert their differentiated phenotypes? To what extent do progeny from differentiated cells cross lineage boundaries during regeneration? How do cell cycle plasticity and lineage plasticity relate to each other? What is the relationship between dedifferentiation of specialized cells and activation of tissue resident stem cells in terms of their contribution to the new limb? Here we highlight these problems through the case of skeletal muscle.

  15. [Regeneration capacity of skeletal muscle].

    PubMed

    Wernig, A

    2003-07-01

    The organotypic stem cell of skeletal muscle has previously been known as satellite cell. They allow muscle fiber growth during ontogenesis, enable fiber hypertrophy and are responsible for the very efficient repair of muscle fibers. This efficient apparatus is to some degree counterbalanced by an enormous use of the satellite cell pool: fiber atrophy probably is accompanied by loss of myonuclei such that every reversal of atrophy is bound to use new myonuclei i.e. satellite cells. How often in life does this occur? Hard to say. Moreover, the potent repair capacity is challenged by an unexpected vulnerability of skeletal muscle fibers: Passive stretching of contracted muscles may cause multiple "microdamage," disruption of contractile elements or tiny areas of true necrosis (focal necrosis). How often does this happen? Well, for many of us at least once per year when we go up and down mountains during vacation time, followed by sour muscles. Others may decide to change his/her (locomotor) behaviour by severe onset of jogging; it may happen that they suffer kidney failure on Monday due to muscle microdamage and the transfer of myoproteins into the serum over weekend. Also 20 minutes of stepping up and down something like a chair will do: There is a remarkable increase in kreatin kinase and other muscle derived proteins which lasts for days and is bound to reflect some muscle damage. How about sportsmen and worker who repeatedly use their muscles in such a way? We don't have answers yet to most of these questions, but considerable amount of information has been collected over the last years both in animal and--less--in human. What is common in all cases of growth and repair is the proliferation of the satellite cells and their consequent incorporation and fusion with the parent fiber. This way focal damage is repaired often without visible reminders. We would run out of satellite cells were they not stem cells: After division one daughter remains a satellite cell

  16. Nonmyogenic cells in skeletal muscle regeneration.

    PubMed

    Paylor, Ben; Natarajan, Anuradha; Zhang, Regan-Heng; Rossi, Fabio

    2011-01-01

    Although classical dogma dictates that satellite cells are the primary cell type involved in skeletal muscle regeneration, alternative cell types such as a variety of inflammatory and stromal cells are also actively involved in this process. A model describing myogenic cells as direct contributors to regeneration and nonmyogenic cells from other developmental sources as important accessories has emerged, with similar systems having been described in numerous other tissues in the body. Increasing evidence supports the notion that inflammatory cells function as supportive accessory cells, and are not merely involved in clearing damage following skeletal muscle injury. Additionally, recent studies have highlighted the role of tissue resident mesenchymal cell populations as playing a central role in regulating regeneration. These "accessory" cell populations are proposed to influence myogenesis via direct cell contact and secretion of paracrine trophic factors. The basic foundations of accessory cell understanding should be recognized as a crucial component to all prospects of regenerative medicine, and this chapter intends to provide a comprehensive background on the current literature describing immune and tissue-resident mesenchymal cells' role in skeletal muscle regeneration.

  17. Satellite Cells and Skeletal Muscle Regeneration.

    PubMed

    Dumont, Nicolas A; Bentzinger, C Florian; Sincennes, Marie-Claude; Rudnicki, Michael A

    2015-07-01

    Skeletal muscles are essential for vital functions such as movement, postural support, breathing, and thermogenesis. Muscle tissue is largely composed of long, postmitotic multinucleated fibers. The life-long maintenance of muscle tissue is mediated by satellite cells, lying in close proximity to the muscle fibers. Muscle satellite cells are a heterogeneous population with a small subset of muscle stem cells, termed satellite stem cells. Under homeostatic conditions all satellite cells are poised for activation by stimuli such as physical trauma or growth signals. After activation, satellite stem cells undergo symmetric divisions to expand their number or asymmetric divisions to give rise to cohorts of committed satellite cells and thus progenitors. Myogenic progenitors proliferate, and eventually differentiate through fusion with each other or to damaged fibers to reconstitute fiber integrity and function. In the recent years, research has begun to unravel the intrinsic and extrinsic mechanisms controlling satellite cell behavior. Nonetheless, an understanding of the complex cellular and molecular interactions of satellite cells with their dynamic microenvironment remains a major challenge, especially in pathological conditions. The goal of this review is to comprehensively summarize the current knowledge on satellite cell characteristics, functions, and behavior in muscle regeneration and in pathological conditions.

  18. Rejuvenating stem cells to restore muscle regeneration in aging

    PubMed Central

    Bengal, Eyal; Perdiguero, Eusebio; Serrano, Antonio L.; Muñoz-Cánoves, Pura

    2017-01-01

    Adult muscle stem cells, originally called satellite cells, are essential for muscle repair and regeneration throughout life. Besides a gradual loss of mass and function, muscle aging is characterized by a decline in the repair capacity, which blunts muscle recovery after injury in elderly individuals. A major effort has been dedicated in recent years to deciphering the causes of satellite cell dysfunction in aging animals, with the ultimate goal of rejuvenating old satellite cells and improving muscle function in elderly people. This review focuses on the recently identified network of cell-intrinsic and -extrinsic factors and processes contributing to the decline of satellite cells in old animals. Some studies suggest that aging-related satellite-cell decay is mostly caused by age-associated extrinsic environmental changes that could be reversed by a “youthful environment”. Others propose a central role for cell-intrinsic mechanisms, some of which are not reversed by environmental changes. We believe that these proposals, far from being antagonistic, are complementary and that both extrinsic and intrinsic factors contribute to muscle stem cell dysfunction during aging-related regenerative decline. The low regenerative potential of old satellite cells may reflect the accumulation of deleterious changes during the life of the cell; some of these changes may be inherent (intrinsic) while others result from the systemic and local environment (extrinsic). The present challenge is to rejuvenate aged satellite cells that have undergone reversible changes to provide a possible approach to improving muscle repair in the elderly. PMID:28163911

  19. Regeneration of frog twitch and slow muscle fibers after mincing.

    PubMed

    Schmidt, H; Emser, W

    1985-10-01

    Iliofibularis muscles of Rana temporaria were minced and allowed to regenerate in the iliofibularis or the sartorius bed of the same frog. Regenerated muscles were examined for the presence of slow muscle fibers using electrophysiologic, histochemical, and contractile parameters. Muscle regeneration from sartorius mince was also studied. Regeneration was more successful from iliofibularis than from sartorius mince, and the iliofibularis bed was more favorable for regeneration than the sartorius bed for both types of muscle. Twitch fibers regenerated within a few months, but slow fibers could not be identified earlier than 14 months after muscle destruction. Slow muscle fibers regenerated only from iliofibularis mince, both orthotopically and heterotopically. All regenerates capable of maintaining a K-contracture contained histochemically identified slow fibers; the membrane properties of electrophysiologically identified slow fibers were normal. It is concluded that slow muscle fibers regenerate only from the remnants of a muscle that contains slow fibers. The results are discussed with respect to the role of innervating nerve fibers.

  20. Stem Cells and Tissue Niche: Two Faces of the Same Coin of Muscle Regeneration

    PubMed Central

    Scicchitano, Bianca Maria; Sica, Gigliola; Musarò, Antonio

    2016-01-01

    Capacity of adult muscle to regenerate in response to injury stimuli represents an important homeostatic process. Regeneration is a highly coordinated program that partially recapitulates the embryonic developmental program. However, muscle regeneration is severely compromised in several pathological conditions. It is likely that the restricted tissue repair program under pathological conditions is due to either progressive loss of stem cell populations or to missing signals that limit the damaged tissues to efficiently activate a regenerative program. It is therefore plausible that loss of control over these cell fates might lead to a pathological cell transdifferentiation, limiting the ability of a pathological muscle to sustain an efficient regenerative process. The critical role of microenvironment on stem cells activity and muscle regeneration is discussed. PMID:28078070

  1. Myoanatomy and anterior muscle regeneration of the fireworm Eurythoe cf. complanata (Annelida: Amphinomidae).

    PubMed

    Weidhase, Michael; Bleidorn, Christoph; Beckers, Patrick; Helm, Conrad

    2016-03-01

    Amphinomidae or so-called "fireworms" are known for their inflammatory substances and their regeneration ability. Recent transcriptome-based molecular analyses revealed that these remarkable annelids are a basal branching taxon outside the annelid main radiation (Pleistoannelida). Although several studies dealing with analyses of the morphology of these annelids have been published, detailed investigations of the anterior muscle regeneration and the musculature in general are largely lacking for amphinomids. Using histology, phalloidin labeling together with subsequent confocal laser scanning microscopy (cLSM), and further light microscopic image acquisition of different regeneration stages, we here present the first morphological study describing the myoanatomy and muscular regeneration. During anterior muscular regeneration, longitudinal muscle bundles develop prior to transverse muscle fibers and segment boundaries. Additionally, Eurythoe cf. complanata develops an independent muscular ring surrounding the mouth opening in an early stage of regeneration. Detailed investigation of adult body wall musculature and the parapodial muscle complex in amphinomids show that E. cf. complanata bears well-developed dorsal and ventral longitudinal muscle bundles as well as outer transverse muscles comparable to the pattern described for several Pleistoannelida. Furthermore, the biramous parapodia possess a complex meshwork of distinct muscle fibers allowing detailed comparisons with other annelid families.

  2. Runx1 Transcription Factor Is Required for Myoblasts Proliferation during Muscle Regeneration

    PubMed Central

    Umansky, Kfir Baruch; Gruenbaum-Cohen, Yael; Tsoory, Michael; Feldmesser, Ester; Goldenberg, Dalia; Brenner, Ori; Groner, Yoram

    2015-01-01

    Following myonecrosis, muscle satellite cells proliferate, differentiate and fuse, creating new myofibers. The Runx1 transcription factor is not expressed in naïve developing muscle or in adult muscle tissue. However, it is highly expressed in muscles exposed to myopathic damage yet, the role of Runx1 in muscle regeneration is completely unknown. Our study of Runx1 function in the muscle’s response to myonecrosis reveals that this transcription factor is activated and cooperates with the MyoD and AP-1/c-Jun transcription factors to drive the transcription program of muscle regeneration. Mice lacking dystrophin and muscle Runx1 (mdx - /Runx1 f/f), exhibit impaired muscle regeneration leading to age-dependent muscle waste, gradual decrease in motor capabilities and a shortened lifespan. Runx1-deficient primary myoblasts are arrested at cell cycle G1 and consequently differentiate. Such premature differentiation disrupts the myoblasts’ normal proliferation/differentiation balance, reduces the number and size of regenerating myofibers and impairs muscle regeneration. Our combined Runx1-dependent gene expression, ChIP-seq, ATAC-seq and histone H3K4me1/H3K27ac modification analyses revealed a subset of Runx1-regulated genes that are co-occupied by MyoD and c-Jun in mdx - /Runx1 f/f muscle. The data provide unique insights into the transcriptional program driving muscle regeneration and implicate Runx1 as an important participant in the pathology of muscle wasting diseases. PMID:26275053

  3. Pericytes: multitasking cells in the regeneration of injured, diseased, and aged skeletal muscle.

    PubMed

    Birbrair, Alexander; Zhang, Tan; Wang, Zhong-Min; Messi, Maria L; Mintz, Akiva; Delbono, Osvaldo

    2014-01-01

    Pericytes are perivascular cells that envelop and make intimate connections with adjacent capillary endothelial cells. Recent studies show that they may have a profound impact in skeletal muscle regeneration, innervation, vessel formation, fibrosis, fat accumulation, and ectopic bone formation throughout life. In this review, we summarize and evaluate recent advances in our understanding of pericytes' influence on adult skeletal muscle pathophysiology. We also discuss how further elucidating their biology may offer new approaches to the treatment of conditions characterized by muscle wasting.

  4. Bex1 knock out mice show altered skeletal muscle regeneration

    SciTech Connect

    Koo, Jae Hyung Smiley, Mark A.; Lovering, Richard M.; Margolis, Frank L.

    2007-11-16

    Bex1 and Calmodulin (CaM) are upregulated during skeletal muscle regeneration. We confirm this finding and demonstrate the novel finding that they interact in a calcium-dependent manner. To study the role of Bex1 and its interaction with CaM in skeletal muscle regeneration, we generated Bex1 knock out (Bex1-KO) mice. These mice appeared to develop normally and are fertile, but displayed a functional deficit in exercise performance compared to wild type (WT) mice. After intramuscular injection of cardiotoxin, which causes extensive and reproducible myotrauma followed by recovery, regenerating muscles of Bex1-KO mice exhibited elevated and prolonged cell proliferation, as well as delayed cell differentiation, compared to WT mice. Thus, our results provide the first evidence that Bex1-KO mice show altered muscle regeneration, and allow us to propose that the interaction of Bex1 with Ca{sup 2+}/CaM may be involved in skeletal muscle regeneration.

  5. Thyroid hormones regulate skeletal muscle regeneration after acute injury.

    PubMed

    Leal, Anna Lúcia R C; Albuquerque, João Paulo C; Matos, Marina S; Fortunato, Rodrigo S; Carvalho, Denise P; Rosenthal, Doris; da Costa, Vânia Maria Corrêa

    2015-02-01

    We evaluated the effects of hypo- and hyperthyroid statuses during the initial phase of skeletal muscle regeneration in rats. To induce hypo- or hyperthyroidism, adult male Wistar rats were treated with methimazole (0.03%) or T4 (10 μg/100 g), respectively, for 10 days. Three days before sacrifice, a crush injury was produced in the solear muscles of one half of the animals, while the other half remained intact. T3, T4, TSH, and leptin serum levels were not affected by the injury. Serum T3 and T4 levels were significantly increased in hyperthyroid and hyper-injury animals. Hypothyroidism was confirmed by the significant increase in serum TSH levels in hypothyroid and hypo-injury animals. Injury increased cell infiltration and macrophage accumulation especially in hyperthyroid animals. Both type 2 and type 3 deiodinases were induced by lesion, and the opposite occurred with the type 1 isoform, at least in the control and hyperthyroid groups. Injury increased both MyoD and myogenin expression in all the studied groups, but only MyoD expression was increased by thyroidal status only at the protein level. We conclude that thyroid hormones modulate skeletal muscle regeneration possibly by regulating the inflammatory process, as well as MyoD and myogenin expression in the injured tissue.

  6. Low Intensity Exercise Training Improves Skeletal Muscle Regeneration Potential

    PubMed Central

    Pietrangelo, Tiziana; Di Filippo, Ester S.; Mancinelli, Rosa; Doria, Christian; Rotini, Alessio; Fanò-Illic, Giorgio; Fulle, Stefania

    2015-01-01

    Purpose: The aim of this study was to determine whether 12 days of low-to-moderate exercise training at low altitude (598 m a.s.l.) improves skeletal muscle regeneration in sedentary adult women. Methods: Satellite cells were obtained from the vastus lateralis skeletal muscle of seven women before and after this exercise training at low altitude. They were investigated for differentiation aspects, superoxide anion production, antioxidant enzymes, mitochondrial potential variation after a depolarizing insult, intracellular Ca2+ concentrations, and micro (mi)RNA expression (miR-1, miR-133, miR-206). Results: In these myogenic populations of adult stem cells, those obtained after exercise training, showed increased Fusion Index and intracellular Ca2+ concentrations. This exercise training also generally reduced superoxide anion production in cells (by 12–67%), although not in two women, where there was an increase of ~15% along with a reduced superoxide dismutase activity. miRNA expression showed an exercise-induced epigenetic transcription profile that was specific according to the reduced or increased superoxide anion production of the cells. Conclusions: The present study shows that low-to-moderate exercise training at low altitude improves the regenerative capacity of skeletal muscle in adult women. The differentiation of cells was favored by increased intracellular calcium concentration and increased the fusion index. This low-to-moderate training at low altitude also depicted the epigenetic signature of cells. PMID:26733888

  7. Molecular circuitry of stem cell fate in skeletal muscle regeneration, ageing, and disease

    PubMed Central

    Almada, Albert E.; Wagers, Amy J.

    2016-01-01

    Satellite cells are adult myogenic stem cells that function to repair damaged muscle. The enduring capacity for muscle regeneration requires efficient satellite cell expansion after injury, differentiation to produce myoblasts that can reconstitute damaged fibers, and self-renewal to replenish the muscle stem cell pool for subsequent rounds of injury and repair. Emerging studies indicate that misregulations of satellite cell fate and function contribute to age-associated muscle dysfunction and influence the severity of muscle diseases, including Duchenne Muscular Dystrophy (DMD). It has also become apparent that satellite cell fate during muscle regeneration, aging, and in the context of DMD is governed by an intricate network of intrinsic and extrinsic regulators. Targeted manipulation of this network may offer unique opportunities for muscle regenerative medicine. PMID:26956195

  8. Planarian Body-Wall Muscle: Regeneration and Function beyond a Simple Skeletal Support.

    PubMed

    Cebrià, Francesc

    2016-01-01

    The body-wall musculature of adult planarians consists of intricately organized muscle fibers, which after amputation are regenerated rapidly and with great precision through the proliferation and differentiation of pluripotent stem cells. These traits make the planarian body-wall musculature a potentially useful model for the study of cell proliferation, differentiation, and pattern formation. Planarian body-wall muscle shows some ambiguous features common to both skeletal and smooth muscle cells. However, its skeletal nature is implied by the expression of skeletal myosin heavy-chain genes and the myogenic transcription factor myoD. Where and when planarian stem cells become committed to the myogenic lineage during regeneration, how the new muscle cells are integrated into the pre-existing muscle net, and the identity of the molecular pathway controlling the myogenic gene program are key aspects of planarian muscle regeneration that need to be addressed. Expression of the conserved transcription factor myoD has been recently demonstrated in putative myogenic progenitors. Moreover, recent studies suggest that differentiated muscle cells may provide positional information to planarian stem cells during regeneration. Here, I review the limited available knowledge on planarian muscle regeneration.

  9. Planarian Body-Wall Muscle: Regeneration and Function beyond a Simple Skeletal Support

    PubMed Central

    Cebrià, Francesc

    2016-01-01

    The body-wall musculature of adult planarians consists of intricately organized muscle fibers, which after amputation are regenerated rapidly and with great precision through the proliferation and differentiation of pluripotent stem cells. These traits make the planarian body-wall musculature a potentially useful model for the study of cell proliferation, differentiation, and pattern formation. Planarian body-wall muscle shows some ambiguous features common to both skeletal and smooth muscle cells. However, its skeletal nature is implied by the expression of skeletal myosin heavy-chain genes and the myogenic transcription factor myoD. Where and when planarian stem cells become committed to the myogenic lineage during regeneration, how the new muscle cells are integrated into the pre-existing muscle net, and the identity of the molecular pathway controlling the myogenic gene program are key aspects of planarian muscle regeneration that need to be addressed. Expression of the conserved transcription factor myoD has been recently demonstrated in putative myogenic progenitors. Moreover, recent studies suggest that differentiated muscle cells may provide positional information to planarian stem cells during regeneration. Here, I review the limited available knowledge on planarian muscle regeneration. PMID:26904543

  10. Regeneration of the Pancreas in Adult Zebrafish

    PubMed Central

    Moss, Jennifer B.; Koustubhan, Punita; Greenman, Melanie; Parsons, Michael J.; Walter, Ingrid; Moss, Larry G.

    2009-01-01

    OBJECTIVE Regenerating organs in diverse biological systems have provided clues to processes that can be harnessed to repair damaged tissue. Adult mammalian β-cells have a limited capacity to regenerate, resulting in diabetes and lifelong reliance on insulin. Zebrafish have been used as a model for the regeneration of many organs. We demonstrate the regeneration of adult zebrafish pancreatic β-cells. This nonmammalian model can be used to define pathways for islet-cell regeneration in humans. RESEARCH DESIGN AND METHODS Adult transgenic zebrafish were injected with a single high dose of streptozotocin or metronidazole and anesthetized at 3, 7, or 14 days or pancreatectomized. Blood glucose measurements were determined and gut sections were analyzed using specific endocrine, exocrine, and duct cell markers as well as markers for dividing cells. RESULTS Zebrafish recovered rapidly without the need for insulin injections, and normoglycemia was attained within 2 weeks. Although few proliferating cells were present in vehicles, ablation caused islet destruction and a striking increase of proliferating cells, some of which were Pdx1 positive. Dividing cells were primarily associated with affected islets and ducts but, with the exception of surgical partial pancreatectomy, were not extensively β-cells. CONCLUSIONS The ability of the zebrafish to regenerate a functional pancreas using chemical, genetic, and surgical approaches enabled us to identify patterns of cell proliferation in islets and ducts. Further study of the origin and contribution of proliferating cells in reestablishing islet function could provide strategies for treating human diseases. PMID:19491207

  11. Smad4 restricts differentiation to promote expansion of satellite cell derived progenitors during skeletal muscle regeneration

    PubMed Central

    Paris, Nicole D; Soroka, Andrew; Klose, Alanna; Liu, Wenxuan; Chakkalakal, Joe V

    2016-01-01

    Skeletal muscle regenerative potential declines with age, in part due to deficiencies in resident stem cells (satellite cells, SCs) and derived myogenic progenitors (MPs); however, the factors responsible for this decline remain obscure. TGFβ superfamily signaling is an inhibitor of myogenic differentiation, with elevated activity in aged skeletal muscle. Surprisingly, we find reduced expression of Smad4, the downstream cofactor for canonical TGFβ superfamily signaling, and the target Id1 in aged SCs and MPs during regeneration. Specific deletion of Smad4 in adult mouse SCs led to increased propensity for terminal myogenic commitment connected to impaired proliferative potential. Furthermore, SC-specific Smad4 disruption compromised adult skeletal muscle regeneration. Finally, loss of Smad4 in aged SCs did not promote aged skeletal muscle regeneration. Therefore, SC-specific reduction of Smad4 is a feature of aged regenerating skeletal muscle and Smad4 is a critical regulator of SC and MP amplification during skeletal muscle regeneration. DOI: http://dx.doi.org/10.7554/eLife.19484.001 PMID:27855784

  12. A developmentally regulated switch from stem cells to dedifferentiation for limb muscle regeneration in newts.

    PubMed

    Tanaka, Hibiki Vincent; Ng, Nathaniel Chuen Yin; Yang Yu, Zhan; Casco-Robles, Martin Miguel; Maruo, Fumiaki; Tsonis, Panagiotis A; Chiba, Chikafumi

    2016-03-30

    The newt, a urodele amphibian, is able to repeatedly regenerate its limbs throughout its lifespan, whereas other amphibians deteriorate or lose their ability to regenerate limbs after metamorphosis. It remains to be determined whether such an exceptional ability of the newt is either attributed to a strategy, which controls regeneration in larvae, or on a novel one invented by the newt after metamorphosis. Here we report that the newt switches the cellular mechanism for limb regeneration from a stem/progenitor-based mechanism (larval mode) to a dedifferentiation-based one (adult mode) as it transits beyond metamorphosis. We demonstrate that larval newts use stem/progenitor cells such as satellite cells for new muscle in a regenerated limb, whereas metamorphosed newts recruit muscle fibre cells in the stump for the same purpose. We conclude that the newt has evolved novel strategies to secure its regenerative ability of the limbs after metamorphosis.

  13. A developmentally regulated switch from stem cells to dedifferentiation for limb muscle regeneration in newts

    PubMed Central

    Tanaka, Hibiki Vincent; Ng, Nathaniel Chuen Yin; Yang Yu, Zhan; Casco-Robles, Martin Miguel; Maruo, Fumiaki; Tsonis, Panagiotis A.; Chiba, Chikafumi

    2016-01-01

    The newt, a urodele amphibian, is able to repeatedly regenerate its limbs throughout its lifespan, whereas other amphibians deteriorate or lose their ability to regenerate limbs after metamorphosis. It remains to be determined whether such an exceptional ability of the newt is either attributed to a strategy, which controls regeneration in larvae, or on a novel one invented by the newt after metamorphosis. Here we report that the newt switches the cellular mechanism for limb regeneration from a stem/progenitor-based mechanism (larval mode) to a dedifferentiation-based one (adult mode) as it transits beyond metamorphosis. We demonstrate that larval newts use stem/progenitor cells such as satellite cells for new muscle in a regenerated limb, whereas metamorphosed newts recruit muscle fibre cells in the stump for the same purpose. We conclude that the newt has evolved novel strategies to secure its regenerative ability of the limbs after metamorphosis. PMID:27026263

  14. Diversity of Cnidarian Muscles: Function, Anatomy, Development and Regeneration

    PubMed Central

    Leclère, Lucas; Röttinger, Eric

    2017-01-01

    The ability to perform muscle contractions is one of the most important and distinctive features of eumetazoans. As the sister group to bilaterians, cnidarians (sea anemones, corals, jellyfish, and hydroids) hold an informative phylogenetic position for understanding muscle evolution. Here, we review current knowledge on muscle function, diversity, development, regeneration and evolution in cnidarians. Cnidarian muscles are involved in various activities, such as feeding, escape, locomotion and defense, in close association with the nervous system. This variety is reflected in the large diversity of muscle organizations found in Cnidaria. Smooth epithelial muscle is thought to be the most common type, and is inferred to be the ancestral muscle type for Cnidaria, while striated muscle fibers and non-epithelial myocytes would have been convergently acquired within Cnidaria. Current knowledge of cnidarian muscle development and its regeneration is limited. While orthologs of myogenic regulatory factors such as MyoD have yet to be found in cnidarian genomes, striated muscle formation potentially involves well-conserved myogenic genes, such as twist and mef2. Although satellite cells have yet to be identified in cnidarians, muscle plasticity (e.g., de- and re-differentiation, fiber repolarization) in a regenerative context and its potential role during regeneration has started to be addressed in a few cnidarian systems. The development of novel tools to study those organisms has created new opportunities to investigate in depth the development and regeneration of cnidarian muscle cells and how they contribute to the regenerative process. PMID:28168188

  15. Diversity of Cnidarian Muscles: Function, Anatomy, Development and Regeneration.

    PubMed

    Leclère, Lucas; Röttinger, Eric

    2016-01-01

    The ability to perform muscle contractions is one of the most important and distinctive features of eumetazoans. As the sister group to bilaterians, cnidarians (sea anemones, corals, jellyfish, and hydroids) hold an informative phylogenetic position for understanding muscle evolution. Here, we review current knowledge on muscle function, diversity, development, regeneration and evolution in cnidarians. Cnidarian muscles are involved in various activities, such as feeding, escape, locomotion and defense, in close association with the nervous system. This variety is reflected in the large diversity of muscle organizations found in Cnidaria. Smooth epithelial muscle is thought to be the most common type, and is inferred to be the ancestral muscle type for Cnidaria, while striated muscle fibers and non-epithelial myocytes would have been convergently acquired within Cnidaria. Current knowledge of cnidarian muscle development and its regeneration is limited. While orthologs of myogenic regulatory factors such as MyoD have yet to be found in cnidarian genomes, striated muscle formation potentially involves well-conserved myogenic genes, such as twist and mef2. Although satellite cells have yet to be identified in cnidarians, muscle plasticity (e.g., de- and re-differentiation, fiber repolarization) in a regenerative context and its potential role during regeneration has started to be addressed in a few cnidarian systems. The development of novel tools to study those organisms has created new opportunities to investigate in depth the development and regeneration of cnidarian muscle cells and how they contribute to the regenerative process.

  16. Nur77 deletion impairs muscle growth during developmental myogenesis and muscle regeneration in mice.

    PubMed

    Cortez-Toledo, Omar; Schnair, Caitlin; Sangngern, Peer; Metzger, Daniel; Chao, Lily C

    2017-01-01

    Muscle atrophy is a prevalent condition in illness and aging. Identifying novel pathways that control muscle mass may lead to therapeutic advancement. We previously identified Nur77 as a transcriptional regulator of glycolysis in skeletal muscle. More recently, we showed that Nur77 expression also controls myofiber size in mice. It was unknown, however, whether Nur77's regulation of muscle size begins during developmental myogenesis or only in adulthood. To determine the importance of Nur77 throughout muscle growth, we examined myofiber size at E18.5, 3 weeks postnatal age, and in young adult mice. Using the global Nur77-/- mice, we showed that Nur77 deficiency reduced myofiber size as early as E18.5. The reduction in myofiber size became more pronounced by 3 weeks of age. We observed comparable reduction in myofiber size in young myofiber-specific Nur77-knockout mice. These findings suggest that Nur77's effect on muscle growth is intrinsic to its expression in differentiating myofibers, and not dependent on its expression in myogenic stem cells. To determine the importance of Nur77 expression in muscle accretion in mature mice, we generated an inducible-, muscle-specific, Nur77-deficient mouse model. We demonstrated that tamoxifen-induced deletion of Nur77 in 3-month-old mice reduced myofiber size. This change was accompanied by increased activity of Smad2 and FoxO3, two negative regulators of muscle mass. The role of Nur77 in muscle growth was further elaborated in the cardiotoxin-induced muscle regeneration model. Compared to wildtype mice, regenerated myofibers were smaller in Nur77-/- mice. However, when normalized to saline-injected muscle, the recovery of sarcoplasmic area was comparable between Nur77-/- and wildtype mice. These findings suggest that Nur77 deficiency compromises myofiber growth, but not the regenerative capacity of myogenic progenitor cells. Collectively, the findings presented here demonstrate Nur77 as an important regulator of muscle

  17. Nur77 deletion impairs muscle growth during developmental myogenesis and muscle regeneration in mice

    PubMed Central

    Cortez-Toledo, Omar; Schnair, Caitlin; Sangngern, Peer; Metzger, Daniel

    2017-01-01

    Muscle atrophy is a prevalent condition in illness and aging. Identifying novel pathways that control muscle mass may lead to therapeutic advancement. We previously identified Nur77 as a transcriptional regulator of glycolysis in skeletal muscle. More recently, we showed that Nur77 expression also controls myofiber size in mice. It was unknown, however, whether Nur77’s regulation of muscle size begins during developmental myogenesis or only in adulthood. To determine the importance of Nur77 throughout muscle growth, we examined myofiber size at E18.5, 3 weeks postnatal age, and in young adult mice. Using the global Nur77-/- mice, we showed that Nur77 deficiency reduced myofiber size as early as E18.5. The reduction in myofiber size became more pronounced by 3 weeks of age. We observed comparable reduction in myofiber size in young myofiber-specific Nur77-knockout mice. These findings suggest that Nur77’s effect on muscle growth is intrinsic to its expression in differentiating myofibers, and not dependent on its expression in myogenic stem cells. To determine the importance of Nur77 expression in muscle accretion in mature mice, we generated an inducible-, muscle-specific, Nur77-deficient mouse model. We demonstrated that tamoxifen-induced deletion of Nur77 in 3-month-old mice reduced myofiber size. This change was accompanied by increased activity of Smad2 and FoxO3, two negative regulators of muscle mass. The role of Nur77 in muscle growth was further elaborated in the cardiotoxin-induced muscle regeneration model. Compared to wildtype mice, regenerated myofibers were smaller in Nur77-/- mice. However, when normalized to saline-injected muscle, the recovery of sarcoplasmic area was comparable between Nur77-/- and wildtype mice. These findings suggest that Nur77 deficiency compromises myofiber growth, but not the regenerative capacity of myogenic progenitor cells. Collectively, the findings presented here demonstrate Nur77 as an important regulator of muscle

  18. Progression of inflammation during immunodeficient mouse skeletal muscle regeneration.

    PubMed

    Grabowska, Iwona; Mazur, Magdalena A; Kowalski, K; Helinska, A; Moraczewski, Jerzy; Stremińska, Władysława; Hoser, Grażyna; Kawiak, Jerzy; Ciemerych, Maria A; Brzoska, Edyta

    2015-12-01

    The skeletal muscle injury triggers the inflammatory response which is crucial for damaged muscle fiber degradation and satellite cell activation. Immunodeficient mice are often used as a model to study the myogenic potential of transplanted human stem cells. Therefore, it is crucial to elucidate whether such model truly reflects processes occurring under physiological conditions. To answer this question we compared skeletal muscle regeneration of BALB/c, i.e. animals producing all types of inflammatory cells, and SCID mice. Results of our study documented that initial stages of muscles regeneration in both strains of mice were comparable. However, lower number of mononucleated cells was noticed in regenerating SCID mouse muscles. Significant differences in the number of CD14-/CD45+ and CD14+/CD45+ cells between BALB/c and SCID muscles were also observed. In addition, we found important differences in M1 and M2 macrophage levels of BALB/c and SCID mouse muscles identified by CD68 and CD163 markers. Thus, our data show that differences in inflammatory response during muscle regeneration, were not translated into significant modifications in muscle regeneration.

  19. Leucine supplementation improves regeneration of skeletal muscles from old rats.

    PubMed

    Pereira, Marcelo G; Silva, Meiricris T; da Cunha, Fernanda M; Moriscot, Anselmo S; Aoki, Marcelo S; Miyabara, Elen H

    2015-12-01

    The decreased regenerative capacity of old skeletal muscles involves disrupted turnover of proteins. This study investigated whether leucine supplementation in old rats could improve muscle regenerative capacity. Young and old male Wistar rats were supplemented with leucine; then, the muscles were cryolesioned and examined after 3 and 10 days. Leucine supplementation attenuated the decrease in the expression of eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1) and eukaryotic translation initiation factor 4E (eIF4E) in young and old muscles on day 3 post-injury and promoted an increase in the cross-sectional area of regenerating myofibers from both young and old soleus muscles on day 10 post-injury. This supplementation decreased the levels of ubiquitinated proteins and increased the proteasome activity in young regenerating muscles, but the opposite effect was observed in old regenerating muscles. Moreover, leucine decreased the inflammation area and induced an increase in the number of proliferating satellite cells in both young and old muscles. Our results suggest that leucine supplementation improves the regeneration of skeletal muscles from old rats, through the preservation of certain biological responses upon leucine supplementation. Such responses comprise the decrease in the inflammation area, increase in the number of proliferating satellite cells and size of regenerating myofibers, combined with the modulation of components of the phosphoinositide 3-kinase/Akt-protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway and ubiquitin-proteasome system.

  20. Diffusion tensor tractography reveals muscle reconnection during axolotl limb regeneration

    PubMed Central

    Wang, Mu-Hui; Chiou, Ling-Ling; Tseng, Wen-Yih Isaac; Lee, Hsuan-Shu

    2017-01-01

    Axolotls have amazing ability to regenerate their lost limbs. Our previous works showed that after amputation the remnant muscle ends remained at their original location whilst sending satellite cells into the regenerating parts to develop into early muscle fibers in the late differentiation stage. The parental and the newly formed muscle fibers were not connected until very late stage. The present study used non-invasive diffusion tensor imaging (DTI) to monitor weekly axolotl upper arm muscles after amputation of their upper arms. DTI tractography showed that the regenerating muscle fibers became visible at 9-wpa (weeks post amputation), but a gap was observed between the regenerating and parental muscles. The gap was filled at 10-wpa, indicating reconnection of the fibers of both muscles. This was confirmed by histology. The DTI results indicate that 23% of the muscle fibers were reconnected at 10-wpa. In conclusion, DTI can be used to visualize axolotls’ skeletal muscles and the results of muscle reconnection were in accordance with our previous findings. This non-invasive technique will allow researchers to identify the timeframe in which muscle fiber reconnection takes place and thus enable the study of the mechanisms underlying this reconnection. PMID:28253344

  1. Diffusion tensor tractography reveals muscle reconnection during axolotl limb regeneration.

    PubMed

    Wu, Cheng-Han; Chen, Yu-Jen; Wang, Mu-Hui; Chiou, Ling-Ling; Tseng, Wen-Yih Isaac; Lee, Hsuan-Shu

    2017-01-01

    Axolotls have amazing ability to regenerate their lost limbs. Our previous works showed that after amputation the remnant muscle ends remained at their original location whilst sending satellite cells into the regenerating parts to develop into early muscle fibers in the late differentiation stage. The parental and the newly formed muscle fibers were not connected until very late stage. The present study used non-invasive diffusion tensor imaging (DTI) to monitor weekly axolotl upper arm muscles after amputation of their upper arms. DTI tractography showed that the regenerating muscle fibers became visible at 9-wpa (weeks post amputation), but a gap was observed between the regenerating and parental muscles. The gap was filled at 10-wpa, indicating reconnection of the fibers of both muscles. This was confirmed by histology. The DTI results indicate that 23% of the muscle fibers were reconnected at 10-wpa. In conclusion, DTI can be used to visualize axolotls' skeletal muscles and the results of muscle reconnection were in accordance with our previous findings. This non-invasive technique will allow researchers to identify the timeframe in which muscle fiber reconnection takes place and thus enable the study of the mechanisms underlying this reconnection.

  2. The CXCR4/SDF1 Axis Improves Muscle Regeneration Through MMP-10 Activity

    PubMed Central

    Bobadilla, Miriam; Sainz, Neira; Abizanda, Gloria; Orbe, Josune; Rodriguez, José Antonio; Páramo, José Antonio; Prósper, Felipe

    2014-01-01

    The CXCR4/SDF1 axis participates in various cellular processes, including cell migration, which is essential for skeletal muscle repair. Although increasing evidence has confirmed the role of CXCR4/SDF1 in embryonic muscle development, the function of this pathway during adult myogenesis remains to be fully elucidated. In addition, a role for CXCR4 signaling in muscle maintenance and repair has only recently emerged. Here, we have demonstrated that CXCR4 and stromal cell-derived factor-1 (SDF1) are up-regulated in injured muscle, suggesting their involvement in the repair process. In addition, we found that notexin-damaged muscles showed delayed muscle regeneration on treatment with CXCR4 agonist (AMD3100). Accordingly, small-interfering RNA-mediated silencing of SDF1 or CXCR4 in injured muscles impaired muscle regeneration, whereas the addition of SDF1 ligand accelerated repair. Furthermore, we identified that CXCR4/SDF1-regulated muscle repair was dependent on matrix metalloproteinase-10 (MMP-10) activity. Thus, our findings support a model in which MMP-10 activity modulates CXCR4/SDF1 signaling, which is essential for efficient skeletal muscle regeneration. PMID:24548137

  3. Sulfs are regulators of growth factor signaling for satellite cell differentiation and muscle regeneration.

    PubMed

    Langsdorf, Aliete; Do, Anh-Tri; Kusche-Gullberg, Marion; Emerson, Charles P; Ai, Xingbin

    2007-11-15

    Heparan sulfate proteoglycans (HSPGs) are required during muscle regeneration for regulating extracellular signaling pathways. HSPGs interact with growth factors and receptors through heparan sulfate (HS) chains. However, the regulatory mechanisms that control HS sulfation to affect the growth factor-dependent proliferation and differentiation of satellite cells are yet unknown. Here we report the essential functions of extracellular HS 6-O-endosulfatases (Sulfs) during muscle regeneration. We show that quiescent and activated satellite cells differentially express mouse Sulf1 (MSulf1) and MSulf2. MSulfs are not required for the formation of skeletal muscles and satellite cells, but they have redundant, essential roles to promote muscle regeneration, as MSulf double mutant mice exhibit delayed myogenic differentiation and prolonged Pax7 expression after cardiotoxin-induced skeletal muscle injury, while single MSulf knockouts regenerate normally. HS structural analysis demonstrates that Sulfs are regulatory HS-modifying enzymes that control HS 6-O-desulfation of activated satellite cells. Mechanistically, we show that MSulfs repress FGF2 signaling in activated satellite cells, leading us to propose that MSulfs are growth factor signaling sensors to control the proliferation to differentiation switch of satellite cells to initiate differentiation during regeneration. Our results establish Sulfs as essential regulators of HS-dependent growth factor signaling in the adult muscle stem cell niche.

  4. Role of pericytes in skeletal muscle regeneration and fat accumulation.

    PubMed

    Birbrair, Alexander; Zhang, Tan; Wang, Zhong-Min; Messi, Maria Laura; Enikolopov, Grigori N; Mintz, Akiva; Delbono, Osvaldo

    2013-08-15

    Stem cells ensure tissue regeneration, while overgrowth of adipogenic cells may compromise organ recovery and impair function. In myopathies and muscle atrophy associated with aging, fat accumulation increases dysfunction, and after chronic injury, the process of fatty degeneration, in which muscle is replaced by white adipocytes, further compromises tissue function and environment. Some studies suggest that pericytes may contribute to muscle regeneration as well as fat formation. This work reports the presence of two pericyte subpopulations in the skeletal muscle and characterizes their specific roles. Skeletal muscle from Nestin-GFP/NG2-DsRed mice show two types of pericytes, Nestin-GFP-/NG2-DsRed+ (type-1) and Nestin-GFP+/NG2-DsRed+ (type-2), in close proximity to endothelial cells. We also found that both Nestin-GFP-/NG2-DsRed+ and Nestin-GFP+/NG2-DsRed+ cells colocalize with staining of two pericyte markers, PDGFRβ and CD146, but only type-1 pericyte express the adipogenic progenitor marker PDGFRα. Type-2 pericytes participate in muscle regeneration, while type-1 contribute to fat accumulation. Transplantation studies indicate that type-1 pericytes do not form muscle in vivo, but contribute to fat deposition in the skeletal muscle, while type-2 pericytes contribute only to the new muscle formation after injury, but not to the fat accumulation. Our results suggest that type-1 and type-2 pericytes contribute to successful muscle regeneration which results from a balance of myogenic and nonmyogenic cells activation.

  5. Systemic Regulators of Skeletal Muscle Regeneration in Obesity

    PubMed Central

    Sinha, Indranil; Sakthivel, Dharaniya; Varon, David E.

    2017-01-01

    Skeletal muscle maintenance is a dynamic process and undergoes constant repair and regeneration. However, skeletal muscle regenerative capacity declines in obesity. In this review, we focus on obesity-associated changes in inflammation, metabolism, and impaired insulin signaling, which are pathologically dysregulated and ultimately result in a loss of muscle mass and function. In addition, we examine the relationships between skeletal muscle, liver, and visceral adipose tissue in an obese state. PMID:28261159

  6. Echinoderms: potential model systems for studies on muscle regeneration.

    PubMed

    García-Arrarás, José E; Dolmatov, Igor Yu

    2010-01-01

    Organisms of the phylum Echinodermata show some of the most impressive regenerative feats within the animal kingdom. Following injury or self-induced autotomy, species in this phylum can regenerate most tissues and organs, being the regeneration of the muscular systems one of the best studied. Even though echinoderms are closely related to chordates, they are little known in the biomedical field, and therefore their uses to study pharmacological effects on muscle formation and/or regeneration have been extremely limited. In order to rectify this lack of knowledge, we describe here the echinoderm muscular systems, particularly the somatic and visceral muscle components. In addition, we provide details of the processes that are known to take place during muscle regeneration, namely dedifferentiation, myogenesis and new muscle formation. Finally, we provide the available information on molecular and pharmacological studies that involve echinoderm muscle regeneration. We expect that by making this information accessible, researchers consider the use of echinoderms as model systems for pharmacological studies in muscle development and regeneration.

  7. Role of muscle stem cells during skeletal regeneration.

    PubMed

    Abou-Khalil, Rana; Yang, Frank; Lieu, Shirley; Julien, Anais; Perry, Jaselle; Pereira, Catia; Relaix, Frédéric; Miclau, Theodore; Marcucio, Ralph; Colnot, Céline

    2015-05-01

    Although the importance of muscle in skeletal regeneration is well recognized clinically, the mechanisms by which muscle supports bone repair have remained elusive. Muscle flaps are often used to cover the damaged bone after traumatic injury yet their contribution to bone healing is not known. Here, we show that direct bone-muscle interactions are required for periosteum activation and callus formation, and that muscle grafts provide a source of stem cells for skeletal regeneration. We investigated the role of satellite cells, the muscle stem cells. Satellite cells loss in Pax7(-/-) mice and satellite cell ablation in Pax7(Cre) (ERT) (2/) (+) ;DTA(f/f) mice impaired bone regeneration. Although satellite cells did not contribute as a large source of cells endogenously, they exhibited a potential to contribute to bone repair after transplantation. The fracture healing phenotype in Pax7(Cre) (ERT) (2/) (+) ;DTA(f/f) mice was associated with decreased bone morphogenetic proteins (BMPs), insulin-like growth factor 1, and fibroblast growth factor 2 expression that are normally upregulated in response to fracture in satellite cells. Exogenous rhBMP2 improved bone healing in Pax7(Cre) (ERT) (2/) (+) ;DTA(f/f) mice further supporting the role of satellite cells as a source of growth factors. These results provide the first functional evidence for a direct contribution of muscle to bone regeneration with important clinical implications as it may impact the use of muscle flaps, muscle stem cells, and growth factors in orthopedic applications.

  8. Roles of nonmyogenic mesenchymal progenitors in pathogenesis and regeneration of skeletal muscle

    PubMed Central

    Uezumi, Akiyoshi; Ikemoto-Uezumi, Madoka; Tsuchida, Kunihiro

    2014-01-01

    Adult skeletal muscle possesses a remarkable regenerative ability that is dependent on satellite cells. However, skeletal muscle is replaced by fatty and fibrous connective tissue in several pathological conditions. Fatty and fibrous connective tissue becomes a major cause of muscle weakness and leads to further impairment of muscle function. Because the occurrence of fatty and fibrous connective tissue is usually associated with severe destruction of muscle, the idea that dysregulation of the fate switch in satellite cells may underlie this pathological change has emerged. However, recent studies identified nonmyogenic mesenchymal progenitors in skeletal muscle and revealed that fatty and fibrous connective tissue originates from these progenitors. Later, these progenitors were also demonstrated to be the major contributor to heterotopic ossification in skeletal muscle. Because nonmyogenic mesenchymal progenitors represent a distinct cell population from satellite cells, targeting these progenitors could be an ideal therapeutic strategy that specifically prevents pathological changes of skeletal muscle, while preserving satellite cell-dependent regeneration. In addition to their roles in pathogenesis of skeletal muscle, nonmyogenic mesenchymal progenitors may play a vital role in muscle regeneration by regulating satellite cell behavior. Conversely, muscle cells appear to regulate behavior of nonmyogenic mesenchymal progenitors. Thus, these cells regulate each other reciprocally and a proper balance between them is a key determinant of muscle integrity. Furthermore, nonmyogenic mesenchymal progenitors have been shown to maintain muscle mass in a steady homeostatic condition. Understanding the nature of nonmyogenic mesenchymal progenitors will provide valuable insight into the pathophysiology of skeletal muscle. In this review, we focus on nonmyogenic mesenchymal progenitors and discuss their roles in muscle pathogenesis, regeneration, and homeostasis. PMID

  9. Bone marrow-derived cell regulation of skeletal muscle regeneration

    PubMed Central

    Sun, Dongxu; Martinez, Carlo O.; Ochoa, Oscar; Ruiz-Willhite, Lourdes; Bonilla, Jose R.; Centonze, Victoria E.; Waite, Lindsay L.; Michalek, Joel E.; McManus, Linda M.; Shireman, Paula K.

    2009-01-01

    Limb regeneration requires the coordination of multiple stem cell populations to recapitulate the process of tissue formation. Therefore, bone marrow (BM) -derived cell regulation of skeletal muscle regeneration was examined in mice lacking the CC chemokine receptor 2 (CCR2). Myofiber size, numbers of myogenic progenitor cells (MPCs), and recruitment of BM-derived cells and macrophages were assessed after cardiotoxin-induced injury of chimeric mice produced by transplanting BM from wild-type (WT) or CCR2−/− mice into irradiated WT or CCR2−/− host mice. Regardless of the host genotype, muscle regeneration and recruitment of BM-derived cells and macrophages were similar in mice replenished with WT BM, whereas BM-derived cells and macrophage accumulation were decreased and muscle regeneration was impaired in all animals receiving CCR2−/− BM. Furthermore, numbers of MPCs (CD34+/Sca-1−/CD45− cells) were significantly increased in mice receiving CCR2−/− BM despite the decreased size of regenerated myofibers. Thus, the expression of CCR2 on BM-derived cells regulated macrophage recruitment into injured muscle, numbers of MPC, and the extent of regenerated myofiber size, all of which were independent of CCR2 expression on host-derived cells. Future studies in regenerative medicine must include consideration of the role of BM-derived cells, possibly macrophages, in CCR2-dependent events that regulate effective skeletal muscle regeneration.—Sun, D., Martinez, C. O., Ochoa, O., Ruiz-Willhite, L., Bonilla, J. R., Centonze, V. E., Waite, L. L., Michalek, J. E., McManus, L. M., Shireman, P. K. Bone marrow-derived cell regulation of skeletal muscle regeneration. PMID:18827026

  10. Preparation of adult muscle fiber-associated stem/precursor cells.

    PubMed

    Conboy, Michael J; Conboy, Irina M

    2010-01-01

    In our studies of muscle regeneration we have developed, modified, and optimized techniques to isolate and study the stem and precursor cells to muscle tissue. Our goals have been to obtain for study muscle fibers in bulk, or the fiber-associated cells, separately from the other cells found in muscle. Using these techniques, myofiber-associated cells may be isolated from neonatal through adult muscle, from resting or from regenerating muscle, thus allowing one to investigate the cellular populations participating during the time course of these events. The protocol is applicable to any age and condition of muscle and may be adapted for other tissues.

  11. Calcium signaling in skeletal muscle development, maintenance and regeneration.

    PubMed

    Tu, Michelle K; Levin, Jacqueline B; Hamilton, Andrew M; Borodinsky, Laura N

    2016-03-01

    Skeletal muscle-specific stem cells are pivotal for tissue development and regeneration. Muscle plasticity, inherent in these processes, is also essential for daily life activities. Great advances and efforts have been made in understanding the function of the skeletal muscle-dedicated stem cells, called muscle satellite cells, and the specific signaling mechanisms that activate them for recruitment in the repair of the injured muscle. Elucidating these signaling mechanisms may contribute to devising therapies for muscular injury or disease. Here we review the studies that have contributed to our understanding of how calcium signaling regulates skeletal muscle development, homeostasis and regeneration, with a focus on the calcium dynamics and calcium-dependent effectors that participate in these processes.

  12. Muscle regeneration in the prolonged absence of myostatin.

    PubMed

    Wagner, Kathryn R; Liu, Xiaosong; Chang, Xiaoli; Allen, Ronald E

    2005-02-15

    Myostatin is an endogenous inhibitor of muscle conserved across diverse species. In the absence of myostatin, there is massive muscle growth in mice, cattle, and humans. Previous studies in the mdx mouse model of muscular dystrophy demonstrate that inhibiting myostatin attenuates several features of dystrophic muscle. These findings have encouraged the development of human therapies to block myostatin. However, little is known of the long-term effects on muscle of myostatin blockade. To evaluate potential sequelae from the prolonged absence of myostatin, senescent myostatin null (mstn-/-) mice were studied. Senescent mstn-/- mice continue to have normal muscle with increased mass and strength relative to controls. Muscles of senescent mstn-/- mice regenerate robustly from both chronic and acute injury. Early markers of regeneration are enhanced in the absence of myostatin, suggesting a mechanism for the attenuation of dystrophic features found in mdx mice lacking myostatin.

  13. FOXO1 delays skeletal muscle regeneration and suppresses myoblast proliferation.

    PubMed

    Yamashita, Atsushi; Hatazawa, Yukino; Hirose, Yuma; Ono, Yusuke; Kamei, Yasutomi

    2016-08-01

    Unloading stress, such as bed rest, inhibits the regenerative potential of skeletal muscles; however, the underlying mechanisms remain largely unknown. FOXO1 expression, which induces the upregulated expression of the cell cycle inhibitors p57 and Gadd45α, is known to be increased in the skeletal muscle under unloading conditions. However, there is no report addressing FOXO1-induced inhibition of myoblast proliferation. Therefore, we induced muscle injury by cardiotoxin in transgenic mice overexpressing FOXO1 in the skeletal muscle (FOXO1-Tg mice) and observed regeneration delay in skeletal muscle mass and cross-sectional area in FOXO1-Tg mice. Increased p57 and Gadd45α mRNA levels, and decreased proliferation capacity were observed in C2C12 myoblasts expressing a tamoxifen-inducible active form of FOXO1. These results suggest that decreased proliferation capacity of myoblasts by FOXO1 disrupts skeletal muscle regeneration under FOXO1-increased conditions, such as unloading.

  14. Angiopoietin-1 enhances skeletal muscle regeneration in mice.

    PubMed

    Mofarrahi, Mahroo; McClung, Joseph M; Kontos, Christopher D; Davis, Elaine C; Tappuni, Bassman; Moroz, Nicolay; Pickett, Amy E; Huck, Laurent; Harel, Sharon; Danialou, Gawiyou; Hussain, Sabah N A

    2015-04-01

    Activation of muscle progenitor cell myogenesis and endothelial cell angiogenesis is critical for the recovery of skeletal muscle from injury. Angiopoietin-1 (Ang-1), a ligand of Tie-2 receptors, enhances angiogenesis and skeletal muscle satellite cell survival; however, its role in skeletal muscle regeneration after injury is unknown. We assessed the effects of Ang-1 on fiber regeneration, myogenesis, and angiogenesis in injured skeletal muscle (tibialis anterior, TA) in mice. We also assessed endogenous Ang-1 levels and localization in intact and injured TA muscles. TA fiber injury was triggered by cardiotoxin injection. Endogenous Ang-1 mRNA levels immediately decreased in response to cardiotoxin then increased during the 2 wk. Ang-1 protein was expressed in satellite cells, both in noninjured and recovering TA muscles. Positive Ang-1 staining was present in blood vessels but not in nerve fibers. Four days after the initiation of injury, injection of adenoviral Ang-1 into injured muscles resulted in significant increases in in situ TA muscle contractility, muscle fiber regeneration, and capillary density. In cultured human skeletal myoblasts, recombinant Ang-1 protein increased survival, proliferation, migration, and differentiation into myotubes. The latter effect was associated with significant upregulation of the expression of the myogenic regulatory factors MyoD and Myogenin and certain genes involved in cell cycle regulation. We conclude that Ang-1 strongly enhances skeletal muscle regeneration in response to fiber injury and that this effect is mediated through induction of the myogenesis program in muscle progenitor cells and the angiogenesis program in endothelial cells.

  15. The PERK arm of the unfolded protein response regulates satellite cell-mediated skeletal muscle regeneration.

    PubMed

    Xiong, Guangyan; Hindi, Sajedah M; Mann, Aman K; Gallot, Yann S; Bohnert, Kyle R; Cavener, Douglas R; Whittemore, Scott R; Kumar, Ashok

    2017-03-23

    Regeneration of skeletal muscle in adults is mediated by satellite stem cells. Accumulation of misfolded proteins triggers endoplasmic reticulum stress that leads to unfolded protein response (UPR). The UPR is relayed to the cell through the activation of PERK, IRE1/XBP1, and ATF6. Here, we demonstrate that levels of PERK and IRE1 are increased in satellite cells upon muscle injury. Inhibition of PERK, but not the IRE1 arm of the UPR in satellite cells inhibits myofiber regeneration in adult mice. PERK is essential for the survival and differentiation of activated satellite cells into the myogenic lineage. Deletion of PERK causes hyper-activation of p38 MAPK during myogenesis. Blocking p38 MAPK activity improves the survival and differentiation of PERK-deficient satellite cells in vitro and muscle formation in vivo. Collectively, our results suggest that the PERK arm of the UPR plays a pivotal role in the regulation of satellite cell homeostasis during regenerative myogenesis.

  16. Barx2 is expressed in satellite cells and is required for normal muscle growth and regeneration.

    PubMed

    Meech, Robyn; Gonzalez, Katie N; Barro, Marietta; Gromova, Anastasia; Zhuang, Lizhe; Hulin, Julie-Ann; Makarenkova, Helen P

    2012-02-01

    Muscle growth and regeneration are regulated through a series of spatiotemporally dependent signaling and transcriptional cascades. Although the transcriptional program controlling myogenesis has been extensively investigated, the full repertoire of transcriptional regulators involved in this process is far from defined. Various homeodomain transcription factors have been shown to play important roles in both muscle development and muscle satellite cell-dependent repair. Here, we show that the homeodomain factor Barx2 is a new marker for embryonic and adult myoblasts and is required for normal postnatal muscle growth and repair. Barx2 is coexpressed with Pax7, which is the canonical marker of satellite cells, and is upregulated in satellite cells after muscle injury. Mice lacking the Barx2 gene show reduced postnatal muscle growth, muscle atrophy, and defective muscle repair. Moreover, loss of Barx2 delays the expression of genes that control proliferation and differentiation in regenerating muscle. Consistent with the in vivo observations, satellite cell-derived myoblasts cultured from Barx2(-/-) mice show decreased proliferation and ability to differentiate relative to those from wild-type or Barx2(+/-) mice. Barx2(-/-) myoblasts show reduced expression of the differentiation-associated factor myogenin as well as cell adhesion and matrix molecules. Finally, we find that mice lacking both Barx2 and dystrophin gene expression have severe early onset myopathy. Together, these data indicate that Barx2 is an important regulator of muscle growth and repair that acts via the control of satellite cell proliferation and differentiation.

  17. Hypoxia induces heart regeneration in adult mice.

    PubMed

    Nakada, Yuji; Canseco, Diana C; Thet, SuWannee; Abdisalaam, Salim; Asaithamby, Aroumougame; Santos, Celio X; Shah, Ajay M; Zhang, Hua; Faber, James E; Kinter, Michael T; Szweda, Luke I; Xing, Chao; Hu, Zeping; Deberardinis, Ralph J; Schiattarella, Gabriele; Hill, Joseph A; Oz, Orhan; Lu, Zhigang; Zhang, Cheng Cheng; Kimura, Wataru; Sadek, Hesham A

    2017-01-12

    The adult mammalian heart is incapable of regeneration following cardiomyocyte loss, which underpins the lasting and severe effects of cardiomyopathy. Recently, it has become clear that the mammalian heart is not a post-mitotic organ. For example, the neonatal heart is capable of regenerating lost myocardium, and the adult heart is capable of modest self-renewal. In both of these scenarios, cardiomyocyte renewal occurs via the proliferation of pre-existing cardiomyocytes, and is regulated by aerobic-respiration-mediated oxidative DNA damage. Therefore, we reasoned that inhibiting aerobic respiration by inducing systemic hypoxaemia would alleviate oxidative DNA damage, thereby inducing cardiomyocyte proliferation in adult mammals. Here we report that, in mice, gradual exposure to severe systemic hypoxaemia, in which inspired oxygen is gradually decreased by 1% and maintained at 7% for 2 weeks, results in inhibition of oxidative metabolism, decreased reactive oxygen species production and oxidative DNA damage, and reactivation of cardiomyocyte mitosis. Notably, we find that exposure to hypoxaemia 1 week after induction of myocardial infarction induces a robust regenerative response with decreased myocardial fibrosis and improvement of left ventricular systolic function. Genetic fate-mapping analysis confirms that the newly formed myocardium is derived from pre-existing cardiomyocytes. These results demonstrate that the endogenous regenerative properties of the adult mammalian heart can be reactivated by exposure to gradual systemic hypoxaemia, and highlight the potential therapeutic role of hypoxia in regenerative medicine.

  18. Regeneration of Zebrafish CNS: Adult Neurogenesis

    PubMed Central

    Ghosh, Sukla; Hui, Subhra Prakash

    2016-01-01

    Regeneration in the animal kingdom is one of the most fascinating problems that have allowed scientists to address many issues of fundamental importance in basic biology. However, we came to know that the regenerative capability may vary across different species. Among vertebrates, fish and amphibians are capable of regenerating a variety of complex organs through epimorphosis. Zebrafish is an excellent animal model, which can repair several organs like damaged retina, severed spinal cord, injured brain and heart, and amputated fins. The focus of the present paper is on spinal cord regeneration in adult zebrafish. We intend to discuss our current understanding of the cellular and molecular mechanism(s) that allows formation of proliferating progenitors and controls neurogenesis, which involve changes in epigenetic and transcription programs. Unlike mammals, zebrafish retains radial glia, a nonneuronal cell type in their adult central nervous system. Injury induced proliferation involves radial glia which proliferate, transcribe embryonic genes, and can give rise to new neurons. Recent technological development of exquisite molecular tools in zebrafish, such as cell ablation, lineage analysis, and novel and substantial microarray, together with advancement in stem cell biology, allowed us to investigate how progenitor cells contribute to the generation of appropriate structures and various underlying mechanisms like reprogramming. PMID:27382491

  19. Pericytes: multitasking cells in the regeneration of injured, diseased, and aged skeletal muscle

    PubMed Central

    Birbrair, Alexander; Zhang, Tan; Wang, Zhong-Min; Messi, Maria L.; Mintz, Akiva; Delbono, Osvaldo

    2014-01-01

    Pericytes are perivascular cells that envelop and make intimate connections with adjacent capillary endothelial cells. Recent studies show that they may have a profound impact in skeletal muscle regeneration, innervation, vessel formation, fibrosis, fat accumulation, and ectopic bone formation throughout life. In this review, we summarize and evaluate recent advances in our understanding of pericytes' influence on adult skeletal muscle pathophysiology. We also discuss how further elucidating their biology may offer new approaches to the treatment of conditions characterized by muscle wasting. PMID:25278877

  20. Role of Pericytes in Skeletal Muscle Regeneration and Fat Accumulation

    PubMed Central

    Birbrair, Alexander; Zhang, Tan; Wang, Zhong-Min; Messi, Maria Laura; Enikolopov, Grigori N.; Mintz, Akiva

    2013-01-01

    Stem cells ensure tissue regeneration, while overgrowth of adipogenic cells may compromise organ recovery and impair function. In myopathies and muscle atrophy associated with aging, fat accumulation increases dysfunction, and after chronic injury, the process of fatty degeneration, in which muscle is replaced by white adipocytes, further compromises tissue function and environment. Some studies suggest that pericytes may contribute to muscle regeneration as well as fat formation. This work reports the presence of two pericyte subpopulations in the skeletal muscle and characterizes their specific roles. Skeletal muscle from Nestin-GFP/NG2-DsRed mice show two types of pericytes, Nestin-GFP-/NG2-DsRed+ (type-1) and Nestin-GFP+/NG2-DsRed+ (type-2), in close proximity to endothelial cells. We also found that both Nestin-GFP-/NG2-DsRed+ and Nestin-GFP+/NG2-DsRed+ cells colocalize with staining of two pericyte markers, PDGFRβ and CD146, but only type-1 pericyte express the adipogenic progenitor marker PDGFRα. Type-2 pericytes participate in muscle regeneration, while type-1 contribute to fat accumulation. Transplantation studies indicate that type-1 pericytes do not form muscle in vivo, but contribute to fat deposition in the skeletal muscle, while type-2 pericytes contribute only to the new muscle formation after injury, but not to the fat accumulation. Our results suggest that type-1 and type-2 pericytes contribute to successful muscle regeneration which results from a balance of myogenic and nonmyogenic cells activation. PMID:23517218

  1. Satellite-like cells contribute to pax7-dependent skeletal muscle repair in adult zebrafish.

    PubMed

    Berberoglu, Michael A; Gallagher, Thomas L; Morrow, Zachary T; Talbot, Jared C; Hromowyk, Kimberly J; Tenente, Inês M; Langenau, David M; Amacher, Sharon L

    2017-04-15

    Satellite cells, also known as muscle stem cells, are responsible for skeletal muscle growth and repair in mammals. Pax7 and Pax3 transcription factors are established satellite cell markers required for muscle development and regeneration, and there is great interest in identifying additional factors that regulate satellite cell proliferation, differentiation, and/or skeletal muscle regeneration. Due to the powerful regenerative capacity of many zebrafish tissues, even in adults, we are exploring the regenerative potential of adult zebrafish skeletal muscle. Here, we show that adult zebrafish skeletal muscle contains cells similar to mammalian satellite cells. Adult zebrafish satellite-like cells have dense heterochromatin, express Pax7 and Pax3, proliferate in response to injury, and show peak myogenic responses 4-5 days post-injury (dpi). Furthermore, using a pax7a-driven GFP reporter, we present evidence implicating satellite-like cells as a possible source of new muscle. In lieu of central nucleation, which distinguishes regenerating myofibers in mammals, we describe several characteristics that robustly identify newly-forming myofibers from surrounding fibers in injured adult zebrafish muscle. These characteristics include partially overlapping expression in satellite-like cells and regenerating myofibers of two RNA-binding proteins Rbfox2 and Rbfoxl1, known to regulate embryonic muscle development and function. Finally, by analyzing pax7a; pax7b double mutant zebrafish, we show that Pax7 is required for adult skeletal muscle repair, as it is in the mouse.

  2. PEDF-derived peptide promotes skeletal muscle regeneration through its mitogenic effect on muscle progenitor cells.

    PubMed

    Ho, Tsung-Chuan; Chiang, Yi-Pin; Chuang, Chih-Kuang; Chen, Show-Li; Hsieh, Jui-Wen; Lan, Yu-Wen; Tsao, Yeou-Ping

    2015-08-01

    In response injury, intrinsic repair mechanisms are activated in skeletal muscle to replace the damaged muscle fibers with new muscle fibers. The regeneration process starts with the proliferation of satellite cells to give rise to myoblasts, which subsequently differentiate terminally into myofibers. Here, we investigated the promotion effect of pigment epithelial-derived factor (PEDF) on muscle regeneration. We report that PEDF and a synthetic PEDF-derived short peptide (PSP; residues Ser(93)-Leu(112)) induce satellite cell proliferation in vitro and promote muscle regeneration in vivo. Extensively, soleus muscle necrosis was induced in rats by bupivacaine, and an injectable alginate gel was used to release the PSP in the injured muscle. PSP delivery was found to stimulate satellite cell proliferation in damaged muscle and enhance the growth of regenerating myofibers, with complete regeneration of normal muscle mass by 2 wk. In cell culture, PEDF/PSP stimulated C2C12 myoblast proliferation, together with a rise in cyclin D1 expression. PEDF induced the phosphorylation of ERK1/2, Akt, and STAT3 in C2C12 myoblasts. Blocking the activity of ERK, Akt, or STAT3 with pharmacological inhibitors attenuated the effects of PEDF/PSP on the induction of C2C12 cell proliferation and cyclin D1 expression. Moreover, 5-bromo-2'-deoxyuridine pulse-labeling demonstrated that PEDF/PSP stimulated primary rat satellite cell proliferation in myofibers in vitro. In summary, we report for the first time that PSP is capable of promoting the regeneration of skeletal muscle. The signaling mechanism involves the ERK, AKT, and STAT3 pathways. These results show the potential utility of this PEDF peptide for muscle regeneration.

  3. Characteristics of locomotion, muscle strength, and muscle tissue in regenerating rat skeletal muscles.

    PubMed

    Iwata, Akira; Fuchioka, Satoshi; Hiraoka, Koichi; Masuhara, Mitsuhiko; Kami, Katsuya

    2010-05-01

    Although numerous studies have aimed to elucidate the mechanisms used to repair the structure and function of injured skeletal muscles, it remains unclear how and when movement recovers following damage. We performed a temporal analysis to characterize the changes in movement, muscle function, and muscle structure after muscle injury induced by the drop-mass technique. At each time-point, movement recovery was determined by ankle kinematic analysis of locomotion, and functional recovery was represented by isometric force. As a histological analysis, the cross-sectional area of myotubes was measured to examine structural regeneration. The dorsiflexion angle of the ankle, as assessed by kinematic analysis of locomotion, increased after injury and then returned to control levels by day 14 post-injury. The isometric force returned to normal levels by day 21 post-injury. However, the size of the myotubes did not reach normal levels, even at day 21 post-injury. These results indicate that recovery of locomotion occurs prior to recovery of isometric force and that functional recovery occurs earlier than structural regeneration. Thus, it is suggested that recovery of the movement and function of injured skeletal muscles might be insufficient as markers for estimating the degree of neuromuscular system reconstitution.

  4. Expression of glucocorticoid receptors in the regenerating human skeletal muscle.

    PubMed

    Filipović, D; Pirkmajer, S; Mis, K; Mars, T; Grubic, Z

    2011-01-01

    Many stress conditions are accompanied by skeletal muscle dysfunction and regeneration, which is essentially a recapitulation of the embryonic development. However, regeneration usually occurs under conditions of hypothalamus-pituitary-adrenal gland axis activation and therefore increased glucocorticoid (GC) levels. Glucocorticoid receptor (GR), the main determinant of cellular responsiveness to GCs, exists in two isoforms (GRalpha and GRbeta) in humans. While the role of GRalpha is well characterized, GRbeta remains an elusive player in GC signalling. To elucidate basic characteristics of GC signalling in the regenerating human skeletal muscle we assessed GRalpha and GRbeta expression pattern in cultured human myoblasts and myotubes and their response to 24-hour dexamethasone (DEX) treatment. There was no difference in GRalpha mRNA and protein expression or DEX-mediated GRalpha down-regulation in myoblasts and myotubes. GRbeta mRNA level was very low in myoblasts and remained unaffected by differentiation and/or DEX. GRbeta protein could not be detected. These results indicate that response to GCs is established very early during human skeletal muscle regeneration and that it remains practically unchanged before innervation is established. Very low GRbeta mRNA expression and inability to detect GRbeta protein suggests that GRbeta is not a major player in the early stages of human skeletal muscle regeneration.

  5. Mesodermal iPSC–derived progenitor cells functionally regenerate cardiac and skeletal muscle

    PubMed Central

    Quattrocelli, Mattia; Swinnen, Melissa; Giacomazzi, Giorgia; Camps, Jordi; Barthélemy, Ines; Ceccarelli, Gabriele; Caluwé, Ellen; Grosemans, Hanne; Thorrez, Lieven; Pelizzo, Gloria; Muijtjens, Manja; Verfaillie, Catherine M.; Blot, Stephane; Janssens, Stefan; Sampaolesi, Maurilio

    2015-01-01

    Conditions such as muscular dystrophies (MDs) that affect both cardiac and skeletal muscles would benefit from therapeutic strategies that enable regeneration of both of these striated muscle types. Protocols have been developed to promote induced pluripotent stem cells (iPSCs) to differentiate toward cardiac or skeletal muscle; however, there are currently no strategies to simultaneously target both muscle types. Tissues exhibit specific epigenetic alterations; therefore, source-related lineage biases have the potential to improve iPSC-driven multilineage differentiation. Here, we determined that differential myogenic propensity influences the commitment of isogenic iPSCs and a specifically isolated pool of mesodermal iPSC-derived progenitors (MiPs) toward the striated muscle lineages. Differential myogenic propensity did not influence pluripotency, but did selectively enhance chimerism of MiP-derived tissue in both fetal and adult skeletal muscle. When injected into dystrophic mice, MiPs engrafted and repaired both skeletal and cardiac muscle, reducing functional defects. Similarly, engraftment into dystrophic mice of canine MiPs from dystrophic dogs that had undergone TALEN-mediated correction of the MD-associated mutation also resulted in functional striatal muscle regeneration. Moreover, human MiPs exhibited the same capacity for the dual differentiation observed in murine and canine MiPs. The findings of this study suggest that MiPs should be further explored for combined therapy of cardiac and skeletal muscles. PMID:26571398

  6. 3-hydroxy 3-methylglutaryl coenzyme A reductase inhibition impairs muscle regeneration.

    PubMed

    Trapani, Laura; Segatto, Marco; La Rosa, Piergiorgio; Fanelli, Francesca; Moreno, Sandra; Marino, Maria; Pallottini, Valentina

    2012-06-01

    Skeletal muscle has the ability to regenerate new muscle fibers after injury. The process of new muscle formation requires that quiescent mononuclear muscle precursor cells (myoblasts) become activated, proliferate, differentiate, and fuse into multinucleated myotubes which, in turn, undergo further differentiation and mature to form functional muscle fibers. Previous data demonstrated the crucial role played by 3-hydroxy 3-methylglutaryl coenzyme A reductase (HMGR), the rate-limiting enzyme of cholesterol biosynthetic pathway, in fetal rat myoblast (L6) differentiation. This finding, along with epidemiological studies assessing the myotoxic effect of statins, HMGR inhibitors, allowed us to speculate that HMGR could be strongly involved in skeletal muscle repair. Thus, our research was aimed at evaluating such involvement: in vitro and in vivo experiments were performed on both mouse adult satellite cell derived myoblasts (SCDM) and mouse muscles injured with cardiotoxin. Results demonstrate that HMGR inhibition by the statin Simvastatin reduces SCDM fusion index, fast MHC protein levels by 60% and slow MHC by 40%. Most importantly, HMGR inhibition delays skeletal muscle regeneration in vivo. Thus, besides complaining of myopathies, patients given Simvastatin could also undergo an impairment in muscle repair.

  7. Muscle Satellite Cell Protein Teneurin-4 Regulates Differentiation During Muscle Regeneration.

    PubMed

    Ishii, Kana; Suzuki, Nobuharu; Mabuchi, Yo; Ito, Naoki; Kikura, Naomi; Fukada, So-Ichiro; Okano, Hideyuki; Takeda, Shin'ichi; Akazawa, Chihiro

    2015-10-01

    Satellite cells are maintained in an undifferentiated quiescent state, but during muscle regeneration they acquire an activated stage, and initiate to proliferate and differentiate as myoblasts. The transmembrane protein teneurin-4 (Ten-4) is specifically expressed in the quiescent satellite cells; however, its cellular and molecular functions remain unknown. We therefore aimed to elucidate the function of Ten-4 in muscle satellite cells. In the tibialis anterior (TA) muscle of Ten-4-deficient mice, the number and the size of myofibers, as well as the population of satellite cells, were reduced with/without induction of muscle regeneration. Furthermore, we found an accelerated activation of satellite cells in the regenerated Ten-4-deficient TA muscle. The cell culture analysis using primary satellite cells showed that Ten-4 suppressed the progression of myogenic differentiation. Together, our findings revealed that Ten-4 functions as a crucial player in maintaining the quiescence of muscle satellite cells.

  8. Cytoglobin modulates myogenic progenitor cell viability and muscle regeneration.

    PubMed

    Singh, Sarvjeet; Canseco, Diana C; Manda, Shilpa M; Shelton, John M; Chirumamilla, Rajendra R; Goetsch, Sean C; Ye, Qiu; Gerard, Robert D; Schneider, Jay W; Richardson, James A; Rothermel, Beverly A; Mammen, Pradeep P A

    2014-01-07

    Mammalian skeletal muscle can remodel, repair, and regenerate itself by mobilizing satellite cells, a resident population of myogenic progenitor cells. Muscle injury and subsequent activation of myogenic progenitor cells is associated with oxidative stress. Cytoglobin is a hemoprotein expressed in response to oxidative stress in a variety of tissues, including striated muscle. In this study, we demonstrate that cytoglobin is up-regulated in activated myogenic progenitor cells, where it localizes to the nucleus and contributes to cell viability. siRNA-mediated depletion of cytoglobin from C2C12 myoblasts increased levels of reactive oxygen species and apoptotic cell death both at baseline and in response to stress stimuli. Conversely, overexpression of cytoglobin reduced reactive oxygen species levels, caspase activity, and cell death. Mice in which cytoglobin was knocked out specifically in skeletal muscle were generated to examine the role of cytoglobin in vivo. Myogenic progenitor cells isolated from these mice were severely deficient in their ability to form myotubes as compared with myogenic progenitor cells from wild-type littermates. Consistent with this finding, the capacity for muscle regeneration was severely impaired in mice deficient for skeletal-muscle cytoglobin. Collectively, these data demonstrate that cytoglobin serves an important role in muscle repair and regeneration.

  9. Reciprocal interaction between TRAF6 and notch signaling regulates adult myofiber regeneration upon injury.

    PubMed

    Hindi, Sajedah M; Paul, Pradyut K; Dahiya, Saurabh; Mishra, Vivek; Bhatnagar, Shephali; Kuang, Shihuan; Choi, Yongwon; Kumar, Ashok

    2012-12-01

    Skeletal muscle is a postmitotic tissue that repairs and regenerates through activation of a population of stem-cell-like satellite cells. However, signaling mechanisms governing adult skeletal muscle regeneration remain less understood. In the present study, we have investigated the role of tumor necrosis factor (TNF) receptor-associated factor 6 (TRAF6), an adaptor protein involved in receptor-mediated activation of multiple signaling pathways in regeneration of adult myofibers. Skeletal muscle-specific depletion of TRAF6 in mice (TRAF6(mko)) improved regeneration of myofibers upon injury with a concomitant increase in the number of satellite cells and activation of the Notch signaling pathway. Ex vivo cultures of TRAF6(mko) myofiber explants demonstrated an increase in the proliferative capacity of myofiber-associated satellite cells accompanied by an upregulation of Notch ligands. Deletion of TRAF6 also inhibited the activity of transcription factor NF-κB and the expression of inflammatory cytokines and augmented the M2c macrophage phenotype in injured muscle tissues. Collectively, our study demonstrates that specific inhibition of TRAF6 improves satellite cell activation and skeletal muscle regeneration through upregulation of Notch signaling and reducing the inflammatory repertoire.

  10. Drug-induced regeneration in adult mice

    PubMed Central

    Zhang, Yong; Strehin, Iossif; Bedelbaeva, Khamilia; Gourevitch, Dmitri; Clark, Lise; Leferovich, John; Messersmith, Phillip B.; Heber-Katz, Ellen

    2015-01-01

    Whereas amphibians regenerate lost appendages spontaneously, mammals generally form scars over the injury site through the process of wound repair. The MRL mouse strain is an exception among mammals because it shows a spontaneous regenerative healing trait and so can be used to investigate proregenerative interventions in mammals. We report that hypoxia-inducible factor 1α (HIF-1α) is a central molecule in the process of regeneration in adult MRL mice. The degradation of HIF-1α protein, which occurs under normoxic conditions, is mediated by prolyl hydroxylases (PHDs). We used the drug 1,4-dihydrophenonthrolin-4-one-3-carboxylic acid (1,4-DPCA), a PHD inhibitor, to stabilize constitutive expression of HIF-1α protein. A locally injectable hydrogel containing 1,4-DPCA was designed to achieve controlled delivery of the drug over 4 to 10 days. Subcutaneous injection of the 1,4-DPCA/hydrogel into Swiss Webster mice that do not show a regenerative phenotype increased stable expression of HIF-1α protein over 5 days, providing a functional measure of drug release in vivo. Multiple peripheral subcutaneous injections of the 1,4-DPCA/hydrogel over a 10-day period led to regenerative wound healing in Swiss Webster mice after ear hole punch injury. Increased expression of the HIF-1α protein may provide a starting point for future studies on regeneration in mammals. PMID:26041709

  11. Electric fish: new insights into conserved processes of adult tissue regeneration.

    PubMed

    Unguez, Graciela A

    2013-07-01

    Biology is replete with examples of regeneration, the process that allows animals to replace or repair cells, tissues and organs. As on land, vertebrates in aquatic environments experience the occurrence of injury with varying frequency and to different degrees. Studies demonstrate that ray-finned fishes possess a very high capacity to regenerate different tissues and organs when they are adults. Among fishes that exhibit robust regenerative capacities are the neotropical electric fishes of South America (Teleostei: Gymnotiformes). Specifically, adult gymnotiform electric fishes can regenerate injured brain and spinal cord tissues and restore amputated body parts repeatedly. We have begun to identify some aspects of the cellular and molecular mechanisms of tail regeneration in the weakly electric fish Sternopygus macrurus (long-tailed knifefish) with a focus on regeneration of skeletal muscle and the muscle-derived electric organ. Application of in vivo microinjection techniques and generation of myogenic stem cell markers are beginning to overcome some of the challenges owing to the limitations of working with non-genetic animal models with extensive regenerative capacity. This review highlights some aspects of tail regeneration in S. macrurus and discusses the advantages of using gymnotiform electric fishes to investigate the cellular and molecular mechanisms that produce new cells during regeneration in adult vertebrates.

  12. Electric fish: new insights into conserved processes of adult tissue regeneration

    PubMed Central

    Unguez, Graciela A.

    2013-01-01

    Summary Biology is replete with examples of regeneration, the process that allows animals to replace or repair cells, tissues and organs. As on land, vertebrates in aquatic environments experience the occurrence of injury with varying frequency and to different degrees. Studies demonstrate that ray-finned fishes possess a very high capacity to regenerate different tissues and organs when they are adults. Among fishes that exhibit robust regenerative capacities are the neotropical electric fishes of South America (Teleostei: Gymnotiformes). Specifically, adult gymnotiform electric fishes can regenerate injured brain and spinal cord tissues and restore amputated body parts repeatedly. We have begun to identify some aspects of the cellular and molecular mechanisms of tail regeneration in the weakly electric fish Sternopygus macrurus (long-tailed knifefish) with a focus on regeneration of skeletal muscle and the muscle-derived electric organ. Application of in vivo microinjection techniques and generation of myogenic stem cell markers are beginning to overcome some of the challenges owing to the limitations of working with non-genetic animal models with extensive regenerative capacity. This review highlights some aspects of tail regeneration in S. macrurus and discusses the advantages of using gymnotiform electric fishes to investigate the cellular and molecular mechanisms that produce new cells during regeneration in adult vertebrates. PMID:23761473

  13. Crustacean muscles: atrophy and regeneration during molting

    SciTech Connect

    Mykles, D.L.; Skinner, D.M.

    1981-01-01

    The ultrastructural basis of atrophy of claw closer muscle of the land crab and the organization of myofibrils and sacroplasmic reticulum during the hydrolysis of protein that occurs during proecdysis was examined. The changes that occur in contractile proteins during claw muscle atrophy and the involvement of Ca/sup 2 +/-dependent proteinases (CDP) in myofilament degradation were investigated. (ACR)

  14. Role of reactive oxygen species in the defective regeneration seen in aging muscle.

    PubMed

    Vasilaki, Aphrodite; Jackson, Malcolm J

    2013-12-01

    The ability of muscles to regenerate successfully following damage diminishes with age and this appears to be a major contributor to the development of muscle weakness and physical frailty. Successful muscle regeneration is dependent on appropriate reinnervation of regenerating muscle. Age-related changes in the interactions between nerve and muscle are poorly understood but may play a major role in the defective regeneration. During aging there is defective redox homeostasis and an accumulation of oxidative damage in nerve and muscle that may contribute to defective regeneration. The aim of this review is to summarise the evidence that abnormal reactive oxygen species (ROS) generation in nerve and/or muscle may be responsible for the defective regeneration that contributes to the degeneration of skeletal muscle observed during aging. Identifying the importance of ROS generation in skeletal muscle during aging could have fundamental implications for interventions to prevent muscle degeneration and treatments to reverse the age-related decline in muscle mass and function.

  15. Regulatory interactions between muscle and the immune system during muscle regeneration.

    PubMed

    Tidball, James G; Villalta, S Armando

    2010-05-01

    Recent discoveries reveal complex interactions between skeletal muscle and the immune system that regulate muscle regeneration. In this review, we evaluate evidence that indicates that the response of myeloid cells to muscle injury promotes muscle regeneration and growth. Acute perturbations of muscle activate a sequence of interactions between muscle and inflammatory cells. The initial inflammatory response is a characteristic Th1 inflammatory response, first dominated by neutrophils and subsequently by CD68(+) M1 macrophages. M1 macrophages can propagate the Th1 response by releasing proinflammatory cytokines and cause further tissue damage through the release of nitric oxide. Myeloid cells in the early Th1 response stimulate the proliferative phase of myogenesis through mechanisms mediated by TNF-alpha and IL-6; experimental prolongation of their presence is associated with delayed transition to the early differentiation stage of myogenesis. Subsequent invasion by CD163(+)/CD206(+) M2 macrophages attenuates M1 populations through the release of anti-inflammatory cytokines, including IL-10. M2 macrophages play a major role in promoting growth and regeneration; their absence greatly slows muscle growth following injury or modified use and inhibits muscle differentiation and regeneration. Chronic muscle injury leads to profiles of macrophage invasion and function that differ from acute injuries. For example, mdx muscular dystrophy yields invasion of muscle by M1 macrophages, but their early invasion is accompanied by a subpopulation of M2a macrophages. M2a macrophages are IL-4 receptor(+)/CD206(+) cells that reduce cytotoxicity of M1 macrophages. Subsequent invasion of dystrophic muscle by M2c macrophages is associated with progression of the regenerative phase in pathophysiology. Together, these findings show that transitions in macrophage phenotype are an essential component of muscle regeneration in vivo following acute or chronic muscle damage.

  16. Regulatory interactions between muscle and the immune system during muscle regeneration

    PubMed Central

    Villalta, S. Armando

    2010-01-01

    Recent discoveries reveal complex interactions between skeletal muscle and the immune system that regulate muscle regeneration. In this review, we evaluate evidence that indicates that the response of myeloid cells to muscle injury promotes muscle regeneration and growth. Acute perturbations of muscle activate a sequence of interactions between muscle and inflammatory cells. The initial inflammatory response is a characteristic Th1 inflammatory response, first dominated by neutrophils and subsequently by CD68+ M1 macrophages. M1 macrophages can propagate the Th1 response by releasing proinflammatory cytokines and cause further tissue damage through the release of nitric oxide. Myeloid cells in the early Th1 response stimulate the proliferative phase of myogenesis through mechanisms mediated by TNF-α and IL-6; experimental prolongation of their presence is associated with delayed transition to the early differentiation stage of myogenesis. Subsequent invasion by CD163+/CD206+ M2 macrophages attenuates M1 populations through the release of anti-inflammatory cytokines, including IL-10. M2 macrophages play a major role in promoting growth and regeneration; their absence greatly slows muscle growth following injury or modified use and inhibits muscle differentiation and regeneration. Chronic muscle injury leads to profiles of macrophage invasion and function that differ from acute injuries. For example, mdx muscular dystrophy yields invasion of muscle by M1 macrophages, but their early invasion is accompanied by a subpopulation of M2a macrophages. M2a macrophages are IL-4 receptor+/CD206+ cells that reduce cytotoxicity of M1 macrophages. Subsequent invasion of dystrophic muscle by M2c macrophages is associated with progression of the regenerative phase in pathophysiology. Together, these findings show that transitions in macrophage phenotype are an essential component of muscle regeneration in vivo following acute or chronic muscle damage. PMID:20219869

  17. PPARδ regulates satellite cell proliferation and skeletal muscle regeneration

    PubMed Central

    2011-01-01

    Peroxisome proliferator-activated receptors (PPARs) are a class of nuclear receptors that play important roles in development and energy metabolism. Whereas PPARδ has been shown to regulate mitochondrial biosynthesis and slow-muscle fiber types, its function in skeletal muscle progenitors (satellite cells) is unknown. Since constitutive mutation of Pparδ leads to embryonic lethality, we sought to address this question by conditional knockout (cKO) of Pparδ using Myf5-Cre/Pparδflox/flox alleles to ablate PPARδ in myogenic progenitor cells. Although Pparδ-cKO mice were born normally and initially displayed no difference in body weight, muscle size or muscle composition, they later developed metabolic syndrome, which manifested as increased body weight and reduced response to glucose challenge at age nine months. Pparδ-cKO mice had 40% fewer satellite cells than their wild-type littermates, and these satellite cells exhibited reduced growth kinetics and proliferation in vitro. Furthermore, regeneration of Pparδ-cKO muscles was impaired after cardiotoxin-induced injury. Gene expression analysis showed reduced expression of the Forkhead box class O transcription factor 1 (FoxO1) gene in Pparδ-cKO muscles under both quiescent and regenerating conditions, suggesting that PPARδ acts through FoxO1 in regulating muscle progenitor cells. These results support a function of PPARδ in regulating skeletal muscle metabolism and insulin sensitivity, and they establish a novel role of PPARδ in muscle progenitor cells and postnatal muscle regeneration. PMID:22040534

  18. Biomimetic Scaffolds for Regeneration of Volumetric Muscle Loss in Skeletal Muscle Injuries

    PubMed Central

    Grasman, Jonathan M.; Zayas, Michelle J.; Page, Ray; Pins, George D.

    2015-01-01

    Skeletal muscle injuries typically result from traumatic incidents such as combat injuries where soft-tissue extremity injuries are present in one of four cases. Further, about 4.5 million reconstructive surgical procedures are performed annually as a result of car accidents, cancer ablation, or cosmetic procedures. These combat- and trauma-induced skeletal muscle injuries are characterized by volumetric muscle loss (VML), which significantly reduces the functionality of the injured muscle. While skeletal muscle has an innate repair mechanism, it is unable to compensate for VML injuries because large amounts of tissue including connective tissue and basement membrane are removed or destroyed. This results in in a significant need to develop off-the-shelf biomimetic scaffolds to direct skeletal muscle regeneration. Here, the structure and organization of native skeletal muscle tissue is described in order to reveal clear design parameters that are necessary for scaffolds to mimic in order to successfully regenerate muscular tissue. We review the literature with respect to the materials and methodologies used to develop scaffolds for skeletal muscle tissue regeneration as well as the limitations of these materials. We further discuss the variety of cell sources and different injury models to provide some context for the multiple approaches used to evaluate these scaffold materials. Recent findings are highlighted to address the state of the field and directions are outlined for future strategies, both in scaffold design and in the use of different injury models to evaluate these materials, for regenerating functional skeletal muscle. PMID:26219862

  19. Abcg2 labels multiple cell types in skeletal muscle and participates in muscle regeneration

    PubMed Central

    Doyle, Michelle J.; Zhou, Sheng; Tanaka, Kathleen Kelly; Pisconti, Addolorata; Farina, Nicholas H.; Sorrentino, Brian P.

    2011-01-01

    Skeletal muscle contains progenitor cells (satellite cells) that maintain and repair muscle. It also contains muscle side population (SP) cells, which express Abcg2 and may participate in muscle regeneration or may represent a source of satellite cell replenishment. In Abcg2-null mice, the SP fraction is lost in skeletal muscle, although the significance of this loss was previously unknown. We show that cells expressing Abcg2 increased upon injury and that muscle regeneration was impaired in Abcg2-null mice, resulting in fewer centrally nucleated myofibers, reduced myofiber size, and fewer satellite cells. Additionally, using genetic lineage tracing, we demonstrate that the progeny of Abcg2-expressing cells contributed to multiple cell types within the muscle interstitium, primarily endothelial cells. After injury, Abcg2 progeny made a minor contribution to regenerated myofibers. Furthermore, Abcg2-labeled cells increased significantly upon injury and appeared to traffic to muscle from peripheral blood. Together, these data suggest an important role for Abcg2 in positively regulating skeletal muscle regeneration. PMID:21949413

  20. Human Satellite Cell Transplantation and Regeneration from Diverse Skeletal Muscles

    PubMed Central

    Xu, Xiaoti; Wilschut, Karlijn J.; Kouklis, Gayle; Tian, Hua; Hesse, Robert; Garland, Catharine; Sbitany, Hani; Hansen, Scott; Seth, Rahul; Knott, P. Daniel; Hoffman, William Y.; Pomerantz, Jason H.

    2015-01-01

    Summary Identification of human satellite cells that fulfill muscle stem cell criteria is an unmet need in regenerative medicine. This hurdle limits understanding how closely muscle stem cell properties are conserved among mice and humans and hampers translational efforts in muscle regeneration. Here, we report that PAX7 satellite cells exist at a consistent frequency of 2–4 cells/mm of fiber in muscles of the human trunk, limbs, and head. Xenotransplantation into mice of 50–70 fiber-associated, or 1,000–5,000 FACS-enriched CD56+/CD29+ human satellite cells led to stable engraftment and formation of human-derived myofibers. Human cells with characteristic PAX7, CD56, and CD29 expression patterns populated the satellite cell niche beneath the basal lamina on the periphery of regenerated fibers. After additional injury, transplanted satellite cells robustly regenerated to form hundreds of human-derived fibers. Together, these findings conclusively delineate a source of bona-fide endogenous human muscle stem cells that will aid development of clinical applications. PMID:26352798

  1. Human Satellite Cell Transplantation and Regeneration from Diverse Skeletal Muscles.

    PubMed

    Xu, Xiaoti; Wilschut, Karlijn J; Kouklis, Gayle; Tian, Hua; Hesse, Robert; Garland, Catharine; Sbitany, Hani; Hansen, Scott; Seth, Rahul; Knott, P Daniel; Hoffman, William Y; Pomerantz, Jason H

    2015-09-08

    Identification of human satellite cells that fulfill muscle stem cell criteria is an unmet need in regenerative medicine. This hurdle limits understanding how closely muscle stem cell properties are conserved among mice and humans and hampers translational efforts in muscle regeneration. Here, we report that PAX7 satellite cells exist at a consistent frequency of 2-4 cells/mm of fiber in muscles of the human trunk, limbs, and head. Xenotransplantation into mice of 50-70 fiber-associated, or 1,000-5,000 FACS-enriched CD56(+)/CD29(+) human satellite cells led to stable engraftment and formation of human-derived myofibers. Human cells with characteristic PAX7, CD56, and CD29 expression patterns populated the satellite cell niche beneath the basal lamina on the periphery of regenerated fibers. After additional injury, transplanted satellite cells robustly regenerated to form hundreds of human-derived fibers. Together, these findings conclusively delineate a source of bona-fide endogenous human muscle stem cells that will aid development of clinical applications.

  2. zebraflash transgenic lines for in vivo bioluminescence imaging of stem cells and regeneration in adult zebrafish.

    PubMed

    Chen, Chen-Hui; Durand, Ellen; Wang, Jinhu; Zon, Leonard I; Poss, Kenneth D

    2013-12-01

    The zebrafish has become a standard model system for stem cell and tissue regeneration research, based on powerful genetics, high tissue regenerative capacity and low maintenance costs. Yet, these studies can be challenged by current limitations of tissue visualization techniques in adult animals. Here we describe new imaging methodology and present several ubiquitous and tissue-specific luciferase-based transgenic lines, which we have termed zebraflash, that facilitate the assessment of regeneration and engraftment in freely moving adult zebrafish. We show that luciferase-based live imaging reliably estimates muscle quantity in an internal organ, the heart, and can longitudinally follow cardiac regeneration in individual animals after major injury. Furthermore, luciferase-based detection enables visualization and quantification of engraftment in live recipients of transplanted hematopoietic stem cell progeny, with advantages in sensitivity and gross spatial resolution over fluorescence detection. Our findings present a versatile resource for monitoring and dissecting vertebrate stem cell and regeneration biology.

  3. Regeneration and myogenic cell proliferation correlate with taurine levels in dystrophin- and MyoD-deficient muscles.

    PubMed

    McIntosh, L M; Garrett, K L; Megeney, L; Rudnicki, M A; Anderson, J E

    1998-10-01

    This study coupled proton magnetic resonance spectroscopy (1H-NMR) and in situ hybridization plus autoradiography in a novel examination of different phenotypes of adult myogenesis that arise from genetic disruptions in mice. Study of muscle extracts from normal and dystrophin-deficient mdx limb and diaphragm muscles confirmed our previous findings linking taurine and muscle regeneration at the peak of damage and repair. 1H-NMR distinguished biochemical differences in regenerating muscles that were consistent with the extent of repair in three strains: mdx dystrophic mice; MyoD(-/-) mice that lack expression of the early myogenic regulatory gene MyoD; and a double-mutant mdx:MyoD(-/-) strain lacking expression of both MyoD and dystrophin. We tested the hypothesis that differences in spectra according to genotype and the regeneration phenotype are related specifically to proliferation by committed myogenic precursor cells. 1H-NMR distinguished the three mutant strains: Taurine was highest in mdx muscles, with the phenotype of most effective regeneration; lowest in MyoD(-/-) muscles, with the least effective formation of new muscle in repair, as reported previously; and intermediate in double-mutant muscles, now reported to show an intermediate repair phenotype. The early and late muscle precursors (mpcs) expressing myf5 and myogenin were examined for proliferation. Eighteen percent of mdx myf5-positive mpcs were proliferative, whereas myf5-positive mpcs did not proliferate in regenerating muscles that lacked MyoD expression. By contrast, whereas 30% of myogenin-positive mpcs were proliferative in mdx muscles, almost none were proliferative in MyoD(-/-) muscles, and 12% were proliferative in double-mutant muscles. Therefore, the extent of accumulated structural regeneration, taurine levels, and proliferation of late mpc (expressing myogenin) were congruent across genotypes. Proliferation by early mpc (expressing myf5) was inhibited by the lack of MyoD expression

  4. Adult motor axons preferentially reinnervate predegenerated muscle nerve.

    PubMed

    Abdullah, M; O'Daly, A; Vyas, A; Rohde, C; Brushart, T M

    2013-11-01

    Preferential motor reinnervation (PMR) is the tendency for motor axons regenerating after repair of mixed nerve to reinnervate muscle nerve and/or muscle rather than cutaneous nerve or skin. PMR may occur in response to the peripheral nerve pathway alone in juvenile rats (Brushart, 1993; Redett et al., 2005), yet the ability to identify and respond to specific pathway markers is reportedly lost in adults (Uschold et al., 2007). The experiments reported here evaluate the relative roles of pathway and end organ in the genesis of PMR in adult rats. Fresh and 2-week predegenerated femoral nerve grafts were transferred in correct or reversed alignment to replace the femoral nerves of previously unoperated Lewis rats. After 8 weeks of regeneration the motoneurons projecting through the grafts to recipient femoral cutaneous and muscle branches and their adjacent end organs were identified by retrograde labeling. Motoneuron counts were subjected to Poisson regression analysis to determine the relative roles of pathway and end organ identity in generating PMR. Transfer of fresh grafts did not result in PMR, whereas substantial PMR was observed when predegenerated grafts were used. Similarly, the pathway through which motoneurons reached the muscle had a significant impact on PMR when grafts were predegenerated, but not when they were fresh. Comparison of the relative roles of pathway and end organ in generating PMR revealed that neither could be shown to be more important than the other. These experiments demonstrate unequivocally that adult muscle nerve and cutaneous nerve differ in qualities that can be detected by regenerating adult motoneurons and that can modify their subsequent behavior. They also reveal that two weeks of Wallerian degeneration modify the environment in the graft from one that provides no modality-specific cues for motor neurons to one that actively promotes PMR.

  5. H19 controls reactivation of the imprinted gene network during muscle regeneration.

    PubMed

    Martinet, Clémence; Monnier, Paul; Louault, Yann; Benard, Matthieu; Gabory, Anne; Dandolo, Luisa

    2016-03-15

    The H19 locus controls fetal growth by regulating expression of several genes from the imprinted gene network (IGN). H19 is fully repressed after birth, except in skeletal muscle. Using loss-of-function H19(Δ3) mice, we investigated the function of H19 in adult muscle. Mutant muscles display hypertrophy and hyperplasia, with increased Igf2 and decreased myostatin (Mstn) expression. Many imprinted genes are expressed in muscle stem cells or satellite cells. Unexpectedly, the number of satellite cells was reduced by 50% in H19(Δ3) muscle fibers. This reduction occurred after postnatal day 21, suggesting a link with their entry into quiescence. We investigated the biological function of these mutant satellite cells in vivo using a regeneration assay induced by multiple injections of cardiotoxin. Surprisingly, despite their reduced number, the self-renewal capacity of these cells is fully retained in the absence of H19. In addition, we observed a better regeneration potential of the mutant muscles, with enhanced expression of several IGN genes and genes from the IGF pathway.

  6. Skeletal muscle hypertrophy and regeneration: interplay between the myogenic regulatory factors (MRFs) and insulin-like growth factors (IGFs) pathways.

    PubMed

    Zanou, Nadège; Gailly, Philippe

    2013-11-01

    Adult skeletal muscle can regenerate in response to muscle damage. This ability is conferred by the presence of myogenic stem cells called satellite cells. In response to stimuli such as injury or exercise, these cells become activated and express myogenic regulatory factors (MRFs), i.e., transcription factors of the myogenic lineage including Myf5, MyoD, myogenin, and Mrf4 to proliferate and differentiate into myofibers. The MRF family of proteins controls the transcription of important muscle-specific proteins such as myosin heavy chain and muscle creatine kinase. Different growth factors are secreted during muscle repair among which insulin-like growth factors (IGFs) are the only ones that promote both muscle cell proliferation and differentiation and that play a key role in muscle regeneration and hypertrophy. Different isoforms of IGFs are expressed during muscle repair: IGF-IEa, IGF-IEb, or IGF-IEc (also known as mechano growth factor, MGF) and IGF-II. MGF is expressed first and is observed in satellite cells and in proliferating myoblasts whereas IGF-Ia and IGF-II expression occurs at the state of muscle fiber formation. Interestingly, several studies report the induction of MRFs in response to IGFs stimulation. Inversely, IGFs expression may also be regulated by MRFs. Various mechanisms are proposed to support these interactions. In this review, we describe the general process of muscle hypertrophy and regeneration and decipher the interactions between the two groups of factors involved in the process.

  7. Fundamental differences in dedifferentiation and stem cell recruitment during skeletal muscle regeneration in two salamander species.

    PubMed

    Sandoval-Guzmán, Tatiana; Wang, Heng; Khattak, Shahryar; Schuez, Maritta; Roensch, Kathleen; Nacu, Eugeniu; Tazaki, Akira; Joven, Alberto; Tanaka, Elly M; Simon, András

    2014-02-06

    Salamanders regenerate appendages via a progenitor pool called the blastema. The cellular mechanisms underlying regeneration of muscle have been much debated but have remained unclear. Here we applied Cre-loxP genetic fate mapping to skeletal muscle during limb regeneration in two salamander species, Notophthalmus viridescens (newt) and Ambystoma mexicanum (axolotl). Remarkably, we found that myofiber dedifferentiation is an integral part of limb regeneration in the newt, but not in axolotl. In the newt, myofiber fragmentation results in proliferating, PAX7(-) mononuclear cells in the blastema that give rise to the skeletal muscle in the new limb. In contrast, myofibers in axolotl do not generate proliferating cells, and do not contribute to newly regenerated muscle; instead, resident PAX7(+) cells provide the regeneration activity. Our results therefore show significant diversity in limb muscle regeneration mechanisms among salamanders and suggest that multiple strategies may be feasible for inducing regeneration in other species, including mammals.

  8. Asymmetric division of clonal muscle stem cells coordinates muscle regeneration in vivo.

    PubMed

    Gurevich, David B; Nguyen, Phong Dang; Siegel, Ashley L; Ehrlich, Ophelia V; Sonntag, Carmen; Phan, Jennifer M N; Berger, Silke; Ratnayake, Dhanushika; Hersey, Lucy; Berger, Joachim; Verkade, Heather; Hall, Thomas E; Currie, Peter D

    2016-07-08

    Skeletal muscle is an example of a tissue that deploys a self-renewing stem cell, the satellite cell, to effect regeneration. Recent in vitro studies have highlighted a role for asymmetric divisions in renewing rare "immortal" stem cells and generating a clonal population of differentiation-competent myoblasts. However, this model currently lacks in vivo validation. We define a zebrafish muscle stem cell population analogous to the mammalian satellite cell and image the entire process of muscle regeneration from injury to fiber replacement in vivo. This analysis reveals complex interactions between satellite cells and both injured and uninjured fibers and provides in vivo evidence for the asymmetric division of satellite cells driving both self-renewal and regeneration via a clonally restricted progenitor pool.

  9. Repression of inactive motor nerve terminals in partially denervated rat muscle after regeneration of active motor axons.

    PubMed Central

    Ribchester, R R; Taxt, T

    1984-01-01

    The fourth deep lumbrical muscle in the hind foot of adult rats was partially denervated by crushing the sural nerve (s.n.). The denervated muscle fibres became completely reinnervated by sprouts from lateral plantar nerve (l.p.n.) motor axons. By about 20 days after the nerve crush, s.n. motor axons started to reinnervate the muscle. In control muscles, a small proportion of the muscle fibres--about 2.5% of the muscle per motor unit--was reinnervated by s.n. motor axons over the following 20 days. Hence the regenerating terminals were able to re-establish functional synapses, despite the fact that all the muscle fibres were functionally innervated by l.p.n. terminals. When nerve impulse conduction in the l.p.n. was blocked with tetrodotoxin for up to 2 weeks, starting from the time when s.n. axons returned to the muscle, s.n. motor axons retrieved a much larger proportion of the muscle fibres--about 6.5% of the muscle per motor unit. There was a concomitant decrease in the tension produced by the sprouted l.p.n. motor axons. Intracellular recordings showed that many muscle fibres became innervated exclusively by regenerated s.n. motor nerve terminals. Measurements of end-plate potentials suggested that l.p.n. sprouts and the original nerve terminals were eliminated non-selectively. These results suggest that regenerating, active motor nerve terminals have an additional competitive advantage in reinnervating innervated muscles, if the intact terminals are inactive. When the l.p.n. was cut, rather than blocked, extensive reinnervation by the s.n. occurred-about 30% of the muscle per motor unit. This suggests that the absence of an intact nerve terminal in the motor end-plate provides a stronger stimulus than inactivity for synapse formation by regenerating motor axons. PMID:6707966

  10. Potential of laryngeal muscle regeneration using induced pluripotent stem cell-derived skeletal muscle cells.

    PubMed

    Dirja, Bayu Tirta; Yoshie, Susumu; Ikeda, Masakazu; Imaizumi, Mitsuyoshi; Nakamura, Ryosuke; Otsuki, Koshi; Nomoto, Yukio; Wada, Ikuo; Hazama, Akihiro; Omori, Koichi

    2016-01-01

    Conclusion Induced pluripotent stem (iPS) cells may be a new potential cell source for laryngeal muscle regeneration in the treatment of vocal fold atrophy after recurrent laryngeal nerve paralysis. Objectives Unilateral vocal fold paralysis can lead to degeneration, atrophy, and loss of force of the thyroarytenoid muscle. At present, there are some treatments such as thyroplasty, arytenoid adduction, and vocal fold injection. However, such treatments cannot restore reduced mass of the thyroarytenoid muscle. iPS cells have been recognized as supplying a potential resource for cell transplantation. The aim of this study was to assess the effectiveness of the use of iPS cells for the regeneration of laryngeal muscle through the evaluation of both in vitro and in vivo experiments. Methods Skeletal muscle cells were generated from tdTomato-labeled iPS cells using embryoid body formation. Differentiation into skeletal muscle cells was analyzed by gene expression and immunocytochemistry. The tdTomato-labeled iPS cell-derived skeletal muscle cells were transplanted into the left atrophied thyroarytenoid muscle. To evaluate the engraftment of these cells after transplantation, immunohistochemistry was performed. Results The tdTomato-labeled iPS cells were successfully differentiated into skeletal muscle cells through an in vitro experiment. These cells survived in the atrophied thyroarytenoid muscle after transplantation.

  11. Efficient muscle regeneration after highly haemorrhagic Bothrops alternatus venom injection.

    PubMed

    Garcia Denegri, María Emilia; Teibler, Gladys P; Maruñak, Silvana L; Hernández, David R; Acosta, Ofelia C; Leiva, Laura C

    2016-11-01

    Bothrops alternatus snake venom is particularly characterized for inducing a prominent haemorrhage and affecting hemostasis as a consequence of 43.1% of metallo-proteinases and less than 10% of PLA2 (almost all non-myotoxic phospholipases) in its venomics. In addition, myonecrosis is the major local effect in viper envenoming which might lead to permanent sequela. Then, the rebuilding of the microvasculature at the local injured site acquires significance since represents one of the pivotal stages for subsequent skeletal muscle regeneration either at morphological or functional aspects. Due to the significance played by vasculature in this process, it is important to study by histology and immunohistochemical techniques, the muscular damage and the sequence of skeletal muscle reconstruction (degree of damage, reconstitution of muscle fibres and capillaries). In this work, we injected intramuscularly 50 or 100 μg per mouse of B. alternatus venom in gastrocnemius muscles. We provided a complete description and characterization of the different stages of myogenesis after mild (50 µg) and severe (100 µg) local injury induced by B. alternatus venom toxins. The regeneration was evaluated 24 h, 3, 7, 14 and 28 days after receiving venom injection. Finally, both doses induced an extended necrosis at the site of injection where, when critical steps in the regenerative process are taking place, an efficient tissue rebuilding is achieved. B. alternatus venom is characterized by the high percentage of exclusively class P-III metalloproteinases, and by the lack of class P-I metalloproteinases in its venom composition. This could explain the effectiveness of muscle regeneration after venom injection despite the severity of the initial phase of envenoming.

  12. Epimorphic regeneration approach to tissue replacement in adult mammals

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Urodeles and fetal mammals are capable of impressive epimorphic regeneration in a variety of tissues, whereas the typical default response to injury in adult mammals consists of inflammation and scar tissue formation. One component of epimorphic regeneration is the recruitment of resident progenitor...

  13. The rejuvenating effect of pregnancy on muscle regeneration.

    PubMed

    Falick Michaeli, Tal; Laufer, Neri; Sagiv, Jitka Yehudit; Dreazen, Avigail; Granot, Zvi; Pikarsky, Eli; Bergman, Yehudit; Gielchinsky, Yuval

    2015-08-01

    Aging is characterized by reduced tissue regenerative capacity attributed to a diminished responsiveness of tissue-specific stem cells. With increasing age, resident precursor cells in muscle tissues show a markedly impaired propensity to proliferate in response to damage. However, exposure to factors present in the serum of young mice restores the regenerative capacity of aged precursor cells. As pregnancy represents a unique biological model of a partially shared blood system between young and old organisms, we hypothesized that pregnancy in aged mice would have a rejuvenating effect on the mother. To test this hypothesis, we assessed muscle regeneration in response to injury in young and aged pregnant and nonpregnant mice. Muscle regeneration in the aged pregnant mice was improved relative to that in age-matched nonpregnant mice. The beneficial effect of pregnancy was transient, lasting up to 2 months after delivery, and appeared to be attributable to activation of satellite cells via the Notch signaling pathway, thus supporting the possibility that pregnancy induces activation of aged dormant muscle progenitor cells.

  14. The rejuvenating effect of pregnancy on muscle regeneration

    PubMed Central

    Falick Michaeli, Tal; Laufer, Neri; Sagiv, Jitka Yehudit; Dreazen, Avigail; Granot, Zvi; Pikarsky, Eli; Bergman, Yehudit; Gielchinsky, Yuval

    2015-01-01

    Aging is characterized by reduced tissue regenerative capacity attributed to a diminished responsiveness of tissue-specific stem cells. With increasing age, resident precursor cells in muscle tissues show a markedly impaired propensity to proliferate in response to damage. However, exposure to factors present in the serum of young mice restores the regenerative capacity of aged precursor cells. As pregnancy represents a unique biological model of a partially shared blood system between young and old organisms, we hypothesized that pregnancy in aged mice would have a rejuvenating effect on the mother. To test this hypothesis, we assessed muscle regeneration in response to injury in young and aged pregnant and nonpregnant mice. Muscle regeneration in the aged pregnant mice was improved relative to that in age-matched nonpregnant mice. The beneficial effect of pregnancy was transient, lasting up to 2 months after delivery, and appeared to be attributable to activation of satellite cells via the Notch signaling pathway, thus supporting the possibility that pregnancy induces activation of aged dormant muscle progenitor cells. PMID:25773509

  15. New function of the myostatin/activin type I receptor (ALK4) as a mediator of muscle atrophy and muscle regeneration

    PubMed Central

    Pasteuning-Vuhman, Svitlana; Boertje-van der Meulen, Johanna W.; van Putten, Maaike; Overzier, Maurice; ten Dijke, Peter; Kiełbasa, Szymon M.; Arindrarto, Wibowo; Wolterbeek, Ron; Lezhnina, Ksenia V.; Ozerov, Ivan V.; Aliper, Aleksandr M.; Hoogaars, Willem M.; Aartsma-Rus, Annemieke; Loomans, Cindy J. M.

    2017-01-01

    Skeletal muscle fibrosis and impaired muscle regeneration are major contributors to muscle wasting in Duchenne muscular dystrophy (DMD). Muscle growth is negatively regulated by myostatin (MSTN) and activins. Blockage of these pathways may improve muscle quality and function in DMD. Antisense oligonucleotides (AONs) were designed specifically to block the function of ALK4, a key receptor for the MSTN/activin pathway in skeletal muscle. AON-induced exon skipping resulted in specific Alk4 down-regulation, inhibition of MSTN activity, and increased myoblast differentiation in vitro. Unexpectedly, a marked decrease in muscle mass (10%) was found after Alk4 AON treatment in mdx mice. In line with in vitro results, muscle regeneration was stimulated, and muscle fiber size decreased markedly. Notably, when Alk4 was down-regulated in adult wild-type mice, muscle mass decreased even more. RNAseq analysis revealed dysregulated metabolic functions and signs of muscle atrophy. We conclude that ALK4 inhibition increases myogenesis but also regulates the tight balance of protein synthesis and degradation. Therefore, caution must be used when developing therapies that interfere with MSTN/activin pathways.—Pasteuning-Vuhman, S., Boertje-van der Meulen, J. W., van Putten, M., Overzier, M., ten Dijke, P., Kiełbasa, S. M., Arindrarto, W., Wolterbeek, R., Lezhnina, K. V., Ozerov, I. V., Aliper, A. M., Hoogaars, W. M., Aartsma-Rus, A., Loomans, C. J. M. New function of the myostatin/activin type I receptor (ALK4) as a mediator of muscle atrophy and muscle regeneration. PMID:27733450

  16. Necdin mediates skeletal muscle regeneration by promoting myoblast survival and differentiation

    PubMed Central

    Deponti, Daniela; François, Stéphanie; Baesso, Silvia; Sciorati, Clara; Innocenzi, Anna; Broccoli, Vania; Muscatelli, Françoise; Meneveri, Raffaella; Clementi, Emilio; Cossu, Giulio; Brunelli, Silvia

    2007-01-01

    Regeneration of muscle fibers that are lost during pathological muscle degeneration or after injuries is sustained by the production of new myofibers. An important cell type involved in muscle regeneration is the satellite cell. Necdin is a protein expressed in satellite cell–derived myogenic precursors during perinatal growth. However, its function in myogenesis is not known. We compare transgenic mice that overexpress necdin in skeletal muscle with both wild-type and necdin null mice. After muscle injury the necdin null mice show a considerable defect in muscle healing, whereas mice that overexpress necdin show a substantial increase in myofiber regeneration. We also find that in muscle, necdin increases myogenin expression, accelerates differentiation, and counteracts myoblast apoptosis. Collectively, these data clarify the function and mechanism of necdin in skeletal muscle and show the importance of necdin in muscle regeneration. PMID:17954612

  17. Muscle wasting and impaired muscle regeneration in a murine model of chronic pulmonary inflammation.

    PubMed

    Langen, Ramon C J; Schols, Annemie M W J; Kelders, Marco C J M; van der Velden, Jos L J; Wouters, Emiel F M; Janssen-Heininger, Yvonne M W

    2006-12-01

    Muscle wasting and increased circulating levels of inflammatory cytokines, including TNF-alpha, are common features of chronic obstructive pulmonary disease. To investigate whether inflammation of the lung is responsible for systemic inflammation and muscle wasting, we adopted a mouse model of pulmonary inflammation resulting from directed overexpression of a TNF-alpha transgene controlled by the surfactant protein C (SP-C) promoter. Compared with wild-type mice, SP-C/TNF-alpha mice exhibited increased levels of TNF-alpha in the circulation and increased endogenous TNF-alpha expression in skeletal muscle, potentially reflecting an amplificatory response to circulating TNF-alpha. Decreased muscle and body weights observed in SP-C/TNF-alpha mice were indicative of muscle wasting. Further evaluation of the SP-C/TNF-alpha mouse musculature revealed a decreased muscle regenerative capacity, shown by attenuated myoblast proliferation and differentiation in response to reloading of disuse-atrophied muscle, which may contribute to skeletal muscle wasting. Importantly, incubation of cultured myoblasts with TNF-alpha also resulted in elevated TNF-alpha mRNA levels and inhibition of myoblast differentiation. Collectively, our results demonstrate that chronic pulmonary inflammation results in muscle wasting and impaired muscle regeneration in SP-C/TNF-alpha mice, possibly as a consequence of an amplificatory TNF-alpha expression circuit extending from the lung to skeletal muscle.

  18. Adult stem cells underlying lung regeneration.

    PubMed

    Xian, Wa; McKeon, Frank

    2012-03-01

    Despite the massive toll in human suffering imparted by degenerative lung disease, including COPD, idiopathic pulmonary fibrosis and ARDS, the scientific community has been surprisingly agnostic regarding the potential of lung tissue, and in particular the alveoli, to regenerate. However, there is circumstantial evidence in humans and direct evidence in mice that ARDS triggers robust regeneration of lung tissue rather than irreversible fibrosis. The stem cells responsible for this remarkable regenerative process has garnered tremendous attention, most recently yielding a defined set of cloned human airway stem cells marked by p63 expression but with distinct commitment to differentiated cell types typical of the upper or lower airways, the latter of which include alveoli-like structures in vitro and in vivo. These recent advances in lung regeneration and distal airway stem cells and the potential of associated soluble factors in regeneration must be harnessed for therapeutic options in chronic lung disease.

  19. Cryotherapy Reduces Inflammatory Response Without Altering Muscle Regeneration Process and Extracellular Matrix Remodeling of Rat Muscle

    PubMed Central

    Vieira Ramos, Gracielle; Pinheiro, Clara Maria; Messa, Sabrina Peviani; Delfino, Gabriel Borges; Marqueti, Rita de Cássia; Salvini, Tania de Fátima; Durigan, Joao Luiz Quagliotti

    2016-01-01

    The application of cryotherapy is widely used in sports medicine today. Cooling could minimize secondary hypoxic injury through the reduction of cellular metabolism and injury area. Conflicting results have also suggested cryotherapy could delay and impair the regeneration process. There are no definitive findings about the effects of cryotherapy on the process of muscle regeneration. The aim of the present study was to evaluate the effects of a clinical-like cryotherapy on inflammation, regeneration and extracellular matrix (ECM) remodeling on the Tibialis anterior (TA) muscle of rats 3, 7 and 14 days post-injury. It was observed that the intermittent application of cryotherapy (three 30-minute sessions, every 2 h) in the first 48 h post-injury decreased inflammatory processes (mRNA levels of TNF-α, NF-κB, TGF-β and MMP-9 and macrophage percentage). Cryotherapy did not alter regeneration markers such as injury area, desmin and Myod expression. Despite regulating Collagen I and III and their growth factors, cryotherapy did not alter collagen deposition. In summary, clinical-like cryotherapy reduces the inflammatory process through the decrease of macrophage infiltration and the accumulation of the inflammatory key markers without influencing muscle injury area and ECM remodeling. PMID:26725948

  20. Cryotherapy Reduces Inflammatory Response Without Altering Muscle Regeneration Process and Extracellular Matrix Remodeling of Rat Muscle.

    PubMed

    Vieira Ramos, Gracielle; Pinheiro, Clara Maria; Messa, Sabrina Peviani; Delfino, Gabriel Borges; Marqueti, Rita de Cássia; Salvini, Tania de Fátima; Durigan, Joao Luiz Quagliotti

    2016-01-04

    The application of cryotherapy is widely used in sports medicine today. Cooling could minimize secondary hypoxic injury through the reduction of cellular metabolism and injury area. Conflicting results have also suggested cryotherapy could delay and impair the regeneration process. There are no definitive findings about the effects of cryotherapy on the process of muscle regeneration. The aim of the present study was to evaluate the effects of a clinical-like cryotherapy on inflammation, regeneration and extracellular matrix (ECM) remodeling on the Tibialis anterior (TA) muscle of rats 3, 7 and 14 days post-injury. It was observed that the intermittent application of cryotherapy (three 30-minute sessions, every 2 h) in the first 48 h post-injury decreased inflammatory processes (mRNA levels of TNF-α, NF-κB, TGF-β and MMP-9 and macrophage percentage). Cryotherapy did not alter regeneration markers such as injury area, desmin and Myod expression. Despite regulating Collagen I and III and their growth factors, cryotherapy did not alter collagen deposition. In summary, clinical-like cryotherapy reduces the inflammatory process through the decrease of macrophage infiltration and the accumulation of the inflammatory key markers without influencing muscle injury area and ECM remodeling.

  1. MicroRNA-155 facilitates skeletal muscle regeneration by balancing pro- and anti-inflammatory macrophages

    PubMed Central

    Nie, M; Liu, J; Yang, Q; Seok, H Y; Hu, X; Deng, Z-L; Wang, D-Z

    2016-01-01

    Skeletal muscle has remarkable regeneration capacity and regenerates in response to injury. Muscle regeneration largely relies on muscle stem cells called satellite cells. Satellite cells normally remain quiescent, but in response to injury or exercise they become activated and proliferate, migrate, differentiate, and fuse to form multinucleate myofibers. Interestingly, the inflammatory process following injury and the activation of the myogenic program are highly coordinated, with myeloid cells having a central role in modulating satellite cell activation and regeneration. Here, we show that genetic deletion of microRNA-155 (miR-155) in mice substantially delays muscle regeneration. Surprisingly, miR-155 does not appear to directly regulate the proliferation or differentiation of satellite cells. Instead, miR-155 is highly expressed in myeloid cells, is essential for appropriate activation of myeloid cells, and regulates the balance between pro-inflammatory M1 macrophages and anti-inflammatory M2 macrophages during skeletal muscle regeneration. Mechanistically, we found that miR-155 suppresses SOCS1, a negative regulator of the JAK-STAT signaling pathway, during the initial inflammatory response upon muscle injury. Our findings thus reveal a novel role of miR-155 in regulating initial immune responses during muscle regeneration and provide a novel miRNA target for improving muscle regeneration in degenerative muscle diseases. PMID:27277683

  2. Neuronal intrinsic barriers for axon regeneration in the adult CNS

    PubMed Central

    Sun, Fang; He, Zhigang

    2010-01-01

    A major reason for the devastating and permanent disabilities after spinal cord and other types of CNS injury is the failure of injured axons to regenerate and to re-build the functional circuits. Thus, a long-standing goal has been to develop strategies that could promote axon regeneration and restore functions. Recent studies revealed that simply removing extracellular inhibitory activities is insufficient for successful axon regeneration in the adult CNS. On the other side, evidence from different species and different models is accumulating to support the notion that diminished intrinsic regenerative ability of mature neurons is a major contributor to regeneration failure. This review will summarize the molecular mechanisms regulating intrinsic axon growth capacity in the adult CNS and discuss potential implications for therapeutic strategies. PMID:20418094

  3. UTX in muscle regeneration — the right dose and the right time

    PubMed Central

    Liu, Ling; Rando, Thomas A.

    2016-01-01

    Precise epigenetic modifications in stem cells control developmental programs and cell fate decisions. In particular, the addition or removal of trimethylation of histone 3 lysine 27 (H3K27me3) at lineage-specific genes has been linked to the repression of gene expression, and a precise balance of methyltransferases and demethylases within cells determines H3K27me3 levels. The demethylase UTX is essential for development and tissue homeostasis; however, a role for UTX in stem cell–mediated tissue regeneration is unknown. In this issue of the JCI, Dilworth and colleagues reveal that UTX and its demethylase activity are required in the muscle stem cell lineage for muscle regeneration in response to injury. Specifically, UTX mediates the removal of H3K27me3 in the promoter of the transcription factor myogenin, which regulates myogenic differentiation. The results of this study provide important insight into the contribution of epigenetic regulation in stem cell–mediated regeneration of adult tissues. PMID:26999609

  4. Fate of 3H-thymidine labelled myogenic cells in regeneration of muscle isografts.

    PubMed

    Gutmann, E; Mares, V; Stichová, J

    1976-03-05

    Intact and denervated extensor digitorum longus (EDL) muscles of 20-day-old inbred Lewis-Wistar rats were labelled with 3H-thymidine. Ninety minutes after the injection of the isotope 4.0% of the nuclei were labelled in the intact (i.e. innervated) and 9.6% in the muscles, denervated 3 days before administration of the isotope. The labelled EDL muscles were grafted into the bed of the previously removed EDL muscles of inbred animals and these isografts were studied 30 days later. In the EDL muscles, regenerated from innervated isografts only occasionally labelled endothelial cells were found whereas in the muscles regenerated from denervated isografts also parenchymal muscle nuclei were regularly labelled. The incidence of labelled nuclei in the regenerated EDL muscles was, however, about 20 times lower than in the donor EDL muscles. The presen experiments provide a direct proof of utilization of donor satelite cell nuclei for regeneration in grafted muscle tissue. With respect to the low incidence of labelled nuclei in regenerated EDL muscles, other sources of cells apparently also contribute to the regeneration process.

  5. Stem Cell-Mediated Regeneration of the Adult Brain

    PubMed Central

    Jessberger, Sebastian

    2016-01-01

    Acute or chronic injury of the adult mammalian brain is often associated with persistent functional deficits as its potential for regeneration and capacity to rebuild lost neural structures is limited. However, the discovery that neural stem cells (NSCs) persist throughout life in discrete regions of the brain, novel approaches to induce the formation of neuronal and glial cells, and recently developed strategies to generate tissue for exogenous cell replacement strategies opened novel perspectives how to regenerate the adult brain. Here, we will review recently developed approaches for brain repair and discuss future perspectives that may eventually allow for developing novel treatment strategies in acute and chronic brain injury. PMID:27781019

  6. Regeneration of optic nerve fibers of adult mammals.

    PubMed

    Watanabe, Masami

    2010-09-01

    The pathway from the retina to the brain in mammals provides a well-defined model system for investigation of not only surviving axotomy but also axonal regeneration of injured neurons. Here I introduce our recent works on axonal regeneration in the optic nerve (OpN) of adult cats. Fibers of retinal ganglion cells (RGCs) extend beyond the crush site of OpN with injections of a macrophage stimulator (oxidized galectin-1) or a Rho kinase (ROCK) inhibitor (Y-39983 or Y-27632) while axonal extension is blocked with injection of saline. Elongation of crushed optic fibers, however, is slowed after 2 weeks. Transplantation of peripheral nerve makes RGCs regenerate their transected axons into a graft but regenerated fibers extend only a few mm in the brain. Effectiveness of combination of the drugs and treatments has to be verified in future.

  7. Macrophages are required for adult salamander limb regeneration.

    PubMed

    Godwin, James W; Pinto, Alexander R; Rosenthal, Nadia A

    2013-06-04

    The failure to replace damaged body parts in adult mammals results from a muted growth response and fibrotic scarring. Although infiltrating immune cells play a major role in determining the variable outcome of mammalian wound repair, little is known about the modulation of immune cell signaling in efficiently regenerating species such as the salamander, which can regrow complete body structures as adults. Here we present a comprehensive analysis of immune signaling during limb regeneration in axolotl, an aquatic salamander, and reveal a temporally defined requirement for macrophage infiltration in the regenerative process. Although many features of mammalian cytokine/chemokine signaling are retained in the axolotl, they are more dynamically deployed, with simultaneous induction of inflammatory and anti-inflammatory markers within the first 24 h after limb amputation. Systemic macrophage depletion during this period resulted in wound closure but permanent failure of limb regeneration, associated with extensive fibrosis and disregulation of extracellular matrix component gene expression. Full limb regenerative capacity of failed stumps was restored by reamputation once endogenous macrophage populations had been replenished. Promotion of a regeneration-permissive environment by identification of macrophage-derived therapeutic molecules may therefore aid in the regeneration of damaged body parts in adult mammals.

  8. FGF2-induced effects on transcriptome associated with regeneration competence in adult human fibroblasts

    PubMed Central

    2013-01-01

    Background Adult human fibroblasts grown in low oxygen and with FGF2 supplementation have the capacity to tip the healing outcome of skeletal muscle injury – by favoring regeneration response in vivo over scar formation. Here, we compare the transcriptomes of control adult human dermal fibroblasts and induced regeneration-competent (iRC) fibroblasts to identify transcriptional changes that may be related to their regeneration competence. Results We identified a unique gene-expression profile that characterizes FGF2-induced iRC fibroblast phenotype. Significantly differentially expressed genes due to FGF2 treatment were identified and analyzed to determine overrepresented Gene Ontology terms. Genes belonging to extracellular matrix components, adhesion molecules, matrix remodelling, cytoskeleton, and cytokines were determined to be affected by FGF2 treatment. Conclusions Transcriptome analysis comparing control adult human fibroblasts with FGF2-treated fibroblasts identified functional groups of genes that reflect transcriptional changes potentially contributing to their regeneration competence. This comparative transcriptome analysis should contribute new insights into genes that characterize cells with greater regenerative potential. PMID:24066673

  9. The CHC22 clathrin-GLUT4 transport pathway contributes to skeletal muscle regeneration.

    PubMed

    Hoshino, Sachiko; Sakamoto, Kazuho; Vassilopoulos, Stéphane; Camus, Stéphane M; Griffin, Christine A; Esk, Christopher; Torres, Jorge A; Ohkoshi, Norio; Ishii, Akiko; Tamaoka, Akira; Funke, Birgit H; Kucherlapati, Raju; Margeta, Marta; Rando, Thomas A; Brodsky, Frances M

    2013-01-01

    Mobilization of the GLUT4 glucose transporter from intracellular storage vesicles provides a mechanism for insulin-responsive glucose import into skeletal muscle. In humans, clathrin isoform CHC22 participates in formation of the GLUT4 storage compartment in skeletal muscle and fat. CHC22 function is limited to retrograde endosomal sorting and is restricted in its tissue expression and species distribution compared to the conserved CHC17 isoform that mediates endocytosis and several other membrane traffic pathways. Previously, we noted that CHC22 was expressed at elevated levels in regenerating rat muscle. Here we investigate whether the GLUT4 pathway in which CHC22 participates could play a role in muscle regeneration in humans and we test this possibility using CHC22-transgenic mice, which do not normally express CHC22. We observed that GLUT4 expression is elevated in parallel with that of CHC22 in regenerating skeletal muscle fibers from patients with inflammatory and other myopathies. Regenerating human myofibers displayed concurrent increases in expression of VAMP2, another regulator of GLUT4 transport. Regenerating fibers from wild-type mouse skeletal muscle injected with cardiotoxin also showed increased levels of GLUT4 and VAMP2. We previously demonstrated that transgenic mice expressing CHC22 in their muscle over-sequester GLUT4 and VAMP2 and have defective GLUT4 trafficking leading to diabetic symptoms. In this study, we find that muscle regeneration rates in CHC22 mice were delayed compared to wild-type mice, and myoblasts isolated from these mice did not proliferate in response to glucose. Additionally, CHC22-expressing mouse muscle displayed a fiber type switch from oxidative to glycolytic, similar to that observed in type 2 diabetic patients. These observations implicate the pathway for GLUT4 transport in regeneration of both human and mouse skeletal muscle, and demonstrate a role for this pathway in maintenance of muscle fiber type. Extrapolating

  10. Dermal fibroblasts participate in the formation of new muscle fibres when implanted into regenerating normal mouse muscle.

    PubMed

    Pye, D; Watt, D J

    2001-02-01

    Both in vitro and in vivo studies have described the conversion of fibroblasts to myogenesis when in the presence of dysfunctional myogenic cells. Myogenic conversion of fibroblasts subjected to a normal, as opposed to a diseased muscle environment has only been reported in vitro. The primary aim of this work was to determine if fibroblasts can convert to a myogenic lineage and contribute to new fibre formation when implanted into the regenerating muscle of a normal mouse. Dermal fibroblasts were prepared from neonatal mouse skin and labelled prior to implantation with the fluorescent nuclear marker 4',6-diamidino-2-phenylindole (DAPI). Cells were implanted into muscles of host mice that had been subjected to either cold/crush or minced muscle injury. Some host muscles were x-irradiated to deplete the muscle of endogenous muscle precursor cells. Muscles were removed at 3 wk postimplantation and analysed both histologically and for the presence of DAPI labelled nuclei. Fibres containing DAPI labelled central nuclei indicated that the implanted cells had participated in the regenerative process. Mouse dermal fibroblasts therefore do contribute to muscle fibre formation in regenerating normal mouse muscle but the extent of their contribution is dependent on the nature of the trauma induced in the host muscle. The study also showed that regeneration was more successful in muscles which had not been irradiated, which is contrary to the previous studies where dermal fibroblasts were introduced into myopathic mouse muscle.

  11. Dermal fibroblasts participate in the formation of new muscle fibres when implanted into regenerating normal mouse muscle

    PubMed Central

    PYE, DEBORAH; WATT, DIANA J.

    2001-01-01

    Both in vitro and in vivo studies have described the conversion of fibroblasts to myogenesis when in the presence of dysfunctional myogenic cells. Myogenic conversion of fibroblasts subjected to a normal, as opposed to a diseased muscle environment has only been reported in vitro. The primary aim of this work was to determine if fibroblasts can convert to a myogenic lineage and contribute to new fibre formation when implanted into the regenerating muscle of a normal mouse. Dermal fibroblasts were prepared from neonatal mouse skin and labelled prior to implantation with the fluorescent nuclear marker 4′,6-diamidino-2-phenylindole (DAPI). Cells were implanted into muscles of host mice that had been subjected to either cold/crush or minced muscle injury. Some host muscles were x-irradiated to deplete the muscle of endogenous muscle precursor cells. Muscles were removed at 3 wk postimplantation and analysed both histologically and for the presence of DAPI labelled nuclei. Fibres containing DAPI labelled central nuclei indicated that the implanted cells had participated in the regenerative process. Mouse dermal fibroblasts therefore do contribute to muscle fibre formation in regenerating normal mouse muscle but the extent of their contribution is dependent on the nature of the trauma induced in the host muscle. The study also showed that regeneration was more successful in muscles which had not been irradiated, which is contrary to the previous studies where dermal fibroblasts were introduced into myopathic mouse muscle. PMID:11273041

  12. Impaired muscle regeneration and myoblast differentiation in mice with a muscle-specific KO of IGF-IR.

    PubMed

    Heron-Milhavet, Lisa; Mamaeva, Daria; LeRoith, Derek; Lamb, Ned J; Fernandez, Anne

    2010-10-01

    IGF-I and its receptor IGF-IR are seen as critical effectors of muscle hypertrophy, a notion recently questioned. Using MKR transgenic mice that express a dominant negative IGF-IR only in skeletal muscle, we have examined the role of the IGF-IR signaling in differentiation and repair of muscle fibers after damage-induced muscle regeneration. This process is impaired in MKR muscle, with incomplete regeneration, persistence of infiltrating cells and sustained expression of differentiation markers. Analysis of MKR and WT muscle-derived progenitor stem cells and myoblasts showed twice as many such cells in MKR muscle and an incomplete in vitro differentiation, that is, despite similar levels of myogenin expression, the level of fusion of MKR myoblasts was significantly reduced in comparison to WT myoblasts. These data show IGF-IR signaling is not only required at early hyperplasia stages of muscle differentiation, but also for late stages of myofiber maturation and hypertrophy.

  13. Regulation of myogenesis and skeletal muscle regeneration: effects of oxygen levels on satellite cell activity.

    PubMed

    Chaillou, Thomas; Lanner, Johanna T

    2016-12-01

    Reduced oxygen (O2) levels (hypoxia) are present during embryogenesis and exposure to altitude and in pathologic conditions. During embryogenesis, myogenic progenitor cells reside in a hypoxic microenvironment, which may regulate their activity. Satellite cells are myogenic progenitor cells localized in a local environment, suggesting that the O2 level could affect their activity during muscle regeneration. In this review, we present the idea that O2 levels regulate myogenesis and muscle regeneration, we elucidate the molecular mechanisms underlying myogenesis and muscle regeneration in hypoxia and depict therapeutic strategies using changes in O2 levels to promote muscle regeneration. Severe hypoxia (≤1% O2) appears detrimental for myogenic differentiation in vitro, whereas a 3-6% O2 level could promote myogenesis. Hypoxia impairs the regenerative capacity of injured muscles. Although it remains to be explored, hypoxia may contribute to the muscle damage observed in patients with pathologies associated with hypoxia (chronic obstructive pulmonary disease, and peripheral arterial disease). Hypoxia affects satellite cell activity and myogenesis through mechanisms dependent and independent of hypoxia-inducible factor-1α. Finally, hyperbaric oxygen therapy and transplantation of hypoxia-conditioned myoblasts are beneficial procedures to enhance muscle regeneration in animals. These therapies may be clinically relevant to treatment of patients with severe muscle damage.-Chaillou, T. Lanner, J. T. Regulation of myogenesis and skeletal muscle regeneration: effects of oxygen levels on satellite cell activity.

  14. Botulinum toxin induces muscle paralysis and inhibits bone regeneration in zebrafish.

    PubMed

    Recidoro, Anthony M; Roof, Amanda C; Schmitt, Michael; Worton, Leah E; Petrie, Timothy; Strand, Nicholas; Ausk, Brandon J; Srinivasan, Sundar; Moon, Randall T; Gardiner, Edith M; Kaminsky, Werner; Bain, Steven D; Allan, Christopher H; Gross, Ted S; Kwon, Ronald Y

    2014-11-01

    Intramuscular administration of Botulinum toxin (BTx) has been associated with impaired osteogenesis in diverse conditions of bone formation (eg, development, growth, and healing), yet the mechanisms of neuromuscular-bone crosstalk underlying these deficits have yet to be identified. Motivated by the emerging utility of zebrafish (Danio rerio) as a rapid, genetically tractable, and optically transparent model for human pathologies (as well as the potential to interrogate neuromuscular-mediated bone disorders in a simple model that bridges in vitro and more complex in vivo model systems), in this study, we developed a model of BTx-induced muscle paralysis in adult zebrafish, and we examined its effects on intramembranous ossification during tail fin regeneration. BTx administration induced rapid muscle paralysis in adult zebrafish in a manner that was dose-dependent, transient, and focal, mirroring the paralytic phenotype observed in animal and human studies. During fin regeneration, BTx impaired continued bone ray outgrowth, morphology, and patterning, indicating defects in early osteogenesis. Further, BTx significantly decreased mineralizing activity and crystalline mineral accumulation, suggesting delayed late-stage osteoblast differentiation and/or altered secondary bone apposition. Bone ray transection proximal to the amputation site focally inhibited bone outgrowth in the affected ray, implicating intra- and/or inter-ray nerves in this process. Taken together, these studies demonstrate the potential to interrogate pathological features of BTx-induced osteoanabolic dysfunction in the regenerating zebrafish fin, define the technological toolbox for detecting bone growth and mineralization deficits in this process, and suggest that pathways mediating neuromuscular regulation of osteogenesis may be conserved beyond established mammalian models of bone anabolic disorders.

  15. Unloading stress disturbs muscle regeneration through perturbed recruitment and function of macrophages.

    PubMed

    Kohno, Shohei; Yamashita, Yui; Abe, Tomoki; Hirasaka, Katsuya; Oarada, Motoko; Ohno, Ayako; Teshima-Kondo, Shigetada; Higashibata, Akira; Choi, Inho; Mills, Edward M; Okumura, Yuushi; Terao, Junji; Nikawa, Takeshi

    2012-05-01

    Skeletal muscle is one of the most sensitive tissues to mechanical loading, and unloading inhibits the regeneration potential of skeletal muscle after injury. This study was designed to elucidate the specific effects of unloading stress on the function of immunocytes during muscle regeneration after injury. We examined immunocyte infiltration and muscle regeneration in cardiotoxin (CTX)-injected soleus muscles of tail-suspended (TS) mice. In CTX-injected TS mice, the cross-sectional area of regenerating myofibers was smaller than that of weight-bearing (WB) mice, indicating that unloading delays muscle regeneration following CTX-induced skeletal muscle damage. Delayed infiltration of macrophages into the injured skeletal muscle was observed in CTX-injected TS mice. Neutrophils and macrophages in CTX-injected TS muscle were presented over a longer period at the injury sites compared with those in CTX-injected WB muscle. Disturbance of activation and differentiation of satellite cells was also observed in CTX-injected TS mice. Further analysis showed that the macrophages in soleus muscles were mainly Ly-6C-positive proinflammatory macrophages, with high expression of tumor necrosis factor-α and interleukin-1β, indicating that unloading causes preferential accumulation and persistence of proinflammatory macrophages in the injured muscle. The phagocytic and myotube formation properties of macrophages from CTX-injected TS skeletal muscle were suppressed compared with those from CTX-injected WB skeletal muscle. We concluded that the disturbed muscle regeneration under unloading is due to impaired macrophage function, inhibition of satellite cell activation, and their cooperation.

  16. Changes in contractile activation characteristics of rat fast and slow skeletal muscle fibres during regeneration

    PubMed Central

    Gregorevic, Paul; Plant, David R; Stupka, Nicole; Lynch, Gordon S

    2004-01-01

    Damaged skeletal muscle fibres are replaced with new contractile units via muscle regeneration. Regenerating muscle fibres synthesize functionally distinct isoforms of contractile and regulatory proteins but little is known of their functional properties during the regeneration process. An advantage of utilizing single muscle fibre preparations is that assessment of their function is based on the overall characteristics of the contractile apparatus and regulatory system and as such, these preparations are sensitive in revealing not only coarse, but also subtle functional differences between muscle fibres. We examined the Ca2+- and Sr2+-activated contractile characteristics of permeabilized fibres from rat fast-twitch (extensor digitorum longus) and slow-twitch (soleus) muscles at 7, 14 and 21 days following myotoxic injury, to test the hypothesis that fibres from regenerating fast and slow muscles have different functional characteristics to fibres from uninjured muscles. Regenerating muscle fibres had ∼10% of the maximal force producing capacity (Po) of control (uninjured) fibres, and an altered sensitivity to Ca2+ and Sr2+ at 7 days post-injury. Increased force production and a shift in Ca2+ sensitivity consistent with fibre maturation were observed during regeneration such that Po was restored to 36–45% of that in control fibres by 21 days, and sensitivity to Ca2+ and Sr2+ was similar to that of control (uninjured) fibres. The findings support the hypothesis that regenerating muscle fibres have different contractile activation characteristics compared with mature fibres, and that they adopt properties of mature fast- or slow-twitch muscle fibres in a progressive manner as the regeneration process is completed. PMID:15181161

  17. Oxytocin is an age-specific circulating hormone that is necessary for muscle maintenance and regeneration.

    PubMed

    Elabd, Christian; Cousin, Wendy; Upadhyayula, Pavan; Chen, Robert Y; Chooljian, Marc S; Li, Ju; Kung, Sunny; Jiang, Kevin P; Conboy, Irina M

    2014-06-10

    The regenerative capacity of skeletal muscle declines with age. Previous studies suggest that this process can be reversed by exposure to young circulation; however, systemic age-specific factors responsible for this phenomenon are largely unknown. Here we report that oxytocin--a hormone best known for its role in lactation, parturition and social behaviours--is required for proper muscle tissue regeneration and homeostasis, and that plasma levels of oxytocin decline with age. Inhibition of oxytocin signalling in young animals reduces muscle regeneration, whereas systemic administration of oxytocin rapidly improves muscle regeneration by enhancing aged muscle stem cell activation/proliferation through activation of the MAPK/ERK signalling pathway. We further show that the genetic lack of oxytocin does not cause a developmental defect in muscle but instead leads to premature sarcopenia. Considering that oxytocin is an FDA-approved drug, this work reveals a potential novel and safe way to combat or prevent skeletal muscle ageing.

  18. Oxytocin is an age-specific circulating hormone that is necessary for muscle maintenance and regeneration

    PubMed Central

    Upadhyayula, Pavan; Chen, Robert Y.; Chooljian, Marc S.; Li, Ju; Kung, Sunny; Jiang, Kevin P.; Conboy, Irina M.

    2014-01-01

    The regenerative capacity of skeletal muscle declines with age. Previous studies suggest that this process can be reversed by exposure to young circulation, but systemic age-specific factors responsible for this phenomenon are largely unknown. Here we report that oxytocin- a hormone best known for its role in lactation, parturition, and social behaviors - is required for proper muscle tissue regeneration and homeostasis, and that plasma levels of oxytocin decline with age. Inhibition of oxytocin signaling in young animals reduces muscle regeneration, whereas systemic administration of oxytocin rapidly improves muscle regeneration by enhancing aged muscle stem cell activation/proliferation throughactivation of the MAPK/ERK signalling pathway. We further show that the genetic lack of oxytocin does not cause a developmental defect in muscle, but instead leads to premature sarcopenia. Considering that oxytocin is an FDA approved drug, this work reveals a potential novel and safe way to combat or prevent skeletal muscle aging. PMID:24915299

  19. HDAC4 promotes Pax7-dependent satellite cell activation and muscle regeneration

    PubMed Central

    Choi, Moon-Chang; Ryu, Soyoung; Hao, Rui; Wang, Bin; Kapur, Meghan; Fan, Chen-Ming; Yao, Tso-Pang

    2014-01-01

    During muscle regeneration, the transcription factor Pax7 stimulates the differentiation of satellite cells (SCs) toward the muscle lineage but restricts adipogenesis. Here, we identify HDAC4 as a regulator of Pax7-dependent muscle regeneration. In HDAC4-deficient SCs, the expression of Pax7 and its target genes is reduced. We identify HDAC4-regulated Lix1 as a Pax7 target gene required for SC proliferation. HDAC4 inactivation leads to defective SC proliferation, muscle regeneration, and aberrant lipid accumulation. Further, expression of the brown adipose master regulator Prdm16 and its inhibitory microRNA-133 are also deregulated. Thus, HDAC4 is a novel regulator of Pax7-dependent SC proliferation and potentially fate determination in regenerating muscle. PMID:25205686

  20. A Twist2-dependent progenitor cell contributes to adult skeletal muscle.

    PubMed

    Liu, Ning; Garry, Glynnis A; Li, Stephen; Bezprozvannaya, Svetlana; Sanchez-Ortiz, Efrain; Chen, Beibei; Shelton, John M; Jaichander, Priscilla; Bassel-Duby, Rhonda; Olson, Eric N

    2017-03-01

    Skeletal muscle possesses remarkable regenerative potential due to satellite cells, an injury-responsive stem cell population located beneath the muscle basal lamina that expresses Pax7. By lineage tracing of progenitor cells expressing the Twist2 (Tw2) transcription factor in mice, we discovered a myogenic lineage that resides outside the basal lamina of adult skeletal muscle. Tw2(+) progenitors are molecularly and anatomically distinct from satellite cells, are highly myogenic in vitro, and can fuse with themselves and with satellite cells. Tw2(+) progenitors contribute specifically to type IIb/x myofibres during adulthood and muscle regeneration, and their genetic ablation causes wasting of type IIb myofibres. We show that Tw2 expression maintains progenitor cells in an undifferentiated state that is poised to initiate myogenesis in response to appropriate cues that extinguish Tw2 expression. Tw2-expressing myogenic progenitors represent a previously unrecognized, fibre-type-specific stem cell involved in postnatal muscle growth and regeneration.

  1. Muscle Stem Cells: A Model System for Adult Stem Cell Biology.

    PubMed

    Cornelison, Ddw; Perdiguero, Eusebio

    2017-01-01

    Skeletal muscle stem cells, originally termed satellite cells for their position adjacent to differentiated muscle fibers, are absolutely required for the process of skeletal muscle repair and regeneration. In the last decade, satellite cells have become one of the most studied adult stem cell systems and have emerged as a standard model not only in the field of stem cell-driven tissue regeneration but also in stem cell dysfunction and aging. Here, we provide background in the field and discuss recent advances in our understanding of muscle stem cell function and dysfunction, particularly in the case of aging, and the potential involvement of muscle stem cells in genetic diseases such as the muscular dystrophies.

  2. The need to more precisely define aspects of skeletal muscle regeneration.

    PubMed

    Grounds, Miranda D

    2014-11-01

    A more precise definition of the term 'skeletal muscle regeneration' is required to reduce confusion and misconceptions. In this paper the term is used only for events that follow myofibre necrosis, to result in myogenesis and new muscle formation: other key events include early inflammation and revascularisation, and later fibrosis and re-innervation. The term 'muscle regeneration' is sometimes used casually for situations that do not involve myonecrosis; such as restoration of muscle mass by hypertrophy after atrophy, and other forms of damage to muscle tissue components. These situations are excluded from the definition in this paper which is focussed on mammalian muscles with the long-term aim of clinical translation to enhance new muscle formation after acute or chronic injury or during surgery to replace whole muscles. The paper briefly outlines the cellular events involved in myogenesis during development and post-natal muscle growth, discusses the role of satellite cells in mature normal muscles, and the likely incidence of myofibre necrosis/regeneration in healthy ageing mammals (even when subjected to exercise). The importance of the various components of regeneration is outlined to emphasise that problems in each of these aspects can influence overall new muscle formation; thus care is needed for correct interpretation of altered kinetics. Various markers used to identify regenerating myofibres are critically discussed and, since these can all occur in other conditions, caution is required for accurate interpretation of these cellular events. Finally, clinical situations are outlined where there is a need to enhance skeletal muscle regeneration: these include acute and chronic injuries or transplantation with bioengineering to form new muscles, therapeutic approaches to muscular dystrophies, and comment on proposed stem cell therapies to reduce age-related loss of muscle mass and function. This article is part of a directed issue entitled: Regenerative

  3. Microcurrent Electrical Neuromuscular Stimulation Facilitates Regeneration of Injured Skeletal Muscle in Mice

    PubMed Central

    Fujiya, Hiroto; Ogura, Yuji; Ohno, Yoshitaka; Goto, Ayumi; Nakamura, Ayane; Ohashi, Kazuya; Uematsu, Daiki; Aoki, Haruhito; Musha, Haruki; Goto, Katsumasa

    2015-01-01

    Conservative therapies, mainly resting care for the damaged muscle, are generally used as a treatment for skeletal muscle injuries (such as muscle fragmentation). Several past studies reported that microcurrent electrical neuromuscular stimulation (MENS) facilitates a repair of injured soft tissues and shortens the recovery period. However, the effects of MENS on the regeneration in injured skeletal muscle are still unclear. The purpose of this study was to investigate the effect of MENS on the regenerative process of injured skeletal muscle and to elucidate whether satellite cells in injured skeletal muscle are activated by MENS by using animal models. Male C57BL/6J mice, aged 7 weeks old, were used (n = 30). Mice were randomly divided into two groups: (1) cardiotoxin (CTX)-injected (CX, n = 15) and (2) CTX-injected with MENS treatment (MX, n=15) groups. CTX was injected into tibialis anterior muscle (TA) of mice in CX and MX groups to initiate the necrosis-regeneration cycle of the muscle. TA was dissected 1, 2, and 3 weeks after the injection. Muscle weight, muscle protein content, the mean cross-sectional areas of muscle fibers, the relative percentage of fibers having central nuclei, and the number of muscle satellite cells were evaluated. MENS facilitated the recovery of the muscle dry weight and protein content relative to body weight, and the mean cross-sectional areas of muscle fibers in CTX-induced injured TA muscle. The number of Pax7-positive muscle satellite cells was increased by MENS during the regenerating period. Decrease in the percentages of fibers with central nuclei after CTX-injection was facilitated by MENS. MENS may facilitate the regeneration of injured skeletal muscles by activating the regenerative potential of skeletal muscles. Key points Microcurrent electrical neuromuscular stimulation (MENS) facilitated the recovery of the relative muscle dry weight, the relative muscle protein content, and the mean cross-sectional areas of muscle

  4. Microcurrent electrical neuromuscular stimulation facilitates regeneration of injured skeletal muscle in mice.

    PubMed

    Fujiya, Hiroto; Ogura, Yuji; Ohno, Yoshitaka; Goto, Ayumi; Nakamura, Ayane; Ohashi, Kazuya; Uematsu, Daiki; Aoki, Haruhito; Musha, Haruki; Goto, Katsumasa

    2015-06-01

    Conservative therapies, mainly resting care for the damaged muscle, are generally used as a treatment for skeletal muscle injuries (such as muscle fragmentation). Several past studies reported that microcurrent electrical neuromuscular stimulation (MENS) facilitates a repair of injured soft tissues and shortens the recovery period. However, the effects of MENS on the regeneration in injured skeletal muscle are still unclear. The purpose of this study was to investigate the effect of MENS on the regenerative process of injured skeletal muscle and to elucidate whether satellite cells in injured skeletal muscle are activated by MENS by using animal models. Male C57BL/6J mice, aged 7 weeks old, were used (n = 30). Mice were randomly divided into two groups: (1) cardiotoxin (CTX)-injected (CX, n = 15) and (2) CTX-injected with MENS treatment (MX, n=15) groups. CTX was injected into tibialis anterior muscle (TA) of mice in CX and MX groups to initiate the necrosis-regeneration cycle of the muscle. TA was dissected 1, 2, and 3 weeks after the injection. Muscle weight, muscle protein content, the mean cross-sectional areas of muscle fibers, the relative percentage of fibers having central nuclei, and the number of muscle satellite cells were evaluated. MENS facilitated the recovery of the muscle dry weight and protein content relative to body weight, and the mean cross-sectional areas of muscle fibers in CTX-induced injured TA muscle. The number of Pax7-positive muscle satellite cells was increased by MENS during the regenerating period. Decrease in the percentages of fibers with central nuclei after CTX-injection was facilitated by MENS. MENS may facilitate the regeneration of injured skeletal muscles by activating the regenerative potential of skeletal muscles. Key pointsMicrocurrent electrical neuromuscular stimulation (MENS) facilitated the recovery of the relative muscle dry weight, the relative muscle protein content, and the mean cross-sectional areas of muscle

  5. High efficiency of muscle regeneration after human myoblast clone transplantation in SCID mice.

    PubMed Central

    Huard, J; Verreault, S; Roy, R; Tremblay, M; Tremblay, J P

    1994-01-01

    SCID mouse tibialis anterior muscles were first irradiated to prevent regeneration by host myoblasts and injected with notexin to damage the muscle fibers and trigger regeneration. The muscles were then injected with roughly 5 million human myoblasts. 1 mo later, 16-33% of the normal number of muscle fibers were present in the injected muscle, because of incomplete regeneration. However, > 90% of these muscle fibers contained human dystrophin. Some newly formed muscle fibers had an accumulation of human dystrophin and desmin on a part of their membrane. Such accumulations have been demonstrated at neuromuscular junctions before suggesting that the new muscle fibers are innervated and functional. The same pool of clones of human myoblasts produced only < or = 4% of muscle fibers containing human dystrophin when injected in nude mice muscles. Several of the human myoblasts did not fuse and remained in interstitial space or tightly associated with muscle fibers suggesting that some of them have formed satellite cells. Moreover, cultures of 98% pure human myoblasts were obtained from transplanted SCID muscles. In some mice where the muscle regeneration was not complete, the muscle fibers containing human dystrophin also expressed uniformly HLA class 1, confirming that the fibers are of human origin. The presence of hybrid muscle fibers containing human dystrophin and mouse MHC was also demonstrated following transplantation. These results establish that in absence of an immune reaction, transplanted human myoblasts participate to the muscle regeneration with a high degree of efficacy even if the animals were killed only 1 mo after the transplantation. Images PMID:8113396

  6. Low intensity laser therapy accelerates muscle regeneration in aged rats

    PubMed Central

    Vatansever, Fatma; Rodrigues, Natalia C.; Assis, Livia L.; Peviani, Sabrina S.; Durigan, Joao L.; Moreira, Fernando M.A.; Hamblin, Michael R.; Parizotto, Nivaldo A.

    2013-01-01

    Background Elderly people suffer from skeletal muscle disorders that undermine their daily activity and quality of life; some of these problems can be listed as but not limited to: sarcopenia, changes in central and peripheral nervous system, blood hypoperfusion, regenerative changes contributing to atrophy, and muscle weakness. Determination, proliferation and differentiation of satellite cells in the regenerative process are regulated by specific transcription factors, known as myogenic regulatory factors (MRFs). In the elderly, the activation of MRFs is inefficient which hampers the regenerative process. Recent studies found that low intensity laser therapy (LILT) has a stimulatory effect in the muscle regeneration process. However, the effects of this therapy when associated with aging are still unknown. Objective This study aimed to evaluate the effects of LILT (λ=830 nm) on the tibialis anterior (TA) muscle of aged rats. Subjects and methods The total of 56 male Wistar rats formed two population sets: old and young, with 28 animals in each set. Each of these sets were randomly divided into four groups of young rats (3 months of age) with n=7 per group and four groups of aged rats (10 months of age) with n=7 per group. These groups were submitted to cryoinjury + laser irradiation, cryoinjury only, laser irradiation only and the control group (no cryoinjury/no laser irradiation). The laser treatment was performed for 5 consecutive days. The first laser application was done 24 h after the injury (on day 2) and on the seventh day, the TA muscle was dissected and removed under anesthesia. After this the animals were euthanized. Histological analyses with toluidine blue as well as hematoxylin-eosin staining (for counting the blood capillaries) were performed for the lesion areas. In addition, MyoD and VEGF mRNA was assessed by quantitative polymerase chain reaction. Results The results showed significant elevation (p<0.05) in MyoD and VEGF genes expression levels

  7. Autophagy regulates cytoplasmic remodeling during cell reprogramming in a zebrafish model of muscle regeneration.

    PubMed

    Saera-Vila, Alfonso; Kish, Phillip E; Louie, Ke'ale W; Grzegorski, Steven J; Klionsky, Daniel J; Kahana, Alon

    2016-10-02

    Cell identity involves both selective gene activity and specialization of cytoplasmic architecture and protein machinery. Similarly, reprogramming differentiated cells requires both genetic program alterations and remodeling of the cellular architecture. While changes in genetic and epigenetic programs have been well documented in dedifferentiating cells, the pathways responsible for remodeling the cellular architecture and eliminating specialized protein complexes are not as well understood. Here, we utilize a zebrafish model of adult muscle regeneration to study cytoplasmic remodeling during cell dedifferentiation. We describe activation of autophagy early in the regenerative response to muscle injury, while blocking autophagy using chloroquine or Atg5 and Becn1 knockdown reduced the rate of regeneration with accumulation of sarcomeric and nuclear debris. We further identify Casp3/caspase 3 as a candidate mediator of cellular reprogramming and Fgf signaling as an important activator of autophagy in dedifferentiating myocytes. We conclude that autophagy plays a critical role in cell reprogramming by regulating cytoplasmic remodeling, facilitating the transition to a less differentiated cell identity.

  8. Plasticity and recovery of skeletal muscle satellite cells during limb regeneration.

    PubMed

    Morrison, Jamie I; Borg, Paula; Simon, András

    2010-03-01

    Salamander limb regeneration depends on local progenitors whose progeny are recruited to the new limb. We previously identified a Pax7(+) cell population in skeletal muscle whose progeny have the potential to contribute to the regenerating limb. However, the plasticity of individual Pax7(+) cells, as well as their recovery within the new limb, was unclear. Here, we show that Pax7(+) cells remain present after multiple rounds of limb amputation/regeneration. Pax7(+) cells are found exclusively within skeletal muscle in the regenerating limb and proliferate where the myofibers are growing. Pax7 is rapidly down-regulated in the blastema, and analyses of clonal derivatives show that Pax7(+) cell progeny are not restricted to skeletal muscle during limb regeneration. Our data suggest that the newt regeneration blastema is not entirely a composite of lineage-restricted progenitors. The results demonstrate that except for a transient and subsequently blunted increase, skeletal muscle satellite cells constitute a stable pool of reserve cells for multiple limb regeneration events.-Morrison, J. I., Borg, P., Simon, A. Plasticity and recovery of skeletal muscle satellite cells during limb regeneration.

  9. Synergist Ablation as a Rodent Model to Study Satellite Cell Dynamics in Adult Skeletal Muscle.

    PubMed

    Kirby, Tyler J; McCarthy, John J; Peterson, Charlotte A; Fry, Christopher S

    2016-01-01

    In adult skeletal muscles, satellite cells are the primary myogenic stem cells involved in myogenesis. Normally, they remain in a quiescent state until activated by a stimulus, after which they proliferate, differentiate, and fuse into an existing myofiber or form a de novo myofiber. To study satellite cell dynamics in adult murine models, most studies utilize regeneration models in which the muscle is severely damaged and requires the participation from satellite cells in order to repair. Here, we describe a model to study satellite cell behavior in muscle hypertrophy that is independent of muscle regeneration.Synergist ablation surgery involves the surgical removal of the gastrocnemius and soleus muscles resulting in functional overload of the remaining plantaris muscle. This functional overload results in myofiber hypertrophy, as well as the activation, proliferation, and fusion of satellite cells into the myofibers. Within 2 weeks of functional overload, satellite cell content increases approximately 275 %, an increase that is accompanied with a ~60 % increase in the number of myonuclei. Therefore, this can be used as an alternative model to study satellite cell behavior in adulthood that is different from regeneration, and capable of revealing new satellite cell functions in regulating muscle adaptation.

  10. PKCε as a novel promoter of skeletal muscle differentiation and regeneration

    PubMed Central

    Di Marcantonio, D; Galli, D; Carubbi, C; Gobbi, G; Queirolo, V; Martini, S; Merighi, S; Vaccarezza, M; Maffulli, N; Sykes, SM; Vitale, M; Mirandola, P

    2016-01-01

    Introduction Satellite cells are muscle resident stem cells and are responsible for muscle regeneration. In this study we investigate the involvement of PKCε during muscle stem cell differentiation in vitro and in vivo. Here, we describe the identification of a previously unrecognized role for the PKCε – HMGA1 signaling axis in myoblast differentiation and regeneration processes. Methods PKCε expression was modulated in the C2C12 cell line and primary murine satellite cells in vitro, as well as in an in vivo model of muscle regeneration. Immunohistochemistry and immunofluorescence, RT-PCR and shRNA silencing techniques were used to determine the role of PKCε and HMGA1 in myogenic differentiation. Results PKCε expression increases and subsequently re-localizes to the nucleus during skeletal muscle cell differentiation. In the nucleus, PKCε blocks Hmga1 expression to promote Myogenin and Mrf4 accumulation and myoblast formation. Following in vivo muscle injury, PKCε accumulates in regenerating, centrally-nucleated myofibers. Pharmacological inhibition of PKCε impairs the expression of two crucial markers of muscle differentiation, namely MyoD and Myogenin, during injury induced muscle regeneration. Conclusion This work identifies the PKCε – HMGA1 signaling axis as a positive regulator of skeletal muscle differentiation. PMID:26431586

  11. Heart regeneration in adult MRL mice

    NASA Astrophysics Data System (ADS)

    Leferovich, John M.; Bedelbaeva, Khamilia; Samulewicz, Stefan; Zhang, Xiang-Ming; Zwas, Donna; Lankford, Edward B.; Heber-Katz, Ellen

    2001-08-01

    The reaction of cardiac tissue to acute injury involves interacting cascades of cellular and molecular responses that encompass inflammation, hormonal signaling, extracellular matrix remodeling, and compensatory adaptation of myocytes. Myocardial regeneration is observed in amphibians, whereas scar formation characterizes cardiac ventricular wound healing in a variety of mammalian injury models. We have previously shown that the MRL mouse strain has an extraordinary capacity to heal surgical wounds, a complex trait that maps to at least seven genetic loci. Here, we extend these studies to cardiac wounds and demonstrate that a severe transmural, cryogenically induced infarction of the right ventricle heals extensively within 60 days, with the restoration of normal myocardium and function. Scarring is markedly reduced in MRL mice compared with C57BL/6 mice, consistent with both the reduced hydroxyproline levels seen after injury and an elevated cardiomyocyte mitotic index of 10-20% for the MRL compared with 1-3% for the C57BL/6. The myocardial response to injury observed in these mice resembles the regenerative process seen in amphibians.

  12. Regeneration of lumbar dorsal root axons into the spinal cord of adult frogs (Rana pipiens), an HRP study.

    PubMed

    Liuzzi, F J; Lasek, R J

    1985-02-22

    Lumbar dorsal roots of adult frogs were crushed or cut and reanastomosed. Following survival times of up to 75 days, the regenerating dorsal roots were recut and anterogradely injury-filled with horseradish peroxidase. This revealed that in the adult frog, regenerating axons re-enter the spinal cord. Comparison of the distribution of these axons with that of normal dorsal root axons showed that there is a partial restoration of the segmental distribution in the gray matter. However, the long ascending sensory tract of the dorsal funiculus was not restored. The dorsal funiculus was markedly gliotic and had relatively few labelled, regenerated axons. The labelled axons that were seen in the dorsal funiculus either extended longitudinally for a distance just beneath the pia, apparently in association with the glia limitans, or traversed the region to enter the dorsal gray matter. Most of the large and small diameter axons that entered the gray matter did so by passing through the region of the dorsolateral fasciculus. Within the gray matter, small diameter, regenerated axons arborized in the region of the dorsal terminal field, a region that has been shown in the normal frog to receive cutaneous afferents only. Many large diameter axons, presumably muscle afferents, arborized in the ventral terminal field, a region shown in the normal frog to receive muscle afferents exclusively. However, many of these large diameter axons had arborizations that extended to both terminal fields, thus suggesting that some abberant connections are made during dorsal root regeneration in the adult frog.

  13. Trbp Is Required for Differentiation of Myoblasts and Normal Regeneration of Skeletal Muscle

    PubMed Central

    Ding, Jian; Nie, Mao; Liu, Jianming; Hu, Xiaoyun; Ma, Lixin; Deng, Zhong-Liang; Wang, Da-Zhi

    2016-01-01

    Global inactivation of Trbp, a regulator of miRNA pathways, resulted in developmental defects and postnatal lethality in mice. Recently, we showed that cardiac-specific deletion of Trbp caused heart failure. However, its functional role(s) in skeletal muscle has not been characterized. Using a conditional knockout model, we generated mice lacking Trbp in the skeletal muscle. Unexpectedly, skeletal muscle specific Trbp mutant mice appear to be phenotypically normal under normal physiological conditions. However, these mice exhibited impaired muscle regeneration and increased fibrosis in response to cardiotoxin-induced muscle injury, suggesting that Trbp is required for muscle repair. Using cultured myoblast cells we further showed that inhibition of Trbp repressed myoblast differentiation in vitro. The impaired myogenesis is associated with reduced expression of muscle-specific miRNAs, miR-1a and miR-133a. Together, our study demonstrated that Trbp participates in the regulation of muscle differentiation and regeneration. PMID:27159388

  14. Hmgb3 Is Regulated by MicroRNA-206 during Muscle Regeneration

    PubMed Central

    Maciotta, Simona; Meregalli, Mirella; Cassinelli, Letizia; Parolini, Daniele; Farini, Andrea; Fraro, Giulia Del; Gandolfi, Francesco; Forcato, Mattia; Ferrari, Sergio; Gabellini, Davide; Bicciato, Silvio; Cossu, Giulio; Torrente, Yvan

    2012-01-01

    Background MicroRNAs (miRNAs) have been recently involved in most of human diseases as targets for potential strategies to rescue the pathological phenotype. Since the skeletal muscle is a spread-wide highly differentiated and organized tissue, rescue of severely compromised muscle still remains distant from nowadays. For this reason, we aimed to identify a subset of miRNAs major involved in muscle remodelling and regeneration by analysing the miRNA-profile of single fibres isolated from dystrophic muscle, which was here considered as a model of chronic damage. Methodology/Principal Findings The miRNA-signature associated to regenerating (newly formed) and remodelling (resting) fibres was investigated in animal models of muscular dystrophies and acute damage, in order to distinguish which miRNAs are primary related to muscle regeneration. In this study we identify fourteen miRNAs associated to dystrophic fibres responsible for muscle regeneration and remodelling, and confirm over-expression of the previously identified regeneration-associated myomiR-206. In particular, a functional binding site for myomiR-206 was identified and validated in the 3′untranslated region (3′UTR) of an X-linked member of a family of sequence independent chromatin-binding proteins (Hmgb3) that is preferentially expressed in hematopoietic stem cells. During regeneration of single muscle fibres, Hmgb3 messenger RNA (mRNA) and protein expression was gradually reduced, concurrent with the up-regulation of miR-206. Conclusion/Significance Our results elucidate a negative feedback circuit in which myomiR-206 represses Hmgb3 expression to modulate the regeneration of single muscle fibres after acute and chronic muscle damage. These findings suggest that myomiR-206 may be a potential therapeutic target in muscle diseases. PMID:22912879

  15. Salamander limb regeneration involves the activation of a multipotent skeletal muscle satellite cell population.

    PubMed

    Morrison, Jamie I; Lööf, Sara; He, Pingping; Simon, András

    2006-01-30

    In contrast to mammals, salamanders can regenerate complex structures after injury, including entire limbs. A central question is whether the generation of progenitor cells during limb regeneration and mammalian tissue repair occur via separate or overlapping mechanisms. Limb regeneration depends on the formation of a blastema, from which the new appendage develops. Dedifferentiation of stump tissues, such as skeletal muscle, precedes blastema formation, but it was not known whether dedifferentiation involves stem cell activation. We describe a multipotent Pax7+ satellite cell population located within the skeletal muscle of the salamander limb. We demonstrate that skeletal muscle dedifferentiation involves satellite cell activation and that these cells can contribute to new limb tissues. Activation of salamander satellite cells occurs in an analogous manner to how the mammalian myofiber mobilizes stem cells during skeletal muscle tissue repair. Thus, limb regeneration and mammalian tissue repair share common cellular and molecular programs. Our findings also identify satellite cells as potential targets in promoting mammalian blastema formation.

  16. Length-tension relationships are altered in regenerating muscles of the rat after bupivacaine injection.

    PubMed

    Plant, David R; Beitzel, Felice; Lynch, Gordon S

    2005-06-01

    Intramuscular injection of bupivacaine causes complete degeneration of fibers in extensor digitorum longus (EDL) muscles of rats, followed by complete regeneration within 60 days. Previous studies have shown that regenerated EDL muscles are protected from contraction-induced injury 60 days after bupivacaine injection. It is possible that these regenerated muscles have altered length-tension relations because of fiber remodeling. We tested the hypothesis that length-tension relations are different in bupivacaine-injected and noninjected control muscles. EDL and soleus muscles of the right hindlimb of deeply anesthetized rats were injected with bupivacaine and then allowed to recover for 7, 14, 21, or 60 days (7D, 14D, 21D, 60D), and isometric contractile properties were assessed. Muscles of the contralateral limb were not injected and served as control. EDL muscles recovered from bupivacaine injection more rapidly than soleus muscles, with mass restored to control levels at 21D, and isometric tetanic force (P(o)) restored to control at 60D. In contrast, mass and P(o) of injected soleus muscles was not restored to control even at 60D. In 7D EDL muscles, length-tension curves were shifted leftward compared with control, but in 21D and 60D EDL muscles length-tension curves were right shifted significantly (treatment x muscle length: P < 0.001). Although no clear shift in the position of the length-tension curve was observed in regenerating soleus muscles, force production was enhanced on the descending limb of the curve in 60D soleus muscles (treatment x relative muscle length: P < 0.01). The rightward shift in the length-tension curve of EDL muscles 60 days after bupivacaine injection is likely to contribute to the mechanism for their previously observed protection from contraction-induced injury.

  17. Characteristics of the Localization of Connexin 43 in Satellite Cells during Skeletal Muscle Regeneration In Vivo.

    PubMed

    Ishido, Minenori; Kasuga, Norikatsu

    2015-04-28

    For myogenesis, new myotubes are formed by the fusion of differentiated myoblasts. In the sequence of events for myotube formation, intercellular communication through gap junctions composed of connexin 43 (Cx43) plays critical roles in regulating the alignment and fusion of myoblasts in advances of myotube formation in vitro. On the other hand, the relationship between the expression patterns of Cx43 and the process of myotube formation in satellite cells during muscle regeneration in vivo remains poorly understood. The present study investigated the relationship between Cx43 and satellite cells in muscle regeneration in vivo. The expression of Cx43 was detected in skeletal muscles on day 1 post-muscle injury, but not in control muscles. Interestingly, the expression of Cx43 was not localized on the inside of the basement membrane of myofibers in the regenerating muscles. Moreover, although the clusters of differentiated satellite cells, which represent a more advanced stage of myotube formation, were observed on the inside of the basement membrane of myofibers in regenerating muscles, the expression of Cx43 was not localized in the clusters of these satellite cells. Therefore, in the present study, it was suggested that Cx43 may not directly contribute to muscle regeneration via satellite cells.

  18. Expansion of revertant fibers in dystrophic mdx muscles reflects activity of muscle precursor cells and serves as an index of muscle regeneration.

    PubMed

    Yokota, Toshifumi; Lu, Qi-Long; Morgan, Jennifer E; Davies, Kay E; Fisher, Rosie; Takeda, Shin'ichi; Partridge, Terence A

    2006-07-01

    Duchenne muscular dystrophy and the mdx mouse myopathies reflect a lack of dystrophin in muscles. However, both contain sporadic clusters of revertant fibers (RFs) that express dystrophin. RF clusters expand in size with age in mdx mice. To test the hypothesis that the expansion of clusters is achieved through the process of muscle degeneration and regeneration, we analyzed muscles of mdx mice in which degeneration and regeneration were inhibited by the expression of micro-dystrophins or utrophin transgenes. Postnatal RF expansion was diminished in direct correlation to the protective effect of the transgene expression. Similarly, expansion of RFs was inhibited when muscle regeneration was blocked by irradiation. However, in irradiated muscles, irradiation-tolerant quiescent muscle precursor cells reactivated by notexin effectively restored RF expansion. Our observations demonstrate that revertant events occur initially within a subset of muscle precursor cells. The proliferation of these cells, as part of the regeneration process, leads to the expansion of RF clusters within degenerating muscles. This expansion of revertant clusters depicts the cumulative history of regeneration, thus providing a useful index for functional evaluation of therapies that counteract muscle degeneration.

  19. Positional identity of adult stem cells in salamander limb regeneration.

    PubMed

    Kumar, Anoop; Gates, Phillip B; Brockes, Jeremy P

    2007-01-01

    Limb regeneration in larval and adult salamanders proceeds from a mound of mesenchymal stem cells called the limb blastema. The blastema gives rise just to those structures distal to its level of origin, and this property of positional identity is reset to more proximal values by treatment with retinoic acid. We have identified a cell surface protein, called Prod1/CD59, which appears to be a determinant of proximodistal identity. Prod1 is expressed in an exponential gradient in an adult limb as determined by detection of both mRNA and immunoreactive protein. Prod1 protein is up-regulated after treatment of distal blastemas with RA and this is particularly marked in cells of the dermis. These cells have previously been implicated in pattern formation during limb regeneration.

  20. In Silico and In Vivo Experiments Reveal M-CSF Injections Accelerate Regeneration Following Muscle Laceration.

    PubMed

    Martin, Kyle S; Kegelman, Christopher D; Virgilio, Kelley M; Passipieri, Julianna A; Christ, George J; Blemker, Silvia S; Peirce, Shayn M

    2017-03-01

    Numerous studies have pharmacologically modulated the muscle milieu in the hopes of promoting muscle regeneration; however, the timing and duration of these interventions are difficult to determine. This study utilized a combination of in silico and in vivo experiments to investigate how inflammation manipulation improves muscle recovery following injury. First, we measured macrophage populations following laceration injury in the rat tibialis anterior (TA). Then we calibrated an agent-based model (ABM) of muscle injury to mimic the observed inflammation profiles. The calibrated ABM was used to simulate macrophage and satellite stem cell (SC) dynamics, and suggested that delivering macrophage colony stimulating factor (M-CSF) prior to injury would promote SC-mediated injury recovery. Next, we performed an experiment wherein 1 day prior to injury, we injected M-CSF into the rat TA muscle. M-CSF increased the number of macrophages during the first 4 days post-injury. Furthermore, treated muscles experienced a swifter increase in the appearance of PAX7(+) SCs and regenerating muscle fibers. Our study suggests that computational models of muscle injury provide novel insights into cellular dynamics during regeneration, and further, that pharmacologically altering inflammation dynamics prior to injury can accelerate the muscle regeneration process.

  1. Suppression of macrophage functions impairs skeletal muscle regeneration with severe fibrosis

    SciTech Connect

    Segawa, Masashi; Fukada, So-ichiro Yamamoto, Yukiko; Yahagi, Hiroshi; Kanematsu, Masanori; Sato, Masaki; Ito, Takahito; Uezumi, Akiyoshi; Hayashi, Shin'ichi; Miyagoe-Suzuki, Yuko; Takeda, Shin'ichi; Tsujikawa, Kazutake; Yamamoto, Hiroshi

    2008-10-15

    When damaged, skeletal muscle regenerates. In the early phases of regeneration, inflammatory cells such as neutrophils/granulocytes and macrophages infiltrate damaged muscle tissue. To reveal the roles of macrophages during skeletal muscle regeneration, we injected an antibody, AFS98 that blocks the binding of M-CSF to its receptor into normal mice that received muscle damages. Anti-M-CSF receptor administration suppressed macrophage but not neutrophil infiltration. Histological study indicated that suppression of macrophages function leads to the incomplete muscle regeneration. In addition FACS and immunohistochemical study showed that the acute lack of macrophages delayed proliferation and differentiation of muscle satellite cells in vivo. Furthermore, mice injected with the anti-M-CSF receptor antibody exhibited not only adipogenesis, but also significant collagen deposition, i.e., fibrosis and continuous high expression of connective tissue growth factor. Finally we indicate that these fibrosis markers were strongly enriched in CD90(+) cells that do not include myogenic cells. These results indicate that macrophages directly affect satellite cell proliferation and that a macrophage deficiency severely impairs skeletal muscle regeneration and causes fibrosis.

  2. Akirin1 (Mighty), a novel promyogenic factor regulates muscle regeneration and cell chemotaxis

    SciTech Connect

    Salerno, Monica Senna; Dyer, Kelly; Bracegirdle, Jeremy; Platt, Leanne; Thomas, Mark; Siriett, Victoria; Kambadur, Ravi; Sharma, Mridula

    2009-07-15

    Akirin1 (Mighty) is a downstream target gene of myostatin and has been shown to be a promyogenic factor. Although expressed in many tissues, akirin1 is negatively regulated by myostatin specifically in skeletal muscle tissue. In this manuscript we have characterized the possible function of akirin1 in postnatal muscle growth. Molecular and immunohistological analyses indicated that while low levels of akirin1 are associated with quiescent satellite cells (SC), higher levels of akirin1 are detected in activated proliferating SC indicating that akirin1 could be associated with satellite cell activation. In addition to SC, macrophages also express akirin1, and increased expression of akirin1 resulted in more efficient chemotaxis of both macrophages and myoblasts. Akirin1 appears to regulate chemotaxis of both macrophages and myoblasts by reorganising actin cytoskeleton, leading to more efficient lamellipodia formation via a PI3 kinase dependent pathway. Expression analysis during muscle regeneration also indicated that akirin1 expression is detected very early (day 2) in regenerating muscle, and expression gradually peaks to coincide the nascent myotube formation stage of muscle regeneration. Based on these results we propose that akirin1 could be acting as a transducer of early signals of muscle regeneration. Thus, we speculate that myostatin regulates key steps of muscle regeneration including chemotaxis of inflammatory cells, SC activation and migration through akirin1.

  3. Brief report: Blockade of Notch signaling in muscle stem cells causes muscular dystrophic phenotype and impaired muscle regeneration.

    PubMed

    Lin, Shuibin; Shen, Huangxuan; Jin, Baofeng; Gu, Yumei; Chen, Zirong; Cao, Chunxia; Hu, Chengbin; Keller, Charles; Pear, Warren S; Wu, Lizi

    2013-04-01

    Muscular dystrophies are a group of devastating diseases characterized by progressive muscle weakness and degeneration, with etiologies including muscle gene mutations and regenerative defects of muscle stem cells. Notch signaling is critical for skeletal myogenesis and has important roles in maintaining the muscle stem cell pool and preventing premature muscle differentiation. To investigate the functional impact of Notch signaling blockade in muscle stem cells, we developed a conditional knock-in mouse model in which endogenous Notch signaling is specifically blocked in muscle stem cell compartment. Mice with Notch signaling inhibition in muscle stem cells showed several muscular dystrophic features and impaired muscle regeneration. Analyses of satellite cells and isolated primary myoblasts revealed that Notch signaling blockade in muscle stem cells caused reduced activation and proliferation of satellite cells but enhanced differentiation of myoblasts. Our data thus indicate that Notch signaling controls processes that are critical to regeneration in muscular dystrophy, suggesting that Notch inhibitor therapies could have potential side effects on muscle functions.

  4. Electrical stimulation of embryonic neurons for 1 hour improves axon regeneration and the number of reinnervated muscles that function.

    PubMed

    Liu, Yang; Grumbles, Robert M; Thomas, Christine K

    2013-07-01

    Motoneuron death after spinal cord injury or disease results in muscle denervation, atrophy, and paralysis. We have previously transplanted embryonic ventral spinal cord cells into the peripheral nerve to reinnervate denervated muscles and to reduce muscle atrophy, but reinnervation was incomplete. Here, our aim was to determine whether brief electrical stimulation of embryonic neurons in the peripheralnerve changes motoneuron survival, axon regeneration, and muscle reinnervation and function because neural depolarization is crucial for embryonic neuron survival and may promote activity-dependent axon growth. At 1 week after denervation by sciatic nerve section, embryonic day 14 to 15 cells were purified for motoneurons, injected into the tibial nerve of adult Fischer rats, and stimulated immediatelyfor up to 1 hour. More myelinated axons were present in tibial nerves 10 weeks after transplantation when transplants had been stimulated acutely at 1 Hz for 1 hour. More muscles were reinnervated if the stimulation treatment lasted for 1 hour. Reinnervation reduced muscle atrophy, with or without the stimulation treatment. These data suggest that brief stimulation of embryonic neurons promotes axon growth, which has a long-term impact on muscle reinnervation and function. Muscle reinnervation is important because it may enable the use of functional electrical stimulation to restore limb movements.

  5. Electrical Stimulation of Embryonic Neurons for 1 Hour Improves Axon Regeneration and the Number of Reinnervated Muscles that Function

    PubMed Central

    Liu, Yang; Grumbles, Robert M.; Thomas, Christine K.

    2013-01-01

    Motoneuron death following spinal cord injury or disease results in muscle denervation, atrophy, and paralysis. We have previously transplanted embryonic ventral spinal cord cells into peripheral nerve to reinnervate denervated muscles and to reduce muscle atrophy, but reinnervation was incomplete. Here, our aim was to determine whether brief electrical stimulation of embryonic neurons in peripheral nerve changes motoneuron survival, axon regeneration, and muscle reinnervation and function because neural depolarization is crucial for embryonic neuron survival and may promote activity-dependent axon growth. At 1 week after denervation by sciatic nerve section, embryonic day 14-15 cells were purified for motoneurons, injected into the tibial nerve of adult Fischer rats, and stimulated immediately for up to 1 hour. More myelinated axons were present in tibial nerves when transplants had been stimulated at 1 Hz for 1 hour at 10 weeks following transplantation. More muscles were reinnervated if the stimulation treatment lasted for 1 hour. Reinnervation reduced muscle atrophy, with or without the stimulation treatment. These data suggest that brief stimulation of embryonic neurons promotes axon growth, which has a long-term impact on muscle reinnervation and function. Muscle reinnervation is important because it may enable the use of functional electrical stimulation to restore limb movements. PMID:23771218

  6. Green tea extract decreases muscle pathology and NF-κB immunostaining in regenerating muscle fibers of mdx mice

    PubMed Central

    Evans, Nicholas P.; Call, Jarrod A.; Bassaganya-Riera, Josep; Robertson, John L.; Grange, Robert W.

    2009-01-01

    BACKGROUND & AIMS Duchenne muscular dystrophy is a debilitating genetic disorder characterized by severe muscle wasting and early death in afflicted boys. The primary cause of this disease is mutations in the dystrophin gene resulting in massive muscle degeneration and inflammation. The purpose of this study was to determine if dystrophic muscle pathology and inflammation were decreased by pre-natal and early dietary intervention with green tea extract. METHODS Mdx breeder mice and pups were fed diets containing 0.25% or 0.5% green tea extract and compared to untreated mdx and C57BL/6J mice. Serum creatine kinase was assessed as a systemic indicator of muscle damage. Quantitative histopathological and immunohistochemical techniques were used to determine muscle pathology, macrophage infiltration, and NF-κB localization. RESULTS Early treatment of mdx mice with green tea extract significantly decreased serum creatine kinase by ~85% at age 42 days (P≤0.05). In these mice, the area of normal fiber morphology was increased by as much as ~32% (P≤0.05). The primary histopathological change was a ~21% decrease in the area of regenerating fibers (P≤0.05). NF-κB staining in regenerating muscle fibers was also significantly decreased in green tea extract-treated mdx mice when compared to untreated mdx mice. CONCLUSION Early treatment with green tea extract decreases dystrophic muscle pathology potentially by regulating NF-κB activity in regenerating muscle fibers. PMID:19897286

  7. Reactive oxygen species generated from skeletal muscles are required for gecko tail regeneration

    PubMed Central

    Zhang, Qing; Wang, Yingjie; Man, Lili; Zhu, Ziwen; Bai, Xue; Wei, Sumei; Liu, Yan; Liu, Mei; Wang, Xiaochuan; Gu, Xiaosong; Wang, Yongjun

    2016-01-01

    Reactive oxygen species (ROS) participate in various physiological and pathological functions following generation from different types of cells. Here we explore ROS functions on spontaneous tail regeneration using gecko model. ROS were mainly produced in the skeletal muscle after tail amputation, showing a temporal increase as the regeneration proceeded. Inhibition of the ROS production influenced the formation of autophagy in the skeletal muscles, and as a consequence, the length of the regenerating tail. Transcriptome analysis has shown that NADPH oxidase (NOX2) and the subunits (p40phox and p47phox) are involved in the ROS production. ROS promoted the formation of autophagy through regulation of both ULK and MAPK activities. Our results suggest that ROS produced by skeletal muscles are required for the successful gecko tail regeneration. PMID:26853930

  8. Reactive oxygen species generated from skeletal muscles are required for gecko tail regeneration.

    PubMed

    Zhang, Qing; Wang, Yingjie; Man, Lili; Zhu, Ziwen; Bai, Xue; Wei, Sumei; Liu, Yan; Liu, Mei; Wang, Xiaochuan; Gu, Xiaosong; Wang, Yongjun

    2016-02-08

    Reactive oxygen species (ROS) participate in various physiological and pathological functions following generation from different types of cells. Here we explore ROS functions on spontaneous tail regeneration using gecko model. ROS were mainly produced in the skeletal muscle after tail amputation, showing a temporal increase as the regeneration proceeded. Inhibition of the ROS production influenced the formation of autophagy in the skeletal muscles, and as a consequence, the length of the regenerating tail. Transcriptome analysis has shown that NADPH oxidase (NOX2) and the subunits (p40(phox) and p47(phox)) are involved in the ROS production. ROS promoted the formation of autophagy through regulation of both ULK and MAPK activities. Our results suggest that ROS produced by skeletal muscles are required for the successful gecko tail regeneration.

  9. Atlas of Cellular Dynamics during Zebrafish Adult Kidney Regeneration

    PubMed Central

    McCampbell, Kristen K.; Springer, Kristin N.; Wingert, Rebecca A.

    2015-01-01

    The zebrafish is a useful animal model to study the signaling pathways that orchestrate kidney regeneration, as its renal nephrons are simple, yet they maintain the biological complexity inherent to that of higher vertebrate organisms including mammals. Recent studies have suggested that administration of the aminoglycoside antibiotic gentamicin in zebrafish mimics human acute kidney injury (AKI) through the induction of nephron damage, but the timing and details of critical phenotypic events associated with the regeneration process, particularly in existing nephrons, have not been characterized. Here, we mapped the temporal progression of cellular and molecular changes that occur during renal epithelial regeneration of the proximal tubule in the adult zebrafish using a platform of histological and expression analysis techniques. This work establishes the timing of renal cell death after gentamicin injury, identifies proliferative compartments within the kidney, and documents gene expression changes associated with the regenerative response of proliferating cells. These data provide an important descriptive atlas that documents the series of events that ensue after damage in the zebrafish kidney, thus availing a valuable resource for the scientific community that can facilitate the implementation of zebrafish research to delineate the mechanisms that control renal regeneration. PMID:26089919

  10. Clones of ectopic stem cells in the regeneration of muscle defects in vivo.

    PubMed

    Yang, Rujing; Chen, Mo; Lee, Chang Hun; Yoon, Richard; Lal, Shan; Mao, Jeremy J

    2010-10-20

    Little is known about whether clones of ectopic, non-muscle stem cells contribute to muscle regeneration. Stem/progenitor cells that are isolated for experimental research or therapeutics are typically heterogeneous. Non-myogenic lineages in a heterogeneous population conceptually may compromise tissue repair. In this study, we discovered that clones of mononucleated stem cells of human tooth pulp fused into multinucleated myotubes that robustly expressed myosin heavy chain in vitro with or without co-culture with mouse skeletal myoblasts (C2C12 cells). Cloned cells were sustainably Oct4+, Nanog+ and Stro1+. The fusion indices of myogenic clones were approximately 16-17 folds greater than their parent, heterogeneous stem cells. Upon infusion into cardio-toxin induced tibialis anterior muscle defects, undifferentiated clonal progenies not only engrafted and colonized host muscle, but also expressed human dystrophin and myosin heavy chain more efficaciously than their parent heterogeneous stem cell populations. Strikingly, clonal progenies yielded ∼9 times more human myosin heavy chain mRNA in regenerating muscles than those infused with their parent, heterogeneous stem cells. The number of human dystrophin positive cells in regenerating muscles infused with clonal progenies was more than ∼3 times greater than muscles infused with heterogeneous stem cells from which clonal progenies were derived. These findings suggest the therapeutic potential of ectopic myogenic clones in muscle regeneration.

  11. Impaired skeletal muscle regeneration in the absence of fibrosis during hibernation in 13-lined ground squirrels.

    PubMed

    Andres-Mateos, Eva; Mejias, Rebeca; Soleimani, Arshia; Lin, Brian M; Burks, Tyesha N; Marx, Ruth; Lin, Benjamin; Zellars, Richard C; Zhang, Yonggang; Huso, David L; Marr, Tom G; Leinwand, Leslie A; Merriman, Dana K; Cohn, Ronald D

    2012-01-01

    Skeletal muscle atrophy can occur as a consequence of immobilization and/or starvation in the majority of vertebrates studied. In contrast, hibernating mammals are protected against the loss of muscle mass despite long periods of inactivity and lack of food intake. Resident muscle-specific stem cells (satellite cells) are known to be activated by muscle injury and their activation contributes to the regeneration of muscle, but whether satellite cells play a role in hibernation is unknown. In the hibernating 13-lined ground squirrel we show that muscles ablated of satellite cells were still protected against atrophy, demonstrating that satellite cells are not involved in the maintenance of skeletal muscle during hibernation. Additionally, hibernating skeletal muscle showed extremely slow regeneration in response to injury, due to repression of satellite cell activation and myoblast differentiation caused by a fine-tuned interplay of p21, myostatin, MAPK, and Wnt signaling pathways. Interestingly, despite long periods of inflammation and lack of efficient regeneration, injured skeletal muscle from hibernating animals did not develop fibrosis and was capable of complete recovery when animals emerged naturally from hibernation. We propose that hibernating squirrels represent a new model system that permits evaluation of impaired skeletal muscle remodeling in the absence of formation of tissue fibrosis.

  12. Impaired Skeletal Muscle Regeneration in the Absence of Fibrosis during Hibernation in 13-Lined Ground Squirrels

    PubMed Central

    Soleimani, Arshia; Lin, Brian M.; Burks, Tyesha N.; Marx, Ruth; Lin, Benjamin; Zellars, Richard C.; Zhang, Yonggang; Huso, David L.; Marr, Tom G.; Leinwand, Leslie A.; Merriman, Dana K.; Cohn, Ronald D.

    2012-01-01

    Skeletal muscle atrophy can occur as a consequence of immobilization and/or starvation in the majority of vertebrates studied. In contrast, hibernating mammals are protected against the loss of muscle mass despite long periods of inactivity and lack of food intake. Resident muscle-specific stem cells (satellite cells) are known to be activated by muscle injury and their activation contributes to the regeneration of muscle, but whether satellite cells play a role in hibernation is unknown. In the hibernating 13-lined ground squirrel we show that muscles ablated of satellite cells were still protected against atrophy, demonstrating that satellite cells are not involved in the maintenance of skeletal muscle during hibernation. Additionally, hibernating skeletal muscle showed extremely slow regeneration in response to injury, due to repression of satellite cell activation and myoblast differentiation caused by a fine-tuned interplay of p21, myostatin, MAPK, and Wnt signaling pathways. Interestingly, despite long periods of inflammation and lack of efficient regeneration, injured skeletal muscle from hibernating animals did not develop fibrosis and was capable of complete recovery when animals emerged naturally from hibernation. We propose that hibernating squirrels represent a new model system that permits evaluation of impaired skeletal muscle remodeling in the absence of formation of tissue fibrosis. PMID:23155423

  13. Annexin A1 Deficiency does not Affect Myofiber Repair but Delays Regeneration of Injured Muscles

    PubMed Central

    Leikina, Evgenia; Defour, Aurelia; Melikov, Kamran; Van der Meulen, Jack H.; Nagaraju, Kanneboyina; Bhuvanendran, Shivaprasad; Gebert, Claudia; Pfeifer, Karl; Chernomordik, Leonid V.; Jaiswal, Jyoti K.

    2015-01-01

    Repair and regeneration of the injured skeletal myofiber involves fusion of intracellular vesicles with sarcolemma and fusion of the muscle progenitor cells respectively. In vitro experiments have identified involvement of Annexin A1 (Anx A1) in both these fusion processes. To determine if Anx A1 contributes to these processes during muscle repair in vivo, we have assessed muscle growth and repair in Anx A1-deficient mouse (AnxA1−/−). We found that the lack of Anx A1 does not affect the muscle size and repair of myofibers following focal sarcolemmal injury and lengthening contraction injury. However, the lack of Anx A1 delayed muscle regeneration after notexin-induced injury. This delay in muscle regeneration was not caused by a slowdown in proliferation and differentiation of satellite cells. Instead, lack of Anx A1 lowered the proportion of differentiating myoblasts that managed to fuse with the injured myofibers by days 5 and 7 after notexin injury as compared to the wild type (w.t.) mice. Despite this early slowdown in fusion of Anx A1−/− myoblasts, regeneration caught up at later times post injury. These results establish in vivo role of Anx A1 in cell fusion required for myofiber regeneration and not in intracellular vesicle fusion needed for repair of myofiber sarcolemma. PMID:26667898

  14. The CHC22 Clathrin-GLUT4 Transport Pathway Contributes to Skeletal Muscle Regeneration

    PubMed Central

    Griffin, Christine A.; Esk, Christopher; Torres, Jorge A.; Ohkoshi, Norio; Ishii, Akiko; Tamaoka, Akira; Funke, Birgit H.; Kucherlapati, Raju; Margeta, Marta; Rando, Thomas A.; Brodsky, Frances M.

    2013-01-01

    Mobilization of the GLUT4 glucose transporter from intracellular storage vesicles provides a mechanism for insulin-responsive glucose import into skeletal muscle. In humans, clathrin isoform CHC22 participates in formation of the GLUT4 storage compartment in skeletal muscle and fat. CHC22 function is limited to retrograde endosomal sorting and is restricted in its tissue expression and species distribution compared to the conserved CHC17 isoform that mediates endocytosis and several other membrane traffic pathways. Previously, we noted that CHC22 was expressed at elevated levels in regenerating rat muscle. Here we investigate whether the GLUT4 pathway in which CHC22 participates could play a role in muscle regeneration in humans and we test this possibility using CHC22-transgenic mice, which do not normally express CHC22. We observed that GLUT4 expression is elevated in parallel with that of CHC22 in regenerating skeletal muscle fibers from patients with inflammatory and other myopathies. Regenerating human myofibers displayed concurrent increases in expression of VAMP2, another regulator of GLUT4 transport. Regenerating fibers from wild-type mouse skeletal muscle injected with cardiotoxin also showed increased levels of GLUT4 and VAMP2. We previously demonstrated that transgenic mice expressing CHC22 in their muscle over-sequester GLUT4 and VAMP2 and have defective GLUT4 trafficking leading to diabetic symptoms. In this study, we find that muscle regeneration rates in CHC22 mice were delayed compared to wild-type mice, and myoblasts isolated from these mice did not proliferate in response to glucose. Additionally, CHC22-expressing mouse muscle displayed a fiber type switch from oxidative to glycolytic, similar to that observed in type 2 diabetic patients. These observations implicate the pathway for GLUT4 transport in regeneration of both human and mouse skeletal muscle, and demonstrate a role for this pathway in maintenance of muscle fiber type. Extrapolating

  15. Pten is necessary for the quiescence and maintenance of adult muscle stem cells

    PubMed Central

    Yue, Feng; Bi, Pengpeng; Wang, Chao; Shan, Tizhong; Nie, Yaohui; Ratliff, Timothy L.; Gavin, Timothy P.; Kuang, Shihuan

    2017-01-01

    Satellite cells (SCs) are myogenic stem cells required for regeneration of adult skeletal muscles. A proper balance among quiescence, activation and differentiation is essential for long-term maintenance of SCs and their regenerative function. Here we show a function of Pten (phosphatase and tensin homologue) in quiescent SCs. Deletion of Pten in quiescent SCs leads to their spontaneous activation and premature differentiation without proliferation, resulting in depletion of SC pool and regenerative failure. However, prior to depletion, Pten-null activated SCs can transiently proliferate upon injury and regenerate injured muscles, but continually decline during regeneration, suggesting an inability to return to quiescence. Mechanistically, Pten deletion increases Akt phosphorylation, which induces cytoplasmic translocation of FoxO1 and suppression of Notch signalling. Accordingly, constitutive activation of Notch1 prevents SC depletion despite Pten deletion. Our findings delineate a critical function of Pten in maintaining SC quiescence and reveal an interaction between Pten and Notch signalling. PMID:28094257

  16. Making Skeletal Muscle from Progenitor and Stem Cells: Development versus Regeneration

    PubMed Central

    Li, Lydia; Rozo, Michelle E.; Lepper, Christoph

    2012-01-01

    For locomotion, vertebrate animals use the force generated by contractile skeletal muscles. These muscles form an actin/myosin-based bio-machinery that is attached to skeletal elements to effect body movement and maintain posture. The mechanics, physiology, and homeostasis of skeletal muscles in normal and disease states are of significant clinical interest. How muscles originate from progenitors during embryogenesis has attracted considerable attention from developmental biologists. How skeletal muscles regenerate and repair themselves after injury by the use of stem cells is an important process to maintain muscle homeostasis throughout lifetime. In recent years, much progress has been made towards uncovering the origins of myogenic progenitors and stem cells as well as the regulation of these cells during development and regeneration. PMID:22737183

  17. Stabilin-2 modulates the efficiency of myoblast fusion during myogenic differentiation and muscle regeneration

    PubMed Central

    Park, Seung-Yoon; Yun, Youngeun; Lim, Jung-Suk; Kim, Mi-Jin; Kim, Sang-Yeob; Kim, Jung-Eun; Kim, In-San

    2016-01-01

    Myoblast fusion is essential for the formation of skeletal muscle myofibres. Studies have shown that phosphatidylserine is necessary for myoblast fusion, but the underlying mechanism is not known. Here we show that the phosphatidylserine receptor stabilin-2 acts as a membrane protein for myoblast fusion during myogenic differentiation and muscle regeneration. Stabilin-2 expression is induced during myogenic differentiation, and is regulated by calcineurin/NFAT signalling in myoblasts. Forced expression of stabilin-2 in myoblasts is associated with increased myotube formation, whereas deficiency of stabilin-2 results in the formation of small, thin myotubes. Stab2-deficient mice have myofibres with small cross-sectional area and few myonuclei and impaired muscle regeneration after injury. Importantly, myoblasts lacking stabilin-2 have reduced phosphatidylserine-dependent fusion. Collectively, our results show that stabilin-2 contributes to phosphatidylserine-dependent myoblast fusion and provide new insights into the molecular mechanism by which phosphatidylserine mediates myoblast fusion during muscle growth and regeneration. PMID:26972991

  18. Regenerating Fish Optic Nerves and a Regeneration-Like Response in Injured Optic Nerves of Adult Rabbits

    NASA Astrophysics Data System (ADS)

    Schwartz, M.; Belkin, M.; Harel, A.; Solomon, A.; Lavie, V.; Hadani, M.; Rachailovich, I.; Stein-Izsak, C.

    1985-05-01

    Regeneration of fish optic nerve (representing regenerative central nervous system) was accompanied by increased activity of regeneration-triggering factors produced by nonneuronal cells. A graft of regenerating fish optic nerve, or a ``wrap-around'' implant containing medium conditioned by it, induced a response associated with regeneration in injured optic nerves of adult rabbits (representing a nonregenerative central nervous system). This response was manifested by an increase of general protein synthesis and of selective polypeptides in the retinas and by the ability of the retina to sprout in culture.

  19. Sporadic inclusion body myositis: morphology, regeneration, and cytoskeletal structure of muscle fibres

    PubMed Central

    Arnardottir, S; Borg, K; Ansved, T

    2004-01-01

    Methods: 14 muscle biopsies from 11 patients with s-IBM were characterised for morphological abnormalities and fibre type composition as well as muscle fibre regeneration and cytoskeletal structure, using histochemical and immunohistochemical techniques. Results: Morphological abnormalities included inflammatory infiltrates and "rimmed vacuoles," and pronounced variation in fibre size. There were no significant differences in fibre type composition between s-IBM patients and controls based on the myofibrillar ATPase staining. A differential effect on muscle fibre sizes was noted, type II fibres being smaller in the s-IBM patients than in the controls. Conversely, the mean type I muscle fibre diameter in the s-IBM patients was larger than in the controls, though this difference was not significant. An ongoing intense regeneration process was present in s-IBM muscle, as indicated by the expression of neonatal myosin heavy chain, vimentin, and CD56 (Leu-19) in most of the muscle fibres. The cytoskeletal proteins dystrophin and desmin were normally expressed in s-IBM muscle fibres that were not undergoing degeneration or regeneration. Conclusions: There are extensive morphological and morphometric alterations in s-IBM, affecting different muscle fibre types in different ways. The cytoskeletal structure of type I and II muscle fibres remains unaffected in different stages of the disease. PMID:15146016

  20. Epimorphic regeneration approach to tissue replacement in adult mammals.

    PubMed

    Agrawal, Vineet; Johnson, Scott A; Reing, Janet; Zhang, Li; Tottey, Stephen; Wang, Gang; Hirschi, Karen K; Braunhut, Susan; Gudas, Lorraine J; Badylak, Stephen F

    2010-02-23

    Urodeles and fetal mammals are capable of impressive epimorphic regeneration in a variety of tissues, whereas the typical default response to injury in adult mammals consists of inflammation and scar tissue formation. One component of epimorphic regeneration is the recruitment of resident progenitor and stem cells to a site of injury. Bioactive molecules resulting from degradation of extracellular matrix (ECM) have been shown to recruit a variety of progenitor and stem cells in vitro in adult mammals. The ability to recruit multipotential cells to the site of injury by in vivo administration of chemotactic ECM degradation products in a mammalian model of digit amputation was investigated in the present study. Adult, 6- to 8-week-old C57/BL6 mice were subjected to midsecond phalanx amputation of the third digit of the right hind foot and either treated with chemotactic ECM degradation products or left untreated. At 14 days after amputation, mice treated with ECM degradation products showed an accumulation of heterogeneous cells that expressed markers of multipotency, including Sox2, Sca1, and Rex1 (Zfp42). Cells isolated from the site of amputation were capable of differentiation along neuroectodermal and mesodermal lineages, whereas cells isolated from control mice were capable of differentiation along only mesodermal lineages. The present findings demonstrate the recruitment of endogenous stem cells to a site of injury, and/or their generation/proliferation therein, in response to ECM degradation products.

  1. Injected matrix stimulates myogenesis and regeneration of mouse skeletal muscle after ischaemic injury.

    PubMed

    Kuraitis, D; Ebadi, D; Zhang, P; Rizzuto, E; Vulesevic, B; Padavan, D T; Al Madhoun, A; McEwan, K A; Sofrenovic, T; Nicholson, K; Whitman, S C; Mesana, T G; Skerjanc, I S; Musarò, A; Ruel, M; Suuronen, E J

    2012-09-12

    Biomaterial-guided regeneration represents a novel approach for the treatment of myopathies. Revascularisation and the intramuscular extracellular matrix are important factors in stimulating myogenesis and regenerating muscle damaged by ischaemia. In this study, we used an injectable collagen matrix, enhanced with sialyl LewisX (sLeX), to guide skeletal muscle differentiation and regeneration. The elastic properties of collagen and sLeX-collagen matrices were similar to those of skeletal muscle, and culture of pluripotent mESCs on the matrices promoted their differentiation into myocyte-like cells expressing Pax3, MHC3, myogenin and Myf5. The regenerative properties of matrices were evaluated in ischaemic mouse hind-limbs. Treatment with the sLeX-matrix augmented the production of myogenic-mediated factors insulin-like growth factor (IGF)-1, and IGF binding protein-2 and -5 after 3 days. This was followed by muscle regeneration, including a greater number of regenerating myofibres and increased transcription of Six1, M-cadherin, myogenin and Myf5 after 10 days. Simultaneously, the sLeX-matrix promoted increased mobilisation and engraftment of bone marrow-derived progenitor cells, the development of larger arterioles and the restoration of tissue perfusion. Both matrix treatments tended to reduce maximal forces of ischaemic solei muscles, but sLeX-matrix lessened this loss of force and also prevented muscle fatigue. Only sLeX-matrix treatment improved mobility of mice on a treadmill. Together, these results suggest a novel approach for regenerative myogenesis, whereby treatment only with a matrix, which possesses an inherent ability to guide myogenic differentiation of pluripotent stem cells, can enhance the endogenous vascular and myogenic regeneration of skeletal muscle, thus holding promise for future clinical use.

  2. Strategies to improve regeneration of the soft palate muscles after cleft palate repair.

    PubMed

    Carvajal Monroy, Paola L; Grefte, Sander; Kuijpers-Jagtman, Anne Marie; Wagener, Frank A D T G; Von den Hoff, Johannes W

    2012-12-01

    Children with a cleft in the soft palate have difficulties with speech, swallowing, and sucking. These patients are unable to separate the nasal from the oral cavity leading to air loss during speech. Although surgical repair ameliorates soft palate function by joining the clefted muscles of the soft palate, optimal function is often not achieved. The regeneration of muscles in the soft palate after surgery is hampered because of (1) their low intrinsic regenerative capacity, (2) the muscle properties related to clefting, and (3) the development of fibrosis. Adjuvant strategies based on tissue engineering may improve the outcome after surgery by approaching these specific issues. Therefore, this review will discuss myogenesis in the noncleft and cleft palate, the characteristics of soft palate muscles, and the process of muscle regeneration. Finally, novel therapeutic strategies based on tissue engineering to improve soft palate function after surgical repair are presented.

  3. Is salamander limb regeneration really perfect? Anatomical and morphogenetic analysis of forelimb muscle regeneration in GFP-transgenic axolotls as a basis for regenerative, developmental, and evolutionary studies.

    PubMed

    Diogo, R; Nacu, E; Tanaka, E M

    2014-06-01

    The axolotl Ambystoma mexicanum is one of the most commonly used model organisms in developmental and regenerative studies because it can reconstitute what is believed to be a completely normal anatomical and functional forelimb/hindlimb after amputation. However, to date it has not been confirmed whether each regenerated forelimb muscle is really a "perfect" copy of the original muscle. This study describes the regeneration of the arm, forearm, hand, and some pectoral muscles (e.g., coracoradialis) in transgenic axolotls that express green fluorescent protein (GFP) in muscle fibers. The observations found that: (1) there were muscle anomalies in 43% of the regenerated forelimbs; (2) however, on average in each regenerated forelimb there are anomalies in only 2.5% of the total number of muscles examined, and there were no significant differences observed in the specific insertion and origin of the other muscles analyzed; (3) one of the most notable and common anomalies (seen in 35% of the regenerated forelimbs) was the presence of a fleshy coracoradialis at the level of the arm; this is a particularly outstanding configuration because in axolotls and in urodeles in general this muscle only has a thin tendon at the level of the arm, and the additional fleshy belly in the regenerated arms is strikingly similar to the fleshy biceps brachii of amniotes, suggesting a remarkable parallel between a regeneration defect and a major phenotypic change that occurred during tetrapod limb evolution; (4) during forelimb muscle regeneration there was a clear proximo-distal and radio-ulnar morphogenetic gradient, as seen in normal development, but also a ventro-dorsal gradient in the order of regeneration, which was not previously described in the literature. These results have broader implications for regenerative, evolutionary, developmental and morphogenetic studies.

  4. Aligned Nanofibers for Regenerating Arteries, Nerves, and Muscles

    NASA Astrophysics Data System (ADS)

    McClendon, Mark Trosper

    annular gap containing PA solution with a rotating rod. Using the shear aligning properties of PA solutions this rotating surface in contact with the PA solution induced a high degree of alignment in the nanofibers which was subsequently locked in place by introducing gelating calcium ions. again say something about what this fabrication procedure entails Cells encapsulated within these tubes responded to the alignment by extending in the circumferential direction mimicking the same cellular alignment observed in native arteries. A similar design strategy was also used to align nanofibers within the core of biopolymer nerve conduits, and these scaffolds were implanted in a rat sciatic nerve model. Histological and behavioral observations confirmed that PA implants sustained regeneration rates comparable to autologous grafts and significantly better than empty biopolymer grafts. Furthermore, these nanofiber gels were used as a vehicle to deliver stem cells into muscle tissue. A specialized injector was designed to introduce aligned PA gels into mouse leg muscles in a 1cm long channel. Bioluminescence and histology showed that stem cell engraftment into myofibers was greatly enhanced when delivered by PA gels compared to saline solution. The final section of this thesis describes a new series of PA molecules designed to degrade upon exposure to UV lightstate here why is this of interest in the context of the work described in the thesis. This was done to understand the degradation behavior of PA nanofibers and provide a controlled approach to changing the rheological properties post gelation.The three PA molecules in this series contained the same peptide sequence V3A3E3, while varying the location of a nitrobenzyl UV-reactive group along the backbone of the molecule. This system allowed for a quick reaction that cleaves the molecule at the reactive nitrobenzyl site without introducing any other reactive molecules. While all three molecules produced nanofibers that remained

  5. Skeletal muscle damage and impaired regeneration due to LPL-mediated lipotoxicity

    PubMed Central

    Tamilarasan, K P; Temmel, H; Das, S K; Al Zoughbi, W; Schauer, S; Vesely, P W; Hoefler, G

    2012-01-01

    According to the concept of lipotoxicity, ectopic accumulation of lipids in non-adipose tissue induces pathological changes. The most prominent effects are seen in fatty liver disease, lipid cardiomyopathy, non-insulin-dependent diabetes mellitus, insulin resistance and skeletal muscle myopathy. We used the MCK(m)-hLPL mouse distinguished by skeletal and cardiac muscle-specific human lipoprotein lipase (hLPL) overexpression to investigate effects of lipid overload in skeletal muscle. We were intrigued to find that ectopic lipid accumulation induced proteasomal activity, apoptosis and skeletal muscle damage. In line with these findings we observed reduced Musculus gastrocnemius and Musculus quadriceps mass in transgenic animals, accompanied by severely impaired physical endurance. We suggest that muscle loss was aggravated by impaired muscle regeneration as evidenced by reduced cross-sectional area of regenerating myofibers after cardiotoxin-induced injury in MCK(m)-hLPL mice. Similarly, an almost complete loss of myogenic potential was observed in C2C12 murine myoblasts upon overexpression of LPL. Our findings directly link lipid overload to muscle damage, impaired regeneration and loss of performance. These findings support the concept of lipotoxicity and are a further step to explain pathological effects seen in muscle of obese patients, patients with the metabolic syndrome and patients with cancer-associated cachexia. PMID:22825472

  6. Wnt/β-catenin signaling promotes regeneration after adult zebrafish spinal cord injury.

    PubMed

    Strand, Nicholas S; Hoi, Kimberly K; Phan, Tien M T; Ray, Catherine A; Berndt, Jason D; Moon, Randall T

    2016-09-02

    Unlike mammals, zebrafish can regenerate their injured spinal cord and regain control of caudal tissues. It was recently shown that Wnt/β-catenin signaling is necessary for spinal cord regeneration in the larval zebrafish. However, the molecular mechanisms of regeneration may or may not be conserved between larval and adult zebrafish. To test this, we assessed the role of Wnt/β-catenin signaling after spinal cord injury in the adult zebrafish. We show that Wnt/β-catenin signaling is increased after spinal cord injury in the adult zebrafish. Moreover, overexpression of Dkk1b inhibited Wnt/β-catenin signaling in the regenerating spinal cord of adult zebrafish. Dkk1b overexpression also inhibited locomotor recovery, axon regeneration, and glial bridge formation in the injured spinal cord. Thus, our data illustrate a conserved role for Wnt/β-catenin signaling in adult and larval zebrafish spinal cord regeneration.

  7. CaMKK2 Suppresses Muscle Regeneration through the Inhibition of Myoblast Proliferation and Differentiation

    PubMed Central

    Ye, Cheng; Zhang, Duo; Zhao, Lei; Li, Yan; Yao, Xiaohan; Wang, Hui; Zhang, Shengjie; Liu, Wei; Cao, Hongchao; Yu, Shuxian; Wang, Yucheng; Jiang, Jingjing; Wang, Hui; Li, Xihua; Ying, Hao

    2016-01-01

    Skeletal muscle has a major role in locomotion and muscle disorders are associated with poor regenerative efficiency. Therefore, a deeper understanding of muscle regeneration is needed to provide a new insight for new therapies. CaMKK2 plays a role in the calcium/calmodulin-dependent kinase cascade; however, its role in skeletal muscle remains unknown. Here, we found that CaMKK2 expression levels were altered under physiological and pathological conditions including postnatal myogensis, freeze or cardiotoxin-induced muscle regeneration, and Duchenne muscular dystrophy. Overexpression of CaMKK2 suppressed C2C12 myoblast proliferation and differentiation, while inhibition of CaMKK2 had opposite effect. We also found that CaMKK2 is able to activate AMPK in C2C12 myocytes. Inhibition of AMPK could attenuate the effect of CaMKK2 overexpression, while AMPK agonist could abrogate the effect of CaMKK2 knockdown on C2C12 cell differentiation and proliferation. These results suggest that CaMKK2 functions as an AMPK kinase in muscle cells and AMPK mediates the effect of CaMKK2 on myoblast proliferation and differentiation. Our data also indicate that CaMKK2 might inhibit myoblast proliferation through AMPK-mediated cell cycle arrest by inducing cdc2-Tyr15 phosphorylation and repress differentiation through affecting PGC1α transcription. Lastly, we show that overexpressing CaMKK2 in the muscle of mice via electroporation impaired the muscle regeneration during freeze-induced injury, indicating that CaMKK2 could serve as a potential target to treat patients with muscle injury or myopathies. Together, our study reveals a new role for CaMKK2 as a negative regulator of myoblast differentiation and proliferation and sheds new light on the molecular regulation of muscle regeneration. PMID:27783047

  8. CX3CR1 deficiency promotes muscle repair and regeneration by enhancing macrophage ApoE production.

    PubMed

    Arnold, Ludovic; Perrin, Hélène; de Chanville, Camille Baudesson; Saclier, Marielle; Hermand, Patricia; Poupel, Lucie; Guyon, Elodie; Licata, Fabrice; Carpentier, Wassila; Vilar, José; Mounier, Rémi; Chazaud, Bénédicte; Benhabiles, Nora; Boissonnas, Alexandre; Combadiere, Béhazine; Combadiere, Christophe

    2015-12-03

    Muscle injury triggers inflammation in which infiltrating mononuclear phagocytes are crucial for tissue regeneration. The interaction of the CCL2/CCR2 and CX3CL1/CX3CR1 chemokine axis that guides phagocyte infiltration is incompletely understood. Here, we show that CX3CR1 deficiency promotes muscle repair and rescues Ccl2(-/-) mice from impaired muscle regeneration as a result of altered macrophage function, not infiltration. Transcriptomic analysis of muscle mononuclear phagocytes reveals that Apolipoprotein E (ApoE) is upregulated in mice with efficient regeneration. ApoE treatment enhances phagocytosis by mononuclear phagocytes in vitro, and restores phagocytic activity and muscle regeneration in Ccl2(-/-) mice. Because CX3CR1 deficiency may compensate for defective CCL2-dependant monocyte recruitment by modulating ApoE-dependent macrophage phagocytic activity, targeting CX3CR1 expressed by macrophages might be a powerful therapeutic approach to improve muscle regeneration.

  9. CX3CR1 deficiency promotes muscle repair and regeneration by enhancing macrophage ApoE production

    PubMed Central

    Arnold, Ludovic; Perrin, Hélène; de Chanville, Camille Baudesson; Saclier, Marielle; Hermand, Patricia; Poupel, Lucie; Guyon, Elodie; Licata, Fabrice; Carpentier, Wassila; Vilar, José; Mounier, Rémi; Chazaud, Bénédicte; Benhabiles, Nora; Boissonnas, Alexandre; Combadiere, Béhazine; Combadiere, Christophe

    2015-01-01

    Muscle injury triggers inflammation in which infiltrating mononuclear phagocytes are crucial for tissue regeneration. The interaction of the CCL2/CCR2 and CX3CL1/CX3CR1 chemokine axis that guides phagocyte infiltration is incompletely understood. Here, we show that CX3CR1 deficiency promotes muscle repair and rescues Ccl2−/− mice from impaired muscle regeneration as a result of altered macrophage function, not infiltration. Transcriptomic analysis of muscle mononuclear phagocytes reveals that Apolipoprotein E (ApoE) is upregulated in mice with efficient regeneration. ApoE treatment enhances phagocytosis by mononuclear phagocytes in vitro, and restores phagocytic activity and muscle regeneration in Ccl2−/− mice. Because CX3CR1 deficiency may compensate for defective CCL2-dependant monocyte recruitment by modulating ApoE-dependent macrophage phagocytic activity, targeting CX3CR1 expressed by macrophages might be a powerful therapeutic approach to improve muscle regeneration. PMID:26632270

  10. Motor activity affects adult skeletal muscle re-innervation acting via tyrosine kinase receptors.

    PubMed

    Sartini, Stefano; Bartolini, Fanny; Ambrogini, Patrizia; Betti, Michele; Ciuffoli, Stefano; Lattanzi, Davide; Di Palma, Michael; Cuppini, Riccardo

    2013-05-01

    Recently, muscle expression of brain-derived neurotrophic factor (BDNF) mRNA and protein under activity control has been reported. BDNF is a neurotrophin known to be involved in axon sprouting in the CNS. Hence, we set out to study the effect of chronic treadmill mid-intensity running on adult rat muscle re-innervation, and to explore the involvement of BDNF and tropomyosin-related kinase (Trk) receptors. After nerve crush, muscle re-innervation was evaluated using intracellular recordings, tension recordings, immunostaining and Western blot analyses. An enhanced muscle multiple innervation was found in running rats that was fully reversed to control values blocking Trk receptors or interrupting the running activity. An increase in muscle multiple innervation was also found in sedentary rats treated with a selective TrkB receptor agonist. The expression of TrkB receptors by intramuscular axons was demonstrated, and increased muscle expression of BDNF was found in running animals. The increase in muscle multiple innervation was consistent with the faster muscle re-innervation that we found in running animals. We conclude that, when regenerating axons contact muscle cells, muscle activity progressively increases modulating BDNF and possibly other growth factors, which in turn, acting via Trk receptors, induce axon sprouting to re-innervate skeletal muscle.

  11. Exercise-induced stem cell activation and its implication for cardiovascular and skeletal muscle regeneration.

    PubMed

    Wahl, Patrick; Brixius, Klara; Bloch, Wilhelm

    2008-01-01

    A number of publications have provided evidence that exercise and physical activity are linked to the activation, mobilization, and differentiation of various types of stem cells. Exercise may improve organ regeneration and function. This review summarizes mechanisms by which exercise contributes to stem cell-induced regeneration in the cardiovascular and the skeletal muscle system. In addition, it discusses whether exercise may improve and support stem cell transplantation in situations of cardiovascular disease or muscular dystrophy.

  12. Matrix metalloproteinase-2 ablation in dystrophin-deficient mdx muscles reduces angiogenesis resulting in impaired growth of regenerated muscle fibers.

    PubMed

    Miyazaki, Daigo; Nakamura, Akinori; Fukushima, Kazuhiro; Yoshida, Kunihiro; Takeda, Shin'ichi; Ikeda, Shu-ichi

    2011-05-01

    Matrix metalloproteases (MMPs) are a family of endopeptidases classified into subgroups based on substrate preference in normal physiological processes such as embryonic development and tissue remodeling, as well as in various disease processes via degradation of extracellular matrix components. Among the MMPs, MMP-9 and MMP-2 have been reported to be up-regulated in skeletal muscles in the lethal X-linked muscle disorder Duchenne muscular dystrophy (DMD), which is caused by loss of dystrophin. A recent study showed that deletion of the MMP9 gene in mdx, a mouse model for DMD, improved skeletal muscle pathology and function; however, the role of MMP-2 in the dystrophin-deficient muscle is not well known. In this study, we aimed at verifying the role of MMP-2 in the dystrophin-deficient muscle by using mdx mice with genetic ablation of MMP-2 (mdx/MMP-2(-/-)). We found impairment of regenerated muscle fiber growth with reduction of angiogenesis in mdx/MMP-2(-/-) mice at 3 months of age. Expression of vascular endothelial growth factor-A (VEGF-A), an important angiogenesis-related factor, decreased in mdx/MMP-2(-/-) mice at 3 months of age. MMP-2 had not a critical role in the degradation of dystrophin-glycoprotein complex (DGC) components such as β-dystroglycan and β-sarcoglycan in the regeneration process of the dystrophic muscle. Accordingly, MMP-2 may be essential for growth of regenerated muscle fibers through VEGF-associated angiogenesis in the dystrophin-deficient skeletal muscle.

  13. Dynamic Phosphorylation of the Myocyte Enhancer Factor 2Cα1 Splice Variant Promotes Skeletal Muscle Regeneration and Hypertrophy.

    PubMed

    Baruffaldi, Fiorenza; Montarras, Didier; Basile, Valentina; De Feo, Luca; Badodi, Sara; Ganassi, Massimo; Battini, Renata; Nicoletti, Carmine; Imbriano, Carol; Musarò, Antonio; Molinari, Susanna

    2017-03-01

    The transcription factor MEF2C (Myocyte Enhancer Factor 2C) plays an established role in the early steps of myogenic differentiation. However, the involvement of MEF2C in adult myogenesis and in muscle regeneration has not yet been systematically investigated. Alternative splicing of mammalian MEF2C transcripts gives rise to two mutually exclusive protein variants: MEF2Cα2 which exerts a positive control of myogenic differentiation, and MEF2Cα1, in which the α1 domain acts as trans-repressor of the MEF2C pro-differentiation activity itself. However, MEF2Cα1 variants are persistently expressed in differentiating cultured myocytes, suggesting a role in adult myogenesis. We found that overexpression of both MEF2Cα1/α2 proteins in a mouse model of muscle injury promotes muscle regeneration and hypertrophy, with each isoform promoting different stages of myogenesis. Besides the ability of MEF2Cα2 to increase differentiation, we found that overexpressed MEF2Cα1 enhances both proliferation and differentiation of primary myoblasts, and activates the AKT/mTOR/S6K anabolic signaling pathway in newly formed myofibers. The multiple activities of MEF2Cα1 are modulated by phosphorylation of Ser98 and Ser110, two amino acid residues located in the α1 domain of MEF2Cα1. These specific phosphorylations allow the interaction of MEF2Cα1 with the peptidyl-prolyl isomerase PIN1, a regulator of MEF2C functions. Overall, in this study we established a novel regulatory mechanism in which the expression and the phosphorylation of MEF2Cα1 are critically required to sustain the adult myogenesis. The described molecular mechanism will represent a new potential target for the development of therapeutical strategies to treat muscle-wasting diseases. Stem Cells 2017;35:725-738.

  14. Muscle fiber type diversification during exercise and regeneration.

    PubMed

    Qaisar, Rizwan; Bhaskaran, Shylesh; Van Remmen, Holly

    2016-09-01

    The plasticity of skeletal muscle can be traced down to extensive metabolic, structural and molecular remodeling at the single fiber level. Skeletal muscle is comprised of different fiber types that are the basis of muscle plasticity in response to various functional demands. Resistance and endurance exercises are two external stimuli that differ in their duration and intensity of contraction and elicit markedly different responses in muscles adaptation. Further, eccentric contractions that are associated with exercise-induced injuries, elicit varied muscle adaptation and regenerative responses. Most adaptive changes are fiber type-specific and are highly influenced by diverse structural, metabolic and functional characteristics of individual fiber types. Regulation of signaling pathways by reactive oxygen species (ROS) and oxidative stress also plays an important role in muscle fiber adaptation during exercise. This review focuses on cellular and molecular responses that regulate the adaptation of skeletal muscle to exercise and exercise-related injuries.

  15. Age-associated repression of type 1 inositol 1, 4, 5-triphosphate receptor impairs muscle regeneration

    PubMed Central

    Lee, Bora; Lee, Seung-Min; Bahn, Young Jae; Lee, Kwang-Pyo; Kang, Moonkyung; Kim, Yeon-Soo; Woo, Sun-Hee; Lim, Jae-Young; Kim, Eunhee; Kwon, Ki-Sun

    2016-01-01

    Skeletal muscle mass and power decrease with age, leading to impairment of mobility and metabolism in the elderly. Ca2+ signaling is crucial for myoblast differentiation as well as muscle contraction through activation of transcription factors and Ca2+-dependent kinases and phosphatases. Ca2+ channels, such as dihydropyridine receptor (DHPR), two-pore channel (TPC) and inositol 1,4,5-triphosphate receptor (ITPR), function to maintain Ca2+ homeostasis in myoblasts. Here, we observed a significant decrease in expression of type 1 IP3 receptor (ITPR1), but not types 2 and 3, in aged mice skeletal muscle and isolated myoblasts, compared with those of young mice. ITPR1 knockdown using shRNA-expressing viruses in C2C12 myoblasts and tibialis anterior muscle of mice inhibited myotube formation and muscle regeneration after injury, respectively, a typical phenotype of aged muscle. This aging phenotype was associated with repression of muscle-specific genes and activation of the epidermal growth factor receptor (EGFR)-Ras-extracellular signal-regulated kinase (ERK) pathway. ERK inhibition by U0126 not only induced recovery of myotube formation in old myoblasts but also facilitated muscle regeneration after injury in aged muscle. The conserved decline in ITPR1 expression in aged human skeletal muscle suggests utility as a potential therapeutic target for sarcopenia, which can be treated using ERK inhibition strategies. PMID:27658230

  16. Regenerating sprouts of axotomized cat muscle afferents express characteristic firing patterns to mechanical stimulation.

    PubMed

    Johnson, R D; Munson, J B

    1991-12-01

    1. In cats, we studied the physiological properties of regenerating sprouts of muscle afferent fibers and compared them with sprouts from cutaneous afferent fibers. 2. Muscle nerves to the triceps surae and cutaneous sural nerves were axotomized in the popliteal fossa, and the proximal ends were inserted into nerve cuffs. Six days later, we recorded action potentials from single Groups I and II muscle and mostly Group II cutaneous afferents driven by mechanostimulation of the cuff. 3. Most muscle afferent sprouts (91%) had a regular slowly adapting discharge in response to sustained mechanical displacement of the cuff, particularly to sustained stretch stimuli, whereas most cutaneous afferents (92%) did not. Muscle afferents were more likely to have a spontaneous discharge and afterdischarge. 4. Group II muscle afferent sprouts had lower stretch thresholds and a higher incidence of spontaneous discharge compared with Group I fiber sprouts, whereas Group I fibers had a higher incidence of high-frequency afterdischarge to mechanical stimuli. 5. We conclude that, 6 days after axotomy, regenerating sprouts of muscle afferents, particularly Group II afferents, have become mechanosensitive in the absence of a receptor target and exhibit physiological properties similar to those found when innervating their native muscle but significantly different from sprouts of cutaneous afferents. Expression of these native muscle afferent firing patterns after the inappropriate reinnervation of hairy skin may be due to inherent properties of the muscle afferent fiber.

  17. Characterization of lymphangiogenesis in a model of adult skin regeneration.

    PubMed

    Rutkowski, Joseph M; Boardman, Kendrick C; Swartz, Melody A

    2006-09-01

    To date, adult lymphangiogenesis is not well understood. In this study we describe the evolution of lymphatic capillaries in regenerating skin and correlate lymphatic migration and organization with the expression of matrix metalloproteinases (MMPs), immune cells, the growth factors VEGF-A and VEGF-C, and the heparan sulfate proteogylcan perlecan, a key component of basement membrane. We show that while lymphatic endothelial cells (LECs) migrate and organize unidirectionally, in the direction of interstitial fluid flow, they do not sprout into the region but rather migrate as single cells that later join together into vessels. Furthermore, in a modified "shunted flow" version of the model, infiltrated LECs fail to organize into functional vessels, indicating that interstitial fluid flow is necessary for lymphatic organization. Perlecan expression on new lymphatic vessels was only observed after vessel organization was complete and also appeared first in the distal region, consistent with the directionality of lymphatic migration and organization. VEGF-C expression peaked at the initiation of lymphangiogenesis but was reduced to lower levels throughout organization and maturation. In mice lacking MMP-9, lymphatics regenerated normally, suggesting that MMP-9 is not required for lymphangiogenesis, at least in mouse skin. This study thus characterizes the process of adult lymphangiogenesis and differentiates it from sprouting blood angiogenesis, verifies its dependence on interstitial fluid flow for vessel organization, and correlates its temporal evolution with those of relevant environmental factors.

  18. Preparation and Culture of Myogenic Precursor Cells/Primary Myoblasts from Skeletal Muscle of Adult and Aged Humans.

    PubMed

    Soriano-Arroquia, Ana; Clegg, Peter D; Molloy, Andrew P; Goljanek-Whysall, Katarzyna

    2017-02-16

    Skeletal muscle homeostasis depends on muscle growth (hypertrophy), atrophy and regeneration. During ageing and in several diseases, muscle wasting occurs. Loss of muscle mass and function is associated with muscle fiber type atrophy, fiber type switching, defective muscle regeneration associated with dysfunction of satellite cells, muscle stem cells, and other pathophysiological processes. These changes are associated with changes in intracellular as well as local and systemic niches. In addition to most commonly used rodent models of muscle ageing, there is a need to study muscle homeostasis and wasting using human models, which due to ethical implications, consist predominantly of in vitro cultures. Despite the wide use of human Myogenic Progenitor Cells (MPCs) and primary myoblasts in myogenesis, there is limited data on using human primary myoblast and myotube cultures to study molecular mechanisms regulating different aspects of age-associated muscle wasting, aiding in the validation of mechanisms of ageing proposed in rodent muscle. The use of human MPCs, primary myoblasts and myotubes isolated from adult and aged people, provides a physiologically relevant model of molecular mechanisms of processes associated with muscle growth, atrophy and regeneration. Here we describe in detail a robust, inexpensive, reproducible and efficient protocol for the isolation and maintenance of human MPCs and their progeny - myoblasts and myotubes from human muscle samples using enzymatic digestion. Furthermore, we have determined the passage number at which primary myoblasts from adult and aged people undergo senescence in an in vitro culture. Finally, we show the ability to transfect these myoblasts and the ability to characterize their proliferative and differentiation capacity and propose their suitability for performing functional studies of molecular mechanisms of myogenesis and muscle wasting in vitro.

  19. Slug is a novel downstream target of MyoD. Temporal profiling in muscle regeneration.

    PubMed

    Zhao, Po; Iezzi, Simona; Carver, Ethan; Dressman, Devin; Gridley, Thomas; Sartorelli, Vittorio; Hoffman, Eric P

    2002-08-16

    Temporal expression profiling was utilized to define transcriptional regulatory pathways in vivo in a mouse muscle regeneration model. Potential downstream targets of MyoD were identified by temporal expression, promoter data base mining, and gel shift assays; Slug and calpain 6 were identified as novel MyoD targets. Slug, a member of the snail/slug family of zinc finger transcriptional repressors critical for mesoderm/ectoderm development, was further shown to be a downstream target by using promoter/reporter constructs and demonstration of defective muscle regeneration in Slug null mice.

  20. Regeneration and Maintenance of Intestinal Smooth Muscle Phenotypes

    NASA Astrophysics Data System (ADS)

    Walthers, Christopher M.

    Tissue engineering is an emerging field of biomedical engineering that involves growing artificial organs to replace those lost to disease or injury. Within tissue engineering, there is a demand for artificial smooth muscle to repair tissues of the digestive tract, bladder, and vascular systems. Attempts to develop engineered smooth muscle tissues capable of contracting with sufficient strength to be clinically relevant have so far proven unsatisfactory. The goal of this research was to develop and sustain mature, contractile smooth muscle. Survival of implanted SMCs is critical to sustain the benefits of engineered smooth muscle. Survival of implanted smooth muscle cells was studied with layered, electrospun polycaprolactone implants with lasercut holes ranging from 0--25% porosity. It was found that greater angiogenesis was associated with increased survival of implanted cells, with a large increase at a threshold between 20% and 25% porosity. Heparan sulfate coatings improved the speed of blood vessel infiltration after 14 days of implantation. With these considerations, thicker engineered tissues may be possible. An improved smooth muscle tissue culture technique was utilized. Contracting smooth muscle was produced in culture by maintaining the native smooth muscle tissue organization, specifically by sustaining intact smooth muscle strips rather than dissociating tissue in to isolated smooth muscle cells. Isolated cells showed a decrease in maturity and contained fewer enteric neural and glial cells. Muscle strips also exhibited periodic contraction and regular fluctuation of intracellular calclium. The muscle strip maturity persisted after implantation in omentum for 14 days on polycaprolactone scaffolds. A low-cost, disposable bioreactor was developed to further improve maturity of cultured smooth muscle cells in an environment of controlled cyclical stress.The bioreactor consistently applied repeated mechanical strain with controllable inputs for strain

  1. Normal and Malignant Muscle Cell Transplantation into Immune Compromised Adult Zebrafish

    PubMed Central

    Moore, John C.; Langenau, David M.

    2014-01-01

    Zebrafish have become a powerful tool for assessing development, regeneration, and cancer. More recently, allograft cell transplantation protocols have been developed that permit engraftment of normal and malignant cells into irradiated, syngeneic, and immune compromised adult zebrafish. These models when coupled with optimized cell transplantation protocols allow for the rapid assessment of stem cell function, regeneration following injury, and cancer. Here, we present a method for cell transplantation of zebrafish adult skeletal muscle and embryonal rhabdomyosarcoma (ERMS), a pediatric sarcoma that shares features with embryonic muscle, into immune compromised adult rag2E450fs homozygous mutant zebrafish. Importantly, these animals lack T cells and have reduced B cell function, facilitating engraftment of a wide range of tissues from unrelated donor animals. Our optimized protocols show that fluorescently labeled muscle cell preparations from α-actin-RFP transgenic zebrafish engraft robustly when implanted into the dorsal musculature of rag2 homozygous mutant fish. We also demonstrate engraftment of fluorescent-transgenic ERMS where fluorescence is confined to cells based on differentiation status. Specifically, ERMS were created in AB-strain myf5-GFP; mylpfa-mCherry double transgenic animals and tumors injected into the peritoneum of adult immune compromised fish. The utility of these protocols extends to engraftment of a wide range of normal and malignant donor cells that can be implanted into dorsal musculature or peritoneum of adult zebrafish. PMID:25591079

  2. S1P lyase in skeletal muscle regeneration and satellite cell activation: exposing the hidden lyase.

    PubMed

    Saba, Julie D; de la Garza-Rodea, Anabel S

    2013-01-01

    Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid whose actions are essential for many physiological processes including angiogenesis, lymphocyte trafficking and development. In addition, S1P serves as a muscle trophic factor that enables efficient muscle regeneration. This is due in part to S1P's ability to activate quiescent muscle stem cells called satellite cells (SCs) that are needed for muscle repair. However, the molecular mechanism by which S1P activates SCs has not been well understood. Further, strategies for harnessing S1P signaling to recruit SCs for therapeutic benefit have been lacking. S1P is irreversibly catabolized by S1P lyase (SPL), a highly conserved enzyme that catalyzes the cleavage of S1P at carbon bond C(2-3), resulting in formation of hexadecenal and ethanolamine-phosphate. SPL enhances apoptosis through substrate- and product-dependent events, thereby regulating cellular responses to chemotherapy, radiation and ischemia. SPL is undetectable in resting murine skeletal muscle. However, we recently found that SPL is dynamically upregulated in skeletal muscle after injury. SPL upregulation occurred in the context of a tightly orchestrated genetic program that resulted in a transient S1P signal in response to muscle injury. S1P activated quiescent SCs via a sphingosine-1-phosphate receptor 2 (S1P2)/signal transducer and activator of transcription 3 (STAT3)-dependent pathway, thereby facilitating skeletal muscle regeneration. Mdx mice, which serve as a model for muscular dystrophy (MD), exhibited skeletal muscle SPL upregulation and S1P deficiency. Pharmacological SPL inhibition raised skeletal muscle S1P levels, enhanced SC recruitment and improved mdx skeletal muscle regeneration. These findings reveal how S1P can activate SCs and indicate that SPL suppression may provide a therapeutic strategy for myopathies. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.

  3. Novel Therapeutic Effects of Non-thermal atmospheric pressure plasma for Muscle Regeneration and Differentiation

    PubMed Central

    Choi, Jae Won; Kang, Sung Un; Kim, Yang Eun; Park, Ju Kyeong; Yang, Sang Sik; Kim, Yeon Soo; Lee, Yun Sang; Lee, Yuijina; Kim, Chul-Ho

    2016-01-01

    Skeletal muscle can repair muscle tissue damage, but significant loss of muscle tissue or its long-lasting chronic degeneration makes injured skeletal muscle tissue difficult to restore. It has been demonstrated that non-thermal atmospheric pressure plasma (NTP) can be used in many biological areas including regenerative medicine. Therefore, we determined whether NTP, as a non-contact biological external stimulator that generates biological catalyzers, can induce regeneration of injured muscle without biomaterials. Treatment with NTP in the defected muscle of a Sprague Dawley (SD) rat increased the number of proliferating muscle cells 7 days after plasma treatment (dapt) and rapidly induced formation of muscle tissue and muscle cell differentiation at 14 dapt. In addition, in vitro experiments also showed that NTP could induce muscle cell proliferation and differentiation of human muscle cells. Taken together, our results demonstrated that NTP promotes restoration of muscle defects through control of cell proliferation and differentiation without biological or structural supporters, suggesting that NTP has the potential for use in muscle tissue engineering and regenerative therapies. PMID:27349181

  4. Rapid release of growth factors regenerates force output in volumetric muscle loss injuries

    PubMed Central

    Grasman, Jonathan M.; Do, Duc M.; Page, Raymond L.; Pins, George D.

    2015-01-01

    A significant challenge in the design and development of biomaterial scaffolds is to incorporate mechanical and biochemical cues to direct organized tissue growth. In this study, we investigated the effect of hepatocyte growth factor (HGF) loaded, crosslinked fibrin (EDCn-HGF) microthread scaffolds on skeletal muscle regeneration in a mouse model of volumetric muscle loss (VML). The rapid, sustained release of HGF significantly enhanced the force production of muscle tissue 60 days after injury, recovering more than 200% of the force output relative to measurements recorded immediately after injury. HGF delivery increased the number of differentiating myoblasts 14 days after injury, and supported an enhanced angiogenic response. The architectural morphology of microthread scaffolds supported the ingrowth of nascent myofibers into the wound site, in contrast to fibrin gel implants which did not support functional regeneration. Together, these data suggest that EDCn-HGF microthreads recapitulate several of the regenerative cues lost in VML injuries, promote remodeling of functional muscle tissue, and enhance the functional regeneration of skeletal muscle. Further, by strategically incorporating specific biochemical factors and precisely tuning the structural and mechanical properties of fibrin microthreads, we have developed a powerful platform technology that may enhance regeneration in other axially aligned tissues. PMID:26344363

  5. Transgenic overexpression of ADAM12 suppresses muscle regeneration and aggravates dystrophy in aged mdx mice.

    PubMed

    Jørgensen, Louise Helskov; Jensen, Charlotte Harken; Wewer, Ulla M; Schrøder, Henrik Daa

    2007-11-01

    Muscular dystrophies are characterized by insufficient restoration and gradual replacement of the skeletal muscle by fat and connective tissue. ADAM12 has previously been shown to alleviate the pathology of young dystrophin-deficient mdx mice, a model for Duchenne muscular dystrophy. The observed effect of ADAM12 was suggested to be mediated via a membrane-stabilizing up-regulation of utrophin, alpha7B integrin, and dystroglycans. Ectopic ADAM12 expression in normal mouse skeletal muscle also improved regeneration after freeze injury, presumably by the same mechanism. Hence, it was suggested that ADAM12 could be a candidate for nonreplacement gene therapy of Duchenne muscular dystrophy. We therefore evaluated the long-term effect of ADAM12 overexpression in muscle. Surprisingly, we observed loss of skeletal muscle and accelerated fibrosis and adipogenesis in 1-year-old mdx mice transgenically overexpressing ADAM12 (ADAM12(+)/mdx mice), even though their utrophin levels were mildly elevated compared with age-matched controls. Thus, membrane stabilization was not sufficient to provide protection during prolonged disease. Consequently, we reinvestigated skeletal muscle regeneration in ADAM12 transgenic mice (ADAM12(+)) after a knife cut lesion and observed that the regeneration process was significantly impaired. ADAM12 seemed to inhibit the satellite cell response and delay myoblast differentiation. These results discourage long-term therapeutic use of ADAM12. They also point to impaired regeneration as a possible factor in development of muscular dystrophy.

  6. Skeletal Muscle Regeneration and Oxidative Stress Are Altered in Chronic Kidney Disease

    PubMed Central

    Chen, Neal X.; Organ, Jason M.; Zarse, Chad; O’Neill, Kalisha; Conway, Richard G.; Konrad, Robert J.; Bacallao, Robert L.; Allen, Matthew R.; Moe, Sharon M.

    2016-01-01

    Skeletal muscle atrophy and impaired muscle function are associated with lower health-related quality of life, and greater disability and mortality risk in those with chronic kidney disease (CKD). However, the pathogenesis of skeletal dysfunction in CKD is unknown. We used a slow progressing, naturally occurring, CKD rat model (Cy/+ rat) with hormonal abnormalities consistent with clinical presentations of CKD to study skeletal muscle signaling. The CKD rats demonstrated augmented skeletal muscle regeneration with higher activation and differentiation signals in muscle cells (i.e. lower Pax-7; higher MyoD and myogenin RNA expression). However, there was also higher expression of proteolytic markers (Atrogin-1 and MuRF-1) in CKD muscle relative to normal. CKD animals had higher indices of oxidative stress compared to normal, evident by elevated plasma levels of an oxidative stress marker, 8-hydroxy-2' -deoxyguanosine (8-OHdG), increased muscle expression of succinate dehydrogenase (SDH) and Nox4 and altered mitochondria morphology. Furthermore, we show significantly higher serum levels of myostatin and expression of myostatin in skeletal muscle of CKD animals compared to normal. Taken together, these data show aberrant regeneration and proteolytic signaling that is associated with oxidative stress and high levels of myostatin in the setting of CKD. These changes likely play a role in the compromised skeletal muscle function that exists in CKD. PMID:27486747

  7. UTX demethylase activity is required for satellite cell–mediated muscle regeneration

    PubMed Central

    Wang, Chaochen; Nakka, Kiran; Benyoucef, Aissa; Sebastian, Soji; Zhuang, Lenan; Chu, Alphonse; Palii, Carmen G.; Camellato, Brendan; Brand, Marjorie

    2016-01-01

    The X chromosome–encoded histone demethylase UTX (also known as KDM6A) mediates removal of repressive trimethylation of histone H3 lysine 27 (H3K27me3) to establish transcriptionally permissive chromatin. Loss of UTX in female mice is embryonic lethal. Unexpectedly, male UTX-null mice escape embryonic lethality due to expression of UTY, a paralog that lacks H3K27 demethylase activity, suggesting an enzyme-independent role for UTX in development and thereby challenging the need for active H3K27 demethylation in vivo. However, the requirement for active H3K27 demethylation in stem cell–mediated tissue regeneration remains untested. Here, we employed an inducible mouse KO that specifically ablates Utx in satellite cells (SCs) and demonstrated that active H3K27 demethylation is necessary for muscle regeneration. Loss of UTX in SCs blocked myofiber regeneration in both male and female mice. Furthermore, we demonstrated that UTX mediates muscle regeneration through its H3K27 demethylase activity, as loss of demethylase activity either by chemical inhibition or knock-in of demethylase-dead UTX resulted in defective muscle repair. Mechanistically, dissection of the muscle regenerative process revealed that the demethylase activity of UTX is required for expression of the transcription factor myogenin, which in turn drives differentiation of muscle progenitors. Thus, we have identified a critical role for the enzymatic activity of UTX in activating muscle-specific gene expression during myofiber regeneration and have revealed a physiological role for active H3K27 demethylation in vivo. PMID:26999603

  8. Correlation of Utrophin Levels with the Dystrophin Protein Complex and Muscle Fibre Regeneration in Duchenne and Becker Muscular Dystrophy Muscle Biopsies

    PubMed Central

    Janghra, Narinder; Morgan, Jennifer E.; Sewry, Caroline A.; Wilson, Francis X.; Davies, Kay E.; Muntoni, Francesco; Tinsley, Jonathon

    2016-01-01

    Duchenne muscular dystrophy is a severe and currently incurable progressive neuromuscular condition, caused by mutations in the DMD gene that result in the inability to produce dystrophin. Lack of dystrophin leads to loss of muscle fibres and a reduction in muscle mass and function. There is evidence from dystrophin-deficient mouse models that increasing levels of utrophin at the muscle fibre sarcolemma by genetic or pharmacological means significantly reduces the muscular dystrophy pathology. In order to determine the efficacy of utrophin modulators in clinical trials, it is necessary to accurately measure utrophin levels and other biomarkers on a fibre by fibre basis within a biopsy section. Our aim was to develop robust and reproducible staining and imaging protocols to quantify sarcolemmal utrophin levels, sarcolemmal dystrophin complex members and numbers of regenerating fibres within a biopsy section. We quantified sarcolemmal utrophin in mature and regenerating fibres and the percentage of regenerating muscle fibres, in muscle biopsies from Duchenne, the milder Becker muscular dystrophy and controls. Fluorescent immunostaining followed by image analysis was performed to quantify utrophin intensity and β-dystrogylcan and ɣ –sarcoglycan intensity at the sarcolemma. Antibodies to fetal and developmental myosins were used to identify regenerating muscle fibres allowing the accurate calculation of percentage regeneration fibres in the biopsy. Our results indicate that muscle biopsies from Becker muscular dystrophy patients have fewer numbers of regenerating fibres and reduced utrophin intensity compared to muscle biopsies from Duchenne muscular dystrophy patients. Of particular interest, we show for the first time that the percentage of regenerating muscle fibres within the muscle biopsy correlate with the clinical severity of Becker and Duchenne muscular dystrophy patients from whom the biopsy was taken. The ongoing development of these tools to quantify

  9. Correlation of Utrophin Levels with the Dystrophin Protein Complex and Muscle Fibre Regeneration in Duchenne and Becker Muscular Dystrophy Muscle Biopsies.

    PubMed

    Janghra, Narinder; Morgan, Jennifer E; Sewry, Caroline A; Wilson, Francis X; Davies, Kay E; Muntoni, Francesco; Tinsley, Jonathon

    2016-01-01

    Duchenne muscular dystrophy is a severe and currently incurable progressive neuromuscular condition, caused by mutations in the DMD gene that result in the inability to produce dystrophin. Lack of dystrophin leads to loss of muscle fibres and a reduction in muscle mass and function. There is evidence from dystrophin-deficient mouse models that increasing levels of utrophin at the muscle fibre sarcolemma by genetic or pharmacological means significantly reduces the muscular dystrophy pathology. In order to determine the efficacy of utrophin modulators in clinical trials, it is necessary to accurately measure utrophin levels and other biomarkers on a fibre by fibre basis within a biopsy section. Our aim was to develop robust and reproducible staining and imaging protocols to quantify sarcolemmal utrophin levels, sarcolemmal dystrophin complex members and numbers of regenerating fibres within a biopsy section. We quantified sarcolemmal utrophin in mature and regenerating fibres and the percentage of regenerating muscle fibres, in muscle biopsies from Duchenne, the milder Becker muscular dystrophy and controls. Fluorescent immunostaining followed by image analysis was performed to quantify utrophin intensity and β-dystrogylcan and ɣ -sarcoglycan intensity at the sarcolemma. Antibodies to fetal and developmental myosins were used to identify regenerating muscle fibres allowing the accurate calculation of percentage regeneration fibres in the biopsy. Our results indicate that muscle biopsies from Becker muscular dystrophy patients have fewer numbers of regenerating fibres and reduced utrophin intensity compared to muscle biopsies from Duchenne muscular dystrophy patients. Of particular interest, we show for the first time that the percentage of regenerating muscle fibres within the muscle biopsy correlate with the clinical severity of Becker and Duchenne muscular dystrophy patients from whom the biopsy was taken. The ongoing development of these tools to quantify

  10. Upregulation of HARP during in vitro myogenesis and rat soleus muscle regeneration.

    PubMed

    Caruelle, Danièle; Mazouzi, Zohra; Husmann, Irene; Delbé, Jean; Duchesnay, Arlette; Gautron, Jean; Martelly, Isabelle; Courty, José

    2004-01-01

    Heparin affin regulatory peptide (HARP) is a heparin binding growth factor that belongs to a family of molecule whose biological function in myogenesis has been suspected without formal demonstration. In the present study, we investigated the expression and the distribution of HARP and its mRNA during soleus muscle regeneration using a crushed-induced regeneration model and also during differentiation of muscle satellite cells in primary cultures. We show that HARP mRNA and protein expression are increased during the regeneration process with a peak at day 5 after muscle crushing when new myotubes are formed. In situ hybridization and immunohistochemical studies showed that activated myoblasts expressed HARP at day two after crushing. Five days after muscle lesion, HARP is localised in newly formed myotubes as well as in prefused activated myoblasts. In regenerated myofibers, 15 days after crushing, expression of HARP was reduced. In vitro experiments using primary cultures of rat satellite cells indicated that HARP expression level increased during the differentiation process and peaked on fusion of myoblasts into myotubes. This is the first study demonstrating the presence of HARP in fusing myogenic cells suggests that this growth factor could play a function in myogenic differentiation.

  11. mTOR is necessary for proper satellite cell activity and skeletal muscle regeneration.

    PubMed

    Zhang, Pengpeng; Liang, Xinrong; Shan, Tizhong; Jiang, Qinyang; Deng, Changyan; Zheng, Rong; Kuang, Shihuan

    The serine/threonine kinase mammalian target of rapamycin (mTOR) is a key regulator of protein synthesis, cell proliferation and energy metabolism. As constitutive deletion of Mtor gene results in embryonic lethality, the function of mTOR in muscle stem cells (satellite cells) and skeletal muscle regeneration remains to be determined. In this study, we established a satellite cell specific Mtor conditional knockout (cKO) mouse model by crossing Pax7(CreER) and Mtor(flox/flox) mice. Skeletal muscle regeneration after injury was severely compromised in the absence of Mtor, indicated by increased number of necrotic myofibers infiltrated by Evans blue dye, and reduced number and size of regenerated myofibers in the Mtor cKO mice compared to wild type (WT) littermates. To dissect the cellular mechanism, we analyzed satellite cell-derived primary myoblasts grown on single myofibers or adhered to culture plates. The Mtor cKO myoblasts exhibited defective proliferation and differentiation kinetics when compared to myoblasts derived from WT littermates. At the mRNA and protein levels, the Mtor cKO myoblasts expressed lower levels of key myogenic determinant genes Pax7, Myf5, Myod, Myog than did the WT myoblasts. These results suggest that mTOR is essential for satellite cell function and skeletal muscle regeneration through controlling the expression of myogenic genes.

  12. Nfix Regulates Temporal Progression of Muscle Regeneration through Modulation of Myostatin Expression.

    PubMed

    Rossi, Giuliana; Antonini, Stefania; Bonfanti, Chiara; Monteverde, Stefania; Vezzali, Chiara; Tajbakhsh, Shahragim; Cossu, Giulio; Messina, Graziella

    2016-03-08

    Nfix belongs to a family of four highly conserved proteins that act as transcriptional activators and/or repressors of cellular and viral genes. We previously showed a pivotal role for Nfix in regulating the transcriptional switch from embryonic to fetal myogenesis. Here, we show that Nfix directly represses the Myostatin promoter, thus controlling the proper timing of satellite cell differentiation and muscle regeneration. Nfix-null mice display delayed regeneration after injury, and this deficit is reversed upon in vivo Myostatin silencing. Conditional deletion of Nfix in satellite cells results in a similar delay in regeneration, confirming the functional requirement for Nfix in satellite cells. Moreover, mice lacking Nfix show reduced myofiber cross sectional area and a predominant slow twitching phenotype. These data define a role for Nfix in postnatal skeletal muscle and unveil a mechanism for Myostatin regulation, thus providing insights into the modulation of its complex signaling pathway.

  13. Nfix Regulates Temporal Progression of Muscle Regeneration through Modulation of Myostatin Expression

    PubMed Central

    Rossi, Giuliana; Antonini, Stefania; Bonfanti, Chiara; Monteverde, Stefania; Vezzali, Chiara; Tajbakhsh, Shahragim; Cossu, Giulio; Messina, Graziella

    2016-01-01

    Summary Nfix belongs to a family of four highly conserved proteins that act as transcriptional activators and/or repressors of cellular and viral genes. We previously showed a pivotal role for Nfix in regulating the transcriptional switch from embryonic to fetal myogenesis. Here, we show that Nfix directly represses the Myostatin promoter, thus controlling the proper timing of satellite cell differentiation and muscle regeneration. Nfix-null mice display delayed regeneration after injury, and this deficit is reversed upon in vivo Myostatin silencing. Conditional deletion of Nfix in satellite cells results in a similar delay in regeneration, confirming the functional requirement for Nfix in satellite cells. Moreover, mice lacking Nfix show reduced myofiber cross sectional area and a predominant slow twitching phenotype. These data define a role for Nfix in postnatal skeletal muscle and unveil a mechanism for Myostatin regulation, thus providing insights into the modulation of its complex signaling pathway. PMID:26923583

  14. Genetic disruption of Smad7 impairs skeletal muscle growth and regeneration

    PubMed Central

    Cohen, Tatiana V; Kollias, Helen D; Liu, Naili; Ward, Christopher W; Wagner, Kathryn R

    2015-01-01

    The transforming growth factor-β (TGF-β) family of growth factors plays an essential role in mediating cellular growth and differentiation. Myostatin is a muscle-specific member of the TGF-β superfamily and a negative regulator of muscle growth. Myostatin inhibitors are currently being pursued as therapeutic options for muscle disorders. Smad7 inhibits intracellular myostatin signalling via Smad2/3, and thus presents a means of regulating myostatin and potentiating muscle growth. We investigated the functional loss of Smad7 on muscle in vivo by examining muscle growth and differentiation in mice deficient in Smad7 (Smad7−/−). Smad7−/− mice showed reduced muscle mass, hypotrophy and hypoplasia of muscle fibres, as well as an increase in oxidative fibre types. Examination of muscle strength showed reduced force generation in vivo and ex vivo compared to wild-type controls. Analysis of muscle regeneration showed a delay in recovery, probably as a result of decreased activation, proliferation and differentiation of satellite cells, as confirmed in vitro. Additionally, myostatin expression was upregulated in Smad7−/− muscle. Our findings suggest that increased Smad2/3 signalling in the absence of Smad7 inhibition impedes muscle growth and regeneration. Taken together, our experiments demonstrate that Smad7 is an important mediator of muscle growth in vivo. Our studies enhance our understanding of in vivo TGF-β pathway modulation and suggest that Smad7 may be an important therapeutic target for muscle disorders. Key points Smad7 is an intracellular antagonist of transforming growth factor-β signalling pathways and modulates muscle growth in vivo. Loss of Smad7 results in decreased muscle mass, reduced force generation, fibre type switching from glycolytic towards oxidative type and delayed recovery from injury. Upregulated Smad2/3 signalling in Smad7−/− muscle results in reduced myoblast proliferation and differentiation. Smad7 is an important

  15. mTOR is necessary for proper satellite cell activity and skeletal muscle regeneration

    SciTech Connect

    Zhang, Pengpeng; Liang, Xinrong; Shan, Tizhong; Jiang, Qinyang; Deng, Changyan; Zheng, Rong; Kuang, Shihuan

    2015-07-17

    The serine/threonine kinase mammalian target of rapamycin (mTOR) is a key regulator of protein synthesis, cell proliferation and energy metabolism. As constitutive deletion of Mtor gene results in embryonic lethality, the function of mTOR in muscle stem cells (satellite cells) and skeletal muscle regeneration remains to be determined. In this study, we established a satellite cell specific Mtor conditional knockout (cKO) mouse model by crossing Pax7{sup CreER} and Mtor{sup flox/flox} mice. Skeletal muscle regeneration after injury was severely compromised in the absence of Mtor, indicated by increased number of necrotic myofibers infiltrated by Evans blue dye, and reduced number and size of regenerated myofibers in the Mtor cKO mice compared to wild type (WT) littermates. To dissect the cellular mechanism, we analyzed satellite cell-derived primary myoblasts grown on single myofibers or adhered to culture plates. The Mtor cKO myoblasts exhibited defective proliferation and differentiation kinetics when compared to myoblasts derived from WT littermates. At the mRNA and protein levels, the Mtor cKO myoblasts expressed lower levels of key myogenic determinant genes Pax7, Myf5, Myod, Myog than did the WT myoblasts. These results suggest that mTOR is essential for satellite cell function and skeletal muscle regeneration through controlling the expression of myogenic genes. - Highlights: • Pax7{sup CreER} was used to delete Mtor gene in satellite cells. • Satellite cell specific deletion of Mtor impairs muscle regeneration. • mTOR is necessary for satellite cell proliferation and differentiation. • Deletion of Mtor leads to reduced expression of key myogenic genes.

  16. BRE facilitates skeletal muscle regeneration by promoting satellite cell motility and differentiation.

    PubMed

    Xiao, Lihai; Lee, Kenneth Ka Ho

    2016-01-06

    The function of the Bre gene in satellite cells was investigated during skeletal muscle regeneration. The tibialis anterior leg muscle was experimentally injured in Bre knockout mutant (BRE-KO) mice. It was established that the accompanying muscle regeneration was impaired as compared with their normal wild-type counterparts (BRE-WT). There were significantly fewer pax7(+) satellite cells and smaller newly formed myofibers present in the injury sites of BRE-KO mice. Bre was required for satellite cell fusion and myofiber formation. The cell fusion index and average length of newly-formed BRE-KO myofibers were found to be significantly reduced as compared with BRE-WT myofibers. It is well established that satellite cells are highly invasive which confers on them the homing ability to reach the muscle injury sites. Hence, we tracked the migratory behavior of these cells using time-lapse microscopy. Image analysis revealed no difference in directionality of movement between BRE-KO and BRE-WT satellite cells but there was a significant decrease in the velocity of BRE-KO cell movement. Moreover, chemotactic migration assays indicated that BRE-KO satellite cells were significantly less responsive to chemoattractant SDF-1α than BRE-WT satellite cells. We also established that BRE normally protects CXCR4 from SDF-1α-induced degradation. In sum, BRE facilitates skeletal muscle regeneration by enhancing satellite cell motility, homing and fusion.

  17. HMGB1 expression and muscle regeneration in idiopathic inflammatory myopathies and degenerative joint diseases.

    PubMed

    Cseri, Karolina; Vincze, János; Cseri, Julianna; Fodor, János; Csernátony, Zoltán; Csernoch, László; Dankó, Katalin

    2015-06-01

    The High-Mobility Group Box 1 protein (HMGB1) is a known nuclear protein which may be released from the nucleus into the cytoplasm and the extracellular space. It is believed that the mobilized HMGB1 plays role in the autoimmune processes as an alarmin, stimulating the immune response. In addition, muscle regeneration and differentiation may also be altered in the inflammatory surroundings. Biopsy specimens derived from patients with idiopathic inflammatory myopathies (IIM) such as polymyositis or dermatomyositis were compared to muscle samples from patients undergoing surgical interventions for coxarthrosis. The biopsy and surgery specimens were used for Western blot analysis, for immunohistochemical detection of HMGB1 in histological preparations and for cell culturing to examine cell proliferation and differentiation. Our data show lower HMGB1 expression, impaired proliferation and slightly altered fusion capacity in the primary cell cultures started from IIM specimens than in cultures of coxarthrotic muscles. The ratio of regenerating muscle fibres with centralised nuclei (myotubes) is lower in the IIM samples than in the coxarthrotic ones but corticosteroid treatment shifts the ratio towards the coxarthrotic value. Our data suggest that the impaired regeneration capacity should also be considered to be behind the muscle weakness in IIM patients. The role of HMGB1 as a pathogenic signal requires further investigation.

  18. In situ regeneration of skeletal muscle tissue through host cell recruitment.

    PubMed

    Ju, Young Min; Atala, Anthony; Yoo, James J; Lee, Sang Jin

    2014-10-01

    Standard reconstructive procedures for restoring normal function after skeletal muscle defects involve the use of existing host tissues such as muscular flaps. In many instances, this approach is not feasible and delays the rehabilitation process and restoration of tissue function. Currently, cell-based tissue engineering strategies have been used for reconstruction; however, donor tissue biopsy and ex vivo cell manipulation are required prior to implantation. The present study aimed to overcome these limitations by demonstrating mobilization of muscle cells into a target-specific site for in situ muscle regeneration. First, we investigated whether host muscle cells could be mobilized into an implanted scaffold. Poly(l-lactic acid) (PLLA) scaffolds were implanted in the tibialis anterior (TA) muscle of rats, and the retrieved scaffolds were characterized by examining host cell infiltration in the scaffolds. The host cell infiltrates, including Pax7+ cells, gradually increased with time. Second, we demonstrated that host muscle cells could be enriched by a myogenic factor released from the scaffolds. Gelatin-based scaffolds containing a myogenic factor were implanted in the TA muscle of rats, and the Pax7+ cell infiltration and newly formed muscle fibers were examined. By the second week after implantation, the Pax7+ cell infiltrates and muscle formation were significantly accelerated within the scaffolds containing insulin-like growth factor 1 (IGF-1). Our data suggest an ability of host stem cells to be recruited into the scaffolds with the capability of differentiating to muscle cells. In addition, the myogenic factor effectively promoted host cell recruitment, which resulted in accelerating muscle regeneration in situ.

  19. Effects of S1P on skeletal muscle repair/regeneration during eccentric contraction.

    PubMed

    Sassoli, Chiara; Formigli, Lucia; Bini, Francesca; Tani, Alessia; Squecco, Roberta; Battistini, Chiara; Zecchi-Orlandini, Sandra; Francini, Fabio; Meacci, Elisabetta

    2011-11-01

    Skeletal muscle regeneration is severely compromised in the case of extended damage. The current challenge is to find factors capable of limiting muscle degeneration and/or potentiating the inherent regenerative program mediated by a specific type of myoblastic cells, the satellite cells. Recent studies from our groups and others have shown that the bioactive lipid, sphingosine 1-phosphate (S1P), promotes myoblast differentiation and exerts a trophic action on denervated skeletal muscle fibres. In the present study, we examined the effects of S1P on eccentric contraction (EC)-injured extensor digitorum longus muscle fibres and resident satellite cells. After EC, skeletal muscle showed evidence of structural and biochemical damage along with significant electrophysiological changes, i.e. reduced plasma membrane resistance and resting membrane potential and altered Na(+) and Ca(2+) current amplitude and kinetics. Treatment with exogenous S1P attenuated the EC-induced tissue damage, protecting skeletal muscle fibre from apoptosis, preserving satellite cell viability and affecting extracellular matrix remodelling, through the up-regulation of matrix metalloproteinase 9 (MMP-9) expression. S1P also promoted satellite cell renewal and differentiation in the damaged muscle. Notably, EC was associated with the activation of sphingosine kinase 1 (SphK1) and with increased endogenous S1P synthesis, further stressing the relevance of S1P in skeletal muscle protection and repair/regeneration. In line with this, the treatment with a selective SphK1 inhibitor during EC, caused an exacerbation of the muscle damage and attenuated MMP-9 expression. Together, these findings are in favour for a role of S1P in skeletal muscle healing and offer new clues for the identification of novel therapeutic approaches to counteract skeletal muscle damage and disease.

  20. Morphology of nerve fiber regeneration along a biodegradable poly (DLLA-epsilon-CL) nerve guide filled with fresh skeletal muscle.

    PubMed

    Varejão, Artur S P; Cabrita, António M; Meek, Marcel F; Fornaro, Michele; Geuna, Stefano; Giacobini-Robecchi, Maria G

    2003-01-01

    Previous morphological and morphometrical studies showed that fresh-skeletal-muscle-enriched vein segments are good conduits for leading peripheral nerve regeneration. In the present study, we investigated the morphological features of peripheral nerve fibers regenerated along a 10-mm-long biodegradable poly (DLLA-epsilon-CL) nerve guide enriched with fresh skeletal muscle, comparing them to nerve fiber regeneration along 10-mm-long phosphate-buffered saline (PBS)-enriched poly (DLLA-epsilon-CL) tubes. Repaired nerves were analyzed at weeks 6 and 24 postoperatively. Structural and ultrastructural observation showed that good nerve fiber regeneration occurred in both PBS-enriched and fresh-skeletal-muscle-enriched nerve guides, and histomorphometrical analysis of regenerated myelinated fibers revealed no statistically significant differences between the two experimental groups at week 24 after surgery. The employment of fresh-muscle-enriched conduits for the repair of nerve defects is critically discussed in the light of these results.

  1. Calpain 3 Expression Pattern during Gastrocnemius Muscle Atrophy and Regeneration Following Sciatic Nerve Injury in Rats

    PubMed Central

    Wu, Ronghua; Yan, Yingying; Yao, Jian; Liu, Yan; Zhao, Jianmei; Liu, Mei

    2015-01-01

    Calpain 3 (CAPN3), also known as p94, is a skeletal muscle-specific member of the calpain family that is involved in muscular dystrophy; however, the roles of CAPN3 in muscular atrophy and regeneration are yet to be understood. In the present study, we attempted to explain the effect of CAPN3 in muscle atrophy by evaluating CAPN3 expression in rat gastrocnemius muscle following reversible sciatic nerve injury. After nerve injury, the wet weight ratio and cross sectional area (CSA) of gastrocnemius muscle were decreased gradually from 1–14 days and then recovery from 14–28 days. The active form of CAPN3 (~62 kDa) protein decreased slightly on day 3 and then increased from day 7 to 14 before a decrease from day 14 to 28. The result of linear correlation analysis showed that expression of the active CAPN3 protein level was negatively correlated with muscle wet weight ratio. CAPN3 knockdown by short interfering RNA (siRNA) injection improved muscle recovery on days 7 and 14 after injury as compared to that observed with control siRNA treatment. Depletion of CAPN3 gene expression could promote myoblast differentiation in L6 cells. Based on these findings, we conclude that the expression pattern of the active CAPN3 protein is linked to muscle atrophy and regeneration following denervation: its upregulation during early stages may promote satellite cell renewal by inhibiting differentiation, whereas in later stages, CAPN3 expression may be downregulated to stimulate myogenic differentiation and enhance recovery. These results provide a novel mechanistic insight into the role of CAPN3 protein in muscle regeneration after peripheral nerve injury. PMID:26569227

  2. Skeletal muscle regeneration is delayed by reduction in Xin expression: consequence of impaired satellite cell activation?

    PubMed

    Nissar, Aliyah A; Zemanek, Bart; Labatia, Rita; Atkinson, Daniel J; van der Ven, Peter F M; Fürst, Dieter O; Hawke, Thomas J

    2012-01-01

    Xin is a striated muscle-specific actin-binding protein whose mRNA expression has been observed in damaged skeletal muscle. Here we demonstrate increased Xin protein expression early postinjury (≤ 12 h) and localization primarily to the periphery of damaged myofibers. At 1 day postinjury, Xin is colocalized with MyoD, confirming expression in activated satellite cells (SCs). By 5 days postinjury, Xin is evident in newly regenerated myofibers, with a return to preinjury levels by 14 days of regeneration. To determine whether the increased Xin expression is functionally relevant, tibialis anterior muscles of wild-type mice were infected with Xin-short hairpin RNA (shRNA) adenovirus, whereas the contralateral tibialis anterior received control adenovirus (Control). Four days postinfection, muscles were harvested or injured with cardiotoxin and collected at 3, 5, or 14 days thereafter. When compared with Control, Xin-shRNA infection attenuated muscle regeneration as demonstrated by Myh3 expression and fiber areas. Given the colocalization of Xin and MyoD, we isolated single myofibers from infected muscles to investigate the effect of silencing Xin on SC function. Relative to Control, SC activation, but not proliferation, was significantly impaired in Xin-shRNA-infected muscles. To determine whether Xin affects the G0-G1 transition, cell cycle reentry was assessed on infected C2C12 myoblasts using a methylcellulose assay. No difference in reentry was noted between groups, suggesting that Xin contributes to SC activation by means other than affecting G0-G1 transition. Together these data demonstrate a critical role for Xin in SC activation and reduction in Xin expression results in attenuated skeletal muscle repair.

  3. Persistent Muscle Fiber Regeneration in Long Term Denervation. Past, Present, Future

    PubMed Central

    Carraro, Ugo; Boncompagni, Simona; Gobbo, Valerio; Rossini, Katia; Zampieri, Sandra; Mosole, Simone; Ravara, Barbara; Nori, Alessandra; Stramare, Roberto; Ambrosio, Francesco; Piccione, Francesco; Masiero, Stefano; Vindigni, Vincenzo; Gargiulo, Paolo; Protasi, Feliciano; Kern, Helmut; Pond, Amber

    2015-01-01

    Despite the ravages of long term denervation there is structural and ultrastructural evidence for survival of muscle fibers in mammals, with some fibers surviving at least ten months in rodents and 3-6 years in humans. Further, in rodents there is evidence that muscle fibers may regenerate even after repeated damage in the absence of the nerve, and that this potential is maintained for several months after denervation. While in animal models permanently denervated muscle sooner or later loses the ability to contract, the muscles may maintain their size and ability to function if electrically stimulated soon after denervation. Whether in mammals, humans included, this is a result of persistent de novo formation of muscle fibers is an open issue we would like to explore in this review. During the past decade, we have studied muscle biopsies from the quadriceps muscle of Spinal Cord Injury (SCI) patients suffering with Conus and Cauda Equina syndrome, a condition that fully and irreversibly disconnects skeletal muscle fibers from their damaged innervating motor neurons. We have demonstrated that human denervated muscle fibers survive years of denervation and can be rescued from severe atrophy by home-based Functional Electrical Stimulation (h-bFES). Using immunohistochemistry with both non-stimulated and the h-bFES stimulated human muscle biopsies, we have observed the persistent presence of muscle fibers which are positive to labeling by an antibody which specifically recognizes the embryonic myosin heavy chain (MHCemb). Relative to the total number of fibers present, only a small percentage of these MHCemb positive fibers are detected, suggesting that they are regenerating muscle fibers and not pre-existing myofibers re-expressing embryonic isoforms. Although embryonic isoforms of acetylcholine receptors are known to be re-expressed and to spread from the end-plate to the sarcolemma of muscle fibers in early phases of muscle denervation, we suggest that the MHCemb

  4. Targeted ablation of IKK2 improves skeletal muscle strength, maintains mass, and promotes regeneration

    PubMed Central

    Mourkioti, Foteini; Kratsios, Paschalis; Luedde, Tom; Song, Yao-Hua; Delafontaine, Patrick; Adami, Raffaella; Parente, Valeria; Bottinelli, Roberto; Pasparakis, Manolis; Rosenthal, Nadia

    2006-01-01

    NF-κB is a major pleiotropic transcription factor modulating immune, inflammatory, cell survival, and proliferative responses, yet the relevance of NF-κB signaling in muscle physiology and disease is less well documented. Here we show that muscle-restricted NF-κB inhibition in mice, through targeted deletion of the activating kinase inhibitor of NF-κB kinase 2 (IKK2), shifted muscle fiber distribution and improved muscle force. In response to denervation, IKK2 depletion protected against atrophy, maintaining fiber type, size, and strength, increasing protein synthesis, and decreasing protein degradation. IKK2-depleted mice with a muscle-specific transgene expressing a local Igf-1 isoform (mIgf-1) showed enhanced protection against muscle atrophy. In response to muscle damage, IKK2 depletion facilitated skeletal muscle regeneration through enhanced satellite cell activation and reduced fibrosis. Our results establish IKK2/NF-κB signaling as an important modulator of muscle homeostasis and suggest a combined role for IKK inhibitors and growth factors in the therapy of muscle diseases. PMID:17080195

  5. Satellite cell activity in muscle regeneration after contusion in rats.

    PubMed

    Srikuea, Ratchakrit; Pholpramool, Chumpol; Kitiyanant, Yindee; Yimlamai, Tossaporn

    2010-11-01

    1. The role of satellite cells in muscle growth during development is well documented, but the involvement of these cells in muscle repair after contusion is less well known. In the present study, we investigated the time-course of satellite cell activity (from 3h to 7days) after contusion of rat gastrocnemius muscle using specific molecular markers for immunofluorescence and real-time polymerase chain reaction (PCR). 2. Inflammation of the injured muscle occurred within 6h, followed by disintegration of the damaged myofibres within 12h. Newly formed myofibres appeared by Day 7. 3. The number of MyoD-positive nuclei (activated satellite cells) in the injured muscle was significantly increased by 6h, reaching a maximum by 12h after contusion. However, the number of MyoD-positive nuclei decreased towards control levels by Day 7. Changes in the number of bromodeoxyuridine-labelled nuclei (proliferating satellite cells) paralleled the changes seen in the number of MyoD-positive nuclei. Conversely, expression of myogenin protein was not apparent until Day 3 and increased further by Day 7. Colabelling of MyoD and myogenin was seen in only a few cells. 4. The time-course of MyoD mRNA expression corresponded with MyoD protein expression. However, there were two peaks in myogenin mRNA expression: 6h and Day 7 after contusion. The second peak coincided with upregulation of myostatin mRNA levels. 5. The results of the present study suggest that contusion activates a homogeneous population of satellite cells to proliferate within 3days, followed by differentiation to form new myofibres. The latter may be regulated, in part, by myostatin.

  6. Local inhibition of nitrergic activity in tenotomized rats accelerates muscle regeneration by increasing fiber area and decreasing central core lesions.

    PubMed

    Seabra, A D; Moraes, S A S; Batista, E J O; Garcia, T B; Souza, M C; Oliveira, K R M; Herculano, A M

    2017-02-20

    Muscular atrophy is a progressive degeneration characterized by muscular proteolysis, loss of mass and decrease in fiber area. Tendon rupture induces muscular atrophy due to an intrinsic functional connection. Local inhibition of nitric oxide synthase (NOS) by Nω-nitro-L-arginine methyl ester (L-NAME) accelerates tendon histological recovery and induces functional improvement. Here we evaluate the effects of such local nitrergic inhibition on the pattern of soleus muscle regeneration after tenotomy. Adult male Wistar rats (240 to 280 g) were divided into four experimental groups: control (n=4), tenotomized (n=6), vehicle (n=6), and L-NAME (n=6). Muscular atrophy was induced by calcaneal tendon rupture in rats. Changes in muscle wet weight and total protein levels were determined by the Bradford method, and muscle fiber area and central core lesion (CCL) occurrence were evaluated by histochemical assays. Compared to tenotomized (69.3±22%) and vehicle groups (68.1%±17%), L-NAME treatment induced an increase in total protein level (108.3±21%) after 21 days post-injury. A reduction in fiber areas was observed in tenotomized (56.3±1.3%) and vehicle groups (53.9±3.9%). However, L-NAME treatment caused an increase in this parameter (69.3±1.6%). Such events were preceded by a remarkable reduction in the number of fibers with CCL in L-NAME-treated animals (12±2%), but not in tenotomized (21±2.5%) and vehicle groups (19.6±2.8%). Altogether, our data reveal that inhibition of tendon NOS contributed to the attenuation of atrophy and acceleration of muscle regeneration.

  7. Local inhibition of nitrergic activity in tenotomized rats accelerates muscle regeneration by increasing fiber area and decreasing central core lesions

    PubMed Central

    Seabra, A.D.; Moraes, S.A.S.; Batista, E.J.O.; Garcia, T.B.; Souza, M.C.; Oliveira, K.R.M.; Herculano, A.M.

    2017-01-01

    Muscular atrophy is a progressive degeneration characterized by muscular proteolysis, loss of mass and decrease in fiber area. Tendon rupture induces muscular atrophy due to an intrinsic functional connection. Local inhibition of nitric oxide synthase (NOS) by Nω-nitro-L-arginine methyl ester (L-NAME) accelerates tendon histological recovery and induces functional improvement. Here we evaluate the effects of such local nitrergic inhibition on the pattern of soleus muscle regeneration after tenotomy. Adult male Wistar rats (240 to 280 g) were divided into four experimental groups: control (n=4), tenotomized (n=6), vehicle (n=6), and L-NAME (n=6). Muscular atrophy was induced by calcaneal tendon rupture in rats. Changes in muscle wet weight and total protein levels were determined by the Bradford method, and muscle fiber area and central core lesion (CCL) occurrence were evaluated by histochemical assays. Compared to tenotomized (69.3±22%) and vehicle groups (68.1%±17%), L-NAME treatment induced an increase in total protein level (108.3±21%) after 21 days post-injury. A reduction in fiber areas was observed in tenotomized (56.3±1.3%) and vehicle groups (53.9±3.9%). However, L-NAME treatment caused an increase in this parameter (69.3±1.6%). Such events were preceded by a remarkable reduction in the number of fibers with CCL in L-NAME-treated animals (12±2%), but not in tenotomized (21±2.5%) and vehicle groups (19.6±2.8%). Altogether, our data reveal that inhibition of tendon NOS contributed to the attenuation of atrophy and acceleration of muscle regeneration. PMID:28225888

  8. Targeting β1-Integrin Signaling Enhances Regeneration in Aged and Dystrophic Muscle in Mice

    PubMed Central

    Rozo, Michelle; Li, Liangji; Fan, Chen-Ming

    2016-01-01

    Interactions between stem cells and their microenvironment, or niche, are essential for stem cell maintenance and function. Our knowledge of the niche for the skeletal muscle stem cell, i.e. the satellite cell (SC), is incomplete. Here we show that β1-integrin is an essential niche molecule that maintains SC homeostasis, and sustains the expansion and self-renewal of this stem cell pool during regeneration. We further show that β1-integrin cooperates with FGF-2, a potent growth factor for SCs, to synergistically activate their common downstream effectors Erk and Akt. Importantly, SCs in aged mice display altered β1-integrin activity and insensitivity to FGF-2. Augmenting β1-integrin activity with a monoclonal antibody restores FGF-2 sensitivity and improves regeneration after experimentally-induced muscle injury. The same treatment also enhances regeneration and function of dystrophic muscles in mdx mice. Therefore, β1-integrin senses the SC niche to maintain responsiveness to FGF-2, and this integrin represents a potential therapeutic target for pathological conditions of the muscle in which the stem cell niche is compromised. PMID:27376575

  9. Satellite Cells CD44 Positive Drive Muscle Regeneration in Osteoarthritis Patients.

    PubMed

    Scimeca, Manuel; Bonanno, Elena; Piccirilli, Eleonora; Baldi, Jacopo; Mauriello, Alessandro; Orlandi, Augusto; Tancredi, Virginia; Gasbarra, Elena; Tarantino, Umberto

    2015-01-01

    Age-related bone diseases, such as osteoarthritis and osteoporosis, are strongly associated with sarcopenia and muscle fiber atrophy. In this study, we analyzed muscle biopsies in order to demonstrate that, in osteoarthritis patients, both osteophytes formation and regenerative properties of muscle stem cells are related to the same factors. In particular, thanks to immunohistochemistry, transmission electron microscopy, and immunogold labeling we investigated the role of BMP-2 in muscle stem cells activity. In patients with osteoarthritis both immunohistochemistry and transmission electron microscopy allowed us to note a higher number of CD44 positive satellite muscle cells forming syncytium. Moreover, the perinuclear and cytoplasmic expression of BMP-2 assessed by in situ molecular characterization of satellite cells syncytia suggest a very strict correlation between BMP-2 expression and muscle regeneration capability. Summing up, the higher BMP-2 expression in osteoarthritic patients could explain the increased bone mineral density as well as decreased muscle atrophy in osteoarthrosic patients. In conclusion, our results suggest that the control of physiological BMP-2 balance between bone and muscle tissues may be considered as a potential pharmacological target in bone-muscle related pathology.

  10. Potential for neural regeneration after neurotoxic injury in the adult mammalian retina

    NASA Astrophysics Data System (ADS)

    Ooto, Sotaro; Akagi, Tadamichi; Kageyama, Ryoichiro; Akita, Joe; Mandai, Michiko; Honda, Yoshihito; Takahashi, Masayo

    2004-09-01

    It has long been believed that the retina of mature mammals is incapable of regeneration. In this study, using the N-methyl-D-aspartate neurotoxicity model of adult rat retina, we observed that some Müller glial cells were stimulated to proliferate in response to a toxic injury and produce bipolar cells and rod photoreceptors. Although these newly produced neurons were limited in number, retinoic acid treatment promoted the number of regenerated bipolar cells. Moreover, misexpression of basic helix-loop-helix and homeobox genes promoted the induction of amacrine, horizontal, and rod photoreceptor specific phenotypes. These findings demonstrated that retinal neurons regenerated even in adult mammalian retina after toxic injury. Furthermore, we could partially control the fate of the regenerated neurons with extrinsic factors or intrinsic genes. The Müller glial cells constitute a potential source for the regeneration of adult mammalian retina and can be a target for drug delivery and gene therapy in retinal degenerative diseases.

  11. MicroRNA-206 is highly expressed in newly formed muscle fibers: implications regarding potential for muscle regeneration and maturation in muscular dystrophy.

    PubMed

    Yuasa, Katsutoshi; Hagiwara, Yasuko; Ando, Masanori; Nakamura, Akinori; Takeda, Shin'ichi; Hijikata, Takao

    2008-01-01

    miR-1, miR-133a, and miR-206 are muscle-specific microRNAs expressed in skeletal muscles and have been shown to contribute to muscle development. To gain insight into the pathophysiological roles of these three microRNAs in dystrophin-deficient muscular dystrophy, their expression in the tibialis anterior (TA) muscles of mdx mice and CXMD(J) dogs were evaluated by semiquantitative RT-PCR and in situ hybridization. Their temporal and spatial expression patterns were also analyzed in C2C12 cells during muscle differentiation and in cardiotoxin (CTX)-injured TA muscles to examine how muscle degeneration and regeneration affect their expression. In dystrophic TA muscles of mdx mice, miR-206 expression was significantly elevated as compared to that in control TA muscles of age-matched B10 mice, whereas there were no differences in miR-1 or miR-133a expression between B10 and mdx TA muscles. On in situ hybridization analysis, intense signals for miR-206 probes were localized in newly formed myotubes with centralized nuclei, or regenerating muscle fibers, but not in intact pre-degenerated fibers or numerous small mononucleated cells, possibly proliferating myoblasts and inflammatory infiltrates. Similar increased expression of miR-206 was also found in C2C12 differentiation and CTX-induced regeneration, in which differentiated myotubes or regenerating fibers showed abundant expression of miR-206. However, CXMD(J) TA muscles contained smaller amounts of miR-206, miR-1, and miR-133a than controls. They exhibited more severe and more progressive degenerative alterations than mdx TA muscles. Taken together, these observations indicated that newly formed myotubes showed markedly increased expression of miR-206, which might reflect active regeneration and efficient maturation of skeletal muscle fibers.

  12. Stimulation of post-traumatic regeneration of skeletal muscles of old rats after x-ray irradiation

    SciTech Connect

    Bulyakova, N.V.; Popova, M.F.

    1987-09-01

    The authors seek a method of stimulating restorative processes in irradiated muscles of old animals. Rats were used in the experiments. Different series of experiments were performed, including complete transverse section of the gastrocnemius muscle after local x-ray irradiation, and laser therapy of the transversly divided gastrocnemius muscle. Post-traumatic regeneration of the gastrocnemius muscle of old rats is illustrated schematically. The experimental data showed that pulsed laser therapy or grafting of minced unirradiated muscle tissue can largely restore the regenerative capacity of the gastrocnemius muscle of old rats when depressed by x-ray irradiation, but the method of grafting minced unirradiated muscle tissue was more effective.

  13. Cardiac muscle plasticity in adult and embryo by heart-derived progenitor cells.

    PubMed

    Oh, Hidemasa; Chi, Xuan; Bradfute, Steven B; Mishina, Yuji; Pocius, Jennifer; Michael, Lloyd H; Behringer, Richard R; Schwartz, Robert J; Entman, Mark L; Schneider, Michael D

    2004-05-01

    The evidence of cardiomyocyte proliferation in damaged heart implied cardiac regeneration might occur by resident or extra cardiac stem cells. However, the specification and origin of these cells remain unknown. Here, we report using fluorescence-activated cell sorting that cardiac progenitor cells resided in adult heart and colocalized with small capillary vessels, within the stem cell antigen (Sca-1) population expressing high telomerase activity. Notably, hematopoietic stem cells capable of efflux Hoechst 33342, termed side population cells, also were identified within the heart-derived cells. The cardiac progenitor cells (CD45(-)/CD34(-)) express neither cardiac muscle nor endothelial cell markers at an undifferentiated stage. The exposure of 5-azacytidine induced cardiac differentiation, which depends, in part, on Bmpr1a, a type IA receptor for bone morphogenetic protein (BMP). The capability of adult Sca1(+) cells to adopt a cardiac muscle in embryogenesis was substantiated by blastocyst injection, using progenitors from the adult hearts of transgenic mice that harbor a bacterial artificial chromosome expressing GFP via the Nkx-2.5 locus. Intravenously injected progenitors, shortly after ischemic/reperfusion, homed and functionally differentiated 3.5% of total left ventricle in the host myocardium. Differentiation included both fusion-independent and fusion-associated components, proved by the Cre/loxP donor/recipient system. Our studies suggest that endogenous cardiac progenitors reside in the adult heart, regenerate cardiomyocytes functionally, and integrate into the existing heart circuitry.

  14. In vitro construction and in vivo regeneration of esophageal bilamellar muscle tissue.

    PubMed

    Hou, Lei; Gong, Changfeng; Zhu, Yabin

    2016-04-01

    In order to induce esophageal muscle cells' orientation, the silicon wafer with prototype 1 and prototype 2 was designed. Prototype 1 has micro-channels of 200 µm width and 30 µm depth with 30 µm wide wall as the interval. Prototype 2 has channels of 100 µm width and 30 µm depth with a discontinuous wall which has 30 µm gap for each 100 µm channel. The poly(ester urethane) scaffolds with pattern prototype 1 and prototype 2 were fabricated using solution casting method and abbreviated as PU1 and PU2, respectively. Silk fibroin was grafted individually on PU1 and PU2 surface (PU1-SF, PU2-SF) using our previous protocol, aiming at improving scaffolds' biocompatibility. The primary esophageal smooth muscle cell was seeded to evaluate the scaffolds' cytocompatibility in vitro. Characterizations like MTT assay, immunocytochemistry, scanning electron microscope, and Western blotting were applied. After that, poly(ester urethane) scaffolds with double patterns, prototype 1 on the exterior, and prototype 2 in the lumen were implanted into the rabbit esophagous to test the regeneration of the muscle tissue. Results from these preliminary tests showed that the growth and differentiation of primary smooth muscle cells were promoted, but also the muscle tissue with endocircular and exolongitudinal architecture was in regenerating, against non-constitution in the animals without the patterned scaffold or with poly(ester urethane) plane membrane at the defaulted sites. This micro-channel pattern together with silk fibroin grafting and vascular endothelial growth factor coating greatly promoted the regeneration of esophageal muscle with normal histological structure.

  15. Comparative Study of Injury Models for Studying Muscle Regeneration in Mice

    PubMed Central

    Hardy, David; Besnard, Aurore; Latil, Mathilde; Jouvion, Grégory; Briand, David; Thépenier, Cédric; Pascal, Quentin; Guguin, Aurélie; Gayraud-Morel, Barbara; Cavaillon, Jean-Marc; Tajbakhsh, Shahragim

    2016-01-01

    Background A longstanding goal in regenerative medicine is to reconstitute functional tissus or organs after injury or disease. Attention has focused on the identification and relative contribution of tissue specific stem cells to the regeneration process. Relatively little is known about how the physiological process is regulated by other tissue constituents. Numerous injury models are used to investigate tissue regeneration, however, these models are often poorly understood. Specifically, for skeletal muscle regeneration several models are reported in the literature, yet the relative impact on muscle physiology and the distinct cells types have not been extensively characterised. Methods We have used transgenic Tg:Pax7nGFP and Flk1GFP/+ mouse models to respectively count the number of muscle stem (satellite) cells (SC) and number/shape of vessels by confocal microscopy. We performed histological and immunostainings to assess the differences in the key regeneration steps. Infiltration of immune cells, chemokines and cytokines production was assessed in vivo by Luminex®. Results We compared the 4 most commonly used injury models i.e. freeze injury (FI), barium chloride (BaCl2), notexin (NTX) and cardiotoxin (CTX). The FI was the most damaging. In this model, up to 96% of the SCs are destroyed with their surrounding environment (basal lamina and vasculature) leaving a “dead zone” devoid of viable cells. The regeneration process itself is fulfilled in all 4 models with virtually no fibrosis 28 days post-injury, except in the FI model. Inflammatory cells return to basal levels in the CTX, BaCl2 but still significantly high 1-month post-injury in the FI and NTX models. Interestingly the number of SC returned to normal only in the FI, 1-month post-injury, with SCs that are still cycling up to 3-months after the induction of the injury in the other models. Conclusions Our studies show that the nature of the injury model should be chosen carefully depending on the

  16. Connective tissue regeneration in skeletal muscle after eccentric contraction-induced injury.

    PubMed

    Mackey, Abigail L; Kjaer, Michael

    2017-03-01

    Human skeletal muscle has the potential to regenerate completely after injury induced under controlled experimental conditions. The events inside the myofibers as they undergo necrosis, followed closely by satellite cell-mediated myogenesis, have been mapped in detail. Much less is known about the adaptation throughout this process of both the connective tissue structures surrounding the myofibers and the fibroblasts, the cells responsible for synthesizing this connective tissue. However, the few studies investigating muscle connective tissue remodeling demonstrate a strong response that appears to be sustained for a long time after the major myofiber responses have subsided. While the use of electrical stimulation to induce eccentric contractions vs. voluntary eccentric contractions appears to lead to a greater extent of myofiber necrosis and regenerative response, this difference is not apparent when the muscle connective tissue responses are compared, although further work is required to confirm this. Pharmacological agents (growth hormone and angiotensin II type I receptor blockers) are considered in the context of accelerating the muscle connective tissue adaptation to loading. Cautioning against this, however, is the association between muscle matrix protein remodeling and protection against reinjury, which suggests that a (so far undefined) period of vulnerability to reinjury may exist during the remodeling phases. The role of individual muscle matrix components and their spatial interaction during adaptation to eccentric contractions is an unexplored field in human skeletal muscle and may provide insight into the optimal timing of rest vs. return to activity after muscle injury.

  17. miR-431 promotes differentiation and regeneration of old skeletal muscle by targeting Smad4

    PubMed Central

    Lee, Kwang-Pyo; Shin, Yeo Jin; Panda, Amaresh C.; Abdelmohsen, Kotb; Kim, Ji Young; Lee, Seung-Min; Bahn, Young Jae; Choi, Jeong Yi; Kwon, Eun-Soo; Baek, Su-Jin; Kim, Seon-Young; Gorospe, Myriam; Kwon, Ki-Sun

    2015-01-01

    The myogenic capacity of myoblasts decreases in skeletal muscle with age. In addition to environmental factors, intrinsic factors are important for maintaining the regenerative potential of muscle progenitor cells, but their identities are largely unknown. Here, comparative analysis of microRNA (miRNA) expression profiles in young and old myoblasts uncovered miR-431 as a novel miRNA showing markedly reduced abundance in aged myoblasts. Importantly, elevating miR-431 improved the myogenic capacity of old myoblasts, while inhibiting endogenous miR-431 lowered myogenesis. Bioinformatic and biochemical analyses revealed that miR-431 directly interacted with the 3′ untranslated region (UTR) of Smad4 mRNA, which encodes one of the downstream effectors of TGF-β signaling. In keeping with the low levels of miR-431 in old myoblasts, SMAD4 levels increased in this myoblast population. Interestingly, in an in vivo model of muscle regeneration following cardiotoxin injury, ectopic miR-431 injection greatly improved muscle regeneration and reduced SMAD4 levels. Consistent with the finding that the mouse miR-431 seed sequence in the Smad4 3′ UTR is conserved in the human SMAD4 3′ UTR, inhibition of miR-431 also repressed the myogenic capacity of human skeletal myoblasts. Taken together, our results suggest that the age-associated miR-431 plays a key role in maintaining the myogenic ability of skeletal muscle with age. PMID:26215566

  18. Effects of age on aneural regeneration of soleus muscle in rat.

    PubMed Central

    Lewis, D M; Schmalbruch, H

    1995-01-01

    1. The ability of autografted soleus muscles to regenerate without innervation was investigated in young (two groups: 17 days or 35 g and 5 weeks or 100 g) and old (10 weeks or 300 g and 19 months or 700 g) rats. 2. Tetanic force and fibre area of the regenerated muscles were followed in 35, 100 and 300 g rats and found to reach a maximum 10-15 days after the operation and then declined. 3. Maximal tetanic force and fibre area were greater in old than in young rats; the largest increase was seen between 100 and 300 g rats. The relaxation phase of the twitch became shorter in the 700 g animals. The force per cross-sectional area appeared to fall with age. The length of the new fibres, inferred from the width of the length-force curve, increased only slightly with age. 4. Ten days after grafting, autophagocytosis of necrotic fibres was completed in young but not in old rats. The new fibres in young rats had one central nucleus per cross-section and fibre size was unimodally distributed; fibres in old rats had multiple internal nuclei and the size distribution was bimodal due to the presence of large fibres. 5. Previous results indicating greater muscle regeneration in young than in old rats may reflect more vigorous reinnervation in young animals rather than a greater myogenic potential. Increased fibre size of regenerated muscles of old compared with young rats may be attributed to the larger amount of necrotic material which is mitogenic for satellite cells, or to age-dependent changes of the expression of cell adhesion molecules. Enhanced lateral fusion of myotubes would give rise to large fibres with multiple internal nuclei. Images Figure 3 Figure 4 PMID:8568686

  19. In vivo Fluorescence Imaging of Muscle Cell Regeneration by Transplanted EGFP-labeled Myoblasts

    PubMed Central

    Xu, Xiaoyin; Yang, Zhong; Liu, Qiang; Wang, Yaming

    2010-01-01

    In vivo fluorescence imaging (FLI) enables monitoring fluorescent protein (FP)-labeled cells and proteins in living organisms noninvasively. Here, we examined whether this modality could reach a sufficient sensitivity to allow evaluation of the regeneration process of enhanced green fluorescent protein (eGFP)-labeled muscle precursors (myoblasts). Using a basic FLI station, we were able to detect clear fluorescence signals generated by 40,000 labeled cells injected into a tibialis anterior (TA) muscle of mouse. We observed that the signal declined to ~25% on the 48 hours of cell injection followed by a recovery starting at the second day and reached a peak of ~45% of the original signal by the 7th day, suggesting that the survived population underwent a limited run of proliferation before differentiation. To assess whether transplanted myoblasts could form satellite cells, we injured the transplanted muscles repeatedly with cardiotoxin. We observed a recovery of fluorescence signal following a disappearance of the signal after each cardiotoxin injection. Histology results showed donor-derived cells located underneath basal membrane and expressing Pax7, confirming that the regeneration observed by imaging was indeed mediated by donor-derived satellite cells. Our results show that FLI is a powerful tool that can extend our ability to unveil complicated biological processes such as stem cell-mediated regeneration. PMID:20125125

  20. High-mobility group box 1 release and redox regulation accompany regeneration and remodeling of skeletal muscle.

    PubMed

    Vezzoli, Michela; Castellani, Patrizia; Corna, Gianfranca; Castiglioni, Alessandra; Bosurgi, Lidia; Monno, Antonella; Brunelli, Silvia; Manfredi, Angelo A; Rubartelli, Anna; Rovere-Querini, Patrizia

    2011-10-15

    High-mobility group box 1 (HMGB1), a damage-associated molecular pattern (DAMP) molecules, favors tissue regeneration via recruitment and activation of leukocytes and stem cells. Here we demonstrate, in a model of acute sterile muscle injury, that regeneration is accompanied by active reactive oxygen species (ROS) production counterbalanced and overcome by the generation of antioxidant moieties. Mitochondria are initially responsible for ROS formation. However, they undergo rapid disruption with almost complete disappearance. Twenty-four hours after injury, we observed a strong induction of MURF1 and atrogin-1 ubiquitin ligases, key signals in activation of the proteasome system and induction of muscle atrophy. At later time points, ROS generation is maintained by nonmitochondrial sources. The antioxidant response occurs in both regenerating fibers and leukocytes that express high levels of free thiols and antioxidant enzymes, such as superoxide dismutase 1 (SOD1) and thioredoxin. HMGB1, a protein thiol, weakly expressed in healthy muscles, increases during regeneration in parallel with the antioxidant response in both fibers and leukocytes. A reduced environment may be important to maintain HMGB1 bioactivity. Indeed, oxidation abrogates both muscle stem cell migration in response to HMGB1 and their ability to differentiate into myofibers in vitro. We propose that the early antioxidant response in regenerating muscle limits HMGB1 oxidation, thus allowing successful muscle regeneration.

  1. Growth and apoptosis during larval forelimb development and adult forelimb regeneration in the newt ( Notophthalmus viridescens).

    PubMed

    Vlaskalin, Tatjana; Wong, Christine J; Tsilfidis, Catherine

    2004-09-01

    Many of the genes involved in the initial development of the limb in higher vertebrates are also expressed during regeneration of the limb in urodeles such as Notophthalmus viridescens. These similarities have led researchers to conclude that the regeneration process is a recapitulation of development, and that patterning of the regenerate mimics pattern formation in development. However, the developing limb and the regenerating limb do not look similar. In developing urodele forelimbs, digits appear sequentially as outgrowths from the limb palette. In regeneration, all the digits appear at once. In this work, we address the issue of whether regeneration and development are similar by examining growth and apoptosis patterns. In contrast to higher vertebrates, forelimb development in the newt, N. viridescens, does not use interdigital apoptosis as the method of digit separation. During adult forelimb regeneration, apoptosis seems to play an important role in wound healing and again during cartilage to bone turnover in the advanced digits and radius/ulna. However, similar to forelimb development, demarcation of the digits in adult forelimb regeneration does not involve interdigital apoptosis. Outgrowth, rather than regression of the interdigital mesenchyme, leads to the individualization of forelimb digits in both newt development and regeneration.

  2. Re-regeneration of lower jaws and the dental lamina in adult urodeles.

    PubMed

    Graver, H T

    1978-09-01

    Transverse amputations were carried out through one-third fully regenerated jaw segments and through normal tissue of the mandible on the same and opposite sides of the jaw in adults of Notophthalmus viridescens. Collectively the results suggest that, in adult urodeles, the mandible and the dental lamina can be replaced in an identical manner more than one time. Although the major histological events are the same in jaw regeneration and re-regeneration, regrowth is more rapid in re-regeneration. It appears that recently differentiated tissues of the regenerate have a higher capacity for regeneration than normal tissues amputated for the first time. Re-regeneration of the jaw occurs by growth of the original regenerate cartilage which has undergone reorganization. In re-regeneration, the skeletal elements exhibit no polarity and regrowth occurs in both directions, while the dental lamina possesses an anterior-posterior polarity and can regrow in an anterior direction only. Information concerning the mechanisms involved in the regenerative events remain to be determined.

  3. Rescue of neuronal function by cross-regeneration of cutaneous afferents into muscle in cats.

    PubMed

    Nishimura, H; Johnson, R D; Munson, J B

    1993-07-01

    1. This study investigates the relation between the peripheral innervation of low-threshold cutaneous afferents and the postsynaptic potentials elicited by electrical stimulation of those afferents. 2. In cats deeply anesthetized with pentobarbital sodium, cord dorsum potentials (CDPs) and postsynaptic potentials (PSPs) in spinal motoneurons were elicited by stimulation of the caudal cutaneous sural nerve (CCS), the lateral cutaneous sural nerve (LCS), and the medial gastrocnemius (MG) muscle nerve. We tested 1) unoperated cats, and cats in which CCS has been 2) chronically axotomized and ligated, 3) cut and self-reunited, 4) cut and cross-united with LCS, or 5) cut and cross-united with the MG. Terminal experiments were performed 3-36 mo after initial surgery. 3. In cats in which the CCS had been self-reunited or cross-united distally with LCS, tactile stimulation of the hairy skin normally innervated by the distal nerve activated afferents in the CCS central to the coaptation, indicating that former CCS afferents had regenerated into native or foreign skin, respectively. 4. In cats in which the CCS had been cross-united distally with the MG, both stretch and contraction of the MG muscle activated the former CCS afferents. 5. In unoperated cats, CDPs elicited by stimulation of CCS and of LCS exhibited a low-threshold N1 wave and a higher-threshold N2 wave. These waves were greatly delayed and appeared to merge after chronic axotomy of CCS. Regeneration of CCS into itself, into LCS, or into MG restored the normal latencies and configurations of these potentials. 6. In unoperated cats, stimulation of CCS, of LCS, and of MG each produced PSPs of characteristic configurations in the various subpopulations of motoneurons of the triceps surae. CDPs and PSPs elicited by the CCS cross-regenerated into LCS or MG were typical of those generated by the normal CCS, i.e., there was no evidence of respecification of central synaptic connections to bring accord between center

  4. Efficient regeneration by activation of neurogenesis in homeostatically quiescent regions of the adult vertebrate brain.

    PubMed

    Berg, Daniel A; Kirkham, Matthew; Beljajeva, Anna; Knapp, Dunja; Habermann, Bianca; Ryge, Jesper; Tanaka, Elly M; Simon, András

    2010-12-01

    In contrast to mammals, salamanders and teleost fishes can efficiently repair the adult brain. It has been hypothesised that constitutively active neurogenic niches are a prerequisite for extensive neuronal regeneration capacity. Here, we show that the highly regenerative salamander, the red spotted newt, displays an unexpectedly similar distribution of active germinal niches with mammals under normal physiological conditions. Proliferation zones in the adult newt brain are restricted to the forebrain, whereas all other regions are essentially quiescent. However, ablation of midbrain dopamine neurons in newts induced ependymoglia cells in the normally quiescent midbrain to proliferate and to undertake full dopamine neuron regeneration. Using oligonucleotide microarrays, we have catalogued a set of differentially expressed genes in these activated ependymoglia cells. This strategy identified hedgehog signalling as a key component of adult dopamine neuron regeneration. These data show that brain regeneration can occur by activation of neurogenesis in quiescent brain regions.

  5. Muscle organizers in Drosophila: the role of persistent larval fibers in adult flight muscle development

    NASA Technical Reports Server (NTRS)

    Farrell, E. R.; Fernandes, J.; Keshishian, H.

    1996-01-01

    In many organisms muscle formation depends on specialized cells that prefigure the pattern of the musculature and serve as templates for myoblast organization and fusion. These include muscle pioneers in insects and muscle organizing cells in leech. In Drosophila, muscle founder cells have been proposed to play a similar role in organizing larval muscle development during embryogenesis. During metamorphosis in Drosophila, following histolysis of most of the larval musculature, there is a second round of myogenesis that gives rise to the adult muscles. It is not known whether muscle founder cells organize the development of these muscles. However, in the thorax specific larval muscle fibers do not histolyze at the onset of metamorphosis, but instead serve as templates for the formation of a subset of adult muscles, the dorsal longitudinal flight muscles (DLMs). Because these persistent larval muscle fibers appear to be functioning in many respects like muscle founder cells, we investigated whether they were necessary for DLM development by using a microbeam laser to ablate them singly and in combination. We found that, in the absence of the larval muscle fibers, DLMs nonetheless develop. Our results show that the persistent larval muscle fibers are not required to initiate myoblast fusion, to determine DLM identity, to locate the DLMs in the thorax, or to specify the total DLM fiber volume. However, they are required to regulate the number of DLM fibers generated. Thus, while the persistent larval muscle fibers are not obligatory for DLM fiber formation and differentiation, they are necessary to ensure the development of the correct number of fibers.

  6. Muscle regeneration during hindlimb unloading results in a reduction in muscle size after reloading

    NASA Technical Reports Server (NTRS)

    Mozdziak, P. E.; Pulvermacher, P. M.; Schultz, E.

    2001-01-01

    The hindlimb-unloading model was used to study the ability of muscle injured in a weightless environment to recover after reloading. Satellite cell mitotic activity and DNA unit size were determined in injured and intact soleus muscles from hindlimb-unloaded and age-matched weight-bearing rats at the conclusion of 28 days of hindlimb unloading, 2 wk after reloading, and 9 wk after reloading. The body weights of hindlimb-unloaded rats were significantly (P < 0.05) less than those of weight-bearing rats at the conclusion of hindlimb unloading, but they were the same (P > 0.05) as those of weight-bearing rats 2 and 9 wk after reloading. The soleus muscle weight, soleus muscle weight-to-body weight ratio, myofiber diameter, number of nuclei per millimeter, and DNA unit size were significantly (P < 0.05) smaller for the injured soleus muscles from hindlimb-unloaded rats than for the soleus muscles from weight-bearing rats at each recovery time. Satellite cell mitotic activity was significantly (P < 0.05) higher in the injured soleus muscles from hindlimb-unloaded rats than from weight-bearing rats 2 wk after reloading, but it was the same (P > 0.05) as in the injured soleus muscles from weight-bearing rats 9 wk after reloading. The injured soleus muscles from hindlimb-unloaded rats failed to achieve weight-bearing muscle size 9 wk after reloading, because incomplete compensation for the decrease in myonuclear accretion and DNA unit size expansion occurred during the unloading period.

  7. A systems-based investigation into vitamin D and skeletal muscle repair, regeneration, and hypertrophy.

    PubMed

    Owens, Daniel J; Sharples, Adam P; Polydorou, Ioanna; Alwan, Nura; Donovan, Timothy; Tang, Jonathan; Fraser, William D; Cooper, Robert G; Morton, James P; Stewart, Claire; Close, Graeme L

    2015-12-15

    Skeletal muscle is a direct target for vitamin D. Observational studies suggest that low 25[OH]D correlates with functional recovery of skeletal muscle following eccentric contractions in humans and crush injury in rats. However, a definitive association is yet to be established. To address this gap in knowledge in relation to damage repair, a randomised, placebo-controlled trial was performed in 20 males with insufficient concentrations of serum 25(OH)D (45 ± 25 nmol/l). Prior to and following 6 wk of supplemental vitamin D3 (4,000 IU/day) or placebo (50 mg of cellulose), participants performed 20 × 10 damaging eccentric contractions of the knee extensors, with peak torque measured over the following 7 days of recovery. Parallel experimentation using isolated human skeletal muscle-derived myoblast cells from biopsies of 14 males with low serum 25(OH)D (37 ± 11 nmol/l) were subjected to mechanical wound injury, which enabled corresponding in vitro studies of muscle repair, regeneration, and hypertrophy in the presence and absence of 10 or 100 nmol 1α,25(OH)2D3. Supplemental vitamin D3 increased serum 25(OH)D and improved recovery of peak torque at 48 h and 7 days postexercise. In vitro, 10 nmol 1α,25(OH)2D3 improved muscle cell migration dynamics and resulted in improved myotube fusion/differentiation at the biochemical, morphological, and molecular level together with increased myotube hypertrophy at 7 and 10 days postdamage. Together, these preliminary data are the first to characterize a role for vitamin D in human skeletal muscle regeneration and suggest that maintaining serum 25(OH)D may be beneficial for enhancing reparative processes and potentially for facilitating subsequent hypertrophy.

  8. Malat1 regulates myogenic differentiation and muscle regeneration through modulating MyoD transcriptional activity

    PubMed Central

    Chen, Xiaona; He, Liangqiang; Zhao, Yu; Li, Yuying; Zhang, Suyang; Sun, Kun; So, Karl; Chen, Fengyuan; Zhou, Liang; Lu, Leina; Wang, Lijun; Zhu, Xihua; Bao, Xichen; Esteban, Miguel A; Nakagawa, Shinichi; Prasanth, Kannanganattu V; Wu, Zhenguo; Sun, Hao; Wang, Huating

    2017-01-01

    Malat1 is one of the most abundant long non-coding RNAs in various cell types; its exact cellular function is still a matter of intense investigation. In this study we characterized the function of Malat1 in skeletal muscle cells and muscle regeneration. Utilizing both in vitro and in vivo assays, we demonstrate that Malat1 has a role in regulating gene expression during myogenic differentiation of myoblast cells. Specifically, we found that knockdown of Malat1 accelerates the myogenic differentiation in cultured cells. Consistently, Malat1 knockout mice display enhanced muscle regeneration after injury and deletion of Malat1 in dystrophic mdx mice also improves the muscle regeneration. Mechanistically, in the proliferating myoblasts, Malat1 recruits Suv39h1 to MyoD-binding loci, causing trimethylation of histone 3 lysine 9 (H3K9me3), which suppresses the target gene expression. Upon differentiation, the pro-myogenic miR-181a is increased and targets the nuclear Malat1 transcripts for degradation through Ago2-dependent nuclear RNA-induced silencing complex machinery; the Malat1 decrease subsequently leads to the destabilization of Suv39h1/HP1β/HDAC1-repressive complex and displacement by a Set7-containing activating complex, which allows MyoD trans-activation to occur. Together, our findings identify a regulatory axis of miR-181a-Malat1-MyoD/Suv39h1 in myogenesis and uncover a previously unknown molecular mechanism of Malat1 action in gene regulation. PMID:28326190

  9. Extraocular muscle degeneration and regeneration after exposure of rats to incandescent radiant energy.

    PubMed

    O'Steen, W K; Shear, C R; Anderson, K V

    1975-06-01

    Exposure of albino rats to incandescent radiant energy for a short period of time in an elevated environmental temperature (39 degrees C) causes degenerative changes in the extraocular muscles. The muscle fibres regenerate and the muscles reorganize if the animals are returned to room lighting and temperature. Extraocular muscles (EOMs) were damaged first near their insertion on the eyeball. All EOMs of both eyes were affected, but the degeneration did not extend the entire length of the muscle. The peripheral fibres of each muscle were damaged before the more central fibres. Mitochondria were swollen and often contained dense bodies. Numerous vesicular profiles, possibly from the sarcotubular system, were present. Myofibrils of the more severely damaged fibres lacked typical Z-disk structures, and I-bands had disappeared by 24 h after the exposure period, a degenerative pattern which seems to be unique for this method of EOM damage. EOM degeneration appeared to be dependent on the interaction between thermal and radiant energy on the orbital contents. However, EOMs were only rarely and very slightly affected when rats were exposed to elevated temperature in the absence of incandescent radiant energy. When an opaque, black, ocular occluder was placed over one eye and the contralateral eye was left unoccluded, EOMs and retinas of occluded eyes were undamaged, while those tissues were severely damaged in unoccluded eyes. Therefore, the most critical single variable in inducing EOM degeneration appears to be exposure to radiant energy.

  10. TNF/p38α/polycomb signaling to Pax7 locus in satellite cells links inflammation to the epigenetic control of muscle regeneration.

    PubMed

    Palacios, Daniela; Mozzetta, Chiara; Consalvi, Silvia; Caretti, Giuseppina; Saccone, Valentina; Proserpio, Valentina; Marquez, Victor E; Valente, Sergio; Mai, Antonello; Forcales, Sonia V; Sartorelli, Vittorio; Puri, Pier Lorenzo

    2010-10-08

    How regeneration cues are converted into the epigenetic information that controls gene expression in adult stem cells is currently unknown. We identified an inflammation-activated signaling in muscle stem (satellite) cells, by which the polycomb repressive complex 2 (PRC2) represses Pax7 expression during muscle regeneration. TNF-activated p38α kinase promotes the interaction between YY1 and PRC2, via threonine 372 phosphorylation of EZH2, the enzymatic subunit of the complex, leading to the formation of repressive chromatin on Pax7 promoter. TNF-α antibodies stimulate satellite cell proliferation in regenerating muscles of dystrophic or normal mice. Genetic knockdown or pharmacological inhibition of the enzymatic components of the p38/PRC2 signaling--p38α and EZH2--invariably promote Pax7 expression and expansion of satellite cells that retain their differentiation potential upon signaling resumption. Genetic knockdown of Pax7 impaired satellite cell proliferation in response to p38 inhibition, thereby establishing the biological link between p38/PRC2 signaling to Pax7 and satellite cell decision to proliferate or differentiate.

  11. A comparison of bone regeneration with human mesenchymal stem cells and muscle-derived stem cells and the critical role of BMP.

    PubMed

    Gao, Xueqin; Usas, Arvydas; Tang, Ying; Lu, Aiping; Tan, Jian; Schneppendahl, Johannes; Kozemchak, Adam M; Wang, Bing; Cummins, James H; Tuan, Rocky S; Huard, Johnny

    2014-08-01

    Adult multipotent stem cells have been isolated from a variety of human tissues including human skeletal muscle, which represent an easily accessible source of stem cells. It has been shown that human skeletal muscle-derived stem cells (hMDSCs) are muscle-derived mesenchymal stem cells capable of multipotent differentiation. Although hMDSCs can undergo osteogenic differentiation and form bone when genetically modified to express BMP2; it is still unclear whether hMDSCs are as efficient as human bone marrow mesenchymal stem cells (hBMMSCs) for bone regeneration. The current study aimed to address this question by performing a parallel comparison between hMDSCs and hBMMSCs to evaluate their osteogenic and bone regeneration capacities. Our results demonstrated that hMDSCs and hBMMSCs had similar osteogenic-related gene expression profiles and had similar osteogenic differentiation capacities in vitro when transduced to express BMP2. Both the untransduced hMDSCs and hBMMSCs formed very negligible amounts of bone in the critical sized bone defect model when using a fibrin sealant scaffold; however, when genetically modified with lenti-BMP2, both populations successfully regenerated bone in the defect area. No significant differences were found in the newly formed bone volumes and bone defect coverage between the hMDSC and hBMMSC groups. Although both cell types formed mature bone tissue by 6 weeks post-implantation, the newly formed bone in the hMDSCs group underwent quicker remodelling than the hBMMSCs group. In conclusion, our results demonstrated that hMDSCs are as efficient as hBMMSCs in terms of their bone regeneration capacity; however, both cell types required genetic modification with BMP in order to regenerate bone in vivo.

  12. Phospholipase D1 facilitates second-phase myoblast fusion and skeletal muscle regeneration.

    PubMed

    Teng, Shuzhi; Stegner, David; Chen, Qin; Hongu, Tsunaki; Hasegawa, Hiroshi; Chen, Li; Kanaho, Yasunori; Nieswandt, Bernhard; Frohman, Michael A; Huang, Ping

    2015-02-01

    Myoblast differentiation and fusion is a well-orchestrated multistep process that is essential for skeletal muscle development and regeneration. Phospholipase D1 (PLD1) has been implicated in the initiation of myoblast differentiation in vitro. However, whether PLD1 plays additional roles in myoblast fusion and exerts a function in myogenesis in vivo remains unknown. Here we show that PLD1 expression is up-regulated in myogenic cells during muscle regeneration after cardiotoxin injury and that genetic ablation of PLD1 results in delayed myofiber regeneration. Myoblasts derived from PLD1-null mice or treated with PLD1-specific inhibitor are unable to form mature myotubes, indicating defects in second-phase myoblast fusion. Concomitantly, the PLD1 product phosphatidic acid is transiently detected on the plasma membrane of differentiating myocytes, and its production is inhibited by PLD1 knockdown. Exogenous lysophosphatidylcholine, a key membrane lipid for fusion pore formation, partially rescues fusion defect resulting from PLD1 inhibition. Thus these studies demonstrate a role for PLD1 in myoblast fusion during myogenesis in which PLD1 facilitates the fusion of mononuclear myocytes with nascent myotubes.

  13. mTOR regulates skeletal muscle regeneration in vivo through kinase-dependent and kinase-independent mechanisms.

    PubMed

    Ge, Yejing; Wu, Ai-Luen; Warnes, Christine; Liu, Jianming; Zhang, Chongben; Kawasome, Hideki; Terada, Naohiro; Boppart, Marni D; Schoenherr, Christopher J; Chen, Jie

    2009-12-01

    Rapamycin-sensitive signaling is required for skeletal muscle differentiation and remodeling. In cultured myoblasts, the mammalian target of rapamycin (mTOR) has been reported to regulate differentiation at different stages through distinct mechanisms, including one that is independent of mTOR kinase activity. However, the kinase-independent function of mTOR remains controversial, and no in vivo studies have examined those mTOR myogenic mechanisms previously identified in vitro. In this study, we find that rapamycin impairs injury-induced muscle regeneration. To validate the role of mTOR with genetic evidence and to probe the mechanism of mTOR function, we have generated and characterized transgenic mice expressing two mutants of mTOR under the control of human skeletal actin (HSA) promoter: rapamycin-resistant (RR) and RR/kinase-inactive (RR/KI). Our results show that muscle regeneration in rapamycin-administered mice is restored by RR-mTOR expression. In the RR/KI-mTOR mice, nascent myofiber formation during the early phase of regeneration proceeds in the presence of rapamycin, but growth of the regenerating myofibers is blocked by rapamycin. Igf2 mRNA levels increase drastically during early regeneration, which is sensitive to rapamycin in wild-type muscles but partially resistant to rapamycin in both RR- and RR/KI-mTOR muscles, consistent with mTOR regulation of Igf2 expression in a kinase-independent manner. Furthermore, systemic ablation of S6K1, a target of mTOR kinase, results in impaired muscle growth but normal nascent myofiber formation during regeneration. Therefore, mTOR regulates muscle regeneration through kinase-independent and kinase-dependent mechanisms at the stages of nascent myofiber formation and myofiber growth, respectively.

  14. V-ATPase Proton Pumping Activity Is Required for Adult Zebrafish Appendage Regeneration

    PubMed Central

    Monteiro, Joana; Aires, Rita; Becker, Jörg D.; Jacinto, António; Certal, Ana C.; Rodríguez-León, Joaquín

    2014-01-01

    The activity of ion channels and transporters generates ion-specific fluxes that encode electrical and/or chemical signals with biological significance. Even though it is long known that some of those signals are crucial for regeneration, only in recent years the corresponding molecular sources started to be identified using mainly invertebrate or larval vertebrate models. We used adult zebrafish caudal fin as a model to investigate which and how ion transporters affect regeneration in an adult vertebrate model. Through the combined use of biophysical and molecular approaches, we show that V-ATPase activity contributes to a regeneration-specific H+ ef`flux. The onset and intensity of both V-ATPase expression and H+ efflux correlate with the different regeneration rate along the proximal-distal axis. Moreover, we show that V-ATPase inhibition impairs regeneration in adult vertebrate. Notably, the activity of this H+ pump is necessary for aldh1a2 and mkp3 expression, blastema cell proliferation and fin innervation. To the best of our knowledge, this is the first report on the role of V-ATPase during adult vertebrate regeneration. PMID:24671205

  15. V-ATPase proton pumping activity is required for adult zebrafish appendage regeneration.

    PubMed

    Monteiro, Joana; Aires, Rita; Becker, Jörg D; Jacinto, António; Certal, Ana C; Rodríguez-León, Joaquín

    2014-01-01

    The activity of ion channels and transporters generates ion-specific fluxes that encode electrical and/or chemical signals with biological significance. Even though it is long known that some of those signals are crucial for regeneration, only in recent years the corresponding molecular sources started to be identified using mainly invertebrate or larval vertebrate models. We used adult zebrafish caudal fin as a model to investigate which and how ion transporters affect regeneration in an adult vertebrate model. Through the combined use of biophysical and molecular approaches, we show that V-ATPase activity contributes to a regeneration-specific H+ ef`flux. The onset and intensity of both V-ATPase expression and H+ efflux correlate with the different regeneration rate along the proximal-distal axis. Moreover, we show that V-ATPase inhibition impairs regeneration in adult vertebrate. Notably, the activity of this H+ pump is necessary for aldh1a2 and mkp3 expression, blastema cell proliferation and fin innervation. To the best of our knowledge, this is the first report on the role of V-ATPase during adult vertebrate regeneration.

  16. p38α MAPK regulates adult muscle stem cell fate by restricting progenitor proliferation during postnatal growth and repair.

    PubMed

    Brien, Patrick; Pugazhendhi, Dhamayanthi; Woodhouse, Samuel; Oxley, David; Pell, Jennifer M

    2013-08-01

    Stem cell function is essential for the maintenance of adult tissue homeostasis. Controlling the balance between self-renewal and differentiation is crucial to maintain a receptive satellite cell pool capable of responding to growth and regeneration cues. The mitogen-activated protein kinase p38α has been implicated in the regulation of these processes but its influence in adult muscle remains unknown. Using conditional satellite cell p38α knockout mice we have demonstrated that p38α restricts excess proliferation in the postnatal growth phase while promoting timely myoblast differentiation. Differentiation was still able to occur in the p38α-null satellite cells, however, but was delayed. An absence of p38α resulted in a postnatal growth defect along with the persistence of an increased reservoir of satellite cells into adulthood. This population was still capable of responding to cardiotoxin-induced injury, resulting in complete, albeit delayed, regeneration, with further enhancement of the satellite cell population. Increased p38γ phosphorylation accompanied the absence of p38α, and inhibition of p38γ ex vivo substantially decreased the myogenic defect. We have used genome-wide transcriptome analysis to characterize the changes in expression that occur between resting and regenerating muscle, and the influence p38α has on these expression profiles. This study provides novel evidence for the fundamental role of p38α in adult muscle homeostasis in vivo.

  17. Muscle niche-driven Insulin-Notch-Myc cascade reactivates dormant Adult Muscle Precursors in Drosophila.

    PubMed

    Aradhya, Rajaguru; Zmojdzian, Monika; Da Ponte, Jean Philippe; Jagla, Krzysztof

    2015-12-09

    How stem cells specified during development keep their non-differentiated quiescent state, and how they are reactivated, remain poorly understood. Here, we applied a Drosophila model to follow in vivo behavior of adult muscle precursors (AMPs), the transient fruit fly muscle stem cells. We report that emerging AMPs send out thin filopodia that make contact with neighboring muscles. AMPs keep their filopodia-based association with muscles throughout their dormant state but also when they start to proliferate, suggesting that muscles could play a role in AMP reactivation. Indeed, our genetic analyses indicate that muscles send inductive dIlp6 signals that switch the Insulin pathway ON in closely associated AMPs. This leads to the activation of Notch, which regulates AMP proliferation via dMyc. Altogether, we report that Drosophila AMPs display homing behavior to muscle niche and that the niche-driven Insulin-Notch-dMyc cascade plays a key role in setting the activated state of AMPs.

  18. Abnormal Skeletal Muscle Regeneration plus Mild Alterations in Mature Fiber Type Specification in Fktn-Deficient Dystroglycanopathy Muscular Dystrophy Mice

    PubMed Central

    Foltz, Steven J.; Modi, Jill N.; Melick, Garrett A.; Abousaud, Marin I.; Luan, Junna; Fortunato, Marisa J.; Beedle, Aaron M.

    2016-01-01

    Glycosylated α-dystroglycan provides an essential link between extracellular matrix proteins, like laminin, and the cellular cytoskeleton via the dystrophin-glycoprotein complex. In secondary dystroglycanopathy muscular dystrophy, glycosylation abnormalities disrupt a complex O-mannose glycan necessary for muscle structural integrity and signaling. Fktn-deficient dystroglycanopathy mice develop moderate to severe muscular dystrophy with skeletal muscle developmental and/or regeneration defects. To gain insight into the role of glycosylated α-dystroglycan in these processes, we performed muscle fiber typing in young (2, 4 and 8 week old) and regenerated muscle. In mice with Fktn disruption during skeletal muscle specification (Myf5/Fktn KO), newly regenerated fibers (embryonic myosin heavy chain positive) peaked at 4 weeks old, while total regenerated fibers (centrally nucleated) were highest at 8 weeks old in tibialis anterior (TA) and iliopsoas, indicating peak degeneration/regeneration activity around 4 weeks of age. In contrast, mature fiber type specification at 2, 4 and 8 weeks old was relatively unchanged. Fourteen days after necrotic toxin-induced injury, there was a divergence in muscle fiber types between Myf5/Fktn KO (skeletal-muscle specific) and whole animal knockout induced with tamoxifen post-development (Tam/Fktn KO) despite equivalent time after gene deletion. Notably, Tam/Fktn KO retained higher levels of embryonic myosin heavy chain expression after injury, suggesting a delay or abnormality in differentiation programs. In mature fiber type specification post-injury, there were significant interactions between genotype and toxin parameters for type 1, 2a, and 2x fibers, and a difference between Myf5/Fktn and Tam/Fktn study groups in type 2b fibers. These data suggest that functionally glycosylated α-dystroglycan has a unique role in muscle regeneration and may influence fiber type specification post-injury. PMID:26751696

  19. Deletion of Mbtps1 (Pcsk8, S1p, Ski-1) Gene in Osteocytes Stimulates Soleus Muscle Regeneration and Increased Size and Contractile Force with Age*

    PubMed Central

    Gorski, Jeff P.; Huffman, Nichole T.; Vallejo, Julian; Brotto, Leticia; Chittur, Sridar V.; Breggia, Anne; Stern, Amber; Huang, Jian; Mo, Chenglin; Seidah, Nabil G.; Bonewald, Lynda; Brotto, Marco

    2016-01-01

    Conditional deletion of Mbtps1 (cKO) protease in bone osteocytes leads to an age-related increase in mass (12%) and in contractile force (30%) in adult slow twitch soleus muscles (SOL) with no effect on fast twitch extensor digitorum longus muscles. Surprisingly, bone from 10–12-month-old cKO animals was indistinguishable from controls in size, density, and morphology except for a 25% increase in stiffness. cKO SOL exhibited increased expression of Pax7, Myog, Myod1, Notch, and Myh3 and 6-fold more centralized nuclei, characteristics of postnatal regenerating muscle, but only in type I myosin heavy chain-expressing cells. Increased expression of gene pathways mediating EGF receptor signaling, circadian exercise, striated muscle contraction, and lipid and carbohydrate oxidative metabolism were also observed in cKO SOL. This muscle phenotype was not observed in 3-month-old mice. Although Mbtps1 mRNA and protein expression was reduced in cKO bone osteocytes, no differences in Mbtps1 or cre recombinase expression were observed in cKO SOL, explaining this age-related phenotype. Understanding bone-muscle cross-talk may provide a fresh and novel approach to prevention and treatment of age-related muscle loss. PMID:26719336

  20. Dietary Flaxseed Mitigates Impaired Skeletal Muscle Regeneration: in Vivo, in Vitro and in Silico Studies

    PubMed Central

    Carotenuto, Felicia; Costa, Alessandra; Albertini, Maria Cristina; Rocchi, Marco Bruno Luigi; Rudov, Alexander; Coletti, Dario; Minieri, Marilena; Di Nardo, Paolo; Teodori, Laura

    2016-01-01

    Background: Diets enriched with n-3 polyunsaturated fatty acids (n-3 PUFAs) have been shown to exert a positive impact on muscle diseases. Flaxseed is one of the richest sources of n-3 PUFA acid α-linolenic acid (ALA). The aim of this study was to assess the effects of flaxseed and ALA in models of skeletal muscle degeneration characterized by high levels of Tumor Necrosis Factor-α (TNF). Methods: The in vivo studies were carried out on dystrophic hamsters affected by muscle damage associated with high TNF plasma levels and fed with a long-term 30% flaxseed-supplemented diet. Differentiating C2C12 myoblasts treated with TNF and challenged with ALA represented the in vitro model. Skeletal muscle morphology was scrutinized by applying the Principal Component Analysis statistical method. Apoptosis, inflammation and myogenesis were analyzed by immunofluorescence. Finally, an in silico analysis was carried out to predict the possible pathways underlying the effects of n-3 PUFAs. Results: The flaxseed-enriched diet protected the dystrophic muscle from apoptosis and preserved muscle myogenesis by increasing the myogenin and alpha myosin heavy chain. Moreover, it restored the normal expression pattern of caveolin-3 thereby allowing protein retention at the sarcolemma. ALA reduced TNF-induced apoptosis in differentiating myoblasts and prevented the TNF-induced inhibition of myogenesis, as demonstrated by the increased expression of myogenin, myosin heavy chain and caveolin-3, while promoting myotube fusion. The in silico investigation revealed that FAK pathways may play a central role in the protective effects of ALA on myogenesis. Conclusions: These findings indicate that flaxseed may exert potent beneficial effects by preserving skeletal muscle regeneration and homeostasis partly through an ALA-mediated action. Thus, dietary flaxseed and ALA may serve as a useful strategy for treating patients with muscle dystrophies. PMID:26941581

  1. Therapies for sarcopenia and regeneration of old skeletal muscles: more a case of old tissue architecture than old stem cells.

    PubMed

    Grounds, Miranda D

    2014-01-01

    Age related loss of skeletal muscle mass and function (sarcopenia) reduces independence and the quality of life for individuals, and leads to falls and fractures with escalating health costs for the rapidly aging human population. Thus there is much interest in developing interventions to reduce sarcopenia. One area that has attracted recent attention is the proposed use of myogenic stem cells to improve regeneration of old muscles. This mini-review challenges the fundamental need for myogenic stem cell therapy for sarcopenia. It presents evidence that demonstrates the excellent capacity of myogenic stem cells from very old rodent and human muscles to form new muscles after experimental myofiber necrosis. The many factors required for successful muscle regeneration are considered with a strong focus on integration of components of old muscle bioarchitecture. The fundamental role of satellite cells in homeostasis of normal aging muscles and the incidence of endogenous regeneration in old muscles is questioned. These issues, combined with problems for clinical myogenic stem cell therapies for severe muscle diseases, raise fundamental concerns about the justification for myogenic stem cell therapy for sarcopenia.

  2. AMP-activated protein kinase stimulates Warburg-like glycolysis and activation of satellite cells during muscle regeneration.

    PubMed

    Fu, Xing; Zhu, Mei-Jun; Dodson, Mike V; Du, Min

    2015-10-30

    Satellite cells are the major myogenic stem cells residing inside skeletal muscle and are indispensable for muscle regeneration. Satellite cells remain largely quiescent but are rapidly activated in response to muscle injury, and the derived myogenic cells then fuse to repair damaged muscle fibers or form new muscle fibers. However, mechanisms eliciting metabolic activation, an inseparable step for satellite cell activation following muscle injury, have not been defined. We found that a noncanonical Sonic Hedgehog (Shh) pathway is rapidly activated in response to muscle injury, which activates AMPK and induces a Warburg-like glycolysis in satellite cells. AMPKα1 is the dominant AMPKα isoform expressed in satellite cells, and AMPKα1 deficiency in satellite cells impairs their activation and myogenic differentiation during muscle regeneration. Drugs activating noncanonical Shh promote proliferation of satellite cells, which is abolished because of satellite cell-specific AMPKα1 knock-out. Taken together, AMPKα1 is a critical mediator linking noncanonical Shh pathway to Warburg-like glycolysis in satellite cells, which is required for satellite activation and muscle regeneration.

  3. Sumoylated α-skeletal muscle actin in the skeletal muscle of adult rats.

    PubMed

    Uda, Munehiro; Kawasaki, Hiroaki; Iizumi, Kyoichi; Shigenaga, Ayako; Baba, Takeshi; Naito, Hisashi; Yoshioka, Toshitada; Yamakura, Fumiyuki

    2015-11-01

    Skeletal muscles are composed of two major muscle fiber types: slow-twitch oxidative fibers and fast-twitch glycolytic fibers. The proteins in these muscle fibers are known to differ in their expression, relative abundance, and post-translational modifications. In this study, we report a previously unreported post-translational modification of α-skeletal muscle actin in the skeletal muscles of adult male F344 rats in vivo. Using two-dimensional electrophoresis (2D-PAGE), we first examined the differences in the protein expression profiles between the soleus and plantaris muscles. We found higher intensity protein spots at approximately 60 kDa and pH 9 on 2D-PAGE for the soleus muscle compared with the plantaris muscle. These spots were identified as α-skeletal muscle actin by liquid chromatography-nanoelectrospray ionization-tandem mass spectrometry and western blot analyses. In addition, we found that the 60 kDa α-skeletal muscle actin is modified by small ubiquitin-like modifier (SUMO) 1, using 2D-PAGE and western blot analyses. Furthermore, we found that α-skeletal muscle actin with larger molecular weight was localized in the nuclear and cytosol of the skeletal muscle, but not in the myofibrillar fraction by the combination of subcellular fractionation and western blot analyses. These results suggest that α-skeletal muscle actin is modified by SUMO-1 in the skeletal muscles, localized in nuclear and cytosolic fractions, and the extent of this modification is much higher in the slow muscles than in the fast muscles. This is the first study to show the presence of SUMOylated actin in animal tissues.

  4. Muscle MRI Findings in Childhood/Adult Onset Pompe Disease Correlate with Muscle Function

    PubMed Central

    Figueroa-Bonaparte, Sebastián; Segovia, Sonia; Llauger, Jaume; Belmonte, Izaskun; Pedrosa, Irene; Alejaldre, Aída; Mayos, Mercè; Suárez-Cuartín, Guillermo; Gallardo, Eduard; Illa, Isabel; Díaz-Manera, Jordi

    2016-01-01

    Objectives Enzyme replacement therapy has shown to be effective for childhood/adult onset Pompe disease (AOPD). The discovery of biomarkers useful for monitoring disease progression is one of the priority research topics in Pompe disease. Muscle MRI could be one possible test but the correlation between muscle MRI and muscle strength and function has been only partially addressed so far. Methods We studied 34 AOPD patients using functional scales (Manual Research Council scale, hand held myometry, 6 minutes walking test, timed to up and go test, time to climb up and down 4 steps, time to walk 10 meters and Motor Function Measure 20 Scale), respiratory tests (Forced Vital Capacity seated and lying, Maximun Inspiratory Pressure and Maximum Expiratory Pressure), daily live activities scales (Activlim) and quality of life scales (Short Form-36 and Individualized Neuromuscular Quality of Life questionnaire). We performed a whole body muscle MRI using T1w and 3-point Dixon imaging centered on thighs and lower trunk region. Results T1w whole body muscle MRI showed a homogeneous pattern of muscle involvement that could also be found in pre-symptomatic individuals. We found a strong correlation between muscle strength, muscle functional scales and the degree of muscle fatty replacement in muscle MRI analyzed using T1w and 3-point Dixon imaging studies. Moreover, muscle MRI detected mild degree of fatty replacement in paraspinal muscles in pre-symptomatic patients. Conclusion Based on our findings, we consider that muscle MRI correlates with muscle function in patients with AOPD and could be useful for diagnosis and follow-up in pre-symptomatic and symptomatic patients under treatment. Take home message Muscle MRI correlates with muscle function in patients with AOPD and could be useful to follow-up patients in daily clinic. PMID:27711114

  5. Intrinsic facilitation of adult peripheral nerve regeneration by the Sonic hedgehog morphogen.

    PubMed

    Martinez, Jose A; Kobayashi, Masaki; Krishnan, Anand; Webber, Christine; Christie, Kimberly; Guo, GuiFang; Singh, Vandana; Zochodne, Douglas W

    2015-09-01

    Intrinsic molecular determinants of neurodevelopmental outcomes assume new, albeit related roles during adult neural regeneration. Here we studied and identified a facilitatory role for Sonic hedgehog protein (Shh), a morphogen that influences motor neuron floor plate architecture, during adult peripheral neuron regeneration. Shh and its receptors were expressed in adult dorsal root ganglia (DRG) neurons, axons and glia and trended toward higher levels following axotomy injury. Knockdown of Shh in adult sensory neurons resulted in decreased outgrowth and branching in vitro, identifying a role for Shh in facilitating outgrowth. The findings argued for an intrinsic action to support neuron regeneration. Support of advancement and turning however, were not identified in adult sensory neuron growth cones in response to local extrinsic gradients of Shh. That intrinsic Shh supported the regrowth of peripheral nerves after injury was confirmed by the analysis of axon regrowth from the proximal stumps of transected sciatic nerves. By exposing regenerating axons to local infusions of Shh siRNA in vivo within a conduit bridging the transected proximal and distal stumps, we achieved local knockdown of Shh. In response, there was attenuated axonal and Schwann cell outgrowth beyond the transection zone. Unlike its role during neurodevelopment, Shh facilitates but does not confer regenerative outgrowth properties to adult neurons alone. Exploring the differing properties of morphogens and related proteins in the adult nervous system identifies new and important roles for them.

  6. Transplanted Endothelial Progenitor Cells Improve Ischemia Muscle Regeneration in Mice by Diffusion Tensor MR Imaging

    PubMed Central

    Bai, Yingying; James, Judy R.; Shlapak, Darya P.

    2016-01-01

    Endothelial progenitor cells (EPCs) play an important role in repairing ischemia tissues. Diffusion tensor imaging (DTI) was applied to detect the architectural organization of skeletal muscle. This study investigated the feasibility and accuracy of using the DTI to evaluate effectiveness of EPCs treatment. Mouse bone marrow-derived EPCs were isolated, cultured, characterized, and transplanted to hindlimb ischemia mice model. DTI was performed on the hindlimb at postischemia time points. The edema regions of diffusion restriction (high signal in diffusion weighted imaging) were decreased in the ischemic muscle of EPCs treated mice after 14 days compared with the controls. These results from DTI show the lower apparent diffusion coefficient and eigenvalues (λ1, λ2, and λ3) and the higher fractional anisotropy and fiber counts of ischemic muscle on 7 and 14 days after EPCs treatment compared to the controls. There was a significant correlation between fiber counts calculated by DTI and survival fibers evaluated by histological section (r = 0.873, P < 0.01). Our study demonstrated that the time frame for muscle fiber regeneration after EPCs transplantation was significantly shortened in vivo. DTI could be a useful tool for noninvasive evaluation of muscle tissue damage and repair in animal models and patient with ischemic diseases. PMID:27656214

  7. Is salamander hindlimb regeneration similar to that of the forelimb? Anatomical and morphogenetic analysis of hindlimb muscle regeneration in GFP-transgenic axolotls as a basis for regenerative and developmental studies

    PubMed Central

    Diogo, R; Murawala, P; Tanaka, E M

    2014-01-01

    The axolotl Ambystoma mexicanum is one of the most used model organisms in developmental and regenerative studies because it is commonly said that it can reconstitute a normal and fully functional forelimb/hindlimb after amputation. However, there is not a publication that has described in detail the regeneration of the axolotl hindlimb muscles. Here we describe and illustrate, for the first time, the regeneration of the thigh, leg and foot muscles in transgenic axolotls that express green fluorescent protein in muscle fibers and compare our results with data obtained by us and by other authors about axolotl forelimb regeneration and about fore-and hindlimb ontogeny in axolotls, frogs and other tetrapods. Our observations and comparisons point out that: (1) there are no muscle anomalies in any regenerated axolotl hindlimbs, in clear contrast to our previous study of axolotl forelimb regeneration, where we found muscle anomalies in 43% of the regenerated forelimbs; (2) during axolotl hindlimb regeneration there is a proximo-distal and a tibio-fibular morphogenetic gradient in the order of muscle regeneration and differentiation, but not a ventro-dorsal gradient, whereas our previous studies showed that in axolotl forelimb muscle regeneration there are proximo-distal, radio-ulnar and ventro-dorsal morphogenetic gradients. We discuss the broader implications of these observations for regenerative, evolutionary, developmental and morphogenetic studies. PMID:24325444

  8. Is salamander hindlimb regeneration similar to that of the forelimb? Anatomical and morphogenetic analysis of hindlimb muscle regeneration in GFP-transgenic axolotls as a basis for regenerative and developmental studies.

    PubMed

    Diogo, R; Murawala, P; Tanaka, E M

    2014-04-01

    The axolotl Ambystoma mexicanum is one of the most used model organisms in developmental and regenerative studies because it is commonly said that it can reconstitute a normal and fully functional forelimb/hindlimb after amputation. However, there is not a publication that has described in detail the regeneration of the axolotl hindlimb muscles. Here we describe and illustrate, for the first time, the regeneration of the thigh, leg and foot muscles in transgenic axolotls that express green fluorescent protein in muscle fibers and compare our results with data obtained by us and by other authors about axolotl forelimb regeneration and about fore- and hindlimb ontogeny in axolotls, frogs and other tetrapods. Our observations and comparisons point out that: (1) there are no muscle anomalies in any regenerated axolotl hindlimbs, in clear contrast to our previous study of axolotl forelimb regeneration, where we found muscle anomalies in 43% of the regenerated forelimbs; (2) during axolotl hindlimb regeneration there is a proximo-distal and a tibio-fibular morphogenetic gradient in the order of muscle regeneration and differentiation, but not a ventro-dorsal gradient, whereas our previous studies showed that in axolotl forelimb muscle regeneration there are proximo-distal, radio-ulnar and ventro-dorsal morphogenetic gradients. We discuss the broader implications of these observations for regenerative, evolutionary, developmental and morphogenetic studies.

  9. Ultrasound modulates the inflammatory response and promotes muscle regeneration in injured muscles.

    PubMed

    Nagata, Kumiko; Nakamura, Tatsuya; Fujihara, Shinji; Tanaka, Eiji

    2013-06-01

    The purpose of this study was to examine the effect of ultrasound on inflammatory skeletal muscle in vitro and in vivo. C2C12 cells were cultured in medium with or without TNF-α or IL-1β. After stimulation with cytokines, the cells received ultrasound or sham exposure. Furthermore, the tibialis anterior (TA) muscle in C57BL/6 mice injured by cardiotoxin (CTX) were dissected after a series of ultrasound treatments and examined. Exposure of C2C12 cells to ultrasound resulted in down-regulation of cyclooxygenase-2 (COX-2) mRNA and protein expression induced by TNF-α or IL-1β, and up-regulated myogenin mRNA and protein depressed by TNF-α or IL-1β. In injured TA muscle induced by CTX, ultrasound caused increase of COX-2 mRNA at 1 day after ultrasound treatment, however, at day 5, reduction of COX-2 mRNA and protein. At day 5, ultrasound caused increase of myogenin mRNA and protein, increase of fast myosin protein, and increase of paired-box transcription factor 7 positive cells. At day 7, the cross-sectional area of myofibers in the ultrasound exposure side was significantly larger than that on the control side. In conclusion, ultrasound stimulation may be a better candidate as a medical remedy to promote myogenesis in inflammatory muscle states, such as muscle injury.

  10. Concise Review: Quiescence in Adult Stem Cells: Biological Significance and Relevance to Tissue Regeneration.

    PubMed

    Rumman, Mohammad; Dhawan, Jyotsna; Kassem, Moustapha

    2015-10-01

    Adult stem cells (ASCs) are tissue resident stem cells responsible for tissue homeostasis and regeneration following injury. In uninjured tissues, ASCs exist in a nonproliferating, reversibly cell cycle-arrested state known as quiescence or G0. A key function of the quiescent state is to preserve stemness in ASCs by preventing precocious differentiation, and thus maintaining a pool of undifferentiated ASCs. Recent evidences suggest that quiescence is an actively maintained state and that excessive or defective quiescence may lead to compromised tissue regeneration or tumorigenesis. The aim of this review is to provide an update regarding the biological mechanisms of ASC quiescence and their role in tissue regeneration.

  11. Myf5 haploinsufficiency reveals distinct cell fate potentials for adult skeletal muscle stem cells.

    PubMed

    Gayraud-Morel, Barbara; Chrétien, Fabrice; Jory, Aurélie; Sambasivan, Ramkumar; Negroni, Elisa; Flamant, Patricia; Soubigou, Guillaume; Coppée, Jean-Yves; Di Santo, James; Cumano, Ana; Mouly, Vincent; Tajbakhsh, Shahragim

    2012-04-01

    Skeletal muscle stem cell fate in adult mice is regulated by crucial transcription factors, including the determination genes Myf5 and Myod. The precise role of Myf5 in regulating quiescent muscle stem cells has remained elusive. Here we show that most, but not all, quiescent satellite cells express Myf5 protein, but at varying levels, and that resident Myf5 heterozygous muscle stem cells are more primed for myogenic commitment compared with wild-type satellite cells. Paradoxically however, heterotypic transplantation of Myf5 heterozygous cells into regenerating muscles results in higher self-renewal capacity compared with wild-type stem cells, whereas myofibre regenerative capacity is not altered. By contrast, Pax7 haploinsufficiency does not show major modifications by transcriptome analysis. These observations provide a mechanism linking Myf5 levels to muscle stem cell heterogeneity and fate by exposing two distinct and opposing phenotypes associated with Myf5 haploinsufficiency. These findings have important implications for how stem cell fates can be modulated by crucial transcription factors while generating a pool of responsive heterogeneous cells.

  12. Robust regeneration of adult zebrafish lateral line hair cells reflects continued precursor pool maintenance

    PubMed Central

    Cruz, Ivan A.; Kappedal, Ryan; Mackenzie, Scott M.; Hailey, Dale W.; Hoffman, Trevor L.; Schilling, Thomas F.; Raible, David W.

    2015-01-01

    We have examined lateral line hair cell and support cell maintenance in adult zebrafish when growth is largely complete. We demonstrate that adult zebrafish not only replenish hair cells after a single instance of hair cell damage, but also maintain hair cells and support cells after multiple rounds of damage and regeneration. We find that hair cells undergo continuous turnover in adult zebrafish in the absence of damage. We identify mitotically-distinct support cell populations and show that hair cells regenerate from underlying support cells in a region-specific manner. Our results demonstrate that there are two distinct support cell populations in the lateral line, which may help explain why zebrafish hair cell regeneration is extremely robust, retained throughout life, and potentially unlimited in regenerative capacity. PMID:25869855

  13. Skeletal muscle regeneration on protein-grafted and microchannel-patterned scaffold for hypopharyngeal tissue engineering.

    PubMed

    Shen, Zhisen; Guo, Shanshan; Ye, Dong; Chen, Jingjing; Kang, Cheng; Qiu, Shejie; Lu, Dakai; Li, Qun; Xu, Kunjie; Lv, Jingjing; Zhu, Yabin

    2013-01-01

    In the field of tissue engineering, polymeric materials with high biocompatibility like polylactic acid and polyglycolic acid have been widely used for fabricating living constructs. For hypopharynx tissue engineering, skeletal muscle is one important functional part of the whole organ, which assembles the unidirectionally aligned myotubes. In this study, a polyurethane (PU) scaffold with microchannel patterns was used to provide aligning guidance for the seeded human myoblasts. Due to the low hydrophilicity of PU, the scaffold was grafted with silk fibroin (PU-SF) or gelatin (PU-Gel) to improve its cell adhesion properties. Scaffolds were observed to degrade slowly over time, and their mechanical properties and hydrophilicities were improved through the surface grafting. Also, the myoblasts seeded on PU-SF had the higher proliferative rate and better differentiation compared with those on the control or PU-Gel. Our results demonstrate that polyurethane scaffolds seeded with myoblasts hold promise to guide hypopharynx muscle regeneration.

  14. Skeletal Muscle Regeneration on Protein-Grafted and Microchannel-Patterned Scaffold for Hypopharyngeal Tissue Engineering

    PubMed Central

    Shen, Zhisen; Guo, Shanshan; Ye, Dong; Chen, Jingjing; Kang, Cheng; Qiu, Shejie; Lu, Dakai; Li, Qun; Xu, Kunjie; Lv, Jingjing

    2013-01-01

    In the field of tissue engineering, polymeric materials with high biocompatibility like polylactic acid and polyglycolic acid have been widely used for fabricating living constructs. For hypopharynx tissue engineering, skeletal muscle is one important functional part of the whole organ, which assembles the unidirectionally aligned myotubes. In this study, a polyurethane (PU) scaffold with microchannel patterns was used to provide aligning guidance for the seeded human myoblasts. Due to the low hydrophilicity of PU, the scaffold was grafted with silk fibroin (PU-SF) or gelatin (PU-Gel) to improve its cell adhesion properties. Scaffolds were observed to degrade slowly over time, and their mechanical properties and hydrophilicities were improved through the surface grafting. Also, the myoblasts seeded on PU-SF had the higher proliferative rate and better differentiation compared with those on the control or PU-Gel. Our results demonstrate that polyurethane scaffolds seeded with myoblasts hold promise to guide hypopharynx muscle regeneration. PMID:24175281

  15. Myogenic-specific ablation of Fgfr1 impairs FGF2-mediated proliferation of satellite cells at the myofiber niche but does not abolish the capacity for muscle regeneration

    PubMed Central

    Yablonka-Reuveni, Zipora; Danoviz, Maria E.; Phelps, Michael; Stuelsatz, Pascal

    2015-01-01

    Skeletal muscle satellite cells (SCs) are Pax7+ myogenic stem cells that reside between the basal lamina and the plasmalemma of the myofiber. In mature muscles, SCs are typically quiescent, but can be activated in response to muscle injury. Depending on the magnitude of tissue trauma, SCs may divide minimally to repair subtle damage within individual myofibers or produce a larger progeny pool that forms new myofibers in cases of overt muscle injury. SC transition through proliferation, differentiation and renewal is governed by the molecular blueprint of the cells as well as by the extracellular milieu at the SC niche. In particular, the role of the fibroblast growth factor (FGF) family in regulating SCs during growth and aging is well recognized. Of the several FGFs shown to affect SCs, FGF1, FGF2, and FGF6 proteins have been documented in adult skeletal muscle. These prototypic paracrine FGFs transmit their mitogenic effect through the FGFRs, which are transmembrane tyrosine kinase receptors. Using the mouse model, we show here that of the four Fgfr genes, only Fgfr1 and Fgfr4 are expressed at relatively high levels in quiescent SCs and their proliferating progeny. To further investigate the role of FGFR1 in adult myogenesis, we have employed a genetic (Cre/loxP) approach for myogenic-specific (MyoDCre-driven) ablation of Fgfr1. Neither muscle histology nor muscle regeneration following cardiotoxin-induced injury were overtly affected in Fgfr1-ablated mice. This suggests that FGFR1 is not obligatory for SC performance in this acute muscle trauma model, where compensatory growth factor/cytokine regulatory cascades may exist. However, the SC mitogenic response to FGF2 is drastically repressed in isolated myofibers prepared from Fgfr1-ablated mice. Collectively, our study indicates that FGFR1 is important for FGF-mediated proliferation of SCs and its mitogenic role is not compensated by FGFR4 that is also highly expressed in SCs. PMID:26074812

  16. Satellite cell proliferation in adult skeletal muscle

    NASA Technical Reports Server (NTRS)

    Booth, Frank W. (Inventor); Thomason, Donald B. (Inventor); Morrison, Paul R. (Inventor); Stancel, George M. (Inventor)

    1995-01-01

    Novel methods of retroviral-mediated gene transfer for the in vivo corporation and stable expression of eukaryotic or prokaryotic foreign genes in tissues of living animals is described. More specifically, methods of incorporating foreign genes into mitotically active cells are disclosed. The constitutive and stable expression of E. coli .beta.-galactosidase gene under the promoter control of the Moloney murine leukemia virus long terminal repeat is employed as a particularly preferred embodiment, by way of example, establishes the model upon which the incorporation of a foreign gene into a mitotically-active living eukaryotic tissue is based. Use of the described methods in therapeutic treatments for genetic diseases, such as those muscular degenerative diseases, is also presented. In muscle tissue, the described processes result in genetically-altered satellite cells which proliferate daughter myoblasts which preferentially fuse to form a single undamaged muscle fiber replacing damaged muscle tissue in a treated animal. The retroviral vector, by way of example, includes a dystrophin gene construct for use in treating muscular dystrophy. The present invention also comprises an experimental model utilizable in the study of the physiological regulation of skeletal muscle gene expression in intact animals.

  17. Axonal regeneration in severed peripheral facial nerve of the rabbit: relation of the number of axonal regenerates to behavioral and evoked muscle activity.

    PubMed

    Spector, J G; Lee, P

    1998-02-01

    The minimum number of regenerating facial nerve myelinated motor axons that are required to innervate and activate the mimetic musculature is not known. We compare rabbit facial nerve regeneration following complete transectional injuries of the buccal division to the evoked and behavioral muscle activities of the quadratus labii superioris muscle of the rabbit in three experimental models: end-to-end direct anastomosis (N = 4), 8-mm autologous nerve grafts (N = 8), and 10-mm silicone chamber implants (N = 40). Data are presented as total numbers of regenerating myelinated axons that traverse the surgical repair and innervate the fascicles of the transected distal nerve stump, as well as the percentage of regenerating neurites, as compared to the preoperative normal controls. Five weeks after neural repair, direct end-to-end anastomosis regained more myelinated axons across the reconstructed defect (2,632 +/- 1,232; 67%) than silicone tube implants (2,006 +/- 445; 51%) or autologous cable graft repairs (1,660 +/- 1,169; 42%). However, only a small percentage of myelinated fibers innervated the intrafascicular region of the distal transected neural stump in direct anastomosis (948 +/- 168; 24%), silicone tube implants (670 +/- 275; 17%), or autologous nerve grafts (445 +/- 120; 12%) in rabbits that regained evoked and behavioral mimetic muscle activity. All rabbits with direct anastomosis and neural cable grafts regained motor activity, despite the fact that 66% of regenerating motor neurites in cable graft repairs and 54% in direct anastomosis were collateral sprouts that did not contribute to effective muscle activity. In 17 rabbits with neural regenerates within the silicone tube implants that did not regain mimetic activity, the mean number of regenerating myelinated motor axons across the defect was 504 +/- 419 (13%), and the mean number of axons that innervated the distal transected nerve stump fascicles was 277 +/- 128 (7%). Therefore, the minimal number of

  18. Myodegeneration with fibrosis and regeneration in the pectoralis major muscle of broilers.

    PubMed

    Sihvo, H-K; Immonen, K; Puolanne, E

    2014-05-01

    A myopathy affecting the pectoralis major muscle of the commercial broiler has emerged creating remarkable economic losses as well as a potential welfare problem of the birds. We here describe the macroscopic and histologic lesions of this myopathy within 10 pectoralis major muscles of 5- to 6-week-old broilers in Finland. Following macroscopic evaluation and palpation of the muscles, a tissue sample of each was fixed in formalin, processed for histology, and histologically evaluated. The muscles that were macroscopically hard, outbulging, pale, and often accompanied with white striping histologically exhibited moderate to severe polyphasic myodegeneration with regeneration as well as a variable amount of interstitial connective tissue accumulation or fibrosis. All affected cases also exhibited perivenular lymphocyte accumulation. The etiology of this myodegenerative lesion remains yet open. Polyphasic myodegeneration is associated with several previously known etiologies, but palpatory hardness focusing on the pectoralis major, together with perivenular lymphocytes, has not been described in relation to them. The results of this study provide the pathological basis for further studies concerning the etiology of the currently described myopathy.

  19. Dose-dependency of low-energy HeNe laser effect in regeneration of skeletal muscle in mice.

    PubMed

    Amaral, A C; Parizotto, N A; Salvini, T F

    2001-01-01

    We evaluated the effect on mice skeletal muscle regeneration of different doses (2.6, 8.4, and 25 J/cm2) of HeNe laser (lambda 632.8 nm; power, 2.6 mW; spot size, 0.007 cm2) applied directly to intact skin of injured muscle. Muscle injury was induced in both right and left Tibialis anterior (TA) muscles by ACL myotoxin (5 mg/kg). Right TA muscles were irradiated daily for 5 days while contralateral muscles received a sham treatment. Only the 2.6 J/cm2 dose resulted in changes such as increased mitochondrial density and muscle fibre in the TA muscles as compared to sham groups (3280 +/- 704 microns 2 versus 2110 +/- 657 microns 2, p = 0.02). We concluded that the HeNe effect on mouse muscle regeneration is dose-specific: only 2.6 J/cm2 increased muscle fibre area and mitochondrial density.

  20. CD8 T cells are involved in skeletal muscle regeneration through facilitating MCP-1 secretion and Gr1(high) macrophage infiltration.

    PubMed

    Zhang, Jing; Xiao, Zhicheng; Qu, Chao; Cui, Wei; Wang, Xiaonan; Du, Jie

    2014-11-15

    Inflammatory microenvironments play a key role in skeletal muscle regeneration. The infiltration of CD8 T cells into injured muscle has been reported. However, the role of CD8 T cells during skeletal muscle regeneration remains unclear. In this study, we used cardiotoxin-induced mouse skeletal muscle injury/regeneration model to investigate the role of CD8 T cells. Muscle regeneration was impaired and matrix deposit was increased in CD8α-deficient mice compared with wild-type (WT) mice whose CD8 T cells were infiltrated into damaged muscle after cardiotoxin injection. Adoptive transfer of CD8 T cells to CD8α-deficient mice improved muscle regeneration and inhibited matrix remodeling. Compared with WT mice, CD8α deficiency limited the recruitment of Gr1(high) macrophages (MPs) into muscle, resulting in the reduction of satellite cell number. The expression of MCP-1 (MCP-1/CCL2), which regulates the migration of Gr1(high) MPs, was reduced in CD8α-deficient mice compared with WT mice. Coculture CD8 T cells with MPs promoted MCP-1 secretion. The i.m. injection of MCP-1 markedly promoted the recruitment of Gr1(high) MPs and improved muscle regeneration in CD8α-deficient mice. We conclude that CD8 T cells are involved in skeletal muscle regeneration by regulating the secretion of MCP-1 to recruit Gr1(high) MPs, which facilitate myoblast proliferation.

  1. Patterns and cellular mechanisms of arm regeneration in adult starfish Asterias rollestoni bell

    NASA Astrophysics Data System (ADS)

    Fan, Tingjun; Fan, Xianyuan; Du, Yutang; Sun, Wenjie; Zhang, Shaofeng; Li, Jiaxin

    2011-09-01

    To understand the mechanisms of starfish regeneration, the arms of adult starfish Asterias rollestoni Bell were amputated and their regeneration patterns and cellular mechanisms were studied. It was found that cells in the outer epidermis and inner parietal peritoneum near the end of the stump began to dedifferentiate 4 d after amputation. The dedifferentiated cells in the outer epidermis proliferated, migrated to the wound site and formed a thickened pre-epidermis which would then re-differentiate gradually into mature epidermis. The new parietal peritoneum formed on the coelomic side of wound might be from the curvely elongated parietal peritoneum, resulting from the dedifferentiated and proliferated cells by extension. Afterwards, the proliferated cells made the outer epidermis and inner parietal peritoneum invaginate into the interior dermis and formed blastema-like structures together with induced dedifferentiated dermal cells. Most interestingly, the arm regeneration in A. rollestoni was achieved synchronously by de novo arm-bud formation and growth, and arm-stump elongation. The crucial aspects of arm-bud formation included cell dedifferentiation, proliferation and migration, while those of arm-stump elongation included cell dedifferentiation, proliferation, invagination, and arm-wall-across blastema-like structure formation. The unique pattern and cellular mechanisms of amputated arm regeneration make it easier to understand the rapid regeneration process of adult starfish. This study may lay solid foundations for the research into molecular mechanisms of echinoderm regeneration.

  2. Perforated microelectrode arrays implanted in the regenerating adult central nervous system.

    PubMed

    Heiduschka, P; Romann, I; Stieglitz, T; Thanos, S

    2001-09-01

    Adult mammalian optic nerve axons are able to regenerate, when provided with the permissive environment of an autologous peripheral nerve graft, which is usually the sciatic nerve. This study demonstrates the ability of adult rat optic nerve axons to regenerate through the preformed perforations of a polyimide electrode carrier implanted at the interface between the proximal stump of the cut optic nerve and the stump of the peripheral nerve piece used for grafting. Evidence that retinal ganglion cells regenerated their axons through the perforated electrode carrier was obtained by retrograde labeling with a fluorescent dye deposited into the sciatic nerve graft beyond the nerve-carrier-nerve junction. The number of regenerating cells could be enhanced by injecting neuroprotective drugs like aurintricarboxylic acid and cortisol intravitreally. A second line of evidence was obtained by immunohistochemical staining with antibodies to neurofilament. Third, electrical activity of the regenerating nerves was recorded after stimulating the retina with a flash of light. The results suggest that a regenerating central nerve tract may serve as an experimental model to implant artificial microdevices to monitor the physiological and topographical properties of neurites passing through the device or to stimulate them, thus interfering with their potential to grow. This study reports for the first time that the optic nerve has unique properties, which aids in the realization of these goals.

  3. Limb Regeneration is Impaired in an Adult Zebrafish Model of Diabetes Mellitus

    PubMed Central

    Olsen, Ansgar S.; Sarras, Michael P.; Intine, Robert V.

    2010-01-01

    The zebrafish (Danio Rerio) is an established model organism for the study of developmental processes, human disease and tissue regeneration. We report that limb regeneration is severely impaired in our newly developed adult zebrafish model of type I diabetes. Intraperitoneal streptozocin injection of adult, wild type zebrafish results in a sustained hyperglycemic state as determined by elevated fasting blood glucose values and increased glycation of serum protein. Serum insulin levels are also decreased and pancreas immunohistochemisty revealed a lesser amount of insulin signal in hyperglycemic fish. Additionally, the diabetic complications of retinal thinning and glomerular basement membrane thickening (early signs of retinopathy and nephropathy) resulting from the hyperglycemic state were evident in streptozocin injected fish at three weeks. Most significantly, limb regeneration, following caudal fin amputation, is severely impaired in diabetic zebrafish. Nonspecific toxic effects outside the pancreas were not found to contribute to impaired limb regeneration. This experimental system using adult zebrafish facilitates a broad spectrum of genetic and molecular approaches to study regeneration in the diabetic background. PMID:20840523

  4. Potential for neural regeneration after neurotoxic injury in the adult mammalian retina

    PubMed Central

    Ooto, Sotaro; Akagi, Tadamichi; Kageyama, Ryoichiro; Akita, Joe; Mandai, Michiko; Honda, Yoshihito; Takahashi, Masayo

    2004-01-01

    It has long been believed that the retina of mature mammals is incapable of regeneration. In this study, using the N-methyl-d-aspartate neurotoxicity model of adult rat retina, we observed that some Müller glial cells were stimulated to proliferate in response to a toxic injury and produce bipolar cells and rod photoreceptors. Although these newly produced neurons were limited in number, retinoic acid treatment promoted the number of regenerated bipolar cells. Moreover, misexpression of basic helix–loop–helix and homeobox genes promoted the induction of amacrine, horizontal, and rod photoreceptor specific phenotypes. These findings demonstrated that retinal neurons regenerated even in adult mammalian retina after toxic injury. Furthermore, we could partially control the fate of the regenerated neurons with extrinsic factors or intrinsic genes. The Müller glial cells constitute a potential source for the regeneration of adult mammalian retina and can be a target for drug delivery and gene therapy in retinal degenerative diseases. PMID:15353594

  5. Clonogenic neoblasts are pluripotent adult stem cells that underlie planarian regeneration.

    PubMed

    Wagner, Daniel E; Wang, Irving E; Reddien, Peter W

    2011-05-13

    Pluripotent cells in the embryo can generate all cell types, but lineage-restricted cells are generally thought to replenish adult tissues. Planarians are flatworms and regenerate from tiny body fragments, a process requiring a population of proliferating cells (neoblasts). Whether regeneration is accomplished by pluripotent cells or by the collective activity of multiple lineage-restricted cell types is unknown. We used ionizing radiation and single-cell transplantation to identify neoblasts that can form large descendant-cell colonies in vivo. These clonogenic neoblasts (cNeoblasts) produce cells that differentiate into neuronal, intestinal, and other known postmitotic cell types and are distributed throughout the body. Single transplanted cNeoblasts restored regeneration in lethally irradiated hosts. We conclude that broadly distributed, adult pluripotent stem cells underlie the remarkable regenerative abilities of planarians.

  6. Isolation and Culture of Individual Myofibers and their Satellite Cells from Adult Skeletal Muscle

    PubMed Central

    Pasut, Alessandra; Jones, Andrew E.; Rudnicki, Michael A.

    2013-01-01

    Muscle regeneration in the adult is performed by resident stem cells called satellite cells. Satellite cells are defined by their position between the basal lamina and the sarcolemma of each myofiber. Current knowledge of their behavior heavily relies on the use of the single myofiber isolation protocol. In 1985, Bischoff described a protocol to isolate single live fibers from the Flexor Digitorum Brevis (FDB) of adult rats with the goal to create an in vitro system in which the physical association between the myofiber and its stem cells is preserved 1. In 1995, Rosenblattmodified the Bischoff protocol such that myofibers are singly picked and handled separately after collagenase digestion instead of being isolated by gravity sedimentation 2, 3. The Rosenblatt or Bischoff protocol has since been adapted to different muscles, age or conditions 3-6. The single myofiber isolation technique is an indispensable tool due its unique advantages. First, in the single myofiber protocol, satellite cells are maintained beneath the basal lamina. This is a unique feature of the protocol as other techniques such as Fluorescence Activated Cell Sorting require chemical and mechanical tissue dissociation 7. Although the myofiber culture system cannot substitute for in vivo studies, it does offer an excellent platform to address relevant biological properties of muscle stem cells. Single myofibers can be cultured in standard plating conditions or in floating conditions. Satellite cells on floating myofibers are subjected to virtually no other influence than the myofiber environment. Substrate stiffness and coating have been shown to influence satellite cells' ability to regenerate muscles 8, 9 so being able to control each of these factors independently allows discrimination between niche-dependent and -independent responses. Different concentrations of serum have also been shown to have an effect on the transition from quiescence to activation. To preserve the quiescence state of

  7. Peroxisome Proliferator Activated Receptor Beta (PPARβ) activity increases the immune response and shortens the early phases of skeletal muscle regeneration.

    PubMed

    Mothe-Satney, Isabelle; Piquet, Jessica; Murdaca, Joseph; Sibille, Brigitte; Grimaldi, Paul A; Neels, Jaap G; Rousseau, Anne-Sophie

    2016-12-07

    Peroxisome Proliferator-Activated Receptor Beta (PPARβ) is a transcription factor playing an important role in both muscle myogenesis and remodeling, and in inflammation. However, its role in the coordination of the transient muscle inflammation and reparation process following muscle injury has not yet been fully determined. We postulated that activation of the PPARβ pathway alters the early phase of the muscle regeneration process, i.e. when immune cells infiltrate in injured muscle. Tibialis anteriors of C57BL6/J mice treated or not with the PPARβ agonist GW0742 were injected with cardiotoxin (or with physiological serum for the contralateral muscle). Muscle regeneration was monitored on days 4, 7, and 14 post-injury. We found that treatment of mice with GW0742 increased, at day 4 post-damage, the recruitment of immune cells (M1 and M2 macrophages) and upregulated the expression of the anti-inflammatory cytokine IL-10 and TGF-β mRNA. Those effects were accompanied by a significant increase at day 4 of myogenic regulatory factors (Pax7, MyoD, Myf5, Myogenin) mRNA in GW0742-treated mice. However, we showed an earlier return (7 days vs 14 days) of Myf5 and Myogenin to basal levels in GW0742- compared to DMSO-treated mice. Differential effects of GW0742 observed during the regeneration were associated with variations of PPARβ pathway activity. Collectively, our findings indicate that PPARβ pathway activity shortens the early phases of skeletal muscle regeneration by increasing the immune response.

  8. Modulation of Stem Cell Differentiation and Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and Regeneration after Injury

    DTIC Science & Technology

    2011-03-01

    07-1-0181 TITLE: Modulation of Stem Cell Differentiation and Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and...views, opinions and/or findings contained in this report are those of the author(s) and should not be construed as an official Department of the Army...Myostatin as an Approach to Counteract Fibrosis in Muscle Dystrophy and Regeneration after Injury Dr. Nestor Gonzalez-Cadavid Charles R. Drew

  9. Stem cells migration during skeletal muscle regeneration - the role of Sdf-1/Cxcr4 and Sdf-1/Cxcr7 axis.

    PubMed

    Kowalski, Kamil; Kołodziejczyk, Aleksandra; Sikorska, Maria; Płaczkiewicz, Jagoda; Cichosz, Paulina; Kowalewska, Magdalena; Stremińska, Władysława; Jańczyk-Ilach, Katarzyna; Koblowska, Marta; Fogtman, Anna; Iwanicka-Nowicka, Roksana; Ciemerych, Maria A; Brzoska, Edyta

    2016-10-13

    The skeletal muscle regeneration occurs due to the presence of tissue specific stem cells - satellite cells. These cells, localized between sarcolemma and basal lamina, are bound to muscle fibers and remain quiescent until their activation upon muscle injury. Due to pathological conditions, such as extensive injury or dystrophy, skeletal muscle regeneration is diminished. Among the therapies aiming to ameliorate skeletal muscle diseases are transplantations of the stem cells. In our previous studies we showed that Sdf-1 (stromal derived factor -1) increased migration of stem cells and their fusion with myoblasts in vitro. Importantly, we identified that Sdf-1 caused an increase in the expression of tetraspanin CD9 - adhesion protein involved in myoblasts fusion. In the current study we aimed to uncover the details of molecular mechanism of Sdf-1 action. We focused at the Sdf-1 receptors - Cxcr4 and Cxcr7, as well as signaling pathways induced by these molecules in primary myoblasts, as well as various stem cells - mesenchymal stem cells and embryonic stem cells, i.e. the cells of different migration and myogenic potential. We showed that Sdf-1 altered actin organization via FAK (focal adhesion kinase), Cdc42 (cell division control protein 42), and Rac-1 (Ras-Related C3 Botulinum Toxin Substrate 1). Moreover, we showed that Sdf-1 modified the transcription profile of genes encoding factors engaged in cells adhesion and migration. As the result, cells such as primary myoblasts or embryonic stem cells, became characterized by more effective migration when transplanted into regenerating muscle.

  10. Liver regeneration after living donor transplantation: adult-to-adult living donor liver transplantation cohort study.

    PubMed

    Olthoff, Kim M; Emond, Jean C; Shearon, Tempie H; Everson, Greg; Baker, Talia B; Fisher, Robert A; Freise, Chris E; Gillespie, Brenda W; Everhart, James E

    2015-01-01

    Adult-to-adult living donors and recipients were studied to characterize patterns of liver growth and identify associated factors in a multicenter study. Three hundred and fifty donors and 353 recipients in the Adult-to-Adult Living Donor Liver Transplantation Cohort Study (A2ALL) receiving transplants between March 2003 and February 2010 were included. Potential predictors of 3-month liver volume included total and standard liver volumes (TLV and SLV), Model for End-Stage Liver Disease (MELD) score (in recipients), the remnant and graft size, remnant-to-donor and graft-to-recipient weight ratios (RDWR and GRWR), remnant/TLV, and graft/SLV. Among donors, 3-month absolute growth was 676 ± 251 g (mean ± SD), and percentage reconstitution was 80% ± 13%. Among recipients, GRWR was 1.3% ± 0.4% (8 < 0.8%). Graft weight was 60% ± 13% of SLV. Three-month absolute growth was 549 ± 267 g, and percentage reconstitution was 93% ± 18%. Predictors of greater 3-month liver volume included larger patient size (donors and recipients), larger graft volume (recipients), and larger TLV (donors). Donors with the smallest remnant/TLV ratios had larger than expected growth but also had higher postoperative bilirubin and international normalized ratio at 7 and 30 days. In a combined donor-recipient analysis, donors had smaller 3-month liver volumes than recipients adjusted for patient size, remnant or graft volume, and TLV or SLV (P = 0.004). Recipient graft failure in the first 90 days was predicted by poor graft function at day 7 (HR = 4.50, P = 0.001) but not by GRWR or graft fraction (P > 0.90 for each). Both donors and recipients had rapid yet incomplete restoration of tissue mass in the first 3 months, and this confirmed previous reports. Recipients achieved a greater percentage of expected total volume. Patient size and recipient graft volume significantly influenced 3-month volumes. Importantly, donor liver volume is a

  11. The immunophilin ligand FK506, but not the P38 kinase inhibitor SB203580, improves function of adult rat muscle reinnervated from transplants of embryonic neurons.

    PubMed

    Grumbles, R M; Casella, G T B; Rudinsky, M J; Godfrey, S; Wood, P M; Thomas, C K

    2005-01-01

    Injury to the adult CNS often involves death of motoneurons, resulting in the paralysis and progressive atrophy of muscle. There is no effective therapy to replace motoneurons in the CNS. Our strategy to replace neurons and to rescue denervated muscles is to transplant dissociated embryonic day 14-15 (E14-15) ventral spinal cord cells into the distal stump of a peripheral nerve near the denervated muscles. Here, we test whether long-term delivery of two pharmacological inhibitors to denervated muscle, FK506 or SB203580, enhances reinnervation of muscle from embryonic cells transplanted in the tibial nerve of adult Fischer rats. FK506, SB203580 (2.5 mg/kg) or saline was delivered under the fascia of the medial gastrocnemius muscle for 4 weeks, beginning when muscles were denervated by section of the sciatic nerve. After 1 week of nerve degeneration, one million E14-15 ventral spinal cord cells were transplanted into the distal tibial nerve stump of each rat in the three treatment groups. Ten weeks later, all cell transplants had neuron-specific nuclear protein (NeuN) positive neurons. Neuron survival and axon regeneration were similar across treatments. An average (+/-S.E.) of 210+/-66, 100+/-36 and 176+/-58 myelinated axons grew distally from the cell transplants of rats with muscles treated with FK506, SB203580 or saline, respectively. Regenerating axons in muscles of all three treatments groups were detected with antibodies against phosphorylated neurofilaments and synaptophysin, and motor end plates were labeled with alpha-bungarotoxin. Muscles of rats that received transplants of media only had no axon growth, indicating that the muscles were denervated. The mean muscle fiber areas of rats that received cell transplants and had long-term delivery of FK506, SB203580 or saline to muscles were significantly larger than those of denervated muscle fibers. Thus, cell transplantation reduced muscle atrophy. Transplantation of embryonic cells also resulted in

  12. Child—Adult Differences in Muscle Activation — A Review

    PubMed Central

    Dotan, Raffy; Mitchell, Cameron; Cohen, Rotem; Klentrou, Panagiota; Gabriel, David; Falk, Bareket

    2013-01-01

    Children differ from adults in many muscular performance attributes such as size-normalized strength and power, endurance, fatigability and the recovery from exhaustive exercise, to name just a few. Metabolic attributes, such as glycolytic capacity, substrate utilization, and VO2 kinetics also differ markedly between children and adults. Various factors, such as dimensionality, intramuscular synchronization, agonist-antagonist coactivation, level of volitional activation, or muscle composition, can explain some, but not all of the observed differences. It is hypothesized that, compared with adults, children are substantially less capable of recruiting or fully employing their higher-threshold, type-II motor units. The review presents and evaluates the wealth of information and possible alternative factors in explaining the observations. Although conclusive evidence is still lacking, only this hypothesis of differential motor-unit activation in children and adults, appears capable of accounting for all observed child—adult differences, whether on its own or in conjunction with other factors. PMID:22433260

  13. Regulation of Müller Glial Dependent Neuronal Regeneration in the Damaged Adult Zebrafish Retina

    PubMed Central

    Gorsuch, Ryne A.; Hyde, David R.

    2013-01-01

    This article examines our current knowledge underlying the mechanisms involved in neuronal regeneration in the adult zebrafish retina. Zebrafish, which has the capacity to regenerate a wide variety of tissues and organs (including the fins, kidney, heart, brain, and spinal cord), has become the premier model system to study retinal regeneration due to the robustness and speed of the response and the variety of genetic tools that can be applied to study this question. It is now well documented that retinal damage induces the resident Müller glia to dedifferentiate and reenter the cell cycle to produce neuronal progenitor cells that continue to proliferate, migrate to the damaged retinal layer and differentiate into the missing neuronal cell types. Increasing our understanding of how these cellular events are regulated and occur in response to neuronal damage may provide critical information that can be applied to stimulating a regeneration response in the mammalian retina. In this review, we will focus on the genes/proteins that regulate zebrafish retinal regeneration and will attempt to critically evaluate how these factors may interact to correctly orchestrate the definitive cellular events that occur during regeneration. PMID:23880528

  14. Regulation of Müller glial dependent neuronal regeneration in the damaged adult zebrafish retina.

    PubMed

    Gorsuch, Ryne A; Hyde, David R

    2014-06-01

    This article examines our current knowledge underlying the mechanisms involved in neuronal regeneration in the adult zebrafish retina. Zebrafish, which has the capacity to regenerate a wide variety of tissues and organs (including the fins, kidney, heart, brain, and spinal cord), has become the premier model system to study retinal regeneration due to the robustness and speed of the response and the variety of genetic tools that can be applied to study this question. It is now well documented that retinal damage induces the resident Müller glia to dedifferentiate and reenter the cell cycle to produce neuronal progenitor cells that continue to proliferate, migrate to the damaged retinal layer and differentiate into the missing neuronal cell types. Increasing our understanding of how these cellular events are regulated and occur in response to neuronal damage may provide critical information that can be applied to stimulating a regeneration response in the mammalian retina. In this review, we will focus on the genes/proteins that regulate zebrafish retinal regeneration and will attempt to critically evaluate how these factors may interact to correctly orchestrate the definitive cellular events that occur during regeneration.

  15. Adult axolotls can regenerate original neuronal diversity in response to brain injury

    PubMed Central

    Amamoto, Ryoji; Huerta, Violeta Gisselle Lopez; Takahashi, Emi; Dai, Guangping; Grant, Aaron K; Fu, Zhanyan; Arlotta, Paola

    2016-01-01

    The axolotl can regenerate multiple organs, including the brain. It remains, however, unclear whether neuronal diversity, intricate tissue architecture, and axonal connectivity can be regenerated; yet, this is critical for recovery of function and a central aim of cell replacement strategies in the mammalian central nervous system. Here, we demonstrate that, upon mechanical injury to the adult pallium, axolotls can regenerate several of the populations of neurons present before injury. Notably, regenerated neurons acquire functional electrophysiological traits and respond appropriately to afferent inputs. Despite the ability to regenerate specific, molecularly-defined neuronal subtypes, we also uncovered previously unappreciated limitations by showing that newborn neurons organize within altered tissue architecture and fail to re-establish the long-distance axonal tracts and circuit physiology present before injury. The data provide a direct demonstration that diverse, electrophysiologically functional neurons can be regenerated in axolotls, but challenge prior assumptions of functional brain repair in regenerative species. DOI: http://dx.doi.org/10.7554/eLife.13998.001 PMID:27156560

  16. Peripheral Axons of the Adult Zebrafish Maxillary Barbel Extensively Remyelinate During Sensory Appendage Regeneration

    PubMed Central

    Moore, Alex C.; Mark, Tiffany E.; Hogan, Ann K.; Topczewski, Jacek; LeClair, Elizabeth E.

    2013-01-01

    Myelination is a cellular adaptation allowing rapid conduction along axons. We have investigated peripheral axons of the zebrafish maxillary barbel (ZMB), an optically clear sensory appendage. Each barbel carries taste buds, solitary chemosensory cells, and epithelial nerve endings, all of which regenerate after amputation (LeClair and Topczewski [2010] PLoS One 5:e8737). The ZMB contains axons from the facial nerve; however, myelination within the barbel itself has not been established. Transcripts of myelin basic protein (mbp) are expressed in normal and regenerating adult barbels, indicating activity in both maintenance and repair. Myelin was confirmed in situ by using toluidine blue, an anti-MBP antibody, and transmission electron microscopy (TEM). The adult ZMB contains ~180 small-diameter axons (<2 μm), approximately 60% of which are myelinated. Developmental myelination was observed via whole-mount immunohistochemistry 4-6 weeks postfertilization, showing myelin sheaths lagging behind growing axons. Early-regenerating axons (10 days postsurgery), having no or few myelin layers, were disorganized within a fibroblast-rich collagenous scar. Twenty-eight days postsurgery, barbel axons had grown out several millimeters and were organized with compact myelin sheaths. Fiber types and axon areas were similar between normal and regenerated tissue; within 4 weeks, regenerating axons restored ~85% of normal myelin thickness. Regenerating barbels express multiple promyelinating transcription factors (sox10, oct6 = pou3f1; krox20a/b = egr2a/b) typical of Schwann cells. These observations extend our understanding of the zebrafish peripheral nervous system within a little-studied sensory appendage. The accessible ZMB provides a novel context for studying axon regeneration, Schwann cell migration, and remyelination in a model vertebrate. PMID:22592645

  17. Acupuncture plus Low-Frequency Electrical Stimulation (Acu-LFES) Attenuates Diabetic Myopathy by Enhancing Muscle Regeneration.

    PubMed

    Su, Zhen; Robinson, Alayna; Hu, Li; Klein, Janet D; Hassounah, Faten; Li, Min; Wang, Haidong; Cai, Hui; Wang, Xiaonan H

    2015-01-01

    Mortality and morbidity are increased in patients with muscle atrophy resulting from catabolic diseases such as diabetes. At present there is no pharmacological treatment that successfully reverses muscle wasting from catabolic conditions. We hypothesized that acupuncture plus low frequency electric stimulation (Acu-LFES) would mimic the impact of exercise and prevent diabetes-induced muscle loss. Streptozotocin (STZ) was used to induce diabetes in mice. The mice were then treated with Acu-LFES for 15 minutes daily for 14 days. Acupuncture points were selected according to the WHO Standard Acupuncture Nomenclature guide. The needles were connected to an SDZ-II electronic acupuncture device delivering pulses at 20Hz and 1mA. Acu-LFES prevented soleus and EDL muscle weight loss and increased hind-limb muscle grip function in diabetic mice. Muscle regeneration capacity was significantly increased by Acu-LFES. The expression of Pax7, MyoD, myogenin and embryo myosin heavy chain (eMyHC) was significantly decreased in diabetic muscle vs. control muscle. The suppressed levels in diabetic muscle were reversed by Acu-LFES. The IGF-1 signaling pathway was also upregulated by Acu-LFES. Phosphorylation of Akt, mTOR and p70S6K were downregulated by diabetes leading to a decline in muscle mass, however, Acu-LFES countered the diabetes-induced decline. In addition, microRNA-1 and -206 were increased by Acu-LFES after 24 days of treatment. We conclude that Acu-LFES is effective in counteracting diabetes-induced skeletal muscle atrophy by increasing IGF-1 and its stimulation of muscle regeneration.

  18. The cardiac stem cell compartment is indispensable for myocardial cell homeostasis, repair and regeneration in the adult.

    PubMed

    Nadal-Ginard, Bernardo; Ellison, Georgina M; Torella, Daniele

    2014-11-01

    Resident cardiac stem cells in embryonic, neonatal and adult mammalian heart have been identified by different membrane markers and transcription factors. However, despite a flurry of publications no consensus has been reached on the identity and actual regenerative effects of the adult cardiac stem cells. Intensive research on the adult mammalian heart's capacity for self-renewal of its muscle cell mass has led to a consensus that new cardiomyocytes (CMs) are indeed formed throughout adult mammalian life albeit at a disputed frequency. The physiological significance of this renewal, the origin of the new CMs, and the rate of adult CM turnover are still highly debated. Myocyte replacement, particularly after injury, was originally attributed to differentiation of a stem cell compartment. More recently, it has been reported that CMs are mainly replaced by the division of pre-existing post-mitotic CMs. These latter results, if confirmed, would shift the target of regenerative therapy toward boosting mature CM cell-cycle re-entry. Despite this controversy, it is documented that the adult endogenous c-kit(pos) cardiac stem cells (c-kit(pos) eCSCs) participate in adaptations to myocardial stress, and, when transplanted into the myocardium, regenerate most cardiomyocytes and microvasculature lost in an infarct. Nevertheless, the in situ myogenic potential of adult c-kit(pos) cardiac cells has been questioned. To revisit the regenerative potential of c-kit(pos) eCSCs, we have recently employed experimental protocols of severe diffuse myocardial damage in combination with several genetic murine models and cell transplantation approaches showing that eCSCs are necessary and sufficient for CM regeneration, leading to complete cellular, anatomical, and functional myocardial recovery. Here we will review the available data on adult eCSC biology and their regenerative potential placing it in the context of the different claimed mechanisms of CM replacement. These data are in

  19. Novel Adult Stem Cells for Peripheral Nerve Regeneration

    DTIC Science & Technology

    2013-09-01

    Symposium on Vascular Tissue Engineering. Holland, 5/2013 o The Joint British Society for Cardiovascular Research/British Atherosclerosis Society meeting... atherosclerosis , involves proliferation and migration of vascular cells1–4. Vascular smooth muscle cells (SMCs) are the predominant cell type in the tunica media...osteogenic cells in diseased vessels remains to be tested using atherosclerosis models. The identification of MVSC in human arteries makes MVSC a

  20. Muscle power failure in mobility-limited adults: preserved single muscle fibre function despite reduced whole muscle size, quality and neuromuscular activiation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study investigated the physiological and gender determinants of the age-related loss of muscle power in 31 healthy middle-aged adults (aged 40-55 years), 28 healthy older adults (70-85 years) and 34 mobility-limited older adults (70-85 years). We hypothesized that leg extensor muscle power woul...

  1. Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps

    PubMed Central

    Broughton, Kathleen; Sussman, Mark A.

    2016-01-01

    Much has changed since our survey of the landscape for myocardial regeneration powered by adult stem cells four years ago (Mohsin et al., Empowering adult stem cells for myocardial regeneration. Circ Res. 2011; 109(12):1415–1428) [1]. The intervening years since that first review has witnessed an explosive expansion of studies that advance both understanding and implementation of adult stem cells in promoting myocardial repair. Painstaking research from innumerable laboratories throughout the world is prying open doors that may lead to restoration of myocardial structure and function in the wake of pathologic injury. This global effort has produced deeper mechanistic comprehension coupled with an evolving appreciation for the complexity of myocardial regeneration in the adult context. Undaunted by both known and (as yet) unknown challenges, pursuit of myocardial regenerative medicine mediated by adult stem cell therapy has gathered momentum fueled by tantalizing clues and visionary goals. This concise review takes a somewhat different perspective than our initial treatise, taking stock of the business sector that has become an integral part of the field while concurrently updating “state of affairs” in cutting edge research. Looking retrospectively at advancement over the years as all reviews eventually must, the fundamental lesson to be learned is best explained by Jonatan Mårtensson: “Success will never be a big step in the future. Success is a small step taken just now.” PMID:26941423

  2. The hypoxia-inducible factors HIF1α and HIF2α are dispensable for embryonic muscle development but essential for postnatal muscle regeneration.

    PubMed

    Yang, Xin; Yang, Shiqi; Wang, Chao; Kuang, Shihuan

    2017-04-07

    Muscle satellite cells are myogenic stem cells whose quiescence, activation, self-renewal, and differentiation are influenced by oxygen supply, an environmental regulator of stem cell activity. Accordingly, stem cell-specific oxygen signaling pathways precisely control the balance between muscle growth and regeneration in response to oxygen fluctuations, and hypoxia-inducible factors (HIFs) are central mediators of these cellular responses. However, the in vivo roles of HIFs in quiescent satellite cells and activated satellite cells (myoblasts) are poorly understood. Using transgenic mouse models for cell-specific HIF expression, we show here that HIF1α and HIF2α are preferentially expressed in pre- and post-differentiation myoblasts, respectively. Interestingly, double knockouts of HIF1α and HIF2α (HIF1α/2α dKO) generated with the MyoD(Cre) system in embryonic myoblasts resulted in apparently normal muscle development and growth. However, HIF1α/2α dKO produced with the tamoxifen-inducible, satellite cell-specific Pax7(CreER) system in postnatal satellite cells delayed injury-induced muscle repair due to a reduced number of myoblasts during regeneration. Analysis of satellite cell dynamics on myofibers confirmed that HIF1α/2α dKO myoblasts exhibit reduced self-renewal but more pronounced differentiation under hypoxic conditions. Mechanistically, the HIF1α/2α dKO blunted hypoxia-induced activation of Notch signaling, a key determinant of satellite cell self-renewal. We conclude that HIF1α and HIF2α are dispensable for muscle stem cell function under normoxia but are required for maintaining satellite cell self-renewal in hypoxic environments. Our insights into a critical mechanism in satellite cell homeostasis during muscle regeneration could help inform research efforts to treat muscle diseases or improve muscle function.

  3. The neonate versus adult mammalian immune system in cardiac repair and regeneration.

    PubMed

    Sattler, Susanne; Rosenthal, Nadia

    2016-07-01

    The immune system is a crucial player in tissue homeostasis and wound healing. A sophisticated cascade of events triggered upon injury ensures protection from infection and initiates and orchestrates healing. While the neonatal mammal can readily regenerate damaged tissues, adult regenerative capacity is limited to specific tissue types, and in organs such as the heart, adult wound healing results in fibrotic repair and loss of function. Growing evidence suggests that the immune system greatly influences the balance between regeneration and fibrotic repair. The neonate mammalian immune system has impaired pro-inflammatory function, is prone to T-helper type 2 responses and has an immature adaptive immune system skewed towards regulatory T cells. While these characteristics make infants susceptible to infection and prone to allergies, it may also provide an immunological environment permissive of regeneration. In this review we will give a comprehensive overview of the immune cells involved in healing and regeneration of the heart and explore differences between the adult and neonate immune system that may explain differences in regenerative ability. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.

  4. Continued Expression of Neonatal Myosin Heavy Chain in Adult Dystrophic Skeletal Muscle

    NASA Astrophysics Data System (ADS)

    Bandman, Everett

    1985-02-01

    The expression of myosin heavy chain isoforms was examined in normal and dystrophic chicken muscle with a monoclonal antibody specific for neonatal myosin. Adult dystrophic muscle continued to contain neonatal myosin long after it disappeared from adult normal muscle. A new technique involving western blotting and peptide mapping demonstrated that the immunoreactive myosin in adult dystrophic muscle was identical to that found in neonatal normal muscle. Immunocytochemistry revealed that all fibers in the dystrophic muscle failed to repress neonatal myosin heavy chain. These studies suggest that muscular dystrophy inhibits the myosin gene switching that normally occurs during muscle maturation.

  5. Identification of cDNAs associated with late dedifferentiation in adult newt forelimb regeneration.

    PubMed

    Vascotto, Sandy G; Beug, Shawn; Liversage, Richard A; Tsilfidis, Catherine

    2005-06-01

    Epimorphic limb regeneration in the adult newt involves the dedifferentiation of differentiated cells to yield a pluripotent blastemal cell. These mesenchymal-like cells proliferate and subsequently respond to patterning and differentiation cues to form a new limb. Understanding the dedifferentiation process requires the selective identification of dedifferentiating cells within the heterogeneous population of cells in the regenerate. In this study, representational differences analysis was used to produce an enriched population of dedifferentiation-associated cDNA fragments. Fifty-nine unique cDNA fragments were identified, sequenced, and analyzed using bioinformatics tools and databases. Some of these clones demonstrate significant similarity to known genes in other species. Other clones can be linked by homology to pathways previously implicated in the dedifferentiation process. These data will form the basis for further analyses to elucidate the role of candidate genes in the dedifferentiation process during newt forelimb regeneration.

  6. Dopamine from the brain promotes spinal motor neuron generation during development and adult regeneration.

    PubMed

    Reimer, Michell M; Norris, Anneliese; Ohnmacht, Jochen; Patani, Rickie; Zhong, Zhen; Dias, Tatyana B; Kuscha, Veronika; Scott, Angela L; Chen, Yu-Chia; Rozov, Stanislav; Frazer, Sarah L; Wyatt, Cameron; Higashijima, Shin-ichi; Patton, E Elizabeth; Panula, Pertti; Chandran, Siddharthan; Becker, Thomas; Becker, Catherina G

    2013-06-10

    Coordinated development of brain stem and spinal target neurons is pivotal for the emergence of a precisely functioning locomotor system. Signals that match the development of these far-apart regions of the central nervous system may be redeployed during spinal cord regeneration. Here we show that descending dopaminergic projections from the brain promote motor neuron generation at the expense of V2 interneurons in the developing zebrafish spinal cord by activating the D4a receptor, which acts on the hedgehog pathway. Inhibiting this essential signal during early neurogenesis leads to a long-lasting reduction of motor neuron numbers and impaired motor responses of free-swimming larvae. Importantly, during successful spinal cord regeneration in adult zebrafish, endogenous dopamine promotes generation of spinal motor neurons, and dopamine agonists augment this process. Hence, we describe a supraspinal control mechanism for the development and regeneration of specific spinal cell types that uses dopamine as a signal.

  7. A new type of Schwann cell graft transplantation to promote optic nerve regeneration in adult rats.

    PubMed

    Fang, Yuan; Mo, Xiaofen; Guo, Wenyi; Zhang, Meng; Zhang, Peihua; Wang, Yan; Rong, Xianfang; Tian, Jie; Sun, Xinghuai

    2010-12-01

    Like other parts of the central nervous system, the adult mammalian optic nerve is difficult to regenerate after injury. Transplantation of the peripheral nerve or a Schwann cell (SC) graft can promote injured axonal regrowth. We tried to develop a new type of tissue-engineered SC graft that consisted of SCs seeded onto a poly(lactic-co-glycolic acid)/chitosan conduit. Meanwhile, SCs were transfected along the ciliary neurotrophic factor (CNTF) gene in vitro by electroporation to increase their neurotrophic effect. Four weeks after transplantation, GAP-43 labelled regenerating axons were found in the SC grafts, and axons in the CNTF-SC graft were longer than those in the SC graft. Tissue-engineered SC grafts can provide a feasible environment for optic nerve regeneration and may become an alternative for bridging damaged nerves and repairing nerve defects in the future.

  8. Stem Cell Differentiation Toward the Myogenic Lineage for Muscle Tissue Regeneration: A Focus on Muscular Dystrophy.

    PubMed

    Ostrovidov, Serge; Shi, Xuetao; Sadeghian, Ramin Banan; Salehi, Sahar; Fujie, Toshinori; Bae, Hojae; Ramalingam, Murugan; Khademhosseini, Ali

    2015-12-01

    Skeletal muscle tissue engineering is one of the important ways for regenerating functionally defective muscles. Among the myopathies, the Duchenne muscular dystrophy (DMD) is a progressive disease due to mutations of the dystrophin gene leading to progressive myofiber degeneration with severe symptoms. Although current therapies in muscular dystrophy are still very challenging, important progress has been made in materials science and in cellular technologies with the use of stem cells. It is therefore useful to review these advances and the results obtained in a clinical point of view. This article focuses on the differentiation of stem cells into myoblasts, and their application in muscular dystrophy. After an overview of the different stem cells that can be induced to differentiate into the myogenic lineage, we introduce scaffolding materials used for muscular tissue engineering. We then described some widely used methods to differentiate different types of stem cell into myoblasts. We highlight recent insights obtained in therapies for muscular dystrophy. Finally, we conclude with a discussion on stem cell technology. We discussed in parallel the benefits brought by the evolution of the materials and by the expansion of cell sources which can differentiate into myoblasts. We also discussed on future challenges for clinical applications and how to accelerate the translation from the research to the clinic in the frame of DMD.

  9. A novel approach to collecting satellite cells from adult skeletal muscles on the basis of their stress tolerance.

    PubMed

    Shigemoto, Taeko; Kuroda, Yasumasa; Wakao, Shohei; Dezawa, Mari

    2013-07-01

    Stem cells are generally collected using flow cytometry, but this method is not applicable when the cell surface marker is not well determined. Satellite cells, which are skeletal muscle stem cells, have the ability to regenerate damaged muscles and are expected to be applicable for treatment of muscle degeneration. Although the transcription factor Pax7 is a known specific marker of satellite cells, it is not located on the cell surface and therefore flow cytometry is not directly applicable. In the present study, we turned our attention to the stress tolerance of adult stem cells, and we propose long-term trypsin incubation (LTT) as a novel approach to collecting satellite cells from mouse and human skeletal muscles. LTT led to a remarkable increase in the ratio of Pax7(+) cells that retain normal myogenic stem cell function. In particular, human Pax7(+) cells made up approximately 30% of primary cultured cells, whereas after LTT, the ratio of Pax7(+) cells increased up to ∼80%, and the ratio of Pax7(+) and/or MyoD(+) myogenic cells increased to ∼95%. Once transplanted, LTT-treated cells contributed to subsequent muscle regeneration following repetitive muscle damage without additional cell transplantation. The stress tolerance of Pax7(+) cells is related to heat shock protein 27 and αB-crystallin, members of the small heat shock protein family. This approach, based on the stress resistance of adult stem cells, is a safe and inexpensive method of efficiently collecting human satellite cells and may also be used for collecting other tissue stem cells whose surface marker is unknown.

  10. From ontogenesis to regeneration: learning how to instruct adult cardiac progenitor cells.

    PubMed

    Chimenti, Isotta; Forte, Elvira; Angelini, Francesco; Giacomello, Alessandro; Messina, Elisa

    2012-01-01

    Since the first observations over two centuries ago by Lazzaro Spallanzani on the extraordinary regenerative capacity of urodeles, many attempts have been made to understand the reasons why such ability has been largely lost in metazoa and whether or how it can be restored, even partially. In this context, important clues can be derived from the systematic analysis of the relevant distinctions among species and of the pathways involved in embryonic development, which might be induced and/or recapitulated in adult tissues. This chapter provides an overview on regeneration and its mechanisms, starting with the lesson learned from lower vertebrates, and will then focus on recent advancements and novel insights concerning regeneration in the adult mammalian heart, including the discovery of resident cardiac progenitor cells (CPCs). Subsequently, it explores all the important pathways involved in regulating differentiation during development and embryogenesis, and that might potentially provide important clues on how to activate and/or modulate regenerative processes in the adult myocardium, including the potential activation of endogenous CPCs. Furthermore the importance of the stem cell niche is discussed, and how it is possible to create in vitro a microenvironment and culture system to provide adult CPCs with the ideal conditions promoting their regenerative ability. Finally, the state of clinical translation of cardiac cell therapy is presented. Overall, this chapter provides a new perspective on how to approach cardiac regeneration, taking advantage of important lessons from development and optimizing biotechnological tools to obtain the ideal conditions for cell-based cardiac regenerative therapy.

  11. Comparative analysis of mesenchymal stem cells from adult mouse adipose, muscle, and fetal muscle.

    PubMed

    Lei, Hulong; Yu, Bing; Huang, Zhiqing; Yang, Xuerong; Liu, Zehui; Mao, Xiangbing; Tian, Gang; He, Jun; Han, Guoquan; Chen, Hong; Mao, Qian; Chen, Daiwen

    2013-02-01

    Recently, increasing evidence supports that adult stem cells are the part of a natural system for tissue growth and repair. This study focused on the differences of mesenchymal stem cells from adult adipose (ADSCs), skeletal muscle (MDSCs) and fetal muscle (FMSCs) in biological characteristics, which is the key to cell therapy success. Stem cell antigen 1 (Sca-1) expression of MDSCs and FMSCs at passage 3 was two times more than that at passage 1 (P < 0.0001). After 28-day myogenic induction, higher expression levels of skeletal muscle-specific genes were observed in MDSCs than FMSCs (P < 0.01), and the lowest expression levels were demonstrated in ADSCs among three cells (P < 0.01). Besides, M-Cad and MyHC expressions in ADSCs were not detected by immunofluorescence or real-time quantitative PCR. Furthermore, after 14 days adipogenic induction, PPARγ2, LPL and aP2 mRNA expressions were higher in ADSCs vs. MDSCs (P < 0.01). Besides, MSCs from adult or fetal muscle expressed higher OCN and OPN than ADSCs after 28 days osteogenic induction (P < 0.01). Taken together, our results suggested that cell source and developmental stage had great impacts on biological properties of mesenchymal stem cells, and proper consideration of all the issues is necessary.

  12. In vivo electroporation of morpholinos into the regenerating adult zebrafish tail fin.

    PubMed

    Hyde, David R; Godwin, Alan R; Thummel, Ryan

    2012-03-29

    Certain species of urodeles and teleost fish can regenerate their tissues. Zebrafish have become a widely used model to study the spontaneous regeneration of adult tissues, such as the heart, retina, spinal cord, optic nerve, sensory hair cells, and fins. The zebrafish fin is a relatively simple appendage that is easily manipulated to study multiple stages in epimorphic regeneration. Classically, fin regeneration was characterized by three distinct stages: wound healing, blastema formation, and fin outgrowth. After amputating part of the fin, the surrounding epithelium proliferates and migrates over the wound. At 33 °C, this process occurs within six hours post-amputation (hpa, Figure 1B). Next, underlying cells from different lineages (ex. bone, blood, glia, fibroblast) re-enter the cell cycle to form a proliferative blastema, while the overlying epidermis continues to proliferate (Figure 1D). Outgrowth occurs as cells proximal to the blastema re-differentiate into their respective lineages to form new tissue (Figure 1E). Depending on the level of the amputation, full regeneration is completed in a week to a month. The expression of a large number of gene families, including wnt, hox, fgf, msx, retinoic acid, shh, notch, bmp, and activin-betaA genes, is up-regulated during specific stages of fin regeneration. However, the roles of these genes and their encoded proteins during regeneration have been difficult to assess, unless a specific inhibitor for the protein exists, a temperature-sensitive mutant exists or a transgenic animal (either overexpressing the wild-type protein or a dominant-negative protein) was generated. We developed a reverse genetic technique to quickly and easily test the function of any gene during fin regeneration. Morpholino oligonucleotides are widely used to study loss of specific proteins during zebrafish, Xenopus, chick, and mouse development. Morpholinos basepair with a complementary RNA sequence to either block pre-mRNA splicing or m

  13. Synthesis and Characterization of a Model Extracellular Matrix that Induces Partial Regeneration of Adult Mammalian Skin

    NASA Astrophysics Data System (ADS)

    Yannas, I. V.; Lee, E.; Orgill, D. P.; Skrabut, E. M.; Murphy, G. F.

    1989-02-01

    Regeneration of the dermis does not occur spontaneously in the adult mammal. The epidermis is regenerated spontaneously provided there is a dermal substrate over which it can migrate. Certain highly porous, crosslinked collagen--glycosaminoglycan copolymers have induced partial morphogenesis of skin when seeded with dermal and epidermal cells and then grafted on standard, full-thickness skin wounds in the adult guinea pig. A mature epidermis and a nearly physiological dermis, which lacked hair follicles but was demonstrably different from scar, were regenerated over areas as large as 16 cm2. These chemical analogs of extracellular matrices were morphogenetically active provided that the average pore diameter ranged between 20 and 125 μ m, the resistance to degradation by collagenase exceeded a critical limit, and the density of autologous dermal and epidermal cells inoculated therein was >5 × 104 cells per cm2 of wound area. Unseeded copolymers with physical structures that were within these limits delayed the onset of wound contraction by about 10 days but did not eventually prevent it. Seeded copolymers not only delayed contraction but eventually arrested and reversed it while new skin was being regenerated. The data identify a model extracellular matrix that acts as if it were an insoluble growth factor with narrowly specified physicochemical structure, functioning as a transient basal lamina during morphogenesis of skin.

  14. Synthesis and characterization of a model extracellular matrix that induces partial regeneration of adult mammalian skin.

    PubMed Central

    Yannas, I V; Lee, E; Orgill, D P; Skrabut, E M; Murphy, G F

    1989-01-01

    Regeneration of the dermis does not occur spontaneously in the adult mammal. The epidermis is regenerated spontaneously provided there is a dermal substrate over which it can migrate. Certain highly porous, crosslinked collagen-glycosaminoglycan copolymers have induced partial morphogenesis of skin when seeded with dermal and epidermal cells and then grafted on standard, full-thickness skin wounds in the adult guinea pig. A mature epidermis and a nearly physiological dermis, which lacked hair follicles but was demonstrably different from scar, were regenerated over areas as large as 16 cm2. These chemical analogs of extracellular matrices were morphogenetically active provided that the average pore diameter ranged between 20 and 125 microns, the resistance to degradation by collagenase exceeded a critical limit, and the density of autologous dermal and epidermal cells inoculated therein was greater than 5 x 10(4) cells per cm2 of wound area. Unseeded copolymers with physical structures that were within these limits delayed the onset of wound contraction by about 10 days but did not eventually prevent it. Seeded copolymers not only delayed contraction but eventually arrested and reversed it while new skin was being regenerated. The data identify a model extracellular matrix that acts as if it were an insoluble growth factor with narrowly specified physiochemical structure, functioning as a transient basal lamina during morphogenesis of skin. Images PMID:2915988

  15. Imaging Beta Cell Regeneration and Interactions with Islet Vasculature in Transparent Adult Zebrafish

    PubMed Central

    Moss, Larry G.; Caplan, Tanner V.

    2013-01-01

    Abstract Blood vessel networks provide nutrients and gaseous exchange that are essential for functions. Pancreatic islet capillaries deliver oxygen to endocrine cells while transporting hormones to organs and peripheral locations throughout the body. We have developed a zebrafish diabetes model in which adult islets can be followed in vivo during beta cell regeneration while calibrating changes in beta cell mass and fasting blood glucose levels. After genetic ablation, beta cells are initially dysfunctional or dying, and blood glucose levels increase fourfold. During a 2-week period, hyperglycemia eventually normalizes as beta cell mass regenerates. We show that mCherry-fluorescent, insulin-positive beta cells re-emerge in close contact with the vascular endothelium. Alterations in the dense vascular network of zebrafish islets were visualized by the expression of green fluorescent protein (GFP) in endothelial cells derived from the Fli transcription factor promoter. The rapid destruction and regeneration of beta cell mass was evaluated in the same animal over time, providing a functional model for investigating the interactions of islet cell types with vascular cells as well as the consequences of hyperglycemia on other tissues. Regenerating adult zebrafish can be utilized as vertebrate, metabolically active models for generating new insights into treatments for type 2 diabetes. PMID:23682836

  16. Regeneration of axotomized olfactory neurons in young and adult locusts quantified by fasciclin I immunofluorescence.

    PubMed

    Wasser, Hannah; Biller, Alexandra; Antonopoulos, Georgios; Meyer, Heiko; Bicker, Gerd; Stern, Michael

    2017-04-01

    The olfactory pathway of the locust Locusta migratoria is characterized by a multiglomerular innervation of the antennal lobe (AL) by olfactory receptor neurons (ORNs). After crushing the antenna and thereby severing ORN axons, changes in the AL were monitored. First, volume changes were measured at different times post-crush with scanning laser optical tomography in 5th instar nymphs. AL volume decreased significantly to a minimum volume at 4 days post-crush, followed by an increase. Second, anterograde labeling was used to visualize details in the AL and antennal nerve (AN) during de- and regeneration. Within 24 h post-crush (hpc) the ORN fragments distal to the lesion degenerated. After 48 hpc, regenerating fibers grew through the crush site. In the AL, labeled ORN projections disappeared completely and reappeared after a few days. A weak topographic match between ORN origin on the antenna and the position of innervated glomeruli that was present in untreated controls did not reappear after regeneration. Third, the cell surface marker fasciclin I that is expressed in ORNs was used for quantifying purposes. Immunofluorescence was measured in the AL during de- and regeneration in adults and 5th instar nymphs: after a rapid but transient, decrease, it reappeared. Both processes happen faster in 5th instar nymphs than in adults.

  17. Novel Adult Stem Cells for Peripheral Nerve Regeneration

    DTIC Science & Technology

    2012-09-01

    Circulation 103, 882–888 (2001). 52. Biernaskie, J. et al. SKPs derive from hair follicle precursors and exhibit properties of adult dermal stem cells...3). * indicates significant difference between indicated groups using Holm’s t- test . (P < 0.01). (l–s) Immunostaining of isolated sm-mHC − cells...and the tissue from which the cells were derived using student’s t- test (P < 0.05). † indicates significant difference between inferior vena cava and

  18. Mending broken hearts: cardiac development as a basis for adult heart regeneration and repair.

    PubMed

    Xin, Mei; Olson, Eric N; Bassel-Duby, Rhonda

    2013-08-01

    As the adult mammalian heart has limited potential for regeneration and repair, the loss of cardiomyocytes during injury and disease can result in heart failure and death. The cellular processes and regulatory mechanisms involved in heart growth and development can be exploited to repair the injured adult heart through 'reawakening' pathways that are active during embryogenesis. Heart function has been restored in rodents by reprogramming non-myocytes into cardiomyocytes, by expressing transcription factors (GATA4, HAND2, myocyte-specific enhancer factor 2C (MEF2C) and T-box 5 (TBX5)) and microRNAs (miR-1, miR-133, miR-208 and miR-499) that control cardiomyocyte identity. Stimulating cardiomyocyte dedifferentiation and proliferation by activating mitotic signalling pathways involved in embryonic heart growth represents a complementary approach for heart regeneration and repair. Recent advances in understanding the mechanistic basis of heart development offer exciting opportunities for effective therapies for heart failure.

  19. Multipotent (adult) and pluripotent stem cells for heart regeneration: what are the pros and cons?

    PubMed

    Liao, Song-Yan; Tse, Hung-Fat

    2013-12-24

    Heart failure after myocardial infarction is the leading cause of mortality and morbidity worldwide. Existing medical and interventional therapies can only reduce the loss of cardiomyocytes during myocardial infarction but are unable to replenish the permanent loss of cardiomyocytes after the insult, which contributes to progressive pathological left ventricular remodeling and progressive heart failure. As a result, cell-based therapies using multipotent (adult) stem cells and pluripotent stem cells (embryonic stem cells or induced pluripotent stem cells) have been explored as potential therapeutic approaches to restore cardiac function in heart failure. Nevertheless, the optimal cell type with the best therapeutic efficacy and safety for heart regeneration is still unknown. In this review, the potential pros and cons of different types of multipotent (adult) stem cells and pluripotent stem cells that have been investigated in preclinical and clinical studies are reviewed, and the future perspective of stem cell-based therapy for heart regeneration is discussed.

  20. Epithelial Wnt ligand secretion is required for adult hair follicle growth and regeneration.

    PubMed

    Myung, Peggy S; Takeo, Makoto; Ito, Mayumi; Atit, Radhika P

    2013-01-01

    β-Catenin, a key transducer molecule of Wnt signaling, is required for adult hair follicle growth and regeneration. However, the cellular source of Wnt ligands required for Wnt/β-catenin activation during anagen induction is unknown. In this study, we genetically deleted Wntless (Wls), a gene required for Wnt ligand secretion by Wnt-producing cells, specifically in the hair follicle epithelium during telogen phase. We show that epithelial Wnt ligands are required for anagen, as loss of Wls in the follicular epithelium resulted in a profound hair cycle arrest. Both the follicular epithelium and dermal papilla showed markedly decreased Wnt/β-catenin signaling during anagen induction compared with control hair follicles. Surprisingly, hair follicle stem cells that are responsible for hair regeneration maintained expression of stem cell markers but exhibited significantly reduced proliferation. Finally, we demonstrate that epidermal Wnt ligands are critical for adult wound-induced de novo hair formation. Collectively, these data show that Wnt ligands secreted by the hair follicle epithelium are required for adult hair follicle regeneration and provide new insight into potential cellular targets for the treatment of hair disorders such as alopecia.

  1. Strategies to Optimize Adult Stem Cell Therapy for Tissue Regeneration.

    PubMed

    Liu, Shan; Zhou, Jingli; Zhang, Xuan; Liu, Yang; Chen, Jin; Hu, Bo; Song, Jinlin; Zhang, Yuanyuan

    2016-06-21

    Stem cell therapy aims to replace damaged or aged cells with healthy functioning cells in congenital defects, tissue injuries, autoimmune disorders, and neurogenic degenerative diseases. Among various types of stem cells, adult stem cells (i.e., tissue-specific stem cells) commit to becoming the functional cells from their tissue of origin. These cells are the most commonly used in cell-based therapy since they do not confer risk of teratomas, do not require fetal stem cell maneuvers and thus are free of ethical concerns, and they confer low immunogenicity (even if allogenous). The goal of this review is to summarize the current state of the art and advances in using stem cell therapy for tissue repair in solid organs. Here we address key factors in cell preparation, such as the source of adult stem cells, optimal cell types for implantation (universal mesenchymal stem cells vs. tissue-specific stem cells, or induced vs. non-induced stem cells), early or late passages of stem cells, stem cells with endogenous or exogenous growth factors, preconditioning of stem cells (hypoxia, growth factors, or conditioned medium), using various controlled release systems to deliver growth factors with hydrogels or microspheres to provide apposite interactions of stem cells and their niche. We also review several approaches of cell delivery that affect the outcomes of cell therapy, including the appropriate routes of cell administration (systemic, intravenous, or intraperitoneal vs. local administration), timing for cell therapy (immediate vs. a few days after injury), single injection of a large number of cells vs. multiple smaller injections, a single site for injection vs. multiple sites and use of rodents vs. larger animal models. Future directions of stem cell-based therapies are also discussed to guide potential clinical applications.

  2. Strategies to Optimize Adult Stem Cell Therapy for Tissue Regeneration

    PubMed Central

    Liu, Shan; Zhou, Jingli; Zhang, Xuan; Liu, Yang; Chen, Jin; Hu, Bo; Song, Jinlin; Zhang, Yuanyuan

    2016-01-01

    Stem cell therapy aims to replace damaged or aged cells with healthy functioning cells in congenital defects, tissue injuries, autoimmune disorders, and neurogenic degenerative diseases. Among various types of stem cells, adult stem cells (i.e., tissue-specific stem cells) commit to becoming the functional cells from their tissue of origin. These cells are the most commonly used in cell-based therapy since they do not confer risk of teratomas, do not require fetal stem cell maneuvers and thus are free of ethical concerns, and they confer low immunogenicity (even if allogenous). The goal of this review is to summarize the current state of the art and advances in using stem cell therapy for tissue repair in solid organs. Here we address key factors in cell preparation, such as the source of adult stem cells, optimal cell types for implantation (universal mesenchymal stem cells vs. tissue-specific stem cells, or induced vs. non-induced stem cells), early or late passages of stem cells, stem cells with endogenous or exogenous growth factors, preconditioning of stem cells (hypoxia, growth factors, or conditioned medium), using various controlled release systems to deliver growth factors with hydrogels or microspheres to provide apposite interactions of stem cells and their niche. We also review several approaches of cell delivery that affect the outcomes of cell therapy, including the appropriate routes of cell administration (systemic, intravenous, or intraperitoneal vs. local administration), timing for cell therapy (immediate vs. a few days after injury), single injection of a large number of cells vs. multiple smaller injections, a single site for injection vs. multiple sites and use of rodents vs. larger animal models. Future directions of stem cell-based therapies are also discussed to guide potential clinical applications. PMID:27338364

  3. Serotonin promotes acinar dedifferentiation following pancreatitis-induced regeneration in the adult pancreas.

    PubMed

    Saponara, Enrica; Grabliauskaite, Kamile; Bombardo, Marta; Buzzi, Raphael; Silva, Alberto B; Malagola, Ermanno; Tian, Yinghua; Hehl, Adrian B; Schraner, Elisabeth M; Seleznik, Gitta M; Zabel, Anja; Reding, Theresia; Sonda, Sabrina; Graf, Rolf

    2015-12-01

    The exocrine pancreas exhibits a distinctive capacity for tissue regeneration and renewal following injury. This regenerative ability has important implications for a variety of disorders, including pancreatitis and pancreatic cancer, diseases associated with high morbidity and mortality. Thus, understanding its underlying mechanisms may help in developing therapeutic interventions. Serotonin has been recognized as a potent mitogen for a variety of cells and tissues. Here we investigated whether serotonin exerts a mitogenic effect in pancreatic acinar cells in three regenerative models, inflammatory tissue injury following pancreatitis, tissue loss following partial pancreatectomy, and thyroid hormone-stimulated acinar proliferation. Genetic and pharmacological techniques were used to modulate serotonin levels in vivo. Acinar dedifferentiation and cell cycle progression during the regenerative phase were investigated over the course of 2 weeks. By comparing acinar proliferation in the different murine models of regeneration, we found that serotonin did not affect the clonal regeneration of mature acinar cells. Serotonin was, however, required for acinar dedifferentiation following inflammation-mediated tissue injury. Specifically, lack of serotonin resulted in delayed up-regulation of progenitor genes and delayed the formation of acinar-to-ductal metaplasia and defective acinar cell proliferation. We identified serotonin-dependent acinar secretion as a key step in progenitor-based regeneration, as it promoted acinar cell dedifferentiation and the recruitment of type 2 macrophages. Finally, we identified a regulatory Hes1-Ptfa axis in the uninjured adult pancreas, activated by zymogen secretion. Our findings indicated that serotonin plays a critical role in the regeneration of the adult pancreas following pancreatitis by promoting the dedifferentiation of acinar cells.

  4. Connective tissue cells, but not muscle cells, are involved in establishing the proximo-distal outcome of limb regeneration in the axolotl.

    PubMed

    Nacu, Eugen; Glausch, Mareen; Le, Huy Quang; Damanik, Febriyani Fiain Rochel; Schuez, Maritta; Knapp, Dunja; Khattak, Shahryar; Richter, Tobias; Tanaka, Elly M

    2013-02-01

    During salamander limb regeneration, only the structures distal to the amputation plane are regenerated, a property known as the rule of distal transformation. Multiple cell types are involved in limb regeneration; therefore, determining which cell types participate in distal transformation is important for understanding how the proximo-distal outcome of regeneration is achieved. We show that connective tissue-derived blastema cells obey the rule of distal transformation. They also have nuclear MEIS, which can act as an upper arm identity regulator, only upon upper arm amputation. By contrast, myogenic cells do not obey the rule of distal transformation and display nuclear MEIS upon amputation at any proximo-distal level. These results indicate that connective tissue cells, but not myogenic cells, are involved in establishing the proximo-distal outcome of regeneration and are likely to guide muscle patterning. Moreover, we show that, similarly to limb development, muscle patterning in regeneration is influenced by β-catenin signalling.

  5. Electrically induced muscle cramps induce hypertrophy of calf muscles in healthy adults

    PubMed Central

    Behringer, M.; Moser, M.; Montag, J.; McCourt, M.; Tenner, D.; Mester, J.

    2015-01-01

    Objectives: Skeletal muscles usually cramp at short lengths, where the tension that can be exerted by muscle fibers is low. Since high tension is an important anabolic stimulus, it is questionable if cramps can induce hypertrophy and strength gains. In the present study we investigated if electrically induced cramps (EIMCs) can elicit these adaptations. Methods: 15 healthy male adults were randomly assigned to an intervention (IG; n=10) and a control group (CG; n=5). The cramp protocol (CP) applied twice a week to one leg of the IG, consisted of 3x6 EIMCs, of 5 s each. Calf muscles of the opposite leg were stimulated equally, but were hindered from cramping by fixating the ankle at 0° plantar flexion (nCP). Results: After six weeks, the cross sectional area of the triceps surae was similarly increased in both the CP (+9.0±3.4%) and the nCP (+6.8±3.7%). By contrast, force of maximal voluntary contractions, measured at 0° and 30° plantar flexion, increased significantly only in nCP (0°: +8.5±8.8%; 30°: 11.7±13.7%). Conclusion: The present data indicate that muscle cramps can induce hypertrophy in calf muscles, though lacking high tension as an important anabolic stimulus. PMID:26032216

  6. Effects of Topical Icing on Inflammation, Angiogenesis, Revascularization, and Myofiber Regeneration in Skeletal Muscle Following Contusion Injury

    PubMed Central

    Singh, Daniel P.; Barani Lonbani, Zohreh; Woodruff, Maria A.; Parker, Tony J.; Steck, Roland; Peake, Jonathan M.

    2017-01-01

    Contusion injuries in skeletal muscle commonly occur in contact sport and vehicular and industrial workplace accidents. Icing has traditionally been used to treat such injuries under the premise that it alleviates pain, reduces tissue metabolism, and modifies vascular responses to decrease swelling. Previous research has examined the effects of icing on inflammation and microcirculatory dynamics following muscle injury. However, whether icing influences angiogenesis, collateral vessel growth, or myofiber regeneration remains unknown. We compared the effects of icing vs. a sham treatment on the presence of neutrophils and macrophages; expression of CD34, von Willebrands factor (vWF), vascular endothelial growth factor (VEGF), and nestin; vessel volume; capillary density; and myofiber regeneration in skeletal after muscle contusion injury in rats. Muscle tissue was collected 1, 3, 7, and 28 d after injury. Compared with uninjured rats, muscles in rats that sustained the contusion injury exhibited major necrosis, inflammation, and increased expression of CD34, vWF, VEGF, and nestin. Compared with the sham treatment, icing attenuated and/or delayed neutrophil and macrophage infiltration; the expression of vWF, VEGF, and nestin; and the change in vessel volume within muscle in the first 7 d after injury (P < 0.05). By contrast, icing did not influence capillary density in muscle 28 d after injury (P = 0.59). The percentage of immature myofibers relative to the total number of fibers was greater in the icing group than in the sham group 28 d after injury (P = 0.026), but myofiber cross-sectional area did not differ between groups after 7 d (P = 0.35) and 28 d (P = 0.30). In conclusion, although icing disrupted inflammation and some aspects of angiogenesis/revascularization, these effects did not result in substantial differences in capillary density or muscle growth. PMID:28326040

  7. The lumbrical muscle: a novel in situ system to evaluate adult skeletal muscle proteolysis and anticatabolic drugs for therapeutic purposes.

    PubMed

    Bergantin, Leandro Bueno; Figueiredo, Leonardo Bruno; Godinho, Rosely Oliveira

    2011-12-01

    The molecular regulation of skeletal muscle proteolysis and the pharmacological screening of anticatabolic drugs have been addressed by measuring tyrosine release from prepubertal rat skeletal muscles, which are thin enough to allow adequate in vitro diffusion of oxygen and substrates. However, the use of muscle at accelerated prepubertal growth has limited the analysis of adult muscle proteolysis or that associated with aging and neurodegenerative diseases. Here we established the adult rat lumbrical muscle (4/hindpaw; 8/rat) as a new in situ experimental model for dynamic measurement of skeletal muscle proteolysis. By incubating lumbrical muscles attached to their individual metatarsal bones in Tyrode solution, we showed that the muscle proteolysis rate of adult and aged rats (3-4 to 24 mo old) is 45-25% of that in prepubertal animals (1 mo old), which makes questionable the usual extrapolation of proteolysis from prepubertal to adult/senile muscles. While acute mechanical injury or 1- to 7-day denervation increased tyrosine release from adult lumbrical muscle by up to 60%, it was reduced by 20-28% after 2-h incubation with β-adrenoceptor agonists, forskolin or phosphodiesterase inhibitor IBMX. Using inhibitors of 26S-proteasome (MG132), lysosome (methylamine), or calpain (E64/leupeptin) systems, we showed that ubiquitin-proteasome is accountable for 40-50% of total lumbrical proteolysis of adult, middle-aged, and aged rats. In conclusion, the lumbrical model allows the analysis of muscle proteolysis rate from prepubertal to senile rats. By permitting eight simultaneous matched measurements per rat, the new model improves similar protocols performed in paired extensor digitorum longus (EDL) muscles from prepubertal rats, optimizing the pharmacological screening of drugs for anticatabolic purposes.

  8. p66(ShcA) and oxidative stress modulate myogenic differentiation and skeletal muscle regeneration after hind limb ischemia.

    PubMed

    Zaccagnini, Germana; Martelli, Fabio; Magenta, Alessandra; Cencioni, Chiara; Fasanaro, Pasquale; Nicoletti, Carmine; Biglioli, Paolo; Pelicci, Pier Giuseppe; Capogrossi, Maurizio C

    2007-10-26

    Oxidative stress plays a pivotal role in ischemic injury, and p66(ShcA)ko mice exhibit both lower oxidative stress and decreased tissue damage following hind limb ischemia. Thus, it was investigated whether tissue regeneration following acute hind limb ischemia was altered in p66(ShcA)ko mice. Upon femoral artery dissection, muscle regeneration started earlier and was completed faster than in wild-type (WT) control. Moreover, faster regeneration was associated with decreased oxidative stress. Unlike ischemia, cardiotoxin injury induced similar skeletal muscle damage in both genotypes. However, p66(ShcA)ko mice regenerated faster, in agreement with the regenerative advantage upon ischemia. Since no difference between p66(ShcA)wt and knock-out (ko) mice was found in blood perfusion recovery after ischemia, satellite cells (SCs), a resident population of myogenic progenitors, were examined. Similar SCs numbers were present in WT and ko mice. However, in vitro cultured p66(ShcA)ko SCs displayed lower oxidative stress levels and higher proliferation rate and differentiated faster than WT. Furthermore, when exposed to sublethal H(2)O(2) doses, p66(ShcA)ko SCs were resistant to H(2)O(2)-induced inhibition of differentiation. Finally, myogenic conversion induced by MyoD overexpression was more efficient in p66(ShcA)ko fibroblasts compared with WT. The present work demonstrates that oxidative stress and p66(ShcA) play a crucial role in the regenerative pathways activated by acute ischemia.

  9. A Tunable Silk Hydrogel Device for Studying Limb Regeneration in Adult Xenopus Laevis

    PubMed Central

    Golding, Anne; Levin, Michael; Kaplan, David L.

    2016-01-01

    In certain amphibian models limb regeneration can be promoted or inhibited by the local wound bed environment. This research introduces a device that can be utilized as an experimental tool to characterize the conditions that promotes limb regeneration in the adult frog (Xenopus laevis) model. In particular, this device was designed to manipulate the local wound environment via a hydrogel insert. Initial characterization of the hydrogel insert revealed that this interaction had a significant influence on mechanical forces to the animal, due to the contraction of the hydrogel. The material and mechanical properties of the hydrogel insert were a factor in the device design in relation to the comfort of the animal and the ability to effectively manipulate the amputation site. The tunable features of the hydrogel were important in determining the pro-regenerative effects in limb regeneration, which was measured by cartilage spike formation and quantified by micro-computed tomography. The hydrogel insert was a factor in the observed morphological outcomes following amputation. Future work will focus on characterizing and optimizing the device’s observed capability to manipulate biological pathways that are essential for limb regeneration. However, the present work provides a framework for the role of a hydrogel in the device and a path forward for more systematic studies. PMID:27257960

  10. Duct Cells Contribute to Regeneration of Endocrine and Acinar Cells Following Pancreatic Damage in Adult Mice

    PubMed Central

    CRISCIMANNA, ANGELA; SPEICHER, JULIE A.; HOUSHMAND, GOLBAHAR; SHIOTA, CHIYO; PRASADAN, KRISHNA; Ji, BAOAN; LOGSDON, CRAIG D.; GITTES, GEORGE K.; ESNI, FARZAD

    2015-01-01

    BACKGROUND & AIMS There have been conflicting results on a cell of origin in pancreatic regeneration. These discrepancies predominantly stem from lack of specific markers for the pancreatic precursors/stem cells, as well as differences in the targeted cells and severity of tissue injury in the experimental models so far proposed. We attempted to create a model that used diphtheria toxin receptor (DTR) to ablate specific cell populations, control the extent of injury, and avoid induction of the inflammatory response. METHODS To target specific types of pancreatic cells, we crossed R26DTR or R26dtR/lacZ mice with transgenic mice that express the Cre recombinase in the pancreas, under control of the Pdx1 (global pancreatic) or elastase (acinar-specific) promoters. RESULTS Exposure of PdxCre;R26DTR mice to diphtheria toxin resulted in extensive ablation of acinar and endocrine tissues but not ductal cells. Surviving cells within the ductal compartment contributed to regeneration of endocrine and acinar cells via recapitulation of the embryonic pancreatic developmental program. However, following selective ablation of acinar tissue in ElaCre-ERT2;R26DTR mice, regeneration likely occurred by reprogramming of ductal cells to acinar lineage. CONCLUSIONS In the pancreas of adult mice, epithelial cells within the ductal compartment contribute to regeneration of endocrine and acinar cells. The severity of injury determines the regenerative mechanisms and cell types that contribute to this process. PMID:21763240

  11. Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps.

    PubMed

    Broughton, Kathleen M; Sussman, Mark A

    2016-03-04

    Much has changed since our survey of the landscape for myocardial regeneration powered by adult stem cells 4 years ago.(1) The intervening years since that first review has witnessed an explosive expansion of studies that advance both understanding and implementation of adult stem cells in promoting myocardial repair. Painstaking research from innumerable laboratories throughout the world is prying open doors that may lead to restoration of myocardial structure and function in the wake of pathological injury. This global effort has produced deeper mechanistic comprehension coupled with an evolving appreciation for the complexity of myocardial regeneration in the adult context. Undaunted by both known and (as yet) unknown challenges, pursuit of myocardial regenerative medicine mediated by adult stem cell therapy has gathered momentum fueled by tantalizing clues and visionary goals. This concise review takes a somewhat different perspective than our initial treatise, taking stock of the business sector that has become an integral part of the field while concurrently updating state of affairs in cutting edge research. Looking retrospectively at advancement over the years as all reviews eventually must, the fundamental lesson to be learned is best explained by Jonatan Mårtensson: "Success will never be a big step in the future. Success is a small step taken just now."

  12. Regulation of neonatal and adult mammalian heart regeneration by the miR-15 family

    PubMed Central

    Porrello, Enzo R.; Mahmoud, Ahmed I.; Simpson, Emma; Johnson, Brett A.; Grinsfelder, David; Canseco, Diana; Mammen, Pradeep P.; Rothermel, Beverly A.; Olson, Eric N.; Sadek, Hesham A.

    2013-01-01

    We recently identified a brief time period during postnatal development when the mammalian heart retains significant regenerative potential after amputation of the ventricular apex. However, one major unresolved question is whether the neonatal mouse heart can also regenerate in response to myocardial ischemia, the most common antecedent of heart failure in humans. Here, we induced ischemic myocardial infarction (MI) in 1-d-old mice and found that this results in extensive myocardial necrosis and systolic dysfunction. Remarkably, the neonatal heart mounted a robust regenerative response, through proliferation of preexisting cardiomyocytes, resulting in full functional recovery within 21 d. Moreover, we show that the miR-15 family of microRNAs modulates neonatal heart regeneration through inhibition of postnatal cardiomyocyte proliferation. Finally, we demonstrate that inhibition of the miR-15 family from an early postnatal age until adulthood increases myocyte proliferation in the adult heart and improves left ventricular systolic function after adult MI. We conclude that the neonatal mammalian heart can regenerate after myocardial infarction through proliferation of preexisting cardiomyocytes and that the miR-15 family contributes to postnatal loss of cardiac regenerative capacity. PMID:23248315

  13. Cellular dynamics of regeneration reveals role of two distinct Pax7 stem cell populations in larval zebrafish muscle repair

    PubMed Central

    Pipalia, Tapan G.; Koth, Jana; Roy, Shukolpa D.; Hammond, Christina L.; Kawakami, Koichi

    2016-01-01

    ABSTRACT Heterogeneity of stem cells or their niches is likely to influence tissue regeneration. Here we reveal stem/precursor cell diversity during wound repair in larval zebrafish somitic body muscle using time-lapse 3D confocal microscopy on reporter lines. Skeletal muscle with incision wounds rapidly regenerates both slow and fast muscle fibre types. A swift immune response is followed by an increase in cells at the wound site, many of which express the muscle stem cell marker Pax7. Pax7+ cells proliferate and then undergo terminal differentiation involving Myogenin accumulation and subsequent loss of Pax7 followed by elongation and fusion to repair fast muscle fibres. Analysis of pax7a and pax7b transgenic reporter fish reveals that cells expressing each of the duplicated pax7 genes are distinctly localised in uninjured larvae. Cells marked by pax7a only or by both pax7a and pax7b enter the wound rapidly and contribute to muscle wound repair, but each behaves differently. Low numbers of pax7a-only cells form nascent fibres. Time-lapse microscopy revealed that the more numerous pax7b-marked cells frequently fuse to pre-existing fibres, contributing more strongly than pax7a-only cells to repair of damaged fibres. pax7b-marked cells are more often present in rows of aligned cells that are observed to fuse into a single fibre, but more rarely contribute to nascent regenerated fibres. Ablation of a substantial portion of nitroreductase-expressing pax7b cells with metronidazole prior to wounding triggered rapid pax7a-only cell accumulation, but this neither inhibited nor augmented pax7a-only cell-derived myogenesis and thus altered the cellular repair dynamics during wound healing. Moreover, pax7a-only cells did not regenerate pax7b cells, suggesting a lineage distinction. We propose a modified founder cell and fusion-competent cell model in which pax7a-only cells initiate fibre formation and pax7b cells contribute to fibre growth. This newly discovered cellular

  14. Skeletal myogenic differentiation of human urine-derived cells as a potential source for skeletal muscle regeneration.

    PubMed

    Chen, Wei; Xie, Minkai; Yang, Bin; Bharadwaj, Shantaram; Song, Lujie; Liu, Guihua; Yi, Shanhong; Ye, Gang; Atala, Anthony; Zhang, Yuanyuan

    2017-02-01

    Stem cells are regarded as possible cell therapy candidates for skeletal muscle regeneration. However, invasive harvesting of those cells can cause potential harvest-site morbidity. The goal of this study was to assess whether human urine-derived stem cells (USCs), obtained through non-invasive procedures, can differentiate into skeletal muscle linage cells (Sk-MCs) and potentially be used for skeletal muscle regeneration. In this study, USCs were harvested from six healthy individuals aged 25-55. Expression profiles of cell-surface markers were assessed by flow cytometry. To optimize the myogenic differentiation medium, we selected two from four different types of myogenic differentiation media to induce the USCs. Differentiated USCs were identified with myogenic markers by gene and protein expression. USCs were implanted into the tibialis anterior muscles of nude mice for 1 month. The results showed that USCs displayed surface markers with positive staining for CD24, CD29, CD44, CD73, CD90, CD105, CD117, CD133, CD146, SSEA-4 and STRO-1, and negative staining for CD14, CD31, CD34 and CD45. After myogenic differentiation, a change in morphology was observed from 'rice-grain'-like cells to spindle-shaped cells. The USCs expressed specific Sk-MC transcripts and protein markers (myf5, myoD, myosin, and desmin) after being induced with different myogenic culture media. Implanted cells expressed Sk-MC markers stably in vivo. Our findings suggest that USCs are able to differentiate into the Sk-MC lineage in vitro and after being implanted in vivo. Thus, they might be a potential source for cell injection therapy in the use of skeletal muscle regeneration. Copyright © 2014 John Wiley & Sons, Ltd.

  15. A subpopulation of adult skeletal muscle stem cells retains all template DNA strands after cell division.

    PubMed

    Rocheteau, Pierre; Gayraud-Morel, Barbara; Siegl-Cachedenier, Irene; Blasco, Maria A; Tajbakhsh, Shahragim

    2012-01-20

    Satellite cells are adult skeletal muscle stem cells that are quiescent and constitute a poorly defined heterogeneous population. Using transgenic Tg:Pax7-nGFP mice, we show that Pax7-nGFP(Hi) cells are less primed for commitment and have a lower metabolic status and delayed first mitosis compared to Pax7-nGFP(Lo) cells. Pax7-nGFP(Hi) can give rise to Pax7-nGFP(Lo) cells after serial transplantations. Proliferating Pax7-nGFP(Hi) cells exhibit lower metabolic activity, and the majority performs asymmetric DNA segregation during cell division, wherein daughter cells retaining template DNA strands express stem cell markers. Using chromosome orientation-fluorescence in situ hybridization, we demonstrate that all chromatids segregate asymmetrically, whereas Pax7-nGFP(Lo) cells perform random DNA segregation. Therefore, quiescent Pax7-nGFP(Hi) cells represent a reversible dormant stem cell state, and during muscle regeneration, Pax7-nGFP(Hi) cells generate distinct daughter cell fates by asymmetrically segregating template DNA strands to the stem cell. These findings provide major insights into the biology of stem cells that segregate DNA asymmetrically.

  16. Influence of Botulinumtoxin A on the Expression of Adult MyHC Isoforms in the Masticatory Muscles in Dystrophin-Deficient Mice (Mdx-Mice)

    PubMed Central

    Todorov, Teodor

    2016-01-01

    The most widespread animal model to investigate Duchenne muscular dystrophy is the mdx-mouse. In contrast to humans, phases of muscle degeneration are replaced by regeneration processes; hence there is only a restricted time slot for research. The aim of the study was to investigate if an intramuscular injection of BTX-A is able to break down muscle regeneration and has direct implications on the gene expression of myosin heavy chains in the corresponding treated and untreated muscles. Therefore, paralysis of the right masseter muscle was induced in adult healthy and dystrophic mice by a specific intramuscular injection of BTX-A. After 21 days the mRNA expression and protein content of MyHC isoforms of the right and left masseter, temporal, and the tongue muscle were determined using quantitative RT-PCR and Western blot technique. MyHC-IIa and MyHC-I-mRNA expression significantly increased in the paralyzed masseter muscle of control-mice, whereas MyHC-IIb and MyHC-IIx/d-mRNA were decreased. In dystrophic muscles no effect of BTX-A could be detected at the level of MyHC. This study suggests that BTX-A injection is a suitable method to simulate DMD-pathogenesis in healthy mice but further investigations are necessary to fully analyse the BTX-A effect and to generate sustained muscular atrophy in mdx-mice. PMID:27689088

  17. Skin incision induces expression of axonal regeneration-related genes in adult rat spinal sensory neurons

    PubMed Central

    Hill, Caitlin E.; Harrison, Benjamin J.; Rau, Kris K.; Hougland, M. Tyler; Bunge, Mary Bartlett; Mendell, Lorne M.; Petruska, Jeffrey C.

    2010-01-01

    Skin incision and nerve injury both induce painful conditions. Incisional and post-surgical pain is believed to arise primarily from inflammation of tissue and the subsequent sensitization of peripheral and central neurons. The role of axonal regeneration-related processes in development of pain has only been considered when there has been injury to the peripheral nerve itself, even though tissue damage likely induces injury of resident axons. We sought to determine if skin incision would affect expression of regeneration-related genes such as activating transcription factor 3 (ATF3) in dorsal root ganglion (DRG) neurons. ATF3 is absent from DRG neurons of the normal adult rodent, but is induced by injury of peripheral nerves and modulates the regenerative capacity of axons. Image analysis of immunolabeled DRG sections revealed that skin incision led to an increase in the number of DRG neurons expressing ATF3. RT-PCR indicated that other regeneration-associated genes (galanin, GAP-43, Gadd45a) were also increased, further suggesting an injury-like response in DRG neurons. Our finding that injury of skin can induce expression of neuronal injury/regeneration-associated genes may impact how clinical post-surgical pain is investigated and treated. Perspective Tissue injury, even without direct nerve injury, may induce a state of enhanced growth capacity in sensory neurons. Axonal regeneration-associated processes should be considered alongside nerve signal conduction and inflammatory/sensitization processes as possible mechanisms contributing to pain, particularly the transition from acute to chronic pain. PMID:20627820

  18. Monoclonal antibodies against muscle actin isoforms: epitope identification and analysis of isoform expression by immunoblot and immunostaining in normal and regenerating skeletal muscle

    PubMed Central

    Chaponnier, Christine; Gabbiani, Giulio

    2016-01-01

    Higher vertebrates (mammals and birds) express six different highly conserved actin isoforms that can be classified in three subgroups: 1) sarcomeric actins, α-skeletal (α-SKA) and α-cardiac (α-CAA), 2) smooth muscle actins (SMAs), α-SMA and γ-SMA, and 3) cytoplasmic actins (CYAs), β-CYA and γ-CYA. The variations among isoactins, in each subgroup, are due to 3-4 amino acid differences located in their acetylated N-decapeptide sequence. The first monoclonal antibody (mAb) against an actin isoform (α-SMA) was produced and characterized in our laboratory in 1986 (Skalli  et al., 1986) . We have further obtained mAbs against the 5 other isoforms. In this report, we focus on the mAbs anti-α-SKA and anti-α-CAA obtained after immunization of mice with the respective acetylated N-terminal decapeptides using the Repetitive Immunizations at Multiple Sites Strategy (RIMMS). In addition to the identification of their epitope by immunoblotting, we describe the expression of the 2 sarcomeric actins in mature skeletal muscle and during muscle repair after micro-lesions. In particular, we analyze the expression of α-CAA, α-SKA and α-SMA by co-immunostaining in a time course frame during the muscle repair process. Our results indicate that a restricted myocyte population expresses α-CAA and suggest a high capacity of self-regeneration in muscle cells. These antibodies may represent a helpful tool for the follow-up of muscle regeneration and pathological changes. PMID:27335638

  19. NGF induces adult stem Leydig cells to proliferate and differentiate during Leydig cell regeneration

    SciTech Connect

    Zhang, Lei; Wang, Huaxi; Yang, Yan; Liu, Hui; Zhang, Qihao; Xiang, Qi; Ge, Renshan; Su, Zhijian; Huang, Yadong

    2013-06-28

    Highlights: •Nerve growth factor has shown significant changes on mRNA levels during Adult Leydig cells regeneration. •We established the organ culture model of rat seminiferous tubules with ethane dimethyl sulphonate (EDS) treatment. •Nerve growth factor has shown proliferation and differentiation-promoting effects on Adult stem Leydig cells. •Nerve growth factor induces progenitor Leydig cells to proliferate and differentiate and immature Leydig cells to proliferate. -- Abstract: Nerve growth factor (NGF) has been reported to be involved in male reproductive physiology. However, few reports have described the activity of NGF during Leydig cell development. The objective of the present study was to examine the role of NGF during stem-Leydig-cell (SLC) regeneration. We investigated the effects of NGF on Leydig-cell (LC) regeneration by measuring mRNA levels in the adult rat testis after ethane dimethanesulfonate (EDS) treatment. Furthermore, we used the established organ culture model of rat seminiferous tubules to examine the regulation of NGF during SLC proliferation and differentiation using EdU staining, real-time PCR and western blotting. Progenitor Leydig cells (PLCs) and immature Leydig cells (ILCs) were also used to investigate the effects of NGF on LCs at different developmental stages. NGF mRNA levels changed significantly during Leydig-cell regeneration in vivo. In vitro, NGF significantly promoted the proliferation of stem Leydig cells and also induced steroidogenic enzyme gene expression and 3β-HSD protein expression. The data from PLCs and ILCs showed that NGF could increase Cyclin D1 and Hsd 17b3 mRNA levels in PLCs and Cyclin D1 mRNA levels in ILCs. These results indicate that NGF may play an important role during LC regeneration by regulating the proliferation and differentiation of LCs at different developmental stages, from SLCs to PLCs and from PLCs to ILCs. The discovery of this effect of NGF on Leydig cells will provide useful

  20. A case of adult-onset reducing body myopathy presenting a novel clinical feature, asymmetrical involvement of the sternocleidomastoid and trapezius muscles.

    PubMed

    Fujii, Takayuki; Hayashi, Shintaro; Kawamura, Nobutoshi; Higuchi, Masa-Aki; Tsugawa, Jun; Ohyagi, Yasumasa; Hayashi, Yukiko K; Nishino, Ichizo; Kira, Jun-Ichi

    2014-08-15

    We herein report a 32-year-old woman with adult-onset reducing body myopathy (RBM) who had a mutation in the four-and-a-half LIM domain 1 gene (FHL1) and showed a marked asymmetrical involvement of sternocleidomastoid and trapezius muscles. At 30 years of age she noticed bilateral foot drop, and over the next two years developed difficulty raising her right arm. At 32 years of age she was admitted to our hospital for a diagnostic evaluation. Neurological examination showed moderate weakness and atrophy of her right sternocleidomastoid muscle, right trapezius muscle, and bilateral upper proximal muscles. There were severe weakness and atrophy of her bilateral tibialis anterior muscles. Her deep tendon reflexes were hypoactive in her upper extremities. Her serum creatine kinase level was mildly increased. Muscle biopsy specimens from the left tibialis anterior muscle revealed marked variation in fiber size, some necrotic or regenerating fibers, and reducing bodies. Gene analysis of FHL1 demonstrated a mutation: a heterozygous missense mutation of c.377G>A (p. C126T) in FHL1. Compared with previous adult-onset RBM cases harboring mutations in FHL1, our case was characterized by asymmetrical atrophy of the sternocleidomastoid and trapezius muscles.

  1. Distinct effects of inflammation on preconditioning and regeneration of the adult zebrafish heart

    PubMed Central

    de Preux Charles, Anne-Sophie; Bise, Thomas; Baier, Felix; Marro, Jan; Jaźwińska, Anna

    2016-01-01

    The adult heart is able to activate cardioprotective programmes and modifies its architecture in response to physiological or pathological changes. While mammalian cardiac remodelling often involves hypertrophic expansion, the adult zebrafish heart exploits hyperplastic growth. This capacity depends on the responsiveness of zebrafish cardiomyocytes to mitogenic signals throughout their entire life. Here, we have examined the role of inflammation on the stimulation of cell cycle activity in the context of heart preconditioning and regeneration. We used thoracotomy as a cardiac preconditioning model and cryoinjury as a model of cardiac infarction in the adult zebrafish. First, we performed a spatio-temporal characterization of leucocytes and cycling cardiac cells after thoracotomy. This analysis revealed a concomitance between the infiltration of inflammatory cells and the stimulation of the mitotic activity. However, decreasing the immune response using clodronate liposome injection, PLX3397 treatment or anti-inflammatory drugs surprisingly had no effect on the re-entry of cardiac cells into the cell cycle. In contrast, reducing inflammation using the same strategies after cryoinjury strongly impaired cardiac cell mitotic activity and the regenerative process. Taken together, our results show that, while the immune response is not necessary to induce cell-cycle activity in intact preconditioned hearts, inflammation is required for the regeneration of injured hearts in zebrafish. PMID:27440424

  2. Trop2 marks transient gastric fetal epithelium and adult regenerating cells after epithelial damage.

    PubMed

    Fernandez Vallone, Valeria; Leprovots, Morgane; Strollo, Sandra; Vasile, Gabriela; Lefort, Anne; Libert, Frederick; Vassart, Gilbert; Garcia, Marie-Isabelle

    2016-05-01

    Mouse fetal intestinal progenitors lining the epithelium prior to villogenesis grow as spheroids when cultured ex vivo and express the transmembrane glycoprotein Trop2 as a marker. Here, we report the characterization of Trop2-expressing cells from fetal pre-glandular stomach, growing as immortal undifferentiated spheroids, and their relationship with gastric development and regeneration. Trop2(+) cells generating gastric spheroids differed from adult glandular Lgr5(+) stem cells, but appeared highly related to fetal intestinal spheroids. Although they shared a common spheroid signature, intestinal and gastric fetal spheroid-generating cells expressed organ-specific transcription factors and were committed to intestinal and glandular gastric differentiation, respectively. Trop2 expression was transient during glandular stomach development, being lost at the onset of gland formation, whereas it persisted in the squamous forestomach. Undetectable under homeostasis, Trop2 was strongly re-expressed in glands after acute Lgr5(+) stem cell ablation or following indomethacin-induced injury. These highly proliferative reactive adult Trop2(+) cells exhibited a transcriptome displaying similarity with that of gastric embryonic Trop2(+) cells, suggesting that epithelium regeneration in adult stomach glands involves the partial re-expression of a fetal genetic program.

  3. Regenerating tail muscles in lizard contain Fast but not Slow Myosin indicating that most myofibers belong to the fast twitch type for rapid contraction.

    PubMed

    Alibardi, L

    2015-10-01

    During tail regeneration in lizards a large mass of muscle tissue is formed in form of segmental myomeres of similar size located under the dermis of the new tail. These muscles accumulate glycogen and a fast form of myosin typical for twitch myofibers as it is shown by light and ultrastructural immunocytochemistry using an antibody directed against a Fast Myosin Heavy Chain. High resolution immunogold labeling shows that an intense labeling for fast myosin is localized over the thick filaments of the numerous myofibrils in about 70% of the regenerated myofibers while the labeling becomes less intense in the remaining muscle fibers. The present observations indicate that at least two subtypes of Fast Myosin containing muscle fibers are regenerated, the prevalent type was of the fast twitch containing few mitochondria, sparse glycogen, numerous smooth endoplasmic reticulum vesicles. The second, and less frequent type was a Fast-Oxidative-Glycolitic twitch fiber containing more mitochondria, a denser cytoplasm and myofibrils. Since their initial differentiation, myoblasts, myotubes and especially the regenerated myofibers do not accumulate any immuno-detectable Slow Myosin Heavy Chain. The study indicates that most of the segmental muscles of the regenerated tail serve for the limited bending of the tail during locomotion and trashing after amputation of the regenerated tail, a phenomenon that facilitates predator escape.

  4. Myogenic Akt signaling attenuates muscular degeneration, promotes myofiber regeneration and improves muscle function in dystrophin-deficient mdx mice

    PubMed Central

    Kim, Michelle H.; Kay, Danielle I.; Rudra, Renuka T.; Chen, Bo Ming; Hsu, Nigel; Izumiya, Yasuhiro; Martinez, Leonel; Spencer, Melissa J.; Walsh, Kenneth; Grinnell, Alan D.; Crosbie, Rachelle H.

    2011-01-01

    Duchenne muscular dystrophy, the most common form of childhood muscular dystrophy, is caused by X-linked inherited mutations in the dystrophin gene. Dystrophin deficiencies result in the loss of the dystrophin–glycoprotein complex at the plasma membrane, which leads to structural instability and muscle degeneration. Previously, we induced muscle-specific overexpression of Akt, a regulator of cellular metabolism and survival, in mdx mice at pre-necrotic (<3.5 weeks) ages and demonstrated upregulation of the utrophin–glycoprotein complex and protection against contractile-induced stress. Here, we found that delaying exogenous Akt treatment of mdx mice after the onset of peak pathology (>6 weeks) similarly increased the abundance of compensatory adhesion complexes at the extrasynaptic sarcolemma. Akt introduction after onset of pathology reverses the mdx histopathological measures, including decreases in blood serum albumin infiltration. Akt also improves muscle function in mdx mice as demonstrated through in vivo grip strength tests and in vitro contraction measurements of the extensor digitorum longus muscle. To further explore the significance of Akt in myofiber regeneration, we injured wild-type muscle with cardiotoxin and found that Akt induced a faster regenerative response relative to controls at equivalent time points. We demonstrate that Akt signaling pathways counteract mdx pathogenesis by enhancing endogenous compensatory mechanisms. These findings provide a rationale for investigating the therapeutic activation of the Akt pathway to counteract muscle wasting. PMID:21245083

  5. Muscle Interstitial Cells: A Brief Field Guide to Non-satellite Cell Populations in Skeletal Muscle.

    PubMed

    Tedesco, Francesco Saverio; Moyle, Louise A; Perdiguero, Eusebio

    2017-01-01

    Skeletal muscle regeneration is mainly enabled by a population of adult stem cells known as satellite cells. Satellite cells have been shown to be indispensable for adult skeletal muscle repair and regeneration. In the last two decades, other stem/progenitor cell populations resident in the skeletal muscle interstitium have been identified as "collaborators" of satellite cells during regeneration. They also appear to have a key role in replacing skeletal muscle with adipose, fibrous, or bone tissue in pathological conditions. Here, we review the role and known functions of these different interstitial skeletal muscle cell types and discuss their role in skeletal muscle tissue homeostasis, regeneration, and disease, including their therapeutic potential for cell transplantation protocols.

  6. GaAs 904-nm laser irradiation improves myofiber mass recovery during regeneration of skeletal muscle previously damaged by crotoxin.

    PubMed

    Silva, Lucila H; Silva, Meiricris T; Gutierrez, Rita M; Conte, Talita C; Toledo, Cláudio A; Aoki, Marcelo S; Liebano, Richard E; Miyabara, Elen H

    2012-09-01

    This work investigated the effect of gallium arsenide (GaAs) irradiation (power: 5 mW; intensity: 77.14 mW/cm(2), spot: 0.07 cm(2)) on regenerating skeletal muscles damaged by crotoxin (CTX). Male C57Bl6 mice were divided into six groups (n = 5 each): control, treated only with laser at doses of 1.5 J or 3 J, CTX-injured and, CTX-injured and treated with laser at doses of 1.5 J or 3 J. The injured groups received a CTX injection into the tibialis anterior (TA) muscle. After 3 days, TA muscles were submitted to GaAs irradiation at doses of 1.5 or 3 J (once a day, during 5 days) and were killed on the eighth day. Muscle histological sections were stained with hematoxylin and eosin (H&E) in order to determine the myofiber cross-sectional area (CSA), the previously injured muscle area (PIMA) and the area density of connective tissue. The gene expression of MyoD and myogenin was detected by real-time PCR. GaAs laser at a dose of 3 J, but not 1.5 J, significantly increased the CSA of regenerating myofibers and reduced the PIMA and the area density of intramuscular connective tissue of CTX-injured muscles. MyoD gene expression increased in the injured group treated with GaAs laser at a dose of 1.5 J. The CTX-injured, 3-J GaAs laser-treated, and the CTX-injured and treated with 3-J laser groups showed an increase in myogenin gene expression when compared to the control group. Our results suggest that GaAs laser treatment at a dose of 3 J improves skeletal muscle regeneration by accelerating the recovery of myofiber mass.

  7. Production of transgenic adult plants from clementine mandarin by enhancing cell competence for transformation and regeneration.

    PubMed

    Cervera, Magdalena; Navarro, Antonio; Navarro, Luis; Peña, Leandro

    2008-01-01

    Genetic transformation of mature trees is difficult because adult tissues are recalcitrant to Agrobacterium tumefaciens infection and transformation and because transgenic mature events are less competent for regeneration. We have shown that reinvigoration allows manipulation of the vegetative phase to increase the potential for transformation and regeneration without loss of competence for flowering and fruiting. To produce transgenic plants from clementine mandarin (Citrus clementina hort. ex Tanaka), we optimized the conditions of the source material both ex vitro and in vitro. Grafting of mature buds on juvenile rootstocks in the spring and preventing multiple bud sprouting by removing all but one bud permitted selection of vigorous first flushes for in vitro culture. Use of additional virulence genes from A. tumefaciens to increase transformation frequency and optimization of culture media and conditions to enhance explant cell competence for T-DNA integration and organogenesis resulted in efficient and reliable transgenic plant production. Transformed regenerants from explants, cultured in media without antibiotics, were identified by a screenable marker (either beta-glucuronidase or green fluorescent protein (GFP)), creating the possibility of generating transgenic clementine plants without antibiotic resistance marker genes. Stable integration of foreign genes was demonstrated by Southern blot analysis, and expression of these foreign genes was confirmed by detection of GFP fluorescence in leaves, floral organs and fruits of the transgenic plants.

  8. Axon-glial relations during regeneration of axons in the adult rat anterior medullary velum.

    PubMed

    Berry, M; Hunter, A S; Duncan, A; Lordan, J; Kirvell, S; Tsang, W L; Butt, A M

    1998-12-01

    The anterior medullary velum (AMV) of adult Wistar rats was lesioned in the midsagittal plane, transecting all decussating axons including those of the central projection of the IVth nerve. At selected times up to 200 days after transection, the degenerative and regenerative responses of axons and glia were analyzed using transmission and scanning electron microscopy and immunohistochemistry. In particular, both the capacity of oligodendrocytes to remyelinate regenerated fibers and the stability of the CNS/PNS junctional zone of the IVth nerve rootlet were documented. Transected central AMV axons exhibited four patterns of fiber regeneration in which fibers grew: rostrocaudally in the reactive paralesion neuropil (Group 1); randomly within the AMV (Group 2); into the ipsilateral IVth nerve rootlet, after turning at the lesion edge and growing recurrently through the old degenerated contralateral central trochlear nerve trajectory (Group 3); and ectopically through paralesion tears in the ependyma onto the surface of the IVth ventricle (Group 4). Group 1-3 axons regenerated unperturbed through degenerating central myelin, reactive astrocytes, oligodendrocytes, microglia, and large accumulations of hematogenous macrophages. Only Group 3 axons survived long term in significant numbers, and all became myelinated by oligodendrocytes, ultimately establishing thin sheaths with relatively normal nodal gaps and intersegmental myelin sheath lengths. Schwann cells at the CNS/PNS junction of the IVth nerve rootlet did not invade the CNS, but astrocyte processes grew across the junction into the PNS portion of the IVth nerve. The basal lamina of the junctional glia limitans remained stable throughout the experimental period.

  9. Effect of vascular endothelial growth factor gene therapy on post-traumatic peripheral nerve regeneration and denervation-related muscle atrophy.

    PubMed

    Moimas, S; Novati, F; Ronchi, G; Zacchigna, S; Fregnan, F; Zentilin, L; Papa, G; Giacca, M; Geuna, S; Perroteau, I; Arnež, Z M; Raimondo, S

    2013-10-01

    Functional recovery after peripheral nerve injury depends on both improvement of nerve regeneration and prevention of denervation-related skeletal muscle atrophy. To reach these goals, in this study we overexpressed vascular endothelial growth factor (VEGF) by means of local gene transfer with adeno-associated virus (AAV). Local gene transfer in the regenerating peripheral nerve was obtained by reconstructing a 1-cm-long rat median nerve defect using a vein segment filled with skeletal muscle fibers that have been previously injected with either AAV2-VEGF or AAV2-LacZ, and the morphofunctional outcome of nerve regeneration was assessed 3 months after surgery. Surprisingly, results showed that overexpression of VEGF in the muscle-vein-combined guide led to a worse nerve regeneration in comparison with AAV-LacZ controls. Local gene transfer in the denervated muscle was obtained by direct injection of either AAV2-VEGF or AAV2-LacZ in the flexor digitorum sublimis muscle after median nerve transection and results showed a significantly lower progression of muscle atrophy in AAV2-VEGF-treated muscles in comparison with muscles treated with AAV2-LacZ. Altogether, our results suggest that local delivery of VEGF by AAV2-VEGF-injected transplanted muscle fibers do not represent a rational approach to promote axonal regeneration along a venous nerve guide. By contrast, AAV2-VEGF direct local injection in denervated skeletal muscle significantly attenuates denervation-related atrophy, thus representing a promising strategy for improving the outcome of post-traumatic neuromuscular recovery after nerve injury and repair.

  10. FOXP3+ T Cells Recruited to Sites of Sterile Skeletal Muscle Injury Regulate the Fate of Satellite Cells and Guide Effective Tissue Regeneration

    PubMed Central

    Castiglioni, Alessandra; Basso, Veronica; Vezzoli, Michela; Monno, Antonella; Almada, Albert E.; Mondino, Anna; Wagers, Amy J.; Manfredi, Angelo A.; Rovere-Querini, Patrizia

    2015-01-01

    Muscle injury induces a classical inflammatory response in which cells of the innate immune system rapidly invade the tissue. Macrophages are prominently involved in this response and required for proper healing, as they are known to be important for clearing cellular debris and supporting satellite cell differentiation. Here, we sought to assess the role of the adaptive immune system in muscle regeneration after acute damage. We show that T lymphocytes are transiently recruited into the muscle after damage and appear to exert a pro-myogenic effect on muscle repair. We observed a decrease in the cross-sectional area of regenerating myofibers after injury in Rag2-/- γ-chain-/- mice, as compared to WT controls, suggesting that T cell recruitment promotes muscle regeneration. Skeletal muscle infiltrating T lymphocytes were enriched in CD4+CD25+FOXP3+ cells. Direct exposure of muscle satellite cells to in vitro induced Treg cells effectively enhanced their expansion, and concurrently inhibited their myogenic differentiation. In vivo, the recruitment of Tregs to acutely injured muscle was limited to the time period of satellite expansion, with possibly important implications for situations in which inflammatory conditions persist, such as muscular dystrophies and inflammatory myopathies. We conclude that the adaptive immune system, in particular T regulatory cells, is critically involved in effective skeletal muscle regeneration. Thus, in addition to their well-established role as regulators of the immune/inflammatory response, T regulatory cells also regulate the activity of skeletal muscle precursor cells, and are instrumental for the proper regeneration of this tissue. PMID:26039259

  11. FOXP3+ T Cells Recruited to Sites of Sterile Skeletal Muscle Injury Regulate the Fate of Satellite Cells and Guide Effective Tissue Regeneration.

    PubMed

    Castiglioni, Alessandra; Corna, Gianfranca; Rigamonti, Elena; Basso, Veronica; Vezzoli, Michela; Monno, Antonella; Almada, Albert E; Mondino, Anna; Wagers, Amy J; Manfredi, Angelo A; Rovere-Querini, Patrizia

    2015-01-01

    Muscle injury induces a classical inflammatory response in which cells of the innate immune system rapidly invade the tissue. Macrophages are prominently involved in this response and required for proper healing, as they are known to be important for clearing cellular debris and supporting satellite cell differentiation. Here, we sought to assess the role of the adaptive immune system in muscle regeneration after acute damage. We show that T lymphocytes are transiently recruited into the muscle after damage and appear to exert a pro-myogenic effect on muscle repair. We observed a decrease in the cross-sectional area of regenerating myofibers after injury in Rag2-/- γ-chain-/- mice, as compared to WT controls, suggesting that T cell recruitment promotes muscle regeneration. Skeletal muscle infiltrating T lymphocytes were enriched in CD4+CD25+FOXP3+ cells. Direct exposure of muscle satellite cells to in vitro induced Treg cells effectively enhanced their expansion, and concurrently inhibited their myogenic differentiation. In vivo, the recruitment of Tregs to acutely injured muscle was limited to the time period of satellite expansion, with possibly important implications for situations in which inflammatory conditions persist, such as muscular dystrophies and inflammatory myopathies. We conclude that the adaptive immune system, in particular T regulatory cells, is critically involved in effective skeletal muscle regeneration. Thus, in addition to their well-established role as regulators of the immune/inflammatory response, T regulatory cells also regulate the activity of skeletal muscle precursor cells, and are instrumental for the proper regeneration of this tissue.

  12. Functional plasticity of regenerated and intact taste receptors in adult rats unmasked by dietary sodium restriction.

    PubMed

    Hill, D L; Phillips, L M

    1994-05-01

    Unilateral chorda tympani nerve sectioning was combined with institution of a sodium-restricted diet in adult rats to determine the role that environment has on the functional properties of regenerating taste receptor cells. Rats receiving chorda tympani sectioning but no dietary manipulation (cut controls) and rats receiving only the dietary manipulation (diet controls) had normal responses to a concentration series of NaCl, sodium acetate (NaAc), and NH4Cl. However, responses from the regenerated nerve in NaCl-restricted rats (40-120 d postsectioning) to NaCl and NaAc were reduced by as much as 30% compared to controls, indicating that regenerating taste receptors are influenced by environmental (dietary) factors. Responses to NH4Cl were normal; therefore, the effect appears specific to sodium salts. Surprisingly, in the same rats, NaCl responses from the contralateral, intact chorda tympani were up to 40% greater than controls. Thus, in the same rat, there was over a twofold difference in sodium responses between the right and left chorda tympani nerves. A study of the time course of the functional alterations in the intact nerve revealed that responses to NaCl were extremely low immediately following sectioning (about 20% of the normal response), and then increased monotonically during the following 50 d until relative response magnitudes became supersensitive. This function occurred even when the cut chorda tympani was prevented from reinnervating lingual epithelia, demonstrating that events related to regeneration do not play a role in the functional properties of the contralateral side of the tongue.(ABSTRACT TRUNCATED AT 250 WORDS)

  13. Embryonic origin of adult stem cells required for tissue homeostasis and regeneration.

    PubMed

    Davies, Erin L; Lei, Kai; Seidel, Christopher W; Kroesen, Amanda E; McKinney, Sean A; Guo, Longhua; Robb, Sofia Mc; Ross, Eric J; Gotting, Kirsten; Alvarado, Alejandro Sánchez

    2017-01-10

    Planarian neoblasts are pluripotent, adult somatic stem cells and lineage-primed progenitors that are required for the production and maintenance of all differentiated cell types, including the germline. Neoblasts, originally defined as undifferentiated cells residing in the adult parenchyma, are frequently compared to embryonic stem cells yet their developmental origin remains obscure. We investigated the provenance of neoblasts during Schmidtea mediterranea embryogenesis, and report that neoblasts arise from an anarchic, cycling piwi-1+ population wholly responsible for production of all temporary and definitive organs during embryogenesis. Early embryonic piwi-1+ cells are molecularly and functionally distinct from neoblasts: they express unique cohorts of early embryo enriched transcripts and behave differently than neoblasts in cell transplantation assays. Neoblast lineages arise as organogenesis begins and are required for construction of all major organ systems during embryogenesis. These subpopulations are continuously generated during adulthood, where they act as agents of tissue homeostasis and regeneration.

  14. The Time Course Effects of Electroacupuncture on Promoting Skeletal Muscle Regeneration and Inhibiting Excessive Fibrosis after Contusion in Rabbits

    PubMed Central

    Wang, Rongguo; Luo, Dan; Xiao, Cheng; Lin, Peng; Liu, Shouyao; Xu, Qianwei; Wang, Yunting

    2013-01-01

    The aim of this study was to investigate the longitudinal effects of electroacupuncture (EA) on Zusanli (ST36) and Ashi acupoints in promoting skeletal muscle regeneration and inhibiting excessive fibrosis after contusion in rabbits. Sixty rabbits were randomly divided into four groups: normal, contusion, EA, and recombinant human insulin-like growth factor-I (rhIGF-I). An acute skeletal muscle contusion was produced on the right gastrocnemius (GM) by an instrument-based drop-mass technique. EA was performed for 15 minutes every two days with 0.4 mA (2 Hz), and GM injections were executed with rhIGF-I (0.25 mL once a week). Rabbits treated with EA had a higher T-SOD and T-AOC serum activities and lower MDA serum level, the blood perfusion of which was also significantly higher. In the EA group, the diameter of the myofibril was uniform and the arrangement was regular, contrary to the contusion group. The number and diameter of regenerative myofibers and MHC expression were increased in the EA group. EA treatment significantly decreased fibrosis formation and reduced both GDF-8 and p-Smad2/3 expressions in injured muscle. Our data indicate that EA may promote myofiber regeneration and reduce excessive fibrosis by improving blood flow and antioxidant capacities. Additionally, EA may regulate signaling factor expression after contusion. PMID:23990848

  15. Chronic hypothyroidism only marginally affects adult-type Leydig cell regeneration after EDS administration.

    PubMed

    Rijntjes, Eddy; van Kesteren-Buiting, Anita; Keijer, Jaap; Teerds, Katja J

    2010-02-01

    Chronic prenatally induced dietary hypothyroidism delays adult-type Leydig cell development, but does not block this process. Using a chemical model to induce hypothyroidism, it was suggested that development of a new population of Leydig cells was completely inhibited following the addition of the cytotoxic compound ethane-1,2-dimethyl sulphonate (EDS). In this study, we used a dietary approach to induce hypothyroidism and reinvestigated the regeneration of the Leydig cell population following EDS administration. Eighty-four day old euthyroid and chronically hypothyroid rats received an injection of EDS and were killed directly before or at regular intervals up to 77 days after EDS. In some control and hypothyroid animals, the first progenitor-type Leydig cells were observed at day 12 after EDS. At day 16, Leydig cell progenitors were present in all rats. The percentage of proliferating Leydig cells peaked in the euthyroid animals at day 21 after EDS. In the hypothyroid testis such a peak was not observed, although the percentage of proliferating regenerating Leydig cells was significantly higher from days 35 to 56 compared with the controls. This suggested that the wave of Leydig cell proliferation was delayed in the hypothyroid animals as compared with the euthyroid controls. On the day of EDS injection, the Leydig/Sertoli cell ratio was 37% lower in the hypothyroid rats compared with the controls. The Leydig/Sertoli cell ratio remained lower in the EDS-treated hypothyroid animals compared with the controls at all time points investigated. At day 77 after EDS, the Leydig cell population had returned to its pre-treatment size in both groups. Plasma testosterone production was reduced to below detectable levels immediately after EDS injection, and started to increase again on day 16, reaching pre-treatment values on day 21 in both groups. Taken together, severely reduced thyroid hormone levels did not block the regeneration of the adult-type Leydig cell population

  16. Constitutive Expression of Yes-Associated Protein (Yap) in Adult Skeletal Muscle Fibres Induces Muscle Atrophy and Myopathy

    PubMed Central

    Judson, Robert N.; Gray, Stuart R.; Walker, Claire; Carroll, Andrew M.; Itzstein, Cecile; Lionikas, Arimantas; Zammit, Peter S.; De Bari, Cosimo; Wackerhage, Henning

    2013-01-01

    The aim of this study was to investigate the function of the Hippo pathway member Yes-associated protein (Yap, gene name Yap1) in skeletal muscle fibres in vivo. Specifically we bred an inducible, skeletal muscle fibre-specific knock-in mouse model (MCK-tTA-hYAP1 S127A) to test whether the over expression of constitutively active Yap (hYAP1 S127A) is sufficient to drive muscle hypertrophy or stimulate changes in fibre type composition. Unexpectedly, after 5–7 weeks of constitutive hYAP1 S127A over expression, mice suddenly and rapidly lost 20–25% body weight and suffered from gait impairments and kyphosis. Skeletal muscles atrophied by 34–40% and the muscle fibre cross sectional area decreased by ≈40% when compared to control mice. Histological analysis revealed evidence of skeletal muscle degeneration and regeneration, necrotic fibres and a NADH-TR staining resembling centronuclear myopathy. In agreement with the histology, mRNA expression of markers of regenerative myogenesis (embryonic myosin heavy chain, Myf5, myogenin, Pax7) and muscle protein degradation (atrogin-1, MuRF1) were significantly elevated in muscles from transgenic mice versus control. No significant changes in fibre type composition were detected using ATPase staining. The phenotype was largely reversible, as a cessation of hYAP1 S127A expression rescued body and muscle weight, restored muscle morphology and prevented further pathological progression. To conclude, high Yap activity in muscle fibres does not induce fibre hypertrophy nor fibre type changes but instead results in a reversible atrophy and deterioration. PMID:23544078

  17. Effects of Estradiol and Methoxychlor on Leydig Cell Regeneration in the Adult Rat Testis

    PubMed Central

    Chen, Bingbing; Chen, Dongxin; Jiang, Zheli; Li, Jingyang; Liu, Shiwen; Dong, Yaoyao; Yao, Wenwen; Akingbemi, Benson; Ge, Renshan; Li, Xiaokun

    2014-01-01

    The objective of the present study is to determine whether methoxychlor (MXC) exposure in adulthood affects rat Leydig cell regeneration and to compare its effects with estradiol (E2). Adult 90-day-old male Sprague-Dawley rats received ethane dimethane sulfonate (EDS) to eliminate the adult Leydig cell population. Subsequently, rats were randomly assigned to four groups and gavaged with corn oil (control), 0.25 mg/kg E2 and 10 or 100 mg/kg MXC daily from days 5 to 30 post-EDS treatment. The results showed that MXC and E2 reduced serum testosterone levels on day 58 post-EDS treatment. qPCR showed Hsd17b3 mRNA levels were downregulated 7–15 fold by E2 and MXC, indicating that development of the new population of Leydig cells was arrested at the earlier stage. This observation was supported by the results of histochemical staining, which demonstrated that Leydig cells in MXC-treated testis on day 58 post-EDS treatment were mostly progenitor Leydig cells. However, Pdgfb mRNA levels were downregulated, while Lif transcript levels were increased by MXC. In contrast, E2 did not affect gene expression for these growth factors. In conclusion, our findings indicated that both MXC and E2 delayed rat Leydig cell regeneration in the EDS-treated model, presumably acting by different mechanisms. PMID:24806340

  18. Mending broken hearts: cardiac development as a basis for adult heart regeneration and repair

    PubMed Central

    Xin, Mei; Olson, Eric N.; Bassel-Duby, Rhonda

    2013-01-01

    As the adult mammalian heart has limited potential for regeneration and repair, the loss of cardiomyocytes during injury and disease can result in heart failure and death. The cellular processes and regulatory mechanisms involved in heart growth and development can be exploited to repair the injured adult heart through ‘reawakening’ pathways that are active during embryogenesis. Heart function has been restored in rodents by reprogramming non-myocytes into cardiomyocytes, by expressing transcription factors (GATA4, HAND2, myocyte-specific enhancer factor 2C (MEF2C) and T-box 5 (TBX5)) and microRNAs (miR-1, miR-133, miR-208 and miR-499) that control cardiomyocyte identity. Stimulating cardiomyocyte dedifferentiation and proliferation by activating mitotic signalling pathways involved in embryonic heart growth represents a complementary approach for heart regeneration and repair. Recent advances in understanding the mechanistic basis of heart development offer exciting opportunities for effective therapies for heart failure. PMID:23839576

  19. TEAD transcription factors are required for normal primary myoblast differentiation in vitro and muscle regeneration in vivo

    PubMed Central

    Joshi, Shilpy; Le Gras, Stéphanie; Watanabe, Shuichi; Braun, Thomas

    2017-01-01

    The TEAD family of transcription factors (TEAD1-4) bind the MCAT element in the regulatory elements of both growth promoting and myogenic differentiation genes. Defining TEAD transcription factor function in myogenesis has proved elusive due to overlapping expression of family members and their functional redundancy. We show that silencing of either Tead1, Tead2 or Tead4 did not effect primary myoblast (PM) differentiation, but that their simultaneous knockdown strongly impaired differentiation. In contrast, Tead1 or Tead4 silencing impaired C2C12 differentiation showing their different contributions in PMs and C2C12 cells. Chromatin immunoprecipitation identified enhancers associated with myogenic genes bound by combinations of Tead4, Myod1 or Myog. Tead4 regulated distinct gene sets in C2C12 cells and PMs involving both activation of the myogenic program and repression of growth and signaling pathways. ChIP-seq from mature mouse muscle fibres in vivo identified a set of highly transcribed muscle cell-identity genes and sites bound by Tead1 and Tead4. Although inactivation of Tead4 in mature muscle fibres caused no obvious phenotype under normal conditions, notexin-induced muscle regeneration was delayed in Tead4 mutants suggesting an important role in myogenic differentiation in vivo. By combining knockdown in cell models in vitro with Tead4 inactivation in muscle in vivo, we provide the first comprehensive description of the specific and redundant roles of Tead factors in myogenic differentiation. PMID:28178271

  20. The circadian clock in skin: implications for adult stem cells, tissue regeneration, cancer, aging, and immunity

    PubMed Central

    Plikus, Maksim V.; Van Spyk, Elyse Noelani; Pham, Kim; Geyfman, Mikhail; Kumar, Vivek; Takahashi, Joseph S.; Andersen, Bogi

    2015-01-01

    Historically work on peripheral circadian clocks has been focused on organs and tissues that have prominent metabolic functions, such as liver, fat and muscle. In recent years, skin is emerging as a model for studying circadian clock regulation of cell proliferation, stem cell functions, tissue regeneration, aging and carcinogenesis. Morphologically skin is complex, containing multiple cell types and structures, and there is evidence for a functional circadian clock in most, if not all, of its cell types. Despite the complexity, skin stem cell populations are well defined, experimentally tractable and exhibit prominent daily cell proliferation cycles. Hair follicle stem cells also participate in recurrent, long-lasting cycles of regeneration -- the hair growth cycles. Among other advantages of skin is a broad repertoire of available genetic tools enabling the creation of cell-type specific circadian mutants. Also, due to the accessibility of the skin, in vivo imaging techniques can be readily applied to study the circadian clock and its outputs in real time, even at the single-cell level. Skin provides the first line of defense against many environmental and stress factors that exhibit dramatic diurnal variations such as solar UV radiation and temperature. Studies have already linked the circadian clock to the control of UVB-induced DNA damage and skin cancers. Due to the important role that skin plays in the defense against microorganisms, it represents a promising model system to further explore the role of the clock in the regulation of the body's immune functions. To that end, recent studies have already linked the circadian clock to psoriasis, one of the most common immune-mediated skin disorders. The skin also provides opportunities to interrogate clock regulation of tissue metabolism in the context of stem cells and regeneration. Furthermore, many animal species feature prominent seasonal hair molt cycles, offering an attractive model for investigating the

  1. The circadian clock in skin: implications for adult stem cells, tissue regeneration, cancer, aging, and immunity.

    PubMed

    Plikus, Maksim V; Van Spyk, Elyse N; Pham, Kim; Geyfman, Mikhail; Kumar, Vivek; Takahashi, Joseph S; Andersen, Bogi

    2015-06-01

    Historically, work on peripheral circadian clocks has been focused on organs and tissues that have prominent metabolic functions, such as the liver, fat, and muscle. In recent years, skin has emerged as a model for studying circadian clock regulation of cell proliferation, stem cell functions, tissue regeneration, aging, and carcinogenesis. Morphologically, skin is complex, containing multiple cell types and structures, and there is evidence for a functional circadian clock in most, if not all, of its cell types. Despite the complexity, skin stem cell populations are well defined, experimentally tractable, and exhibit prominent daily cell proliferation cycles. Hair follicle stem cells also participate in recurrent, long-lasting cycles of regeneration: the hair growth cycles. Among other advantages of skin is a broad repertoire of available genetic tools enabling the creation of cell type-specific circadian mutants. Also, due to the accessibility of skin, in vivo imaging techniques can be readily applied to study the circadian clock and its outputs in real time, even at the single-cell level. Skin provides the first line of defense against many environmental and stress factors that exhibit dramatic diurnal variations such as solar ultraviolet (UV) radiation and temperature. Studies have already linked the circadian clock to the control of UVB-induced DNA damage and skin cancers. Due to the important role that skin plays in the defense against microorganisms, it also represents a promising model system to further explore the role of the clock in the regulation of the body's immune functions. To that end, recent studies have already linked the circadian clock to psoriasis, one of the most common immune-mediated skin disorders. Skin also provides opportunities to interrogate the clock regulation of tissue metabolism in the context of stem cells and regeneration. Furthermore, many animal species feature prominent seasonal hair molt cycles, offering an attractive model

  2. Effects of neutrophil depletion in the local pathological alterations and muscle regeneration in mice injected with Bothrops jararaca snake venom

    PubMed Central

    Teixeira, Catarina F P; Chaves, Fernando; Zamunér, Stella R; Fernandes, Cristina M; Zuliani, Juliana P; Cruz-Hofling, María Alice; Fernandes, Irene; Gutiérrez, José María

    2005-01-01

    In order to study the role of neutrophils in the acute local pathological alterations induced by Bothrops jararaca snake venom, and in the process of skeletal muscle regeneration that follows, an experimental model was developed in mice pretreated with either an anti-mouse granulocyte rat monoclonal immunoglobulin G, which induces a profound neutropenia, or an isotype-matched control antibody. B. jararaca venom induced prominent haemorrhage and oedema, but only a moderate myonecrosis. No significant differences were observed in the extent of local haemorrhage, oedema and myonecrosis between neutropenic and control mice, suggesting that neutrophils do not play a determinant role in the acute pathological alterations induced by B. jararaca venom in this experimental model. Moreover, no differences were observed in skeletal muscle regeneration between these two experimental groups. In both the cases, limited areas of myonecrosis were associated with a drastic damage to the microvasculature and a scarce inflammatory infiltrate, with the consequent lack of removal of necrotic debris during the first week, resulting in a poor regenerative response at this time interval. Subsequently, a similar regenerative process occurred in both groups, and by 30 days, necrotic areas were substituted by groups of small regenerating muscle fibres. It is suggested that the drastic effect exerted by B. jararaca venom in the microvasculature precludes an effective access of inflammatory cells to necrotic areas, thereby compromising an effective removal of necrotic debris; this explains the poor regenerative response observed during the first week and the fact that there were no differences between neutropenic and control mice. As neutropenia in this model lasted only 7 days, the successful regenerative process observed at 30 days is associated with revascularization of necrotic regions and with a successful removal by phagocytes of necrotic debris in both groups. PMID:15810982

  3. Regeneration of organs and tissues in lower vertebrates during and after space flight

    NASA Astrophysics Data System (ADS)

    Mitashov, V. I.; Brushlinskaya, N. V.; Grigoryan, E. N.; Tuchkova, S. Ya.; Anton, H. J.

    In this paper most important data obtained in studies on the effect of space flight conditions on regeneration in the adult newt are summarized. We demonstrate a phenomenon of synchronization of limb and lens regeneration and increase in its rate during and after space flight. We also describe a peculiarities of cell proliferation in lens, limb and tail regenerates and of the process of minced muscle regeneration.

  4. Small Fractions of Muscular Dystrophy Embryonic Stem Cells Yield Severe Cardiac and Skeletal Muscle Defects in Adult Mouse Chimeras.

    PubMed

    Gonzalez, J Patrick; Kyrychenko, Sergii; Kyrychenko, Viktoriia; Schneider, Joel S; Granier, Celine J; Himelman, Eric; Lahey, Kevin C; Zhao, Qingshi; Yehia, Ghassan; Tao, Yuan-Xiang; Bhaumik, Mantu; Shirokova, Natalia; Fraidenraich, Diego

    2017-03-01

    Duchenne muscular dystrophy (DMD) is characterized by the loss of the protein dystrophin, leading to muscle fragility, progressive weakening, and susceptibility to mechanical stress. Although dystrophin-negative mdx mouse models have classically been used to study DMD, phenotypes appear mild compared to patients. As a result, characterization of muscle pathology, especially in the heart, has proven difficult. We report that injection of mdx embryonic stem cells (ESCs) into Wild Type blastocysts produces adult mouse chimeras with severe DMD phenotypes in the heart and skeletal muscle. Inflammation, regeneration and fibrosis are observed at the whole organ level, both in dystrophin-negative and dystrophin-positive portions of the chimeric tissues. Skeletal and cardiac muscle function are also decreased to mdx levels. In contrast to mdx heterozygous carriers, which show no significant phenotypes, these effects are even observed in chimeras with low levels of mdx ESC incorporation (10%-30%). Chimeric mice lack typical compensatory utrophin upregulation, and show pathological remodeling of Connexin-43. In addition, dystrophin-negative and dystrophin-positive isolated cardiomyocytes show augmented calcium response to mechanical stress, similar to mdx cells. These global effects highlight a novel role of mdx ESCs in triggering muscular dystrophy even when only low amounts are present. Stem Cells 2017;35:597-610.

  5. Mechanisms of Cardiac Regeneration

    PubMed Central

    Uygur, Aysu; Lee, Richard T.

    2016-01-01

    Adult humans fail to regenerate their hearts following injury, and this failure to regenerate myocardium is a leading cause of heart failure and death worldwide. Although all adult mammals appear to lack significant cardiac regeneration potential, some vertebrates can regenerate myocardium throughout life. In addition, new studies indicate that mammals have cardiac regeneration potential during development and very soon after birth. The mechanisms of heart regeneration among model organisms, including neonatal mice, appear remarkably similar. Orchestrated waves of inflammation, matrix deposition and remodeling, and cardiomyocyte proliferation are commonly seen in heart regeneration models. Understanding why adult mammals develop extensive scarring instead of regeneration is a crucial goal for regenerative biology. PMID:26906733

  6. Lentiviral-mediated transfer of CDNF promotes nerve regeneration and functional recovery after sciatic nerve injury in adult rats

    SciTech Connect

    Cheng, Lei; Liu, Yi; Zhao, Hua; Zhang, Wen; Guo, Ying-Jun; Nie, Lin

    2013-10-18

    Highlights: •CDNF was successfully transfected by a lentiviral vector into the distal sciatic nerve. •CDNF improved S-100, NF200 expression and nerve regeneration after sciatic injury. •CDNF improved the remyelination and thickness of the regenerated sciatic nerve. •CDNF improved gastrocnemius muscle weight and sciatic functional recovery. -- Abstract: Peripheral nerve injury is often followed by incomplete and unsatisfactory functional recovery and may be associated with sensory and motor impairment of the affected limb. Therefore, a novel method is needed to improve the speed of recovery and the final functional outcome after peripheral nerve injuries. This report investigates the effect of lentiviral-mediated transfer of conserved dopamine neurotrophic factor (CDNF) on regeneration of the rat peripheral nerve in a transection model in vivo. We observed notable overexpression of CDNF protein in the distal sciatic nerve after recombinant CDNF lentiviral vector application. We evaluated sciatic nerve regeneration after surgery using light and electron microscopy and the functional recovery using the sciatic functional index and target muscle weight. HE staining revealed better ordered structured in the CDNF-treated group at 8 weeks post-surgery. Quantitative analysis of immunohistochemistry of NF200 and S-100 in the CDNF group revealed significant improvement of axonal and Schwann cell regeneration compared with the control groups at 4 weeks and 8 weeks after injury. The thickness of the myelination around the axons in the CDNF group was significantly higher than in the control groups at 8 weeks post-surgery. The CDNF group displayed higher muscle weights and significantly increased sciatic nerve index values. Our findings suggest that CDNF gene therapy could provide durable and stable CDNF protein concentration and has the potential to enhance peripheral nerve regeneration, morphological and functional recovery following nerve injury, which suggests a

  7. Asynchronous Inflammation and Myogenic Cell Migration Limit Muscle Tissue Regeneration Mediated by a Cellular Scaffolds

    DTIC Science & Technology

    2015-02-11

    muscle with volumetric muscle loss injury following the transplantation of muscle-ECM. Biomaterials 2013; 34:3324-3335. 4. Mase VJ, Jr., Hsu JR, Wolf... Biomaterials 2009; 30:1482-1491. 9. Wolf MT, Dearth CL, Sonnenberg SB, Loboa EGBadylak SF. Naturally derived and synthetic scaffolds for skeletal muscle...labeled porcine dermis biologic scaffold. Biomaterials 2014; 35:8297-8304. 11. Gilbert TW, Stewart-Akers AMBadylak SF. A quantitative method for

  8. Effects of Human Mesenchymal Stem Cells Isolated from Wharton's Jelly of the Umbilical Cord and Conditioned Media on Skeletal Muscle Regeneration Using a Myectomy Model

    PubMed Central

    Pereira, T.; Armada-da Silva, P. A. S.; Amorim, I.; Rêma, A.; Caseiro, A. R.; Gärtner, A.; Rodrigues, M.; Lopes, M. A.; Bártolo, P. J.; Santos, J. D.; Luís, A. L.; Maurício, A. C.

    2014-01-01

    Skeletal muscle has good regenerative capacity, but the extent of muscle injury and the developed fibrosis might prevent complete regeneration. The in vivo application of human mesenchymal stem cells (HMSCs) of the umbilical cord and the conditioned media (CM) where the HMSCs were cultured and expanded, associated with different vehicles to induce muscle regeneration, was evaluated in a rat myectomy model. Two commercially available vehicles and a spherical hydrogel developed by our research group were used. The treated groups obtained interesting results in terms of muscle regeneration, both in the histological and in the functional assessments. A less evident scar tissue, demonstrated by collagen type I quantification, was present in the muscles treated with HMSCs or their CM. In terms of the histological evaluation performed by ISO 10993-6 scoring, it was observed that HMSCs apparently have a long-term negative effect, since the groups treated with CM presented better scores. CM could be considered an alternative to the in vivo transplantation of these cells, as it can benefit from the local tissue response to secreted molecules with similar results in terms of muscular regeneration. Searching for an optimal vehicle might be the key point in the future of skeletal muscle tissue engineering. PMID:25379040

  9. Effects of Human Mesenchymal Stem Cells Isolated from Wharton's Jelly of the Umbilical Cord and Conditioned Media on Skeletal Muscle Regeneration Using a Myectomy Model.

    PubMed

    Pereira, T; Armada-da Silva, P A S; Amorim, I; Rêma, A; Caseiro, A R; Gärtner, A; Rodrigues, M; Lopes, M A; Bártolo, P J; Santos, J D; Luís, A L; Maurício, A C

    2014-01-01

    Skeletal muscle has good regenerative capacity, but the extent of muscle injury and the developed fibrosis might prevent complete regeneration. The in vivo application of human mesenchymal stem cells (HMSCs) of the umbilical cord and the conditioned media (CM) where the HMSCs were cultured and expanded, associated with different vehicles to induce muscle regeneration, was evaluated in a rat myectomy model. Two commercially available vehicles and a spherical hydrogel developed by our research group were used. The treated groups obtained interesting results in terms of muscle regeneration, both in the histological and in the functional assessments. A less evident scar tissue, demonstrated by collagen type I quantification, was present in the muscles treated with HMSCs or their CM. In terms of the histological evaluation performed by ISO 10993-6 scoring, it was observed that HMSCs apparently have a long-term negative effect, since the groups treated with CM presented better scores. CM could be considered an alternative to the in vivo transplantation of these cells, as it can benefit from the local tissue response to secreted molecules with similar results in terms of muscular regeneration. Searching for an optimal vehicle might be the key point in the future of skeletal muscle tissue engineering.

  10. Vertebrate-like regeneration in the invertebrate chordate amphioxus

    PubMed Central

    Somorjai, Ildikó M. L.; Garcia-Fernàndez, Jordi; Escrivà, Hector

    2012-01-01

    An important question in biology is why some animals are able to regenerate, whereas others are not. The basal chordate amphioxus is uniquely positioned to address the evolution of regeneration. We report here the high regeneration potential of the European amphioxus Branchiostoma lanceolatum. Adults regenerate both anterior and posterior structures, including neural tube, notochord, fin, and muscle. Development of a classifier based on tail regeneration profiles predicts the assignment of young and old adults to their own class with >94% accuracy. The process involves loss of differentiated characteristics, formation of an msx-expressing blastema, and neurogenesis. Moreover, regeneration is linked to the activation of satellite-like Pax3/7 progenitor cells, the extent of which declines with size and age. Our results provide a framework for understanding the evolution and diversity of regeneration mechanisms in vertebrates. PMID:22203957

  11. Stromal derived factor‐1 and granulocyte‐colony stimulating factor treatment improves regeneration of Pax7−/− mice skeletal muscles

    PubMed Central

    Kowalski, Kamil; Archacki, Rafał; Archacka, Karolina; Stremińska, Władysława; Paciorek, Anna; Gołąbek, Magdalena; Ciemerych, Maria A.

    2015-01-01

    Abstract Background The skeletal muscle has the ability to regenerate after injury. This process is mediated mainly by the muscle specific stem cells, that is, satellite cells. In case of extensive damage or under pathological conditions, such as muscular dystrophy, the process of muscle reconstruction does not occur properly. The aim of our study was to test whether mobilized stem cells, other than satellite cells, could participate in skeletal muscle reconstruction. Methods Experiments were performed on wild‐type mice and mice lacking the functional Pax7 gene, that is, characterized by the very limited satellite cell population. Gastrocnemius mice muscles were injured by cardiotoxin injection, and then the animals were treated by stromal derived factor‐1 (Sdf‐1) with or without granulocyte‐colony stimulating factor (G‐CSF) for 4 days. The muscles were subjected to thorough assessment of the tissue regeneration process using histological and in vitro methods, as well as evaluation of myogenic factors' expression at the transcript and protein levels. Results Stromal derived factor‐1 alone and Sdf‐1 in combination with G‐CSF significantly improved the regeneration of Pax7−/− skeletal muscles. The Sdf‐1 and G‐CSF treatment caused an increase in the number of mononucleated cells associated with muscle fibres. Further analysis showed that Sdf‐1 and G‐CSF treatment led to the rise in the number of CD34+ and Cxcr4+ cells and expression of Cxcr7. Conclusions Stromal derived factor‐1 and G‐CSF stimulated regeneration of the skeletal muscles deficient in satellite cells. We suggest that mobilized CD34+, Cxcr4+, and Cxcr7+ cells can efficiently participate in the skeletal muscle reconstruction and compensate for the lack of satellite cells. PMID:27239402

  12. Conditioned medium derived from umbilical cord mesenchymal stem cells regenerates atrophied muscles.

    PubMed

    Kim, Mi Jin; Kim, Z-Hun; Kim, Sun-Mi; Choi, Yong-Soo

    2016-10-01

    We investigated the regenerative effects and regulatory mechanisms of human umbilical cord mesenchymal stem cells (UC-MSCs)-derived conditioned medium (CM) in atrophied muscles using an in vivo model. To determine the appropriate harvest point of UC-CM, active factor content was analyzed in the secretome over time. A muscle atrophy model was induced in rats by hindlimb suspension (HS) for 2 weeks. Next, UC-CM was injected directly into the soleus muscle of both hind legs to assess its regenerative efficacy on atrophy-related factors after 1 week of HS. During HS, muscle mass and muscle fiber size were significantly reduced by over 2-fold relative to untreated controls. Lactate accumulation within the muscles was similarly increased. By contrast, all of the above analytical factors were significantly improved in HS-induced rats by UC-CM injection compared with saline injection. Furthermore, the expression levels of desmin and skeletal muscle actin were significantly elevated by UC-CM treatment. Importantly, UC-CM effectively suppressed expression of the atrophy-related ubiquitin E3-ligases, muscle ring finger 1 and muscle atrophy F-box by 2.3- and 2.1-fold, respectively. UC-CM exerted its actions by stimulating the phosphoinositol-3-kinase (PI3K)/Akt signaling cascade. These findings suggest that UC-CM provides an effective stimulus to recover muscle status and function in atrophied muscles.

  13. Growth and respiration of regenerating tissues of the axolotl tail.

    PubMed

    Vladimirova, I G

    1975-01-01

    Changes in the weight and oxygen consumption were studied during regeneration of the tail in adult axolotls and larvae. The curve of the increase in weight of the regenerating tail in both age groups is S-shaped. The intensity of respiration of the regenerating tail increases in adult axolotls and in larvae at the blastema stage; in adult axolotls there is also a second increase in the intensity of respiration of the regenerating tail during differentiation of the muscles. The relationship between weight and the rate of respiration was compared during regeneration of the tail in axolotl and the normal growth of the animals. Whereas growth of the animals was characterized by the relationship QO2 equals aPk with a constant value of k, during regeneration the various stages of this process have their own corresponding values of k.

  14. Possibility of leg muscle hypertrophy by ambulation in older adults: a brief review.

    PubMed

    Ozaki, Hayao; Loenneke, Jeremy P; Thiebaud, Robert S; Stager, Joel M; Abe, Takashi

    2013-01-01

    It is known that ambulatory exercises such as brisk walking and jogging are potent stimuli for improving aerobic capacity, but it is less understood whether ambulatory exercise can increase leg muscle size and function. The purpose of this brief review is to discuss whether or not ambulatory exercise elicits leg muscle hypertrophy in older adults. Daily ambulatory activity with moderate (>3 metabolic equivalents [METs], which is defined as the ratio of the work metabolic rate to the resting metabolic rate) intensity estimated by accelerometer is positively correlated with lower body muscle size and function in older adults. Although there is conflicting data on the effects of short-term training, it is possible that relatively long periods of walking, jogging, or intermittent running for over half a year can increase leg muscle size among older adults. In addition, slow-walk training with a combination of leg muscle blood flow restriction elicits muscle hypertrophy only in the blood flow restricted leg muscles. Competitive marathon running and regular high intensity distance running in young and middle-aged adults may not produce leg muscle hypertrophy due to insufficient recovery from the damaging running bout, although there have been no studies that have investigated the effects of running on leg muscle morphology in older subjects. It is clear that skeletal muscle hypertrophy can occur independently of exercise mode and load.

  15. Bone Regeneration Mediated by BMP4-Expressing Muscle-Derived Stem Cells Is Affected by Delivery System

    PubMed Central

    Usas, Arvydas; Ho, Andrew M.; Cooper, Gregory M.; Olshanski, Anne; Peng, Hairong

    2009-01-01

    This study investigated the delivery of bone morphogenetic protein (BMP)4-secreting muscle-derived stem cells (MDSC-B4) capable of inducing bone formation in mice using collagen gel (CG), fibrin sealant (FS), and gelatin sponge carriers. After implanting these various cell-loaded scaffolds intramuscularly or into critical-size skull defects, we measured the extent of heterotopic ossification and calvarial defect healing over a 6-week period via radiographic, radiomorphometric, histological, and micro-computed tomography analyses. As expected, in the absence of MDSC-B4, there was no ectopic ossification and only minimal calvarial regeneration using each type of scaffold. Although CG and gelatin sponges loaded with BMP4-secreting cells produced the most ectopic bone, FS constructs produced bone with comparably less mineralization. In the mouse calvaria, we observed MDSC-B4-loaded scaffolds able to promote bone defect healing to a variable degree, but there were differences between these implants in the volume, shape, and morphology of regenerated bone. MDSC-B4 delivery in a gelatin sponge produced hypertrophic bone, whereas delivery in a CG and FS healed the defect with bone that closely resembled the quantity and configuration of native calvarium. In summary, hydrogels are suitable carriers for osteocompetent MDSCs in promoting bone regeneration, especially at craniofacial injury sites. PMID:19061430

  16. Long-term engraftment of myogenic progenitors from adipose-derived stem cells and muscle regeneration in dystrophic mice.

    PubMed

    Zhang, Yu; Zhu, Yuling; Li, Yaqin; Cao, Jiqing; Zhang, Huili; Chen, Menglong; Wang, Liang; Zhang, Cheng

    2015-11-01

    Stem cell therapy is a promising approach for treating Duchenne muscular dystrophy (DMD); however, its application is hindered by poor cell engraftment. There have been no reports to date describing the efficient generation of myogenic progenitors from adipose-derived stem cells (ADSCs) that can contribute to muscle regeneration. In this study, we examined the in vivo myogenic potential of progenitors differentiated from ADSCs using forskolin, basic fibroblast growth factor, the glycogen synthase kinase 3β inhibitor 6-bromoindirubin-3'-oxime as well as the supernatant of ADSC cultures. The results indicate that a proliferative population of myogenic progenitors can be derived from ADSCs that have characteristics similar to muscle satellite cells and are capable of terminal differentiation into multinucleated myotubes. When transplanted into DMD model mdx mice either by intramuscular injection or systemic delivery, progenitors were successfully engrafted in skeletal muscle for up to 12 weeks, and generated new muscle fibers, restored dystrophin expression and contributed to the satellite cell compartment. These findings highlight the potential application of myogenic progenitors derived from ADSCs to the treatment of muscular dystrophy.

  17. Clearance of Cell Remnants and Regeneration of Injured Muscle Depend on Soluble Pattern Recognition Receptor PTX3

    PubMed Central

    Vezzoli, Michela; Sciorati, Clara; Campana, Lara; Monno, Antonella; Doglio, Maria Giulia; Rigamonti, Elena; Corna, Gianfranca; Touvier, Thierry; Castiglioni, Alessandra; Capobianco, Annalisa; Mantovani, Alberto; Manfredi, Angelo A; Garlanda, Cecilia; Rovere-Querini, Patrizia

    2016-01-01

    The signals causing resolution of muscle inflammation are only partially characterized. The long pentraxin PTX3, which modulates leukocyte recruitment and activation, could contribute. We analyzed the expression of PTX3 after muscle injury and verified whether hematopoietic precursors are a source of the protein. The kinetics of regeneration and leukocyte infiltration and the accumulation of cell remnants and anti-histidyl-t-RNA synthetase autoantibodies were compared in wild-type and PTX3-deficient mice. PTX3 expression was upregulated 3 d to 5 d after injury and restricted to the extracellular matrix. Cellular debris and leukocytes persisted in the muscle of PTX3-deficient mice for a long time after wild-type animals had healed. PTX3-deficient macrophages expressed receptors involved in apoptotic cell clearance and engulfed dead cells in vitro. Accumulation of cell debris in a proinflammatory microenvironment was not sufficient to elicit autoantibodies. We concluded that PTX3 generated in response to muscle injury prompts clearance of debris and termination of the inflammatory response. PMID:27900389

  18. Duchenne Muscular Dystrophy Gene Expression in Normal and Diseased Human Muscle

    NASA Astrophysics Data System (ADS)

    Oronzi Scott, M.; Sylvester, J. E.; Heiman-Patterson, T.; Shi, Y.-J.; Fieles, W.; Stedman, H.; Burghes, A.; Ray, P.; Worton, R.; Fischbeck, K. H.

    1988-03-01

    A probe for the 5' end of the Duchenne muscular dystrophy (DMD) gene was used to study expression of the gene in normal human muscle, myogenic cell cultures, and muscle from patients with DMD. Expression was found in RNA from normal fetal muscle, adult cardiac and skeletal muscle, and cultured muscle after myoblast fusion. In DMD muscle, expression of this portion of the gene was also revealed by in situ RNA hybridization, particularly in regenerating muscle fibers.

  19. Streptomycin ototoxicity and hair cell regeneration in the adult pigeon utricle

    NASA Technical Reports Server (NTRS)

    Frank, T. C.; Dye, B. J.; Newlands, S. D.; Dickman, J. D.

    1999-01-01

    OBJECTIVE: The purpose of this study was to develop a technique to investigate the regeneration of utricular hair cells in the adult pigeon (Columba livia) following complete hair cell loss through administration of streptomycin. STUDY DESIGN: Experimental animal study. METHODS: Animals were divided into four groups. Group 1 received 10 to 15 days of systemic streptomycin injections. Animals in Groups 2 and 3 received a single direct placement of a 1-, 2-, 4-, or 8-mg streptomycin dose into the perilymphatic space. Animals in Groups 1 and 2 were analyzed within 1 week from injection to investigate hair cell destruction, whereas Group 3 was investigated at later dates to study hair cell recovery. Group 4 animals received a control injection of saline into the perilymphatic space. Damage and recovery were quantified by counting hair cells in isolated utricles using scanning electron microscopy. RESULTS: Although systemic injections failed to reliably achieve complete utricular hair cell destruction, a single direct placement of a 2-, 4-, or 8-mg streptomycin dose caused complete destruction within the first week. Incomplete hair cell loss was observed with the 1-mg dose. Over the long term, regeneration of the hair cells was seen with the 2-mg dose but not the 8-mg dose. Control injections of saline into the perilymphatic space caused no measurable hair cell loss. CONCLUSIONS: Direct placement of streptomycin into the perilymph is an effective, reliable method for complete destruction of utricular hair cells while preserving the regenerative potential of the neuroepithelium.

  20. A Novel Light Damage Paradigm for Use in Retinal Regeneration Studies in Adult Zebrafish

    PubMed Central

    Thomas, Jennifer L.; Thummel, Ryan

    2013-01-01

    Light-induced retinal degeneration (LIRD) is commonly used in both rodents and zebrafish to damage rod and cone photoreceptors. In adult zebrafish, photoreceptor degeneration triggers Müller glial cells to re-enter the cell cycle and produce transient-amplifying progenitors. These progenitors continue to proliferate as they migrate to the damaged area, where they ultimately give rise to new photoreceptors. Currently, there are two widely-used LIRD paradigms, each of which results in varying degrees of photoreceptor loss and corresponding differences in the regeneration response. As more genetic and pharmacological tools are available to test the role of individual genes of interest during regeneration, there is a need to develop a robust LIRD paradigm. Here we describe a LIRD protocol that results in widespread and consistent loss of both rod and cone photoreceptors in which we have combined the use of two previously established LIRD techniques. Furthermore, this protocol can be extended for use in pigmented animals, which eliminates the need to maintain transgenic lines of interest on the albino background for LIRD studies. PMID:24192580

  1. DLL4 promotes continuous adult intestinal lacteal regeneration and dietary fat transport

    PubMed Central

    Bernier-Latmani, Jeremiah; Cisarovsky, Christophe; Demir, Cansaran Saygili; Bruand, Marine; Jaquet, Muriel; Davanture, Suzel; Ragusa, Simone; Siegert, Stefanie; Dormond, Olivier; Benedito, Rui; Radtke, Freddy; Luther, Sanjiv A.; Petrova, Tatiana V.

    2015-01-01

    The small intestine is a dynamic and complex organ that is characterized by constant epithelium turnover and crosstalk among various cell types and the microbiota. Lymphatic capillaries of the small intestine, called lacteals, play key roles in dietary fat absorption and the gut immune response; however, little is known about the molecular regulation of lacteal function. Here, we performed a high-resolution analysis of the small intestinal stroma and determined that lacteals reside in a permanent regenerative, proliferative state that is distinct from embryonic lymphangiogenesis or quiescent lymphatic vessels observed in other tissues. We further demonstrated that this continuous regeneration process is mediated by Notch signaling and that the expression of the Notch ligand delta-like 4 (DLL4) in lacteals requires activation of VEGFR3 and VEGFR2. Moreover, genetic inactivation of Dll4 in lymphatic endothelial cells led to lacteal regression and impaired dietary fat uptake. We propose that such a slow lymphatic regeneration mode is necessary to match a unique need of intestinal lymphatic vessels for both continuous maintenance, due to the constant exposure to dietary fat and mechanical strain, and efficient uptake of fat and immune cells. Our work reveals how lymphatic vessel responses are shaped by tissue specialization and uncover a role for continuous DLL4 signaling in the function of adult lymphatic vasculature. PMID:26529256

  2. [The role of the psoas muscle: apropos of the dissection of the muscles from 10 adults and 10 newborn infants].

    PubMed

    Le Floch-Prigent, P

    1983-06-01

    The action of the iliopsoas muscle (Musculus iliopsoas) on movements of the hip is studied by direct traction on fresh cadavers (10 still-born and 10 adults). The psoas muscle is a powerful flexor of the hip but also an external rotator. The action of external rotation is moderate but obvious in every position of the femur (Os femoris) and more important if previously in abduction and internal rotation.

  3. Muscle performance and physical function are associated with voluntary rate of neuromuscular activation in older adults

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Participants were recruited to three experimental groups: middle-aged healthy adults (MH), older healthy adults (OH), and older adults with mobility limitations (OML). OH and OML were primarily differentiated by performance on the Short Physical Performance Battery (SPPB). Muscle performance (accele...

  4. Correlated NOS-Imu and myf5 expression by satellite cells in mdx mouse muscle regeneration during NOS manipulation and deflazacort treatment.

    PubMed

    Anderson, Judy E; Vargas, Cinthya

    2003-06-01

    Satellite cells, muscle precursor cells in skeletal muscle, are normally quiescent and become activated by disease or injury. A lack of dystrophin and changes in the expression or activity of neuronal nitric oxide synthase (NOS-I) affect the timing of activation in vivo. Nitric oxide synthase inhibition delays muscle repair in normal mice, and worsens muscular dystrophy in the mdx mouse, a genetic homologue of Duchenne muscular dystrophy. However, the potential role of activation and repair events mediated by nitric oxide in determining the outcome of steroid or other treatments for muscular dystrophy is not clear. We tested the hypothesis that the extent of repair in dystrophic muscles of mdx mice is partly dependent on NOS-Imu expression and activity. Myotube formation in regenerating muscle was promoted by deflazacort treatment of mdx dystrophic mice (P<0.05), and improved by combination with the nitric oxide synthase substrate, L-arginine, especially in the diaphragm. NOS-Imu mRNA expression and activity were present in satellite cells and very new myotubes of regenerating and dystrophic muscle. Deflazacort treatment resulted in increased NOS-Imu expression in regenerating muscles in a strong and specific correlation with myf5 expression (r=0.95, P<0.01), a marker for muscle repair. Nitric oxide synthase inhibition prevented the deflazacort-induced rise in NOS-Imu and myf5 expression in the diaphragm without affecting the diameter of non-regenerating fibres. These in vivo studies suggest that gains in NOS-Imu expression and nitric oxide synthase activity in satellite cells can increase the extent and speed of repair, even in the absence of dystrophin in muscle fibres. NOS-Imu may be a useful therapeutic target to augment the effects of steroidal or other treatments of muscular dystrophy.

  5. Regulation of jaw-specific isoforms of myosin-binding protein-C and tropomyosin in regenerating cat temporalis muscle innervated by limb fast and slow motor nerves.

    PubMed

    Kang, Lucia H D; Hoh, Joseph F Y

    2010-11-01

    Cat jaw-closing muscles are a distinct muscle allotype characterized by the expression of masticatory-specific myofibrillar proteins. Transplantation studies showed that expression of masticatory myosin heavy chain (m-MyHC) is promoted by fast motor nerves, but suppressed by slow motor nerves. We investigated whether masticatory myosin-binding protein-C (m-MBP-C) and masticatory tropomyosin (m-Tm) are similarly regulated. Temporalis muscle strips were transplanted into limb muscle beds to allow innervation by fast or slow muscle nerve during regeneration. Regenerated muscles were examined postoperatively up to 168 days by peroxidase IHC using monoclonal antibodies to m-MyHC, m-MBP-C, and m-Tm. Regenerates in both muscle beds expressed fetal and slow MyHCs, m-MyHC, m-MBP-C, and m-Tm during the first 4 weeks. Longer-term regenerates innervated by fast nerve suppressed fetal and slow MyHCs, retaining m-MyHC, m-MBP-C, and m-Tm, whereas fibers innervated by slow nerve suppressed fetal MyHCs and the three masticatory-specific proteins, induced slow MyHC, and showed immunohistochemical characteristics of jaw-slow fibers. We concluded that expression of m-MBP-C and m-Tm is coregulated by m-MyHC and that neural impulses to limb slow muscle are capable of suppressing masticatory-specific proteins and to channel gene expression along the jaw-slow phenotype unique to jaw-closing muscle.

  6. Mobilized adult pituitary stem cells contribute to endocrine regeneration in response to physiological demand.

    PubMed

    Rizzoti, Karine; Akiyama, Haruhiko; Lovell-Badge, Robin

    2013-10-03

    Pituitary hormone deficiencies, with Growth Hormone deficiency being most frequent (1 in 3,500-10,000 births), cause significant morbidity. Regeneration of missing endocrine cells would be a significant improvement over hormone replacement therapies, which incur side effects and do not mimic physiological secretion patterns. Recent in vitro studies have identified a population of adult pituitary progenitors that express the HMG box transcription factors SOX2 and SOX9. Here, we apply cell-lineage tracing analysis to demonstrate that SOX2- and SOX9-expressing progenitors can self-renew and give rise to endocrine cells in vivo, suggesting that they are tissue stem cells. Moreover, we show that they can become mobilized and differentiate into the appropriate endocrine cell types in response to physiological stress. Our results highlight the pituitary as a model for exploring how physiological changes influence stem cell behavior and suggest that manipulation of endogenous pituitary stem cells is a potential therapeutic strategy for pituitary deficiencies.

  7. OnabotulinumtoxinA muscle injection patterns in adult spasticity: a systematic literature review

    PubMed Central

    2013-01-01

    Background OnabotulinumtoxinA has demonstrated significant benefit in adult focal spasticity. This study reviews the injection patterns (i.e., muscle distribution, dosing) of onabotulinumtoxinA for treatment of adult spasticity, as reported in published studies. Methods A systematic review of clinical trials and observational studies published between 1990 and 2011 reporting data on muscles injected with onabotulinumtoxinA in adult patients treated for any cause of spasticity. Results 28 randomized, 5 nonrandomized, and 37 single-arm studies evaluating 2,163 adult patients were included. The most frequently injected upper-limb muscles were flexor carpi radialis (64.0% of patients), flexor carpi ulnaris (59.1%), flexor digitorum superficialis (57.2%), flexor digitorum profundus (52.5%), and biceps brachii (38.8%). The most frequently injected lower-limb muscles were the gastrocnemius (66.1% of patients), soleus (54.7%), and tibialis posterior (50.5%). The overall dose range reported was 5–200 U for upper-limb muscles and 10–400 U for lower-limb muscles. Conclusions The reviewed evidence indicates that the muscles most frequently injected with onabotulinumtoxinA in adults with spasticity were the wrist, elbow, and finger flexors and the ankle plantar flexors. OnabotulinumtoxinA was injected over a broad range of doses per muscle among the studies included in this review, but individual practitioners should be mindful of local regulatory approvals and regulations. PMID:24011236

  8. Intraocular elevation of cyclic AMP potentiates ciliary neurotrophic factor-induced regeneration of adult rat retinal ganglion cell axons.

    PubMed

    Cui, Qi; Yip, Henry K; Zhao, Robert C H; So, Kwok-Fai; Harvey, Alan R

    2003-01-01

    In vitro, cyclic AMP (cAMP) elevation alters neuronal responsiveness to diffusible growth factors and myelin-associated inhibitory molecules. Here we used an established in vivo model of adult central nervous system injury to investigate the effects of elevated cAMP on neuronal survival and axonal regeneration. We studied the effects of intraocular injections of neurotrophic factors and/or a cAMP analogue (CPT-cAMP) on the regeneration of axotomized rat retinal ganglion cell (RGC) axons into peripheral nerve autografts. Elevation of cAMP alone did not significantly increase RGC survival or the number of regenerating RGCs. Ciliary neurotrophic factor increased RGC viability and axonal regrowth, the latter effect substantially enhanced by coapplication with CPT-cAMP. Under these conditions over 60% of surviving RGCs regenerated their axons. Neurotrophin-4/5 injections also increased RGC viability, but there was reduced long-distance axonal regrowth into grafts, an effect partially ameliorated by cAMP elevation. Thus, cAMP can act cooperatively with appropriate neurotrophic factors to promote axonal regeneration in the injured adult mammalian central nervous system.

  9. Long-term delivery of FGF-6 changes the fiber type and fatigability of muscle reinnervated from embryonic neurons transplanted into adult rat peripheral nerve.

    PubMed

    Grumbles, Robert M; Casella, Gizelda T B; Rudinsky, Michelle J; Wood, Patrick M; Sesodia, Sanjay; Bent, Melissa; Thomas, Christine K

    2007-07-01

    Motoneuron death leads to muscle denervation and atrophy. Transplantation of embryonic neurons into peripheral nerves results in reinnervation and provides a strategy to rescue muscles from atrophy independent of neuron replacement in a damaged or diseased spinal cord. But the count of regenerating axons always exceeds the number of motor units in this model, so target-derived trophic factor levels may limit reinnervation. Our aim was to examine whether long-term infusion of fibroblast growth factor-6 (FGF-6) into denervated medial gastrocnemius muscles improved the function of muscles reinnervated from neurons transplanted into nerve of adult Fischer rats. Factor delivery (10 microg, 4 weeks) began after sciatic nerve transection. After a week of nerve degeneration, 1 million embryonic day 14-15 ventral spinal cord cells were transplanted into the distal tibial stump as a neuron source. Ten weeks later, neurons that expressed motoneuron markers survived in the nerves. More myelinated axons were in nerves to saline-treated muscles than in FGF-6-treated muscles. However, each group showed comparable reductions in muscle fiber atrophy because of reinnervation. Mean reinnervated fiber area was 43%-51% of non-denervated fibers. Denervated fiber area averaged 11%. FGF-6-treated muscles were more fatigable than other reinnervated muscles but had stronger motor units and fewer type I fibers than did saline-treated muscles. FGF-6 thus influenced function by changing the type of fiber reinnervated by transplanted neurons. Deficits in FGF-6 may also contribute to the increase in type I fibers in muscles reinnervated from peripheral axons, suggesting that the effects of FGF-6 on fiber type are independent of the neuron source used for reinnervation.

  10. Sensory deprivation disrupts homeostatic regeneration of newly generated olfactory sensory neurons after injury in adult mice.

    PubMed

    Kikuta, Shu; Sakamoto, Takashi; Nagayama, Shin; Kanaya, Kaori; Kinoshita, Makoto; Kondo, Kenji; Tsunoda, Koichi; Mori, Kensaku; Yamasoba, Tatsuya

    2015-02-11

    Although it is well known that injury induces the generation of a substantial number of new olfactory sensory neurons (OSNs) in the adult olfactory epithelium (OE), it is not well understood whether olfactory sensory input influences the survival and maturation of these injury-induced OSNs in adults. Here, we investigated whether olfactory sensory deprivation affected the dynamic incorporation of newly generated OSNs 3, 7, 14, and 28 d after injury in adult mice. Mice were unilaterally deprived of olfactory sensory input by inserting a silicone tube into their nostrils. Methimazole, an olfactotoxic drug, was also injected intraperitoneally to bilaterally ablate OSNs. The OE was restored to its preinjury condition with new OSNs by day 28. No significant differences in the numbers of olfactory marker protein-positive mature OSNs or apoptotic OSNs were observed between the deprived and nondeprived sides 0-7 d after injury. However, between days 7 and 28, the sensory-deprived side showed markedly fewer OSNs and mature OSNs, but more apoptotic OSNs, than the nondeprived side. Intrinsic functional imaging of the dorsal surface of the olfactory bulb at day 28 revealed that responses to odor stimulation were weaker in the deprived side compared with those in the nondeprived side. Furthermore, prevention of cell death in new neurons 7-14 d after injury promoted the recovery of the OE. These results indicate that, in the adult OE, sensory deprivation disrupts compensatory OSN regeneration after injury and that newly generated OSNs have a critical time window for sensory-input-dependent survival 7-14 d after injury.

  11. Long-distance axonal regeneration in the filum terminale of adult rats is regulated by ependymal cells.

    PubMed

    Kwiecien, Jacek M; Avram, Ronen

    2008-03-01

    Studies of regeneration of transected adult central nervous system (CNS) axons are difficult due to lack of appropriate in vivo models. In adult rats, we described filum terminale (FT), a caudal slender extension of the sacral spinal cord and an integral part of the central nervous system (CNS), to use it as a model of spinal cord injury. FT is more than 3 cm long, encompasses a central canal lined with ependymal cells surrounded by a narrow band of axons interspersed with oligodendrocytes and astrocytes but not neurons. Two weeks after the crush of FT, histological, ultrastructural, and axonal tracing studies revealed long distance descending axonal regeneration uniquely in close proximity of the ependymal cells of the central canal. Ependymal cells extended basal processes to form channels encompassing axons apparently regenerating at a rate of more than 2 mm a day. Remarkable increase of axonal sprouting was observed in the sacral spinal cord of Long Evans Shaker (LES) rats with crushed FT. FT offers an excellent model to study mechanisms of axonal regeneration regulated by ependymal cells in the adult CNS.

  12. How to mend a broken heart: adult and induced pluripotent stem cell therapy for heart repair and regeneration.

    PubMed

    Wegener, Marie; Bader, Augustinus; Giri, Shibashish

    2015-06-01

    The recently developed ability to differentiate primary adult stem cells and induced pluripotent stem cells (iPSCs) into cardiomyocytes is providing unprecedented opportunities to produce an unlimited supply of cardiomyocytes for use in patients with heart disease. Here, we examine the evidence for the preclinical use of such cells for successful heart regeneration. We also describe advances in the identification of new cardiac molecular and cellular targets to induce proliferation of cardiomyocytes for heart regeneration. Such new advances are paving the way for a new innovative drug development process for the treatment of heart disease.

  13. Embryonic origin of adult stem cells required for tissue homeostasis and regeneration

    PubMed Central

    Davies, Erin L; Lei, Kai; Seidel, Christopher W; Kroesen, Amanda E; McKinney, Sean A; Guo, Longhua; Robb, Sofia MC; Ross, Eric J; Gotting, Kirsten; Alvarado, Alejandro Sánchez

    2017-01-01

    Planarian neoblasts are pluripotent, adult somatic stem cells and lineage-primed progenitors that are required for the production and maintenance of all differentiated cell types, including the germline. Neoblasts, originally defined as undifferentiated cells residing in the adult parenchyma, are frequently compared to embryonic stem cells yet their developmental origin remains obscure. We investigated the provenance of neoblasts during Schmidtea mediterranea embryogenesis, and report that neoblasts arise from an anarchic, cycling piwi-1+ population wholly responsible for production of all temporary and definitive organs during embryogenesis. Early embryonic piwi-1+ cells are molecularly and functionally distinct from neoblasts: they express unique cohorts of early embryo enriched transcripts and behave differently than neoblasts in cell transplantation assays. Neoblast lineages arise as organogenesis begins and are required for construction of all major organ systems during embryogenesis. These subpopulations are continuously generated during adulthood, where they act as agents of tissue homeostasis and regeneration. DOI: http://dx.doi.org/10.7554/eLife.21052.001 PMID:28072387

  14. Effect of Loquat Leaf Extract on Muscle Strength, Muscle Mass, and Muscle Function in Healthy Adults: A Randomized, Double-Blinded, and Placebo-Controlled Trial

    PubMed Central

    Choe, Sangmin; Lee, Chang-Hyung; Shin, Jin-Hong

    2016-01-01

    Ursolic acid (UA) is the major active component of the loquat leaf extract (LLE) and several previous studies have indicated that UA may have the ability to prevent skeletal muscle atrophy. Therefore, we conducted a randomized, double-blind, and placebo-controlled study to investigate the effects of the LLE on muscle strength, muscle mass, muscle function, and metabolic markers in healthy adults; the safety of the compound was also evaluated. We examined the peak torque/body weight at 60°/s knee extension, handgrip strength, skeletal muscle mass, physical performance, and metabolic parameters at baseline, as well as after 4 and 12 weeks of intervention. Either 500 mg of LLE (50.94 mg of UA) or a placebo was administered to fifty-four healthy adults each day for 12 weeks; no differences in muscle strength, muscle mass, and physical performance were observed between the two groups. However, the right-handgrip strength of female subjects in the LLE group was found to be significantly better than that of subjects in the control group (P = 0.047). Further studies are required to determine the optimal dose and duration of LLE supplementation to confirm the first-stage study results for clinical application. ClinicalTrials.gov Identifier is NCT02401113. PMID:27999607

  15. Methods for promoting wound healing and muscle regeneration with the cell signaling protein Nell1

    SciTech Connect

    Culiat, Cymbeline T

    2014-11-04

    The present invention provides methods for promoting wound healing and treating muscle atrophy in a mammal in need. The method comprises administering to the mammal a Nell1 protein or a Nell1 nucleic acid molecule.

  16. Methods for promoting wound healing and muscle regeneration with the cell signaling protein Nell1

    SciTech Connect

    Culiat, Cymbeline T

    2011-03-22

    The present invention provides methods for promoting wound healing and treating muscle atrophy in a mammal in need. The method comprises administering to the mammal a Nell1 protein or a Nell1 nucleic acid molecule.

  17. Magnetic targeting of human peripheral blood CD133+ cells for skeletal muscle regeneration.

    PubMed

    Ohkawa, Shingo; Kamei, Naosuke; Kamei, Goki; Shi, Ming; Adachi, Nobuo; Deie, Masataka; Ochi, Mitsuo

    2013-08-01

    Skeletal muscle injuries often leave lasting functional damage or pain. Muscle injuries are routinely treated conservatively, but the most effective treatment to promote the repair of injured muscles has not yet been established. Our previous report demonstrated that human peripheral blood-derived CD133(+) cell transplantation to rat skeletal muscle injury models inhibited fibrosis and enhanced myogenesis after injury. However, the acquisition of a sufficient number of cells remains the limitation for clinical application, as the CD133(+) population is rare in human blood. In this study, we applied a magnetic cell targeting system to accumulate transplanted cells in the muscle injury site and to enhance the regenerative effects of CD133(+) cell transplantation, focusing on the fact that CD133(+) cells are labeled with a magnetic bead for isolation. For the magnetic cell targeting, the magnet field generator was set up to adjust the peak of the magnetic gradient to the injury site of the tibialis anterior muscle, and 1×10(4) human peripheral blood CD133(+) cells were locally injected into the injury site. This cell number is 10% of that used in the previous study. In another group, the same number of CD133(+) cells was injected without magnetic force. The CD133(+) cells transplanted with the magnetic force were more accumulated in the muscle injury site compared with the CD133(+) cells transplanted without the magnetic force. In addition, the transplantation of CD133(+) cells under the magnetic control inhibited fibrous scar formation and promoted angiogenesis and myogenesis, and also upregulated the mRNA expression of myogenic transcription factors, including Pax7, MyoD1 and Myogenin. However, the transplantation of CD133(+) cells without the magnetic force failed to demonstrate these effects. Thus, our magnetic cell targeting system enables transplantation of a limited number of CD133(+) cells to promote the repair of skeletal muscle injury.

  18. Functional Assessment of Skeletal Muscle Regeneration Utilizing Homologous Extracellular Matrix as Scaffolding

    DTIC Science & Technology

    2010-01-01

    should also serve as a scaffold and allow the incorporation of regenerat- ing muscle, connective, nerve , and blood vessel tissues. Several types of...To close the wound, the biceps femoris muscle was reattached to the tibia using simple, interrupted, polypropylene suture (Prolene 5-0; Ethicon, San...from the superficial skin and biceps femoris as well as the deep soleus and plan- taris. The nerve branch innervating the medial GAS was transected to

  19. [Reinnervation of a mixed muscle in the frog Rana temporaria with a regenerating homogeneous nerve].

    PubMed

    Radziukevich, T L

    1995-01-01

    Mixed muscle m. iliofibularis from the frog Rana temporaria, consisting of monosynaptically innervated phasic and polysynaptically innervated postural muscle fibers, was reinnervated by homogeneous tailor's nerve having no axons of tonic motor system in its composition. Within 2-7 months after nerves binding treatment phasic muscle fibers were easily identified by subneural apparatus structure revealed at coloration of synaptic acetylcholinesterase. Presynaptic part of neuromuscular apparatus of these fibers after the impregnation by protargol was presented by immature nervous terminals. The identification of tonic Muscular fibers was difficult especially at the late stages of reinnervation as subneural apparatus structure typical for tonic fibers was not revealed. Nonmyelinizated nerve fibers without features of terminal branch were observed in individual regions of nonidentified muscle fibers. The results obtained show that subneural apparatus of tonic muscle fibers depends to a great extent on the influence of inherent tonic motor system. Axons of phasic motor system even at distant reinnervation periods and in the absence of competitive influences of tonic motor system do not form typical "phasic" terminal picture of innervation under the contact with tonic muscle fibers.

  20. Polymeric scaffold aided stem cell therapeutics for cardiac muscle repair and regeneration.

    PubMed

    Lakshmanan, Rajesh; Krishnan, Uma Maheswari; Sethuraman, Swaminathan

    2013-09-01

    The constantly expanding repository of novel polymers and stem cells has opened up new vistas in the field of cardiac tissue engineering. Successful regeneration of the complex cardiac tissue mainly centres on the appropriate scaffold material with topographical features that mimic the native environment. The integration of stem cells on these scaffolds is expected to enhance the regeneration potential. This review elaborates on the interplay of these vital factors in achieving the functional cardiac tissue. The recent advances in polymers, nanocomposites, and stem cells from different sources are highlighted. Special emphasis is laid on the clinical trials involving stem cells and the state-of-the-art materials to obtain a balanced perspective on the translational potential of this strategy.

  1. The effect of superficial trunk muscle exercise and deep trunk muscle exercise on the foot pressure of healthy adults

    PubMed Central

    Kim, Suzy; Shim, Jemyung; Kim, Sungjoong; Namkoong, Seung; Kim, Hwanhee

    2015-01-01

    [Purpose] The purpose of this study was to analyze the effect of superficial trunk muscle exercise and deep trunk muscle exercise on the foot pressure of healthy adults. [Subjects] The subjects were 30 healthy females and males who agreed to participate in this study. There were two groups, a superficial trunk muscle exercise group and a deep trunk muscle exercise group, with 15 participants in each. [Methods] The exercises were conducted 5 times a week for 4 weeks for both groups. A gait analyzer was used to measure foot plantar pressure while walking on a plate. Participants were measured before starting the exercise and after 4 weeks. The paired t-test was used to analyze the pre-and post-test results. [Results] There were no significant differences in foot pressure in any region in the superficial trunk muscle exercise group. In the deep trunk muscle exercise group, there were statistically significant increase in F1, F4, F5, R1 and R3. In addition, there were significant decreases in R2 and R4. [Conclusion] After the 4-week deep trunk muscle exercise group decreases in foot pressure on the inner foot and increases on the outside of the feet indicate normal and overall even distribution of body weight on the feet. PMID:25931714

  2. Adult-Derived Human Liver Stem/Progenitor Cells Infused 3 Days Postsurgery Improve Liver Regeneration in a Mouse Model of Extended Hepatectomy.

    PubMed

    Herrero, Astrid; Prigent, Julie; Lombard, Catherine; Rosseels, Valérie; Daujat-Chavanieu, Martine; Breckpot, Karine; Najimi, Mustapha; Deblandre, Gisèle; Sokal, Etienne M

    2017-02-16

    There is growing evidence that cell therapy constitutes a promising strategy for liver regenerative medicine. In the setting of hepatic cancer treatments, cell therapy could prove a useful therapeutic approach for managing the acute liver failure that occurs following extended hepatectomy. In this study, we examined the influence of delivering adult-derived human liver stem/progenitor cells (ADHLSCs) at two different early time points in an immunodeficient mouse model (Rag2-/-IL2Rγ-/-) that had undergone a 70% hepatectomy procedure. The hepatic mesenchymal cells were intrasplenically infused either immediately after surgery (n = 26) or following a critical 3-day period (n = 26). We evaluated the cells' capacity to engraft at day 1 and day 7 following transplantation by means of human Alu qPCR quantification, along with histological assessment of human albumin and α-smooth muscle actin. In addition, cell proliferation (anti-mouse and human Ki-67 staining) and murine liver weight were measured in order to evaluate liver regeneration. At day 1 posttransplantation, the ratio of human to mouse cells was similar in both groups, whereas 1 week posttransplantation this ratio was significantly improved (p < 0.016) in mice receiving ADHLSC injection at day 3 posthepatectomy (1.7%), compared to those injected at the time of surgery (1%). On the basis of liver weight, mouse liver regeneration was more extensive 1 week posttransplantation in mice transplanted with ADHLSCs (+65.3%) compared to that of mice from the sham vehicle group (+42.7%). In conclusion, infusing ADHLSCs 3 days after extensive hepatectomy improves the cell engraftment and murine hepatic tissue regeneration, thereby confirming that ADHLSCs could be a promising cell source for liver cell therapy and hepatic tissue repair.

  3. EFFECTS OF AGE AND ACUTE MUSCLE FATIGUE ON REACTIVE POSTURAL CONTROL IN HEALTHY ADULTS

    PubMed Central

    Papa, Evan V.; Foreman, K. Bo; Dibble, Lee E.

    2015-01-01

    BACKGROUND Falls can cause moderate to severe injuries such as hip fractures and head trauma in older adults. While declines in muscle strength and sensory function contribute to increased falls in older adults, skeletal muscle fatigue is often overlooked as an additional contributor to fall risk. The purpose of this investigation was to examine the effects of acute lower extremity muscle fatigue and age on reactive postural control in healthy adults. METHODS A sample of 16 individuals participated in this study (8 healthy older adults and 8 healthy young persons). Whole body kinematic and kinetic data were collected during anterior and posterior reproducible fall tests before (T0) and immediately after (T1) eccentric muscle fatiguing exercise, as well as after 15-minutes (T15) and 30-minutes (T30) of rest. FINDINGS Lower extremity joint kinematics of the stepping limb during the support (landing) phase of the anterior fall were significantly altered by the presence of acute muscle fatigue. Step velocity was significantly decreased during the anterior falls. Statistically significant main effects of age were found for step length in both fall directions. Effect sizes for all outcomes were small. No statistically significant interaction effects were found. INTERPRETATION Muscle fatigue has a measurable effect on lower extremity joint kinematics during simulated falls. These alterations appear to resolve within 15 minutes of recovery. The above deficits, coupled with a reduced step length, may help explain the increased fall risk in older adults. PMID:26351001

  4. Do muscle mass, muscle density, strength and physical function similarly influence risk of hospitalization in older adults?

    PubMed Central

    Cawthon, Peggy Mannen; Fox, Kathleen M.; Gandra, Shravanthi. R.; Delmonico, Matthew J.; Chiou, Chiun-Fang; Anthony, Mary S.; Sewall, Ase; Goodpaster, Bret; Satterfield, Suzanne; Cummings, Steven R.; Harris, Tamara B.

    2012-01-01

    Objectives To examine the association between strength, function, lean mass, muscle density and risk of hospitalization. Design Prospective cohort stud Setting Two U.S. clinical centers Participants Adults aged 70 – 80 years (N=3,011) from the Health, Aging and Body Composition Study. Measurements Measures included grip strength; knee extension strength; lean mass; walking speed; chair stand pace. Thigh computed tomography scans assessed muscle area and density (a proxy for muscle fat infiltration). Hospitalizations were confirmed by local review of medical records. Negative binomial regression models estimated incident rate ratios (IRRs) of hospitalization for race/sex specific quartiles of each muscle/function parameter separately. Multivariate models adjusted for age, body mass index, health status and coexisting medical conditions. Results During an average 4.7 years of follow-up, 1,678 (55.7%) participants experienced ≥1 hospitalization. Participants in the lowest quartile of muscle density were more likely to be subsequently hospitalized (multivariate IRR: 1.47, 95% CI: 1.24, 1.73) compared to the highest quartile. Similarly, participants with the weakest grip strength were at increased risk of hospitalization (MIRR: 1.52, 95% CI: 1.30, 1.78, Q1 vs. Q4). Comparable results were seen for knee strength, walking pace and chair stands pace. Lean mass and muscle area were not associated with risk of hospitalization. Conclusion Weak strength, poor function and low muscle density, but not muscle size or lean mass, were associated with an increased risk of hospitalization. Interventions to reduce the disease burden associated with sarcopenia should focus on increasing muscle strength and improving physical function rather than simply increasing lean mass. PMID:19682143

  5. Delayed Onset Muscle Soreness After Inspiratory Threshold Loading in Healthy Adults

    PubMed Central

    Mathur, Sunita; Sheel, A. William; Road, Jeremy D.; Reid, W. Darlene

    2010-01-01

    Purpose: Skeletal muscle damage occurs following high-intensity or unaccustomed exercise; however, it is difficult to monitor damage to the respiratory muscles, particularly in humans. The aim of this study was to use clinical measures to investigate the presence of skeletal muscle damage in the inspiratory muscles. Methods: Ten healthy subjects underwent 60 minutes of voluntary inspiratory threshold loading (ITL) at 70% of maximal inspiratory pressure. Maximal inspiratory and expiratory mouth pressures, delayed onset muscle soreness on a visual analogue scale and plasma creatine kinase were measured prior to ITL, and at repeated time points after ITL (4, 24 and 48 hours post-ITL). Results: Delayed onset muscle soreness was present in all subjects 24 hours following ITL (intensity = 22 ± 6 mm; significantly higher than baseline p = 0.02). Muscle soreness was reported primarily in the anterior neck region, and was correlated to the amount of work done by the inspiratory muscles during ITL (r = 0.72, p = 0.02). However, no significant change was observed in maximal inspiratory or expiratory pressures or creatine kinase. Conclusions: These findings suggest that an intense bout of ITL results in muscle soreness primarily in the accessory muscles of inspiration, however, may be insufficient to cause significant muscle damage in healthy adults. PMID:20467514

  6. Relationships between metabolic rate, muscle electromyograms, and swim performance of adult chinook salmon

    SciTech Connect

    Geist, David R. ); Brown, Richard S. ); Cullinan, Valerie I. ); Mesa, Matthew G.; VanderKooi, S P.; McKinstry, Craig A. )

    2003-10-01

    We measured oxygen consumption rates of adult spring Chinook salmon and compared these values to other species of Pacific salmon. Our results indicated that adult salmon achieve their maximum level of oxygen consumption at about their upper critical swim speed. It is also at this speed that the majority of the energy supplied to the swimming fish switches from red muscle (powered by aerobic metabolism) to white muscle (powered by anaerobic metabolism). Determining the swimming performance of adult salmon will assist managers in developing fishways and other means to safely pass fish over hydroelectric dams and other man-made structures.

  7. Fetal and adult liver stem cells for liver regeneration and tissue engineering.

    PubMed

    Fiegel, H C; Lange, Claudia; Kneser, U; Lambrecht, W; Zander, A R; Rogiers, X; Kluth, D

    2006-01-01

    For the development of innovative cell-based liver directed therapies, e.g. liver tissue engineering, the use of stem cells might be very attractive to overcome the limitation of donor liver tissue. Liver specific differentiation of embryonic, fetal or adult stem cells is currently under investigation. Different types of fetal liver (stem) cells during development were identified, and their advantageous growth potential and bipotential differentiation capacity were shown. However, ethical and legal issues have to be addressed before using fetal cells. Use of adult stem cells is clinically established, e.g. transplantation of hematopoietic stem cells. Other bone marrow derived liver stem cells might be mesenchymal stem cells (MSC). However, the transdifferentiation potential is still in question due to the observation of cellular fusion in several in vivo experiments. In vitro experiments revealed a crucial role of the environment (e.g. growth factors and extracellular matrix) for specific differentiation of stem cells. Co-cultured liver cells also seemed to be important for hepatic gene expression of MSC. For successful liver cell transplantation, a novel approach of tissue engineering by orthotopic transplantation of gel-immobilized cells could be promising, providing optimal environment for the injected cells. Moreover, an orthotopic tissue engineering approach using bipotential stem cells could lead to a repopulation of the recipients liver with healthy liver and biliary cells, thus providing both hepatic functions and biliary excretion. Future studies have to investigate, which stem cell and environmental conditions would be most suitable for the use of stem cells for liver regeneration or tissue engineering approaches.

  8. Dystrophy of the diaphragmatic muscles in adult Meuse-Rhine-Yssel cattle: electromyographical and histological findings.

    PubMed

    Goedegebuure, S A; Hartman, W; Hoebe, H P

    1983-01-01

    Electromyographic investigation of diaphragmatic muscles of Meuse-Rhine-Yssel cattle revealed a significant decreased duration of action potentials, while the number of polyphasic potentials was increased in four of seven cows. Histologically, the diaphragmatic muscles in all cows were affected severely, as characterized by variation in size of individual fibers, abundant vacuolar and hyaline degeneration with occasional fragmentation and phagocytosis, fiber splitting, apparent increase in internal nuclei, vesicular nuclei, chains of central nuclei, absence of regeneration, and proliferation of endomysial and perimysial connective tissue. Core-like structures seemed to be a hallmark of the disease. The intercostal muscles in all cows had core-like structures and some variation in fiber size; degenerative lesions did occur, but were less severe than in diaphragmatic muscles. In other skeletal muscles and cardiac muscles, core-like structures were present predominantly, indicating a generalized muscle disorder. No lesions were detected in the peripheral or central nervous systems. The muscular alterations were classified as a progressive muscular dystrophy, with a suspicion of hereditary transmission. This dystrophy may be an important animal model.

  9. Fibroadipogenic progenitors mediate the ability of HDAC inhibitors to promote regeneration in dystrophic muscles of young, but not old Mdx mice.

    PubMed

    Mozzetta, Chiara; Consalvi, Silvia; Saccone, Valentina; Tierney, Matthew; Diamantini, Adamo; Mitchell, Kathryn J; Marazzi, Giovanna; Borsellino, Giovanna; Battistini, Luca; Sassoon, David; Sacco, Alessandra; Puri, Pier Lorenzo

    2013-04-01

    HDAC inhibitors (HDACi) exert beneficial effects in mdx mice, by promoting endogenous regeneration; however, the cellular determinants of HDACi activity on dystrophic muscles have not been determined. We show that fibroadipogenic progenitors (FAP) influence the regeneration potential of satellite cells during disease progression in mdx mice and mediate HDACi ability to selectively promote regeneration at early stages of disease. FAPs from young mdx mice promote, while FAPs from old mdx mice repress, satellite cell-mediated formation of myotubes. In young mdx mice HDACi inhibited FAP adipogenic potential, while enhancing their ability to promote differentiation of adjacent satellite cells, through upregulation of the soluble factor follistatin. By contrast, FAPs from old mdx mice were resistant to HDACi-mediated inhibition of adipogenesis and constitutively repressed satellite cell-mediated formation of myotubes. We show that transplantation of FAPs from regenerating young muscles restored HDACi ability to increase myofibre size in old mdx mice. These results reveal that FAPs are key cellular determinants of disease progression in mdx mice and mediate a previously unappreciated stage-specific beneficial effect of HDACi in dystrophic muscles.

  10. Substrate availability limits human skeletal muscle oxidative ATP regeneration at the onset of ischemic exercise.

    PubMed Central

    Timmons, J A; Gustafsson, T; Sundberg, C J; Jansson, E; Hultman, E; Kaijser, L; Chwalbinska-Moneta, J; Constantin-Teodosiu, D; Macdonald, I A; Greenhaff, P L

    1998-01-01

    We have demonstrated previously that dichloroacetate can attenuate skeletal muscle fatigue by up to 35% in a canine model of peripheral ischemia (Timmons, J.A., S.M. Poucher, D. Constantin-Teodosiu, V. Worrall, I.A. Macdonald, and P.L. Greenhaff. 1996. J. Clin. Invest. 97:879-883). This was thought to be a consequence of dichloroacetate increasing acetyl group availability early during contraction. In this study we characterized the metabolic effects of dichloroacetate in a human model of peripheral muscle ischemia. On two separate occasions (control-saline or dichloroacetate infusion), nine subjects performed 8 min of single-leg knee extension exercise at an intensity aimed at achieving volitional exhaustion in approximately 8 min. During exercise each subject's lower limbs were exposed to 50 mmHg of positive pressure, which reduces blood flow by approximately 20%. Dichloroacetate increased resting muscle pyruvate dehydrogenase complex activation status by threefold and elevated acetylcarnitine concentration by fivefold. After 3 min of exercise, phosphocreatine degradation and lactate accumulation were both reduced by approximately 50% after dichloroacetate pretreatment, when compared with control conditions. However, after 8 min of exercise no differences existed between treatments. Therefore, it would appear that dichloroacetate can delay the accumulation of metabolites which lead to the development of skeletal muscle fatigue during ischemia but does not alter the metabolic profile when a maximal effort is approached. PMID:9421469

  11. The insulin-like growth factor 1 receptor is essential for axonal regeneration in adult central nervous system neurons.

    PubMed

    Dupraz, Sebastián; Grassi, Diego; Karnas, Diana; Nieto Guil, Alvaro F; Hicks, David; Quiroga, Santiago

    2013-01-01

    Axonal regeneration is an essential condition to re-establish functional neuronal connections in the injured adult central nervous system (CNS), but efficient regrowth of severed axons has proven to be very difficult to achieve. Although significant progress has been made in identifying the intrinsic and extrinsic mechanisms involved, many aspects remain unresolved. Axonal development in embryonic CNS (hippocampus) requires the obligate activation of the insulin-like growth factor 1 receptor (IGF-1R). Based on known similarities between axonal growth in fetal compared to mature CNS, we decided to examine the expression of the IGF-1R, using an antibody to the βgc subunit or a polyclonal anti-peptide antibody directed to the IGF-R (C20), in an in vitro model of adult CNS axonal regeneration, namely retinal ganglion cells (RGC) derived from adult rat retinas. Expression of both βgc and the β subunit recognized by C20 antibody were low in freshly isolated adult RGC, but increased significantly after 4 days in vitro. As in embryonic axons, βgc was localised to distal regions and leading growth cones in RGC. IGF-1R-βgc co-localised with activated p85 involved in the phosphatidylinositol-3 kinase (PI3K) signaling pathway, upon stimulation with IGF-1. Blocking experiments using either an antibody which neutralises IGF-1R activation, shRNA designed against the IGF-1R sequence, or the PI3K pathway inhibitor LY294002, all significantly reduced axon regeneration from adult RGC in vitro (∼40% RGC possessed axons in controls vs 2-8% in the different blocking studies). Finally, co-transfection of RGC with shRNA to silence IGF-1R together with a vector containing a constitutively active form of downstream PI3K (p110), fully restored axonal outgrowth in vitro. Hence these data demonstrate that axonal regeneration in adult CNS neurons requires re-expression and activation of IGF-1R, and targeting this system may offer new therapeutic approaches to enhancing axonal

  12. Fgfr4 is required for effective muscle regeneration in vivo. Delineation of a MyoD-Tead2-Fgfr4 transcriptional pathway.

    PubMed

    Zhao, Po; Caretti, Giuseppina; Mitchell, Stephanie; McKeehan, Wallace L; Boskey, Adele L; Pachman, Lauren M; Sartorelli, Vittorio; Hoffman, Eric P

    2006-01-06

    Fgfr4 has been shown to be important for appropriate muscle development in chick limb buds; however, Fgfr4 null mice show no phenotype. Here, we show that staged induction of muscle regeneration in Fgfr4 null mice becomes highly abnormal at the time point when Fgfr4 is normally expressed. By 7 days of regeneration, differentiation of myotubes became poorly coordinated and delayed by both histology and embryonic myosin heavy chain staining. By 14 days much of the muscle was replaced by fat and calcifications. To begin to dissect the molecular pathways involving Fgfr4, we queried the promoter sequences for transcriptional factor binding sites and tested candidate regulators in a 27-time point regeneration series. The Fgfr4 promoter region contained a Tead protein binding site (M-CAT 5'-CATTCCT-3'), and Tead2 showed induction during regeneration commensurate with Fgfr4 regulation. Co-transfection of Tead2 and Fgfr4 promoter reporter constructs into C2C12 myotubes showed Tead2 to activate Fgfr4, and mutation of the M-CAT motif in the Fgfr4 promoter abolished these effects. Immunostaining for Tead2 showed timed expression in myotube nuclei consistent with the mRNA data. Query of the expression timing and genomic sequences of Tead2 suggested direct regulation by MyoD, and consistent with this, MyoD directly bound to two strong E-boxes in the first intron of Tead2 by chromatin immunoprecipitation assay. Moreover, co-transfection of MyoD and Tead2 intron reporter constructs into 10T1/2 cells activated reporter activity in a dose-dependent manner. This activation was greatly reduced when the two E-boxes were mutated. Our data suggest a novel MyoD-Tead2-Fgfr4 pathway important for effective muscle regeneration.

  13. MASH1/Ascl1a leads to GAP43 expression and axon regeneration in the adult CNS.

    PubMed

    Williams, Ryan R; Venkatesh, Ishwariya; Pearse, Damien D; Udvadia, Ava J; Bunge, Mary Bartlett

    2015-01-01

    Unlike CNS neurons in adult mammals, neurons in fish and embryonic mammals can regenerate their axons after injury. These divergent regenerative responses are in part mediated by the growth-associated expression of select transcription factors. The basic helix-loop-helix (bHLH) transcription factor, MASH1/Ascl1a, is transiently expressed during the development of many neuronal subtypes and regulates the expression of genes that mediate cell fate determination and differentiation. In the adult zebrafish (Danio rerio), Ascl1a is also transiently expressed in retinal ganglion cells (RGCs) that regenerate axons after optic nerve crush. Utilizing transgenic zebrafish with a 3.6 kb GAP43 promoter that drives expression of an enhanced green fluorescent protein (EGFP), we observed that knock-down of Ascl1a expression reduces both regenerative gap43 gene expression and axonal growth after injury compared to controls. In mammals, the development of noradrenergic brainstem neurons requires MASH1 expression. In contrast to zebrafish RGCs, however, MASH1 is not expressed in the mammalian brainstem after spinal cord injury (SCI). Therefore, we utilized adeno-associated viral (AAV) vectors to overexpress MASH1 in four month old rat (Rattus norvegicus) brainstem neurons in an attempt to promote axon regeneration after SCI. We discovered that after complete transection of the thoracic spinal cord and implantation of a Schwann cell bridge, animals that express MASH1 exhibit increased noradrenergic axon regeneration and improvement in hindlimb joint movements compared to controls. Together these data demonstrate that MASH1/Ascl1a is a fundamental regulator of axonal growth across vertebrates and can induce modifications to the intrinsic state of neurons to promote functional regeneration in response to CNS injury.

  14. MASH1/Ascl1a Leads to GAP43 Expression and Axon Regeneration in the Adult CNS

    PubMed Central

    Pearse, Damien D.; Udvadia, Ava J.; Bunge, Mary Bartlett

    2015-01-01

    Unlike CNS neurons in adult mammals, neurons in fish and embryonic mammals can regenerate their axons after injury. These divergent regenerative responses are in part mediated by the growth-associated expression of select transcription factors. The basic helix-loop-helix (bHLH) transcription factor, MASH1/Ascl1a, is transiently expressed during the development of many neuronal subtypes and regulates the expression of genes that mediate cell fate determination and differentiation. In the adult zebrafish (Danio rerio), Ascl1a is also transiently expressed in retinal ganglion cells (RGCs) that regenerate axons after optic nerve crush. Utilizing transgenic zebrafish with a 3.6 kb GAP43 promoter that drives expression of an enhanced green fluorescent protein (EGFP), we observed that knock-down of Ascl1a expression reduces both regenerative gap43 gene expression and axonal growth after injury compared to controls. In mammals, the development of noradrenergic brainstem neurons requires MASH1 expression. In contrast to zebrafish RGCs, however, MASH1 is not expressed in the mammalian brainstem after spinal cord injury (SCI). Therefore, we utilized adeno-associated viral (AAV) vectors to overexpress MASH1 in four month old rat (Rattus norvegicus) brainstem neurons in an attempt to promote axon regeneration after SCI. We discovered that after complete transection of the thoracic spinal cord and implantation of a Schwann cell bridge, animals that express MASH1 exhibit increased noradrenergic axon regeneration and improvement in hindlimb joint movements compared to controls. Together these data demonstrate that MASH1/Ascl1a is a fundamental regulator of axonal growth across vertebrates and can induce modifications to the intrinsic state of neurons to promote functional regeneration in response to CNS injury. PMID:25751153

  15. Human Adipose Tissue Derived Stem Cells as a Source of Smooth Muscle Cells in the Regeneration of Muscular Layer of Urinary Bladder Wall

    PubMed Central

    SALEM, Salah Abood; HWIE, Angela Ng Min; SAIM, Aminuddin; CHEE KONG, Christopher Ho; SAGAP, Ismail; SINGH, Rajesh; YUSOF, Mohd Reusmaazran; MD ZAINUDDIN, Zulkifili; HJ IDRUS, Ruszymah

    2013-01-01

    Background: Adipose tissue provides an abundant source of multipotent cells, which represent a source of cell-based regeneration strategies for urinary bladder smooth muscle repair. Our objective was to confirm that adipose-derived stem cells (ADSCs) can be differentiated into smooth muscle cells. Methods: In this study, adipose tissue samples were digested with 0.075% collagenase, and the resulting ADSCs were cultured and expanded in vitro. ADSCs at passage two were differentiated by incubation in smooth muscle inductive media (SMIM) consisting of MCDB I31 medium, 1% FBS, and 100 U/mL heparin for three and six weeks. ADSCs in non-inductive media were used as controls. Characterisation was performed by cell morphology and gene and protein expression. Result: The differentiated cells became elongated and spindle shaped, and towards the end of six weeks, sporadic cell aggregation appeared that is typical of smooth muscle cell culture. Smooth muscle markers (i.e. alpha smooth muscle actin (ASMA), calponin, and myosin heavy chain (MHC)) were used to study gene expression. Expression of these genes was detected by PCR after three and six weeks of differentiation. At the protein expression level, ASMA, MHC, and smoothelin were expressed after six weeks of differentiation. However, only ASMA and smoothelin were expressed after three weeks of differentiation. Conclusion: Adipose tissue provides a possible source of smooth muscle precursor cells that possess the potential capability of smooth muscle differentiation. This represents a promising alternative for urinary bladder smooth muscle repair. PMID:24044001

  16. Exposure to microgravity for 30 days onboard Bion M1 caused muscle atrophy and decreased regeneration in the mouse femoral Quadriceps

    NASA Astrophysics Data System (ADS)

    Grigoryan, Eleonora; Radugina, Elena A.; Almeida, Eduardo; Blaber, Elizabeth; Poplinskaya, Valentina; Markitantova, Yulia

    Mechanical unloading of muscle during spaceflight in microgravity is known to cause muscular atrophy, changes in muscle fiber type composition, gene expression, and reductions in regenerative muscle growth. Although limited data exists for long-term effects of microgravity in human muscle, these processes have mostly been studied in rodents for short periods of time, up to two weeks of spaceflight. Here we report on how 30-day, long-term, mechanical unloading in microgravity affects mouse muscle of the femoral Quadriceps group. To conduct these studies we used muscle tissue from 6 mice from the NASA Biospecimen Sharing Program conducted in collaboration with the Institute for Biomedical Problems of the Russian Academy of Sciences, during the Russian Bion M1 biosatellite mission in 2013. Muscle morphology observed in histological sections shows signs of extensive atrophy and regenerative hypoplasia. Specifically, we observed a two-fold decrease in the number of myonuclei and low density of myofibrils, their separation and fragmentation. Despite obvious atrophy, muscle regeneration nevertheless appears to have continued after 30 days in microgravity as evidenced by thin and short newly formed muscle fibers. Many of them however showed evidence of apoptosis and degradation of synthesized fibrils, suggesting long-term unloading in microgravity affects late stages of myofiber differentiation. Ground asynchronous and vivarium control animals showed normal, well-developed tissue structure with sufficient blood and nerve supply and evidence of regenerative formation of new muscle fibers free of apoptotic nuclei. Myofiber nuclei stress responses in spaceflight animals was detected by positive nuclear immunolocalization of c-jun and c-myc proteins. Regenerative activity of satellite cells in muscle was localized with pax-7, MyoD and MCad immunostaining, and did not appear altered in microgravity. In summary, long-term spaceflight in microgravity causes significant atrophy

  17. A case of adult Langerhans cell histiocytosis showing successfully regenerated osseous tissue of the skull after chemotherapy.

    PubMed

    Suzuki, Takahiro; Izutsu, Koji; Kako, Shinichi; Ohta, Satoshi; Hangaishi, Akira; Kanda, Yoshinobu; Motokura, Toru; Chiba, Shigeru; Kurokawa, Mineo

    2008-04-01

    Langerhans cell histiocytosis (LCH) is a proliferative disorder of Langerhans cells and extremely rare in adults. Adult LCH is often associated with osteolytic bone lesions, but large bone-defective lesions have been rarely reported. We report an adult case of LCH accompanied by large osteolytic lesions in the skull that successfully responded to chemotherapy. A 47-year-old woman with LCH who had multiple, large osteolytic areas of more than 3 cm in diameter in the skull was admitted to our hospital. She was treated with systemic chemotherapy consisting of prednisolone, vinblastine, and 6-mercaptopurine. Twelve months later, when she completed the treatment, osteolytic areas were covered with hard osseous tissue, and X-ray examination confirmed regeneration of the bone. This case indicates that chemotherapy can be effective even for the treatment of large osteolytic lesions in adult LCH patients.

  18. Sequential identification of a degradable phosphate glass scaffold for skeletal muscle regeneration.

    PubMed

    Shah, Rishma; Ready, Derren; Knowles, Jonathan C; Hunt, Nigel P; Lewis, Mark P

    2014-10-01

    Tissue engineering has the potential to overcome limitations associated with current management of skeletal muscle defects. This study aimed to sequentially identify a degradable phosphate glass scaffold for the restoration of muscle defects. A series of glass compositions were investigated for the potential to promote bacterial growth. Thereafter, the response of human craniofacial muscle-derived cells was determined. Glass compositions containing Fe4- and 5 mol% did not promote greater Staphylococcus aureus and Staphylococcus epidermidis growth compared to the control (p > 0.05). Following confirmation of myogenicity, further studies assessed the biocompatibility of glasses containing Fe5 mol%. Cells seeded on collagen-coated disks demonstrated comparable cellular metabolic activity to control. Upregulation of genes encoding for myogenic regulatory factors (MRFs) confirmed myofibre formation and there was expression of developmental MYH genes. The use of 3-D aligned fibre scaffolds supported unidirectional cell alignment and upregulation of MRF and developmental MYH genes. Compared to the 2-D disks, there was also expression of MYH2 and MYH7 genes, indicating further myofibre maturation on the 3-D scaffolds and confirming the importance of key biophysical cues.

  19. Adult stem cells and mammalian epimorphic regeneration-insights from studying annual renewal of deer antlers.

    PubMed

    Li, Chunyi; Yang, Fuhe; Sheppard, Allan

    2009-09-01

    Mammalian organ regeneration is the "Holy Grail" of modern regenerative biology and medicine. The most dramatic organ replacement is known as epimorphic regeneration. To date our knowledge of epimorphic regeneration has come from studies of amphibians. Notably, these animals have the ability to reprogram phenotypically committed cells at the amputation plane toward an embryonic-like cell phenotype (dedifferentiation). The capability of mammals to initiate analogous regeneration, and whether similar mechanisms would be involved if it were to occur, remain unclear. Deer antlers are the only mammalian appendages capable of full renewal, and therefore offer a unique opportunity to explore how nature has solved the problem of mammalian epimorphic regeneration. Following casting of old hard antlers, new antlers regenerate from permanent bony protuberances, known as pedicles. Studies through morphological and histological examinations, tissue deletion and transplantation, and cellular and molecular techniques have demonstrated that antler renewal is markedly different from that of amphibian limb regeneration (dedifferentiation-based), being a stem cell-based epimorphic process. Antler stem cells reside in the pedicle periosteum. We envisage that epimorphic regeneration of mammalian appendages, other than antler, could be made possible by recreating comparable milieu to that which supports the elaboration of that structure from the pedicle periosteum.

  20. Notch regulates blastema proliferation and prevents differentiation during adult zebrafish fin regeneration.

    PubMed

    Münch, Juliane; González-Rajal, Alvaro; de la Pompa, José Luis

    2013-04-01

    Zebrafish have the capacity to regenerate several organs, including the heart and fins. Fin regeneration is epimorphic, involving the formation at the amputation plane of a mass of undifferentiated, proliferating mesenchymal progenitor-like cells, called blastema. This tissue provides all the cell types that form the fin, so that after damage or amputation the fin pattern and structure are fully restored. How blastema cells remain in this progenitor-like state is poorly understood. Here, we show that the Notch pathway plays an essential role during fin regeneration. Notch signalling is activated during blastema formation and remains active throughout the regeneration process. Chemical inhibition or morpholino-mediated knockdown of Notch signalling impairs fin regeneration via decreased proliferation accompanied by reduced expression of Notch target genes in the blastema. Conversely, overexpression of a constitutively active form of the Notch1 receptor (N1ICD) in the regenerating fin leads to increased proliferation and to the expansion of the blastema cell markers msxe and msxb, as well as increased expression of the proliferation regulator aldh1a2. This blastema expansion prevents regenerative fin outgrowth, as indicated by the reduction in differentiating osteoblasts and the inhibition of bone regeneration. We conclude that Notch signalling maintains blastema cells in a plastic, undifferentiated and proliferative state, an essential requirement for fin regeneration.