Thomas Rando, MD, PhD

All Publications

• Exercise reprograms the inflammatory landscape of multiple stem cell compartments during mammalian aging Cell Stem Cell Liu, L., Kim, S., Buckley, M., Reyes, J., Kang, J., Tian, L., Wang, M., Lieu, A., Mao, M., Mateo, C., Ishak, H., Jeong, M., Wu, J., Goodell, M., Brunet, A., Rando, T. 2023; 30 (1-17)

  View details for DOI 10.1016/j.stem.2023.03.016

• Tubastatin A maintains adult skeletal muscle stem cells in a quiescent state ex vivo and improves their engraftment ability in vivo. Stem cell reports Arjona, M., Goshayeshi, A., Rodriguez-Mateo, C., Brett, J. O., Both, P., Ishak, H., Rando, T. A. 2022; 17 (1): 82-95

  Abstract

  Adult skeletal muscle stem cells (MuSCs) are important for muscle regeneration and constitute a potential source of cell therapy. However, upon isolation, MuSCs rapidly exit quiescence and lose transplantation potency. Maintenance of the quiescent state in vitro preserves MuSC transplantation efficiency and provides an opportunity to study the biology of quiescence. Here we show that Tubastatin A (TubA), an Hdac6 inhibitor, prevents primary cilium resorption, maintains quiescence, and enhances MuSC survival ex vivo. Phenotypic characterization and transcriptomic analysis of TubA-treated cells revealed that TubA maintains most of the biological features and molecular signatures of quiescence. Furthermore, TubA-treated MuSCs showed improved engraftment ability upon transplantation. TubA also induced a return to quiescence and improved engraftment of cycling MuSCs, revealing a potentially expanded application for MuSC therapeutics. Altogether, these studies demonstrate the ability of TubA to maintain MuSC quiescence ex vivo and to enhance the therapeutic potential of MuSCs and their progeny.

  View details for DOI 10.1016/j.stemcr.2021.11.012

  View details for PubMedID 35021050

• Exercise plasma boosts memory and dampens brain inflammation via clusterin. Nature De Miguel, Z., Khoury, N., Betley, M. J., Lehallier, B., Willoughby, D., Olsson, N., Yang, A. C., Hahn, O., Lu, N., Vest, R. T., Bonanno, L. N., Yerra, L., Zhang, L., Saw, N. L., Fairchild, J. K., Lee, D., Zhang, H., McAlpine, P. L., Contrepois, K., Shamloo, M., Elias, J. E., Rando, T. A., Wyss-Coray, T. 2021

  Abstract

  Physical exercise is generally beneficial to all aspects of human and animal health, slowing cognitive ageing and neurodegeneration1. The cognitive benefits of physical exercise are tied to an increased plasticity and reduced inflammation within the hippocampus2-4, yet little is known about the factors and mechanisms that mediate these effects. Here we show that 'runner plasma', collected from voluntarily running mice and infused into sedentary mice, reduces baseline neuroinflammatory gene expression and experimentally induced brain inflammation. Plasma proteomic analysis revealed a concerted increase in complement cascade inhibitors including clusterin (CLU). Intravenously injected CLU binds to brain endothelial cells and reduces neuroinflammatory gene expression in a mouse model of acute brain inflammation and a mouse model of Alzheimer's disease. Patients with cognitive impairment who participated in structured exercise for 6 months had higher plasma levels of CLU. These findings demonstrate the existence of anti-inflammatory exercise factors that are transferrable, target the cerebrovasculature and benefit the brain, and are present in humans who engage in exercise.

  View details for DOI 10.1038/s41586-021-04183-x

  View details for PubMedID 34880498

• Cells, scaffolds, and bioactive factors: Engineering strategies for improving regeneration following volumetric muscle loss. Biomaterials Eugenis, I., Wu, D., Rando, T. A. 2021; 278: 121173

  Abstract

  Severe traumatic skeletal muscle injuries, such as volumetric muscle loss (VML), result in the obliteration of large amounts of skeletal muscle and lead to permanent functional impairment. Current clinical treatments are limited in their capacity to regenerate damaged muscle and restore tissue function, promoting the need for novel muscle regeneration strategies. Advances in tissue engineering, including cell therapy, scaffold design, and bioactive factor delivery, are promising solutions for VML therapy. Herein, we review tissue engineering strategies for regeneration of skeletal muscle, development of vasculature and nerve within the damaged muscle, and achievements in immunomodulation following VML. In addition, we discuss the limitations of current state of the art technologies and perspectives of tissue-engineered bioconstructs for muscle regeneration and functional recovery following VML.

  View details for DOI 10.1016/j.biomaterials.2021.121173

  View details for PubMedID 34619561

• Hairless regulates heterochromatin maintenance and muscle stem cell function as a histone demethylase antagonist. Proceedings of the National Academy of Sciences of the United States of America Liu, L., Rodriguez-Mateo, C., Huang, P., Huang, A., Lieu, A., Mao, M., Chung, M., Yang, S., Yu, K., Charville, G. W., Gan, Q., Rando, T. A. 2021; 118 (37)

  Abstract

  Skeletal muscle possesses remarkable regenerative ability because of the resident muscle stem cells (MuSCs). A prominent feature of quiescent MuSCs is a high content of heterochromatin. However, little is known about the mechanisms by which heterochromatin is maintained in MuSCs. By comparing gene-expression profiles from quiescent and activated MuSCs, we found that the mammalian Hairless (Hr) gene is expressed in quiescent MuSCs and rapidly down-regulated upon MuSC activation. Using a mouse model in which Hr can be specifically ablated in MuSCs, we demonstrate that Hr expression is critical for MuSC function and muscle regeneration. In MuSCs, loss of Hr results in reduced trimethylated Histone 3 Lysine 9 (H3K9me3) levels, reduced heterochromatin, increased susceptibility to genotoxic stress, and the accumulation of DNA damage. Deletion of Hr leads to an acceleration of the age-related decline in MuSC numbers. We have also demonstrated that despite the fact that Hr is homologous to a family of histone demethylases and binds to di- and trimethylated H3K9, the expression of Hr does not lead to H3K9 demethylation. In contrast, we show that the expression of Hr leads to the inhibition of the H3K9 demethylase Jmjd1a and an increase in H3K9 methylation. Taking these data together, our study has established that Hr is a H3K9 demethylase antagonist specifically expressed in quiescent MuSCs.

  View details for DOI 10.1073/pnas.2025281118

  View details for PubMedID 34493660

• RNA splicing programs define tissue compartments and cell types at single-cell resolution ELIFE Olivieri, J., Dehghannasiri, R., Wang, P. L., Jang, S., de Morree, A., Tan, S. Y., Ming, J., Wu, A., Consortium, T., Quake, S. R., Krasnow, M. A., Salzman, J. 2021; 10

  View details for DOI 10.7554/eLife.70692.sa2

  View details for Web of Science ID 000715795700001

• Regeneration, Rejuvenation, and Replacement: Turning Back the Clock on Tissue Aging. Cold Spring Harbor perspectives in biology Rando, T. A., Jones, D. L. 2021

  Abstract

  While some animals, such as planaria and hydra, appear to be capable of seemingly endless cycles of regeneration, most animals experience a gradual decline in fitness and ultimately die. The progressive loss of cell and tissue function, leading to senescence and death, is generally referred to as aging. Adult ("tissue") stem cells maintain tissue homeostasis and facilitate repair; however, age-related changes in stem cell function over time are major contributors to loss of organ function or disease in older individuals. Therefore, considerable effort is being invested in restoring stem cell function to counter degenerative diseases and age-related tissue dysfunction. Here, we will review strategies that could be used to restore stem cell function, including the use of environmental interventions, such as diet and exercise, heterochronic approaches, and cellular reprogramming. Maintaining optimal stem cell function and tissue homeostasis into late life will likely extend the amount of time older adults are able to be independent and lead healthy lives.

  View details for DOI 10.1101/cshperspect.a040907

  View details for PubMedID 34187808

• Context-dependent modulation of aggressiveness of pediatric tumors by individual oncogenic RAS isoforms. Oncogene Bauer, J., Cuvelier, N., Ragab, N., Simon-Keller, K., Nitzki, F., Geyer, N., Botermann, D. S., Elmer, D. P., Rosenberger, A., Rando, T. A., Biressi, S., Fagin, J. A., Saur, D., Dullin, C., Schildhaus, H., Schulz-Schaeffer, W., Aberger, F., Uhmann, A., Hahn, H. 2021

  Abstract

  A prototypic pediatric cancer that frequently shows activation of RAS signaling is embryonal rhabdomyosarcoma (ERMS). ERMS also show aberrant Hedgehog (HH)/GLI signaling activity and can be driven by germline mutations in this pathway. We show, that in ERMS cell lines derived from sporadic tumors i.e. from tumors not caused by an inherited genetic variant, HH/GLI signaling plays a subordinate role, because oncogenic mutations in HRAS, KRAS, or NRAS (collectively named oncRAS) inhibit the main HH target GLI1 via the MEK/ERK-axis, but simultaneously increase proliferation and tumorigenicity. oncRAS also modulate expression of stem cell markers in an isoform- and context-dependent manner. In Hh-driven murine ERMS that are caused by a Patched mutation, oncHRAS and mainly oncKRAS accelerate tumor development, whereas oncNRAS induces a more differentiated phenotype. These features occur when the oncRAS mutations are induced at the ERMS precursor stage, but not when induced in already established tumors. Moreover, in contrast to what is seen in human cell lines, oncRAS mutations do not alter Hh signaling activity and marginally affect expression of stem cell markers. Together, all three oncRAS mutations seem to be advantageous for ERMS cell lines despite inhibition of HH signaling and isoform-specific modulation of stem cell markers. In contrast, oncRAS mutations do not inhibit Hh-signaling in Hh-driven ERMS. In this model, oncRAS mutations seem to be advantageous for specific ERMS populations that occur within a specific time window during ERMS development. In addition, this window may be different for individual oncRAS isoforms, at least in the mouse.

  View details for DOI 10.1038/s41388-021-01904-4

  View details for PubMedID 34172934

• Electrical stimulation of human neural stem cells via conductive polymer nerve guides enhances peripheral nerve recovery. Biomaterials Song, S., McConnell, K. W., Amores, D., Levinson, A., Vogel, H., Quarta, M., Rando, T. A., George, P. M. 2021; 275: 120982

  Abstract

  Severe peripheral nerve injuries often result in permanent loss of function of the affected limb. Current treatments are limited by their efficacy in supporting nerve regeneration and behavioral recovery. Here we demonstrate that electrical stimulation through conductive nerve guides (CNGs) enhances the efficacy of human neural progenitor cells (hNPCs) in treating a sciatic nerve transection in rats. Electrical stimulation strengthened the therapeutic potential of NPCs by upregulating gene expression of neurotrophic factors which are critical in augmenting synaptic remodeling, nerve regeneration, and myelination. Electrically-stimulated hNPC-containing CNGs are significantly more effective in improving sensory and motor functions starting at 1-2 weeks after treatment than either treatment alone. Electrophysiology and muscle assessment demonstrated successful re-innervation of the affected target muscles in this group. Furthermore, histological analysis highlighted an increased number of regenerated nerve fibers with thicker myelination in electrically-stimulated hNPC-containing CNGs. The elevated expression of tyrosine kinase receptors (Trk) receptors, known to bind to neurotrophic factors, indicated the long-lasting effect from electrical stimulation on nerve regeneration and distal nerve re-innervation. These data suggest that electrically-enhanced stem cell-based therapy provides a regenerative rehabilitative approach to promote peripheral nerve regeneration and functional recovery.

  View details for DOI 10.1016/j.biomaterials.2021.120982

  View details for PubMedID 34214785

• Asynchronous, contagious and digital aging. Nature aging Rando, T. A., Wyss-Coray, T. 2021; 1 (1): 29-35

  Abstract

  Aging has largely been defined by analog measures of organ and organismal dysfunction. This has led to the characterization of aging processes at the molecular and cellular levels that underlie these gradual changes. However, current knowledge does not fully explain the growing body of data emerging from large epidemiological, systems biology, and single cell studies of entire organisms pointing to varied rates of aging between individuals (different functionality and lifespan), across lifespan (asynchronous aging), and within an organism at the tissue and organ levels (aging mosaicism). Here we consider these inhomogeneities in the broader context of the rate of aging and from the perspective of underlying cellular changes. These changes reflect genetic, environmental, and stochastic factors that cells integrate to adopt new homeostatic, albeit less functional, states, such as cellular senescence. In this sense, cellular aging can be viewed, at least in part, as a quantal process we refer to as "digital aging". Nevertheless, analog declines of tissue dysfunction and organ failure with age could be the sum of underlying digital events. Importantly, cellular aging, digital or otherwise, is not uniform across time or space within the organism or between organisms of the same species. Certain tissues may exhibit earliest signs of cellular aging, acting as drivers for organismal aging as signals from those driver cells within those tissues may accelerate the aging of other cells locally or even systemically. Advanced methodologies at the systems level and at the single cell level are likely to continue to refine our understanding to the processes of how cells and tissues age and how the integration of those processes leads to the complexities of individual, organismal aging.

  View details for DOI 10.1038/s43587-020-00015-1

  View details for PubMedID 34223194

• Targeting microRNA-mediated gene repression limits adipogenic conversion of skeletal muscle mesenchymal stromal cells. Cell stem cell Wosczyna, M. N., Perez Carbajal, E. E., Wagner, M. W., Paredes, S. n., Konishi, C. T., Liu, L. n., Wang, T. T., Walsh, R. A., Gan, Q. n., Morrissey, C. S., Rando, T. A. 2021

  Abstract

  Intramuscular fatty deposits, which are seen in muscular dystrophies and with aging, negatively affect muscle function. The cells of origin of adipocytes constituting these fatty deposits are mesenchymal stromal cells, fibroadipogenic progenitors (FAPs). We uncover a molecular fate switch, involving miR-206 and the transcription factor Runx1, that controls FAP differentiation to adipocytes. Mice deficient in miR-206 exhibit increased adipogenesis following muscle injury. Adipogenic differentiation of FAPs is abrogated by miR-206 mimics. Using a labeled microRNA (miRNA) pull-down and sequencing (LAMP-seq), we identified Runx1 as a miR-206 target, with miR-206 repressing Runx1 translation. In the absence of miR-206 in FAPs, Runx1 occupancy near transcriptional start sites of adipogenic genes and expression of these genes increase. We demonstrate that miR-206 mimicry in vivo limits intramuscular fatty infiltration. Our results provide insight into the underlying molecular mechanisms of FAP fate determination and formation of harmful fatty deposits in skeletal muscle.

  View details for DOI 10.1016/j.stem.2021.04.008

  View details for PubMedID 33945794

• Computational modeling of malignant ascites reveals CCL5-SDC4 interaction in the immune microenvironment of ovarian cancer. Molecular carcinogenesis Kim, S. n., Han, Y. n., Kim, S. I., Lee, J. n., Jo, H. n., Wang, W. n., Cho, U. n., Park, W. Y., Rando, T. A., Dhanasekaran, D. N., Song, Y. S. 2021

  Abstract

  Fluid accumulation in the abdominal cavity is commonly found in advanced-stage ovarian cancer patients, which creates a specialized tumor microenvironment for cancer progression. Using single-cell RNA sequencing (scRNA-seq) of ascites cells from five patients with ovarian cancer, we identified seven cell types, including heterogeneous macrophages and ovarian cancer cells. We resolved a distinct polarization state of macrophages by MacSpectrum analysis and observed subtype-specific enrichment of pathways associated with their functions. The communication between immune and cancer cells was predicted through a putative ligand-receptor pair analysis using NicheNet. We found that CCL5, a chemotactic ligand, is enriched in immune cells (T cells and NK cells) and mediates ovarian cancer cell survival in the ascites, possibly through SDC4. Moreover, SDC4 expression correlated with poor overall survival in ovarian cancer patients. Our study highlights the potential role of T cells and NK cells in long-term survival patients with ovarian cancer, indicating SDC4 as a potential prognostic marker in ovarian cancer patients.

  View details for DOI 10.1002/mc.23289

  View details for PubMedID 33721368

• Functional redundancy of type I and type II receptors in the regulation of skeletal muscle growth by myostatin and activin A. Proceedings of the National Academy of Sciences of the United States of America Lee, S., Lehar, A., Liu, Y., Ly, C. H., Pham, Q., Michaud, M., Rydzik, R., Youngstrom, D. W., Shen, M. M., Kaartinen, V., Germain-Lee, E. L., Rando, T. A. 2020

  Abstract

  Myostatin (MSTN) is a transforming growth factor-beta (TGF-beta) family member that normally acts to limit muscle growth. The function of MSTN is partially redundant with that of another TGF-beta family member, activin A. MSTN and activin A are capable of signaling through a complex of type II and type I receptors. Here, we investigated the roles of two type II receptors (ACVR2 and ACVR2B) and two type I receptors (ALK4 and ALK5) in the regulation of muscle mass by these ligands by genetically targeting these receptors either alone or in combination specifically in myofibers in mice. We show that targeting signaling in myofibers is sufficient to cause significant increases in muscle mass, showing that myofibers are the direct target for signaling by these ligands in the regulation of muscle growth. Moreover, we show that there is functional redundancy between the two type II receptors as well as between the two type I receptors and that all four type II/type I receptor combinations are utilized in vivo. Targeting signaling specifically in myofibers also led to reductions in overall body fat content and improved glucose metabolism in mice fed either regular chow or a high-fat diet, demonstrating that these metabolic effects are the result of enhanced muscling. We observed no effect, however, on either bone density or muscle regeneration in mice in which signaling was targeted in myofibers. The latter finding implies that MSTN likely signals to other cells, such as satellite cells, in addition to myofibers to regulate muscle homeostasis.

  View details for DOI 10.1073/pnas.2019263117

  View details for PubMedID 33219121

• Transplantation of insulin-like growth factor-1 laden scaffolds combined with exercise promotes neuroregeneration and angiogenesis in a preclinical muscle injury model BIOMATERIALS SCIENCE Alcazar, C. A., Hu, C., Rando, T. A., Huang, N. F., Nakayama, K. H. 2020; 8 (19): 5376–89

  Abstract

  The regeneration of skeletal muscle can be permanently impaired by traumatic injuries, despite the high regenerative capacity of native muscle. An attractive therapeutic approach for treating severe muscle inuries is the implantation of off-the-shelf engineered biomimetic scaffolds into the site of tissue damage to enhance muscle regeneration. Anisotropic nanofibrillar scaffolds provide spatial patterning cues to create organized myofibers, and growth factors such as insulin-like growth factor-1 (IGF-1) are potent inducers of both muscle regeneration as well as angiogenesis. The aim of this study was to test the therapeutic efficacy of anisotropic IGF-1-releasing collagen scaffolds combined with voluntary exercise for the treatment of acute volumetric muscle loss, with a focus on histomorphological effects. To enhance the angiogenic and regenerative potential of injured murine skeletal muscle, IGF-1-laden nanofibrillar scaffolds with aligned topography were fabricated using a shear-mediated extrusion approach, followed by growth factor adsorption. Individual scaffolds released a cumulative total of 1244 ng ± 153 ng of IGF-1 over the course of 21 days in vitro. To test the bioactivity of IGF-1-releasing scaffolds, the myotube formation capacity of murine myoblasts was quantified. On IGF-1-releasing scaffolds seeded with myoblasts, the resulting myotubes formed were 1.5-fold longer in length and contained 2-fold greater nuclei per myotube, when compared to scaffolds without IGF-1. When implanted into the ablated murine tibialis anterior muscle, the IGF-1-laden scaffolds, in conjunction with voluntary wheel running, significantly increased the density of perfused microvessels by greater than 3-fold, in comparison to treatment with scaffolds without IGF-1. Enhanced myogenesis was also observed in animals treated with the IGF-1-laden scaffolds combined with exercise, compared to control scaffolds transplanted into mice that did not receive exercise. Furthermore, the abundance of mature neuromuscular junctions was greater by approximately 2-fold in muscles treated with IGF-1-laden scaffolds, when paired with exercise, in comparison to the same treatment without exercise. These findings demonstrate that voluntary exercise improves the regenerative effect of growth factor-laden scaffolds by augmenting neurovascular regeneration, and have important translational implications in the design of off-the-shelf therapeutics for the treatment of traumatic muscle injury.

  View details for DOI 10.1039/d0bm00990c

  View details for Web of Science ID 000573820100008

  View details for PubMedID 32996916

  View details for PubMedCentralID PMC7531607

• Exercise rejuvenates quiescent skeletal muscle stem cells in old mice through restoration of Cyclin D1. Nature metabolism Brett, J. O., Arjona, M., Ikeda, M., Quarta, M., de Morrée, A., Egner, I. M., Perandini, L. A., Ishak, H. D., Goshayeshi, A., Benjamin, D. I., Both, P., Rodríguez-Mateo, C., Betley, M. J., Wyss-Coray, T., Rando, T. A. 2020; 2 (4): 307-317

  Abstract

  Aging impairs tissue repair. This is pronounced in skeletal muscle, whose regeneration by muscle stem cells (MuSCs) is robust in young adult animals but inefficient in older organisms. Despite this functional decline, old MuSCs are amenable to rejuvenation through strategies that improve the systemic milieu, such as heterochronic parabiosis. One such strategy, exercise, has long been appreciated for its benefits on healthspan, but its effects on aged stem cell function in the context of tissue regeneration are incompletely understood. Here we show that exercise in the form of voluntary wheel running accelerates muscle repair in old animals and improves old MuSC function. Through transcriptional profiling and genetic studies, we discovered that the restoration of old MuSC activation ability hinges on restoration of Cyclin D1, whose expression declines with age in MuSCs. Pharmacologic studies revealed that Cyclin D1 maintains MuSC activation capacity by repressing TGFβ signaling. Taken together, these studies demonstrate that voluntary exercise is a practicable intervention for old MuSC rejuvenation. Furthermore, this work highlights the distinct role of Cyclin D1 in stem cell quiescence.

  View details for DOI 10.1038/s42255-020-0190-0

  View details for PubMedID 32601609

  View details for PubMedCentralID PMC7323974

• A single-cell transcriptomic atlas characterizes ageing tissues in the mouse. Nature 2020

  Abstract

  Ageing is characterized by a progressive loss of physiological integrity, leading to impaired function and increased vulnerability to death1. Despite rapid advances over recent years, many of the molecular and cellular processes that underlie the progressive loss of healthy physiology are poorly understood2. To gain a better insight into these processes, here we generate a single-cell transcriptomic atlas across the lifespan of Mus musculus that includes data from 23 tissues and organs. We found cell-specific changes occurring across multiple cell types and organs, as well as age-related changes in the cellular composition of different organs. Using single-cell transcriptomic data, we assessed cell-type-specific manifestations of different hallmarks of ageing-such as senescence3, genomic instability4 and changes in the immune system2. This transcriptomic atlas-which we denote Tabula Muris Senis, or 'Mouse Ageing Cell Atlas'-provides molecular information about how the most important hallmarks of ageing are reflected in a broad range of tissues and cell types.

  View details for DOI 10.1038/s41586-020-2496-1

  View details for PubMedID 32669714

• Stem cell therapy for muscular dystrophies. The Journal of clinical investigation Biressi, S. n., Filareto, A. n., Rando, T. A. 2020

  Abstract

  Muscular dystrophies are a heterogeneous group of genetic diseases, characterized by progressive degeneration of skeletal and cardiac muscle. Despite the intense investigation of different therapeutic options, a definitive treatment has not been yet developed for this debilitating class of pathologies. Cell-based therapies in muscular dystrophies have been pursued experimentally for the last three decades. Several cell types with different characteristics and tissues of origin, including myogenic stem and progenitor cells, stromal cells, and pluripotent stem cells, have been investigated over the years and have recently entered in the clinical arena with mixed results. In this review, we will do a roundup of the past attempts and describe the updated status of cell-based therapies aimed at counteracting the skeletal and cardiac myopathy present in dystrophic patients. We will present current challenges, summarize recent progresses, and make recommendations for future research and clinical trials.

  View details for DOI 10.1172/JCI142031

  View details for PubMedID 32946430

• Aging of the skeletal muscle extracellular matrix drives a stem cell fibrogenic conversion Stearns-Reider, K. M., D'Amore, A., Beezhold, K., Rothrauff, B., Cavalli, L., Wagner, W. R., Vorp, D. A., Tsamis, A., Shinde, S., Zhang, C., Barchowsky, A., Rando, T. A., Tuan, R. S., Ambrosio, F. WILEY. 2020: 189

  View details for Web of Science ID 000619403800040

• ARDD 2020: from aging mechanisms to interventions. Aging Mkrtchyan, G. V., Abdelmohsen, K. n., Andreux, P. n., Bagdonaite, I. n., Barzilai, N. n., Brunak, S. n., Cabreiro, F. n., de Cabo, R. n., Campisi, J. n., Cuervo, A. M., Demaria, M. n., Ewald, C. Y., Fang, E. F., Faragher, R. n., Ferrucci, L. n., Freund, A. n., Silva-García, C. G., Georgievskaya, A. n., Gladyshev, V. N., Glass, D. J., Gorbunova, V. n., de Grey, A. n., He, W. W., Hoeijmakers, J. n., Hoffmann, E. n., Horvath, S. n., Houtkooper, R. H., Jensen, M. K., Jensen, M. B., Kane, A. n., Kassem, M. n., de Keizer, P. n., Kennedy, B. n., Karsenty, G. n., Lamming, D. W., Lee, K. F., MacAulay, N. n., Mamoshina, P. n., Mellon, J. n., Molenaars, M. n., Moskalev, A. n., Mund, A. n., Niedernhofer, L. n., Osborne, B. n., Pak, H. H., Parkhitko, A. n., Raimundo, N. n., Rando, T. A., Rasmussen, L. J., Reis, C. n., Riedel, C. G., Franco-Romero, A. n., Schumacher, B. n., Sinclair, D. A., Suh, Y. n., Taub, P. R., Toiber, D. n., Treebak, J. T., Valenzano, D. R., Verdin, E. n., Vijg, J. n., Young, S. n., Zhang, L. n., Bakula, D. n., Zhavoronkov, A. n., Scheibye-Knudsen, M. n. 2020; 12

  Abstract

  Aging is emerging as a druggable target with growing interest from academia, industry and investors. New technologies such as artificial intelligence and advanced screening techniques, as well as a strong influence from the industry sector may lead to novel discoveries to treat age-related diseases. The present review summarizes presentations from the 7th Annual Aging Research and Drug Discovery (ARDD) meeting, held online on the 1st to 4th of September 2020. The meeting covered topics related to new methodologies to study aging, knowledge about basic mechanisms of longevity, latest interventional strategies to target the aging process as well as discussions about the impact of aging research on society and economy. More than 2000 participants and 65 speakers joined the meeting and we already look forward to an even larger meeting next year. Please mark your calendars for the 8th ARDD meeting that is scheduled for the 31st of August to 3rd of September, 2021, at Columbia University, USA.

  View details for DOI 10.18632/aging.202454

  View details for PubMedID 33378272

• Transient non-integrative expression of nuclear reprogramming factors promotes multifaceted amelioration of aging in human cells. Nature communications Sarkar, T. J., Quarta, M. n., Mukherjee, S. n., Colville, A. n., Paine, P. n., Doan, L. n., Tran, C. M., Chu, C. R., Horvath, S. n., Qi, L. S., Bhutani, N. n., Rando, T. A., Sebastiano, V. n. 2020; 11 (1): 1545

  Abstract

  Aging is characterized by a gradual loss of function occurring at the molecular, cellular, tissue and organismal levels. At the chromatin level, aging associates with progressive accumulation of epigenetic errors that eventually lead to aberrant gene regulation, stem cell exhaustion, senescence, and deregulated cell/tissue homeostasis. Nuclear reprogramming to pluripotency can revert both the age and the identity of any cell to that of an embryonic cell. Recent evidence shows that transient reprogramming can ameliorate age-associated hallmarks and extend lifespan in progeroid mice. However, it is unknown how this form of rejuvenation would apply to naturally aged human cells. Here we show that transient expression of nuclear reprogramming factors, mediated by expression of mRNAs, promotes a rapid and broad amelioration of cellular aging, including resetting of epigenetic clock, reduction of the inflammatory profile in chondrocytes, and restoration of youthful regenerative response to aged, human muscle stem cells, in each case without abolishing cellular identity.

  View details for DOI 10.1038/s41467-020-15174-3

  View details for PubMedID 32210226

• Taking the Next Steps in Regenerative Rehabilitation: Establishment of a New Interdisciplinary Field. Archives of physical medicine and rehabilitation Willett, N. J., Boninger, M. L., Miller, L. J., Alvarez, L. n., Aoyama, T. n., Bedoni, M. n., Brix, K. A., Chisari, C. n., Christ, G. n., Dearth, C. L., Dyson-Hudson, T. A., Evans, C. H., Goldman, S. M., Gregory, K. n., Gualerzi, A. n., Hart, J. n., Ito, A. n., Kuroki, H. n., Loghmani, M. T., Mack, D. L., Malanga, G. A., Noble-Haeusslein, L. n., Pasquina, P. n., Roche, J. A., Rose, L. n., Stoddart, M. J., Tajino, J. n., Terzic, C. n., Topp, K. S., Wagner, W. R., Warden, S. J., Wolf, S. L., Xie, H. n., Rando, T. A., Ambrosio, F. n. 2020

  Abstract

  The growing field of Regenerative Rehabilitation has great potential to improve clinical outcomes for individuals with disabilities. However, the science to elucidate the specific biological underpinnings of Regenerative Rehabilitation-based approaches is still in its infancy and critical questions regarding clinical translation and implementation still exist. In a recent roundtable discussion from International Consortium for Regenerative Rehabilitation (ICRR) stakeholders, key challenges to progress in the field were identified. The goal of this white paper is to summarize those discussions and to initiate a broader discussion among clinicians and scientists across the fields of regenerative medicine and rehabilitation science to ultimately progress Regenerative Rehabilitation from an emerging field to an established interdisciplinary one. Strategies and case studies from Consortium institutions-including interdisciplinary research centers, formalized courses, degree programs, international symposia, and collaborative grants-are presented. We propose that these strategic directions have the potential to engage and train clinical practitioners and basic scientists, transform clinical practice and, ultimately, optimize patient outcomes.

  View details for DOI 10.1016/j.apmr.2020.01.007

  View details for PubMedID 32035141

• Ageing hallmarks exhibit organ-specific temporal signatures. Nature Schaum, N. n., Lehallier, B. n., Hahn, O. n., Pálovics, R. n., Hosseinzadeh, S. n., Lee, S. E., Sit, R. n., Lee, D. P., Losada, P. M., Zardeneta, M. E., Fehlmann, T. n., Webber, J. T., McGeever, A. n., Calcuttawala, K. n., Zhang, H. n., Berdnik, D. n., Mathur, V. n., Tan, W. n., Zee, A. n., Tan, M. n., Pisco, A. O., Karkanias, J. n., Neff, N. F., Keller, A. n., Darmanis, S. n., Quake, S. R., Wyss-Coray, T. n. 2020

  Abstract

  Ageing is the single greatest cause of disease and death worldwide, and understanding the associated processes could vastly improve quality of life. Although major categories of ageing damage have been identified-such as altered intercellular communication, loss of proteostasis and eroded mitochondrial function1-these deleterious processes interact with extraordinary complexity within and between organs, and a comprehensive, whole-organism analysis of ageing dynamics has been lacking. Here we performed bulk RNA sequencing of 17 organs and plasma proteomics at 10 ages across the lifespan of Mus musculus, and integrated these findings with data from the accompanying Tabula Muris Senis2-or 'Mouse Ageing Cell Atlas'-which follows on from the original Tabula Muris3. We reveal linear and nonlinear shifts in gene expression during ageing, with the associated genes clustered in consistent trajectory groups with coherent biological functions-including extracellular matrix regulation, unfolded protein binding, mitochondrial function, and inflammatory and immune response. Notably, these gene sets show similar expression across tissues, differing only in the amplitude and the age of onset of expression. Widespread activation of immune cells is especially pronounced, and is first detectable in white adipose depots during middle age. Single-cell RNA sequencing confirms the accumulation of T cells and B cells in adipose tissue-including plasma cells that express immunoglobulin J-which also accrue concurrently across diverse organs. Finally, we show how gene expression shifts in distinct tissues are highly correlated with corresponding protein levels in plasma, thus potentially contributing to the ageing of the systemic circulation. Together, these data demonstrate a similar yet asynchronous inter- and intra-organ progression of ageing, providing a foundation from which to track systemic sources of declining health at old age.

  View details for DOI 10.1038/s41586-020-2499-y

  View details for PubMedID 32669715

• Adult stem cells and regenerative medicine-a symposium report. Annals of the New York Academy of Sciences Cable, J., Fuchs, E., Weissman, I., Jasper, H., Glass, D., Rando, T. A., Blau, H., Debnath, S., Oliva, A., Park, S., Passegue, E., Kim, C., Krasnow, M. A. 2019

  Abstract

  Adult stem cells are rare, undifferentiated cells found in all tissues of the body. Although normally kept in a quiescent, nondividing state, these cells can proliferate and differentiate to replace naturally dying cells within their tissue and to repair its wounds in response to injury. Due to their proliferative nature and ability to regenerate tissue, adult stem cells have the potential to treat a variety of degenerative diseases as well as aging. In addition, since stem cells are often thought to be the source of malignant tumors, understanding the mechanisms that keep their proliferative abilities in check can pave the way for new cancer therapies. While adult stem cells have had limited practical and clinical applications to date, several clinical trials of stem cell-based therapies are underway. This report details recent research presented at the New York Academy of Sciences on March 14, 2019 on understanding the factors that regulate stem cell activity and differentiation, with the hope of translating these findings into the clinic.

  View details for DOI 10.1111/nyas.14243

  View details for PubMedID 31655007

• mTORC1 underlies age-related muscle fiber damage and loss by inducing oxidative stress and catabolism AGING CELL Tang, H., Inoki, K., Brooks, S. V., Okazawa, H., Lee, M., Wang, J., Kim, M., Kennedy, C. L., Macpherson, P. D., Ji, X., Van Roekel, S., Fraga, D. A., Wang, K., Zhu, J., Wang, Y., Sharp, Z. D., Miller, R. A., Rando, T. A., Goldman, D., Guan, K., Shrager, J. B. 2019; 18 (3)

  View details for DOI 10.1111/acel.12943

  View details for Web of Science ID 000467861100044

• Mesenchymal Stromal Cells Are Required for Regeneration and Homeostatic Maintenance of Skeletal Muscle CELL REPORTS Wosczyna, M. N., Konishi, C. T., Carbajal, E., Wang, T. T., Walsh, R. A., Gan, Q., Wagner, M. W., Rando, T. A. 2019; 27 (7): 2029-+

  View details for DOI 10.1016/j.celrep.2019.04.074

  View details for Web of Science ID 000467811500009

• Treatment of volumetric muscle loss in mice using nanofibrillar scaffolds enhances vascular organization and integration COMMUNICATIONS BIOLOGY Nakayama, K. H., Quarta, M., Paine, P., Alcazar, C., Karakikes, I., Garcia, V., Abilez, O. J., Calvo, N. S., Simmons, C. S., Rando, T. A., Huang, N. F. 2019; 2

  View details for DOI 10.1038/s42003-019-0416-4

  View details for Web of Science ID 000467214900002

• Treatment of volumetric muscle loss in mice using nanofibrillar scaffolds enhances vascular organization and integration. Communications biology Nakayama, K. H., Quarta, M., Paine, P., Alcazar, C., Karakikes, I., Garcia, V., Abilez, O. J., Calvo, N. S., Simmons, C. S., Rando, T. A., Huang, N. F. 2019; 2: 170

  Abstract

  Traumatic skeletal muscle injuries cause irreversible tissue damage and impaired revascularization. Engineered muscle is promising for enhancing tissue revascularization and regeneration in injured muscle. Here we fabricated engineered skeletal muscle composed of myotubes interspersed with vascular endothelial cells using spatially patterned scaffolds that induce aligned cellular organization, and then assessed their therapeutic benefit for treatment of murine volumetric muscle loss. Murine skeletal myoblasts co-cultured with endothelial cells in aligned nanofibrillar scaffolds form endothelialized and aligned muscle with longer myotubes, more synchronized contractility, and more abundant secretion of angiogenic cytokines, compared to endothelialized engineered muscle formed from randomly-oriented scaffolds. Treatment of traumatically injured muscle with endothelialized and aligned skeletal muscle promotes the formation of highly organized myofibers and microvasculature, along with greater vascular perfusion, compared to treatment of muscle derived from randomly-oriented scaffolds. This work demonstrates the potential of endothelialized and aligned engineered skeletal muscle to promote vascular regeneration following transplantation.

  View details for DOI 10.1038/s42003-019-0416-4

  View details for PubMedID 31098403

  View details for PubMedCentralID PMC6505043

• mTORC1 underlies age-related muscle fiber damage and loss by inducing oxidative stress and catabolism. Aging cell Tang, H., Inoki, K., Brooks, S. V., Okazawa, H., Lee, M., Wang, J., Kim, M., Kennedy, C. L., Macpherson, P. C., Ji, X., Van Roekel, S., Fraga, D. A., Wang, K., Zhu, J., Wang, Y., Sharp, Z. D., Miller, R. A., Rando, T. A., Goldman, D., Guan, K., Shrager, J. B. 2019: e12943

  Abstract

  Aging leads to skeletal muscle atrophy (i.e., sarcopenia), and muscle fiber loss is a critical component of this process. The mechanisms underlying these age-related changes, however, remain unclear. We show here that mTORC1 signaling is activated in a subset of skeletal muscle fibers in aging mouse and human, colocalized with fiber damage. Activation of mTORC1 in TSC1 knockout mouse muscle fibers increases the content of morphologically abnormal mitochondria and causes progressive oxidative stress, fiber damage, and fiber loss over the lifespan. Transcriptomic profiling reveals that mTORC1's activation increases the expression of growth differentiation factors (GDF3, 5, and 15), and of genes involved in mitochondrial oxidative stress and catabolism. We show that increased GDF15 is sufficient to induce oxidative stress and catabolic changes, and that mTORC1 increases the expression of GDF15 via phosphorylation of STAT3. Inhibition of mTORC1 in aging mouse decreases the expression of GDFs and STAT3's phosphorylation in skeletal muscle, reducing oxidative stress and muscle fiber damage and loss. Thus, chronically increased mTORC1 activity contributes to age-related muscle atrophy, and GDF signaling is a proposed mechanism.

  View details for PubMedID 30924297

• Models and Studies of Aging: Executive Summary of a Report from the U13 Conference Series JOURNAL OF THE AMERICAN GERIATRICS SOCIETY Hurria, A., Carpenter, C. R., McFarland, F., Lundebjerg, N. E., de Cabo, R., Ferrucci, L., Studenski, S. A., Barzilai, N., Briggs, J. P., Ix, J. H., Kitzman, D. W., Kuchel, G. A., Musi, N., Newman, J. C., Rando, T. A., Smith, A. K., Walston, J. D., Kirkland, J. L., Yung, R. 2019; 67 (3): 428–33

  View details for DOI 10.1111/jgs.15788

  View details for Web of Science ID 000461567700004

• Stem Cell Quiescence: Dynamism, Restraint, and Cellular Idling CELL STEM CELL van Velthoven, C. J., Rando, T. A. 2019; 24 (2): 213–25

  View details for DOI 10.1016/j.stem.2019.01.001

  View details for Web of Science ID 000458027300009

• Stem Cell Quiescence: Dynamism, Restraint, and Cellular Idling. Cell stem cell van Velthoven, C. T., Rando, T. A. 2019; 24 (2): 213–25

  Abstract

  Stem cells can reside in a state of reversible growth arrest, or quiescence, for prolonged periods of time. Although quiescence has long been viewed as a dormant, low-activity state, increasing evidence suggests that quiescence represents states of poised potential and active restraint, as stem cells "idle" in anticipation of activation, proliferation, and differentiation. Improved understanding of quiescent stem cell dynamics is leading to novel approaches to enhance maintenance and repair of aged or diseased tissues. In this Review, we discuss recent advances in our understanding of stem cell quiescence and techniques enabling more refined analyses of quiescence invivo.

  View details for PubMedID 30735649

• Models and Studies of Aging: Executive Summary of a Report from the U13 Conference Series. Journal of the American Geriatrics Society Hurria, A., Carpenter, C. R., McFarland, F., Lundebjerg, N. E., de Cabo, R., Ferrucci, L., Studenski, S. A., Barzilai, N., Briggs, J. P., Ix, J. H., Kitzman, D. W., Kuchel, G. A., Musi, N., Newman, J. C., Rando, T. A., Smith, A. K., Walston, J. D., Kirkland, J. L., Yung, R. 2019

  Abstract

  The American Geriatrics Society convened a conference in Bethesda, Maryland, to explore models and studies of aging. This was the second of three conferences, supported by a U13 grant from the National Institute on Aging, to aid recipients of Grants for Early Medical/Surgical Specialists Transition to Aging Research (GEMSSTAR) in integrating geriatrics into their specialties. Recognizing that aging is the largest risk factor for multiple chronic diseases and age-related loss of resilience, the conference organizers focused scientific sessions on how targeting age-related mechanisms can delay, prevent, or reverse geriatric syndromes, age-related chronic diseases, and loss of resilience. The rationale for studying models of aging as well as study designs, strategies, and challenges of studying human aging were reviewed. This article provides a summary of the full conference report, Models and Studies of Aging: Report from the U13 Conference Series, and summarizes key take-home messages that were designed to support GEMSSTAR awardees in developing their research careers focused on aging research (see supplementary text for the full report).

  View details for PubMedID 30693953

• Angiotensin receptor blockade mimics the effect of exercise on recovery after orthopaedic trauma by decreasing pain and improving muscle regeneration. The Journal of physiology Tawfik, V. L., Quarta, M. n., Paine, P. n., Forman, T. E., Pajarinen, J. n., Takemura, Y. n., Goodman, S. B., Rando, T. A., Clark, J. D. 2019

  Abstract

  Our tibial fracture orthopaedic injury model in mice recapitulates the major manifestations of complex trauma including nociceptive sensitization, bone fracture, muscle fibrosis and muscle fibre loss. Delayed exercise after complex orthopaedic trauma results in decreased muscle fibrosis and improved pain Losartan, an angiotensin-receptor blocker with antifibrotic abilities, recapitulates the effect of exercise on post-injury recovery and may provide an enhanced recovery option for those who are unable to exercise after injury ABSTRACT: Chronic pain and disability after limb injury are major public health problems. Early mobilization after injury improves functional outcomes for patients but when and how to implement rehabilitation strategies remains a clinical challenge. Additionally, whether the beneficial effects of exercise can be reproduced using pharmacological tools remains unknown and may benefit patients who are unable to exercise due to immobilization. We developed a murine model of orthopaedic trauma combining tibia fracture and pin fixation with muscle damage. Behavioral measures included mechanical nociceptive thresholds and distances run on exercise wheels. Bone healing was quantified using microCT scanning, and muscle fibre size distribution and fibrosis were followed using immunohistochemistry. We found that the model provided robust mechanical allodynia, fibrosis and a shift to smaller average muscle fibre size lasting up to 5 weeks from injury. We also observed that allowing "late" (weeks 1-2) rather than "early" (weeks 0-1) exercise after injury resulted in greater overall running activity and greater reversal of allodynia. In parallel, the late running paradigm was associated with reduced muscle fibrosis, earlier increase in muscle fibre diameter and a short-term benefit in reducing callus volume. Providing the anti-fibrotic angiotensin receptor blocker losartan to mice in drinking water reduced both allodynia and muscle fibrosis. Combining losartan and late exercise provided no additional benefit. We conclude that early healing after orthopaedic trauma must be allowed prior to the initiation of exercise to achieve optimal pain, functional and physiological outcomes and that losartan is a viable candidate for translational studies. This article is protected by copyright. All rights reserved.

  View details for DOI 10.1113/JP278991

  View details for PubMedID 31784993

• Chronic inflammation in the etiology of disease across the life span. Nature medicine Furman, D. n., Campisi, J. n., Verdin, E. n., Carrera-Bastos, P. n., Targ, S. n., Franceschi, C. n., Ferrucci, L. n., Gilroy, D. W., Fasano, A. n., Miller, G. W., Miller, A. H., Mantovani, A. n., Weyand, C. M., Barzilai, N. n., Goronzy, J. J., Rando, T. A., Effros, R. B., Lucia, A. n., Kleinstreuer, N. n., Slavich, G. M. 2019; 25 (12): 1822–32

  Abstract

  Although intermittent increases in inflammation are critical for survival during physical injury and infection, recent research has revealed that certain social, environmental and lifestyle factors can promote systemic chronic inflammation (SCI) that can, in turn, lead to several diseases that collectively represent the leading causes of disability and mortality worldwide, such as cardiovascular disease, cancer, diabetes mellitus, chronic kidney disease, non-alcoholic fatty liver disease and autoimmune and neurodegenerative disorders. In the present Perspective we describe the multi-level mechanisms underlying SCI and several risk factors that promote this health-damaging phenotype, including infections, physical inactivity, poor diet, environmental and industrial toxicants and psychological stress. Furthermore, we suggest potential strategies for advancing the early diagnosis, prevention and treatment of SCI.

  View details for DOI 10.1038/s41591-019-0675-0

  View details for PubMedID 31806905

• Alternative polyadenylation of Pax3 controls muscle stem cell fate and muscle function. Science (New York, N.Y.) de Morree, A. n., Klein, J. D., Gan, Q. n., Farup, J. n., Urtasun, A. n., Kanugovi, A. n., Bilen, B. n., van Velthoven, C. T., Quarta, M. n., Rando, T. A. 2019; 366 (6466): 734–38

  Abstract

  Adult stem cells are essential for tissue homeostasis. In skeletal muscle, muscle stem cells (MuSCs) reside in a quiescent state, but little is known about the mechanisms that control homeostatic turnover. Here we show that, in mice, the variation in MuSC activation rate among different muscles (for example, limb versus diaphragm muscles) is determined by the levels of the transcription factor Pax3. We further show that Pax3 levels are controlled by alternative polyadenylation of its transcript, which is regulated by the small nucleolar RNA U1. Isoforms of the Pax3 messenger RNA that differ in their 3' untranslated regions are differentially susceptible to regulation by microRNA miR206, which results in varying levels of the Pax3 protein in vivo. These findings highlight a previously unrecognized mechanism of the homeostatic regulation of stem cell fate by multiple RNA species.

  View details for DOI 10.1126/science.aax1694

  View details for PubMedID 31699935

• Mesenchymal Stromal Cells Are Required for Regeneration and Homeostatic Maintenance of Skeletal Muscle. Cell reports Wosczyna, M. N., Konishi, C. T., Perez Carbajal, E. E., Wang, T. T., Walsh, R. A., Gan, Q. n., Wagner, M. W., Rando, T. A. 2019; 27 (7): 2029–35.e5

  Abstract

  The necessity of mesenchymal stromal cells, called fibroadipogenic progenitors (FAPs), in skeletal muscle regeneration and maintenance remains unestablished. We report the generation of a PDGFRαCreER knockin mouse model that provides a specific means of labeling and targeting FAPs. Depletion of FAPs using Cre-dependent diphtheria toxin expression results in loss of expansion of muscle stem cells (MuSCs) and CD45+ hematopoietic cells after injury and impaired skeletal muscle regeneration. Furthermore, FAP-depleted mice under homeostatic conditions exhibit muscle atrophy and loss of MuSCs, revealing that FAPs are required for the maintenance of both skeletal muscle and the MuSC pool. We also report that local tamoxifen metabolite delivery to target CreER activity in a single muscle, removing potentially confounding systemic effects of ablating PDGFRα+ cells distantly, also causes muscle atrophy. These data establish a critical role of FAPs in skeletal muscle regeneration and maintenance.

  View details for PubMedID 31091443

• Honey bee Royalactin unlocks conserved pluripotency pathway in mammals NATURE COMMUNICATIONS Wan, D. C., Morgan, S. L., Spencley, A. L., Mariano, N., Chang, E. Y., Shankar, G., Luo, Y., Li, T. H., Huh, D., Huynh, S. K., Garcia, J. M., Dovey, C. M., Lumb, J., Liu, L., Brown, K. V., Bermudez, A., Luong, R., Zeng, H., Mascetti, V. L., Pitteri, S. J., Wang, J., Tu, H., Quarta, M., Sebastiano, V., Nusse, R., Rando, T. A., Carette, J. E., Bazan, J., Wang, K. C. 2018; 9

  View details for DOI 10.1038/s41467-018-06256-4

  View details for Web of Science ID 000452040500001

• Honey bee Royalactin unlocks conserved pluripotency pathway in mammals. Nature communications Wan, D. C., Morgan, S. L., Spencley, A. L., Mariano, N., Chang, E. Y., Shankar, G., Luo, Y., Li, T. H., Huh, D., Huynh, S. K., Garcia, J. M., Dovey, C. M., Lumb, J., Liu, L., Brown, K. V., Bermudez, A., Luong, R., Zeng, H., Mascetti, V. L., Pitteri, S. J., Wang, J., Tu, H., Quarta, M., Sebastiano, V., Nusse, R., Rando, T. A., Carette, J. E., Bazan, J. F., Wang, K. C. 2018; 9 (1): 5078

  Abstract

  Royal jelly is the queen-maker for the honey bee Apis mellifera, and has cross-species effects on longevity, fertility, and regeneration in mammals. Despite this knowledge, how royal jelly or its components exert their myriad effects has remained poorly understood. Using mouse embryonic stem cells as a platform, here we report that through its major protein component Royalactin, royal jelly can maintain pluripotency by activating a ground-state pluripotency-like gene network. We further identify Regina, a mammalian structural analog of Royalactin that also induces a naive-like state in mouse embryonic stem cells. This reveals an important innate program for stem cell self-renewal with broad implications in understanding the molecular regulation of stem cell fate across species.

  View details for PubMedID 30510260

• Age-related declines in alpha-Klotho drive progenitor cell mitochondrial dysfunction and impaired muscle regeneration NATURE COMMUNICATIONS Sahu, A., Mamiya, H., Shinde, S. N., Cheikhi, A., Winter, L. L., Vo, N. V., Stolz, D., Roginskaya, Tang, W. Y., Croix, C., Sanders, L. H., Franti, M., Van Houten, B., Rando, T. A., Barchowsky, A., Ambrosio, F. 2018; 9

  View details for DOI 10.1038/s41467-018-07253-3

  View details for Web of Science ID 000450405900007

• Age-related declines in alpha-Klotho drive progenitor cell mitochondrial dysfunction and impaired muscle regeneration. Nature communications Sahu, A., Mamiya, H., Shinde, S. N., Cheikhi, A., Winter, L. L., Vo, N. V., Stolz, D., Roginskaya, V., Tang, W. Y., St Croix, C., Sanders, L. H., Franti, M., Van Houten, B., Rando, T. A., Barchowsky, A., Ambrosio, F. 2018; 9 (1): 4859

  Abstract

  While young muscle is capable of restoring the original architecture of damaged myofibers, aged muscle displays a markedly reduced regeneration. We show that expression of the "anti-aging" protein, alpha-Klotho, is up-regulated within young injured muscle as a result of transient Klotho promoter demethylation. However, epigenetic control of the Klotho promoter is lost with aging. Genetic inhibition of alpha-Klotho in vivo disruptedmuscle progenitor cell (MPC) lineage progression and impairedmyofiber regeneration, revealing a critical role for alpha-Klotho in the regenerative cascade. Genetic silencing of Klotho in young MPCs drovemitochondrial DNA (mtDNA) damage and decreased cellular bioenergetics. Conversely, supplementation with alpha-Klotho restoredmtDNA integrity and bioenergetics of aged MPCs to youthful levels in vitro and enhancedfunctional regeneration of aged muscle in vivo in a temporally-dependent manner. These studies identify a role for alpha-Klotho in the regulation of MPC mitochondrial function and implicate alpha-Klotho declines as a driver of impaired muscle regeneration with age.

  View details for PubMedID 30451844

• The regenerative rehabilitation collection: a forum for an emerging field NPJ REGENERATIVE MEDICINE Ambrosio, F., Rando, T. A. 2018; 3

  View details for DOI 10.1038/s41536-018-0058-z

  View details for Web of Science ID 000461464900001

• Biomechanics show stem cell necessity for effective treatment of volumetric muscle loss using bioengineered constructs NPJ REGENERATIVE MEDICINE Quarta, M., Lear, M., Blonigan, J., Paine, P., Chacon, R., Rando, T. A. 2018; 3

  View details for DOI 10.1038/s41536-018-0057-0

  View details for Web of Science ID 000461464700001

• Impaired Notch Signaling Leads to a Decrease in p53 Activity and Mitotic Catastrophe in Aged Muscle Stem Cells CELL STEM CELL Liu, L., Charville, G. W., Cheung, T. H., Yoo, B., Santos, P. J., Schroeder, M., Rando, T. A. 2018; 23 (4): 544-+

  View details for DOI 10.1016/j.stem.2018.08.019

  View details for Web of Science ID 000446342800014

• Rehabilitative exercise and spatially patterned nanofibrillar scaffolds enhance vascularization and innervation following volumetric muscle loss NPJ REGENERATIVE MEDICINE Nakayama, K. H., Alcazar, C., Yang, G., Quarta, M., Paine, P., Doan, L., Davies, A., Rando, T. A., Huang, N. F. 2018; 3: 16

  Abstract

  Muscle regeneration can be permanently impaired by traumatic injuries, despite the high regenerative capacity of skeletal muscle. Implantation of engineered biomimetic scaffolds to the site of muscle ablation may serve as an attractive off-the-shelf therapeutic approach. The objective of the study was to histologically assess the therapeutic benefit of a three-dimensional spatially patterned collagen scaffold, in conjunction with rehabilitative exercise, for treatment of volumetric muscle loss. To mimic the physiologic organization of skeletal muscle, which is generally composed of myofibers aligned in parallel, three-dimensional parallel-aligned nanofibrillar collagen scaffolds were fabricated. When implanted into the ablated murine tibialis anterior muscle, the aligned nanofibrillar scaffolds, in conjunction with voluntary caged wheel exercise, significantly improved the density of perfused microvessels, in comparison to treatments of the randomly oriented nanofibrillar scaffold, decellularized scaffold, or in the untreated control group. The abundance of neuromuscular junctions was 19-fold higher when treated with aligned nanofibrillar scaffolds in conjunction with exercise, in comparison to treatment of aligned scaffold without exercise. Although, the density of de novo myofibers was not significantly improved by aligned scaffolds, regardless of exercise activity, the cross-sectional area of regenerating myofibers was increased by > 60% when treated with either aligned and randomly oriented scaffolds, in comparison to treatment of decellularized scaffold or untreated controls. These findings demonstrate that voluntary exercise improved the regenerative effect of aligned scaffolds by augmenting neurovascularization, and have important implications in the design of engineered biomimetic scaffolds for treatment of traumatic muscle injury.

  View details for PubMedID 30245849

• Impaired Notch Signaling Leads to a Decrease in p53 Activity and Mitotic Catastrophe in Aged Muscle Stem Cells. Cell stem cell Liu, L., Charville, G. W., Cheung, T. H., Yoo, B., Santos, P. J., Schroeder, M., Rando, T. A. 2018

  Abstract

  The decline of tissue regenerative potential with age correlates with impaired stem cell function. However, limited strategies are available for therapeutic modulation of stem cell function during aging. Using skeletal muscle stem cells (MuSCs) as a model system, we identify cell death by mitotic catastrophe as a cause of impaired stem cell proliferative expansion in aged animals. The mitotic cell death is caused by a deficiency in Notch activators in the microenvironment. We discover that ligand-dependent stimulation of Notch activates p53 in MuSCs via inhibition of Mdm2 expression through Hey transcription factors during normal muscle regeneration and that this pathway is impaired in aged animals. Pharmacologic activation of p53 promotes the expansion of aged MuSCs invivo. Altogether, these findings illuminate a Notch-p53 signaling axis that plays an important role in MuSC survival during activation and is dysregulated during aging, contributing to the age-related decline in muscle regenerative potential.

  View details for PubMedID 30244867

• Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle MOLECULAR THERAPY-METHODS & CLINICAL DEVELOPMENT Paulk, N. K., Pekrun, K., Charville, G. W., Maguire-Nguyen, K., Wosczyna, M. N., Xu, J., Zhang, Y., Lisowski, L., Yoo, B., Vilches-Moure, J. G., Lee, G. K., Shrager, J. B., Rando, T. A., Kay, M. A. 2018; 10: 144–55

  View details for DOI 10.1016/j.omtm.2018.06.001

  View details for Web of Science ID 000451115100014

• Monitoring disease activity noninvasively in the mdx model of Duchenne muscular dystrophy PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Filareto, A., Maguire-Nguyen, K., Gan, Q., Aldanondo, G., Machado, L., Chamberlain, J. S., Rando, T. A. 2018; 115 (30): 7741–46

  View details for DOI 10.1073/pnas.1802425115

  View details for Web of Science ID 000439574700053

• A Muscle Stem Cell Support Group: Coordinated Cellular Responses in Muscle Regeneration. Developmental cell Wosczyna, M. N., Rando, T. A. 2018; 46 (2): 135–43

  Abstract

  Skeletal muscle has an extraordinary regenerative capacity due to the activity of tissue-specific muscle stem cells. Consequently, these cells have received the most attention in studies investigating the cellular processes of skeletal muscle regeneration. However, efficient capacity to rebuild this tissue also depends on additional cells in the local milieu, as disrupting their normal contributions often leads to incomplete regeneration. Here, we review these additional cells that contribute to the regenerative process. Understanding the complex interactions between and among these cell populations has the potential to lead to therapies that will help promote normal skeletal muscle regeneration under conditions in which this process is suboptimal.

  View details for PubMedID 30016618

• A Muscle Stem Cell Support Group: Coordinated Cellular Responses in Muscle Regeneration DEVELOPMENTAL CELL Wosczyna, M. N., Rando, T. A. 2018; 46 (2): 135–43

  View details for DOI 10.1016/j.devcel.2018.06.018

  View details for Web of Science ID 000438893000005

• Monitoring disease activity noninvasively in the mdx model of Duchenne muscular dystrophy. Proceedings of the National Academy of Sciences of the United States of America Filareto, A., Maguire-Nguyen, K., Gan, Q., Aldanondo, G., Machado, L., Chamberlain, J. S., Rando, T. A. 2018

  Abstract

  Duchenne muscular dystrophy (DMD) is a rare, muscle degenerative disease resulting from the absence of the dystrophin protein. DMD is characterized by progressive loss of muscle fibers, muscle weakness, and eventually loss of ambulation and premature death. Currently, there is no cure for DMD and improved methods of disease monitoring are crucial for the development of novel treatments. In this study, we describe a new method of assessing disease progression noninvasively in the mdx model of DMD. The reporter mice, which we term the dystrophic Degeneration Reporter strains, contain an inducible CRE-responsive luciferase reporter active in mature myofibers. In these mice, muscle degeneration is reflected in changes in the level of luciferase expression, which can be monitored using noninvasive, bioluminescence imaging. We monitored the natural history and disease progression in these dystrophic report mice and found that decreases in luciferase signals directly correlated with muscle degeneration. We further demonstrated that this reporter strain, as well as a previously reported Regeneration Reporter strain, successfully reveals the effectiveness of a gene therapy treatment following systemic administration of a recombinant adeno-associated virus-6 (rAAV-6) encoding a microdystrophin construct. Our data demonstrate the value of these noninvasive imaging modalities for monitoring disease progression and response to therapy in mouse models of muscular dystrophy.

  View details for PubMedID 29987034

• Lysosome activation clears aggregates and enhances quiescent neural stem cell activation during aging SCIENCE Leeman, D. S., Hebestreit, K., Ruetz, T., Webb, A. E., McKay, A., Pollina, E. A., Dulken, B. W., Zhao, X., Yeo, R. W., Ho, T. T., Mahmoudi, S., Devarajan, K., Passegue, E., Rando, T. A., Frydman, J., Brunet, A. 2018; 359 (6381): 1277–82

  Abstract

  In the adult brain, the neural stem cell (NSC) pool comprises quiescent and activated populations with distinct roles. Transcriptomic analysis revealed that quiescent and activated NSCs exhibited differences in their protein homeostasis network. Whereas activated NSCs had active proteasomes, quiescent NSCs contained large lysosomes. Quiescent NSCs from young mice accumulated protein aggregates, and many of these aggregates were stored in large lysosomes. Perturbation of lysosomal activity in quiescent NSCs affected protein-aggregate accumulation and the ability of quiescent NSCs to activate. During aging, quiescent NSCs displayed defects in their lysosomes, increased accumulation of protein aggregates, and reduced ability to activate. Enhancement of the lysosome pathway in old quiescent NSCs cleared protein aggregates and ameliorated the ability of quiescent NSCs to activate, allowing them to regain a more youthful state.

  View details for PubMedID 29590078

  View details for PubMedCentralID PMC5915358

• Regenerative Rehabilitation: Applied Biophysics Meets Stem Cell Therapeutics CELL STEM CELL Rando, T. A., Ambrosio, F. 2018; 22 (3): 306–9

  Abstract

  The emerging field of regenerative rehabilitation integrates biological and bioengineering advances in regenerative medicine with rehabilitative sciences. Here we highlight recent stem cell-based examples of the regenerative rehabilitation paradigm to promote tissue repair and regeneration, and we discuss remaining challenges and future directions for the field.

  View details for PubMedID 29499150

  View details for PubMedCentralID PMC5931336

• Inhibition of Methyltransferase Setd7 Allows the In Vitro Expansion of Myogenic Stem Cells with Improved Therapeutic Potential CELL STEM CELL Judson, R. N., Quarta, M., Oudhoff, M. J., Soliman, H., Yi, L., Chang, C., Loi, G., Werff, R., Cait, A., Hamer, M., Blonigan, J., Paine, P., Doan, L. N., Groppa, E., He, W., Su, L., Zhang, R. H., Xu, P., Eisner, C., Low, M., Barta, I., Lewis, C. B., Zaph, C., Karimi, M. M., Rando, T. A., Rossi, F. M. 2018; 22 (2): 177-+

  Abstract

  The development of cell therapy for repairing damaged or diseased skeletal muscle has been hindered by the inability to significantly expand immature, transplantable myogenic stem cells (MuSCs) in culture. To overcome this limitation, a deeper understanding of the mechanisms regulating the transition between activated, proliferating MuSCs and differentiation-primed, poorly engrafting progenitors is needed. Here, we show that methyltransferase Setd7 facilitates such transition by regulating the nuclear accumulation of β-catenin in proliferating MuSCs. Genetic or pharmacological inhibition of Setd7 promotes in vitro expansion of MuSCs and increases the yield of primary myogenic cell cultures. Upon transplantation, both mouse and human MuSCs expanded with a Setd7 small-molecule inhibitor are better able to repopulate the satellite cell niche, and treated mouse MuSCs show enhanced therapeutic potential in preclinical models of muscular dystrophy. Thus, Setd7 inhibition may help bypass a key obstacle in the translation of cell therapy for muscle disease.

  View details for PubMedID 29395054

• Single-cell transcriptomics of 20 mouse organs creates a Tabula Muris. Nature 2018; 562 (7727): 367–72

  Abstract

  Here we present a compendium of single-cell transcriptomic data from the model organism Mus musculus that comprises more than 100,000 cells from 20 organs and tissues. These data represent a new resource for cell biology, reveal gene expression in poorly characterized cell populations and enable the direct and controlled comparison of gene expression in cell types that are shared between tissues, such as T lymphocytes and endothelial cells from different anatomical locations. Two distinct technical approaches were used for most organs: one approach, microfluidic droplet-based 3'-end counting, enabled the survey of thousands of cells at relatively low coverage, whereas the other, full-length transcript analysis based on fluorescence-activated cell sorting, enabled the characterization of cell types with high sensitivity and coverage. The cumulative data provide the foundation for an atlas of transcriptomic cell biology.

  View details for DOI 10.1038/s41586-018-0590-4

  View details for PubMedID 30283141

• The regenerative rehabilitation collection: a forum for an emerging field. NPJ Regenerative medicine Ambrosio, F., Rando, T. A. 2018; 3: 20

  View details for PubMedID 30374410

• Biomechanics show stem cell necessity for effective treatment of volumetric muscle loss using bioengineered constructs. NPJ Regenerative medicine Quarta, M., Cromie Lear, M. J., Blonigan, J., Paine, P., Chacon, R., Rando, T. A. 2018; 3: 18

  Abstract

  Despite the regenerative capacity of muscle, tissue volume is not restored after volumetric muscle loss (VML), perhaps due to a loss-of-structural extracellular matrix. We recently demonstrated the structural and functional restoration of muscle tissue in a mouse model of VML using an engineered "bioconstruct," comprising an extracellular matrix scaffold (decellularized muscle), muscle stem cells (MuSCs), and muscle-resident cells (MRCs). To test the ability of the cell-based bioconstruct to restore whole-muscle biomechanics, we measured biomechanical parameters in uninjured muscles, muscles injured to produce VML lesions, and in muscles that were injured and then treated by implanting either the scaffolds alone or with bioconstructs containing the scaffolds, MuSCs, and MRCs. We measured the active and passive forces over a range of lengths, viscoelastic force relaxation, optimal length, and twitch dynamics. Injured muscles showed a narrowed length-tension curve or lower force over a narrower range of muscle lengths, and increased passive force. When treated with bioconstructs, but not with scaffolds alone, injured muscles showed active and passive length-tension relationships that were not different from uninjured muscles. Moreover, injured muscles treated with bioconstructs exhibited reduced fibrosis compared to injured muscles either untreated or treated with scaffolds alone. The cell-based bioconstruct is a promising treatment approach for future translational efforts to restore whole-muscle biomechanics in muscles with VML lesions.

  View details for PubMedID 30323949

• Bioengineered Viral Platform for Intramuscular Passive Vaccine Delivery to Human Skeletal Muscle. Molecular therapy. Methods & clinical development Paulk, N. K., Pekrun, K. n., Charville, G. W., Maguire-Nguyen, K. n., Wosczyna, M. N., Xu, J. n., Zhang, Y. n., Lisowski, L. n., Yoo, B. n., Vilches-Moure, J. G., Lee, G. K., Shrager, J. B., Rando, T. A., Kay, M. A. 2018; 10: 144–55

  Abstract

  Skeletal muscle is ideal for passive vaccine administration as it is easily accessible by intramuscular injection. Recombinant adeno-associated virus (rAAV) vectors are in consideration for passive vaccination clinical trials for HIV and influenza. However, greater human skeletal muscle transduction is needed for therapeutic efficacy than is possible with existing serotypes. To bioengineer capsids with therapeutic levels of transduction, we utilized a directed evolution approach to screen libraries of shuffled AAV capsids in pools of surgically resected human skeletal muscle cells from five patients. Six rounds of evolution were performed in various muscle cell types, and evolved variants were validated against existing muscle-tropic serotypes rAAV1, 6, and 8. We found that evolved variants NP22 and NP66 had significantly increased primary human and rhesus skeletal muscle fiber transduction from surgical explants ex vivo and in various primary and immortalized myogenic lines in vitro. Importantly, we demonstrated reduced seroreactivity compared to existing serotypes against normal human serum from 50 adult donors. These capsids represent powerful tools for human skeletal muscle expression and secretion of antibodies from passive vaccines.

  View details for PubMedID 30101152

• mTORC1 Activation during Repeated Regeneration Impairs Somatic Stem Cell Maintenance CELL STEM CELL Haller, S., Kapuria, S., Riley, R. R., O'Leary, M. N., Schreiber, K. H., Andersen, J. K., Melov, S., Que, J., Rando, T. A., Rock, J., Kennedy, B. K., Rodgers, J. T., Jasper, H. 2017; 21 (6): 806-+

  Abstract

  The balance between self-renewal and differentiation ensures long-term maintenance of stem cell (SC) pools in regenerating epithelial tissues. This balance is challenged during periods of high regenerative pressure and is often compromised in aged animals. Here, we show that target of rapamycin (TOR) signaling is a key regulator of SC loss during repeated regenerative episodes. In response to regenerative stimuli, SCs in the intestinal epithelium of the fly and in the tracheal epithelium of mice exhibit transient activation of TOR signaling. Although this activation is required for SCs to rapidly proliferate in response to damage, repeated rounds of damage lead to SC loss. Consistently, age-related SC loss in the mouse trachea and in muscle can be prevented by pharmacologic or genetic inhibition, respectively, of mammalian target of rapamycin complex 1 (mTORC1) signaling. These findings highlight an evolutionarily conserved role of TOR signaling in SC function and identify repeated rounds of mTORC1 activation as a driver of age-related SC decline.

  View details for PubMedID 29220665

  View details for PubMedCentralID PMC5823264

• Transcriptional Profiling of Quiescent Muscle Stem Cells In Vivo CELL REPORTS van Velthoven, C. J., de Morree, A., Egner, I. M., Brett, J. O., Rando, T. A. 2017; 21 (7): 1994–2004

  Abstract

  Muscle stem cells (MuSCs) persist in a quiescent state and activate in response to specific stimuli. The quiescent state is both actively maintained and dynamically regulated. However, analyses of quiescence have come primarily from cells removed from their niche. Although these cells are still quiescent, biochemical changes certainly occur during the isolation process. Here, we analyze the transcriptome of MuSCs in vivo utilizing MuSC-specific labeling of RNA. Notably, labeling transcripts during the isolation procedure revealed very active transcription of specific subsets of genes. However, using the transcription inhibitor α-amanitin, we show that the ex vivo transcriptome remains largely reflective of the in vivo transcriptome. Together, these data provide perspective on the molecular regulation of the quiescent state at the transcriptional level, demonstrate the utility of these tools for probing transcriptional dynamics in vivo, and provide an invaluable resource for understanding stem cell state transitions.

  View details for PubMedID 29141228

• Smad3 initiates oxidative stress and proteolysis that underlies diaphragm dysfunction during mechanical ventilation SCIENTIFIC REPORTS Tang, H., Kennedy, C. L., Lee, M., Gao, Y., Xia, H., Olguin, F., Fraga, D. A., Ayers, K., Choi, S., Kim, M., Tehrani, A., Sowb, Y. A., Rando, T. A., Shrager, J. B. 2017; 7: 14530

  Abstract

  Prolonged use of mechanical ventilation (MV) leads to atrophy and dysfunction of the major inspiratory muscle, the diaphragm, contributing to ventilator dependence. Numerous studies have shown that proteolysis and oxidative stress are among the major effectors of ventilator-induced diaphragm muscle dysfunction (VIDD), but the upstream initiator(s) of this process remain to be elucidated. We report here that periodic diaphragm contraction via phrenic nerve stimulation (PNS) substantially reduces MV-induced proteolytic activity and oxidative stress in the diaphragm. We show that MV rapidly induces phosphorylation of Smad3, and PNS nearly completely prevents this effect. In cultured cells, overexpressed Smad3 is sufficient to induce oxidative stress and protein degradation, whereas inhibition of Smad3 activity suppresses these events. In rats subjected to MV, inhibition of Smad3 activity by SIS3 suppresses oxidative stress and protein degradation in the diaphragm and prevents the reduction in contractility that is induced by MV. Smad3's effect appears to link to STAT3 activity, which we previously identified as a regulator of VIDD. Inhibition of Smad3 suppresses STAT3 signaling both in vitro and in vivo. Thus, MV-induced diaphragm inactivity initiates catabolic changes via rapid activation of Smad3 signaling. An early intervention with PNS and/or pharmaceutical inhibition of Smad3 may prevent clinical VIDD.

  View details for PubMedID 29109401

• Staufen1 inhibits MyoD translation to actively maintain muscle stem cell quiescence PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA de Morree, A., van Velthoven, C. J., Gan, Q., Salvi, J. S., Klein, J., Akimenko, I., Quarta, M., Biressi, S., Rando, T. A. 2017; 114 (43): E8996–E9005

  Abstract

  Tissue regeneration depends on the timely activation of adult stem cells. In skeletal muscle, the adult stem cells maintain a quiescent state and proliferate upon injury. We show that muscle stem cells (MuSCs) use direct translational repression to maintain the quiescent state. High-resolution single-molecule and single-cell analyses demonstrate that quiescent MuSCs express high levels of Myogenic Differentiation 1 (MyoD) transcript in vivo, whereas MyoD protein is absent. RNA pulldowns and costainings show that MyoD mRNA interacts with Staufen1, a potent regulator of mRNA localization, translation, and stability. Staufen1 prevents MyoD translation through its interaction with the MyoD 3'-UTR. MuSCs from Staufen1 heterozygous (Staufen1+/-) mice have increased MyoD protein expression, exit quiescence, and begin proliferating. Conversely, blocking MyoD translation maintains the quiescent phenotype. Collectively, our data show that MuSCs express MyoD mRNA and actively repress its translation to remain quiescent yet primed for activation.

  View details for PubMedID 29073096

• Bioengineered constructs combined with exercise enhance stem cell-mediated treatment of volumetric muscle loss NATURE COMMUNICATIONS Quarta, M., Cromie, M., Chacon, R., Blonigan, J., Garcia, V., Akimenko, I., Hamer, M., Paine, P., Stok, M., Shrager, J. B., Rando, T. A. 2017; 8: 15613

  Abstract

  Volumetric muscle loss (VML) is associated with loss of skeletal muscle function, and current treatments show limited efficacy. Here we show that bioconstructs suffused with genetically-labelled muscle stem cells (MuSCs) and other muscle resident cells (MRCs) are effective to treat VML injuries in mice. Imaging of bioconstructs implanted in damaged muscles indicates MuSCs survival and growth, and ex vivo analyses show force restoration of treated muscles. Histological analysis highlights myofibre formation, neovascularisation, but insufficient innervation. Both innervation and in vivo force production are enhanced when implantation of bioconstructs is followed by an exercise regimen. Significant improvements are also observed when bioconstructs are used to treat chronic VML injury models. Finally, we demonstrate that bioconstructs made with human MuSCs and MRCs can generate functional muscle tissue in our VML model. These data suggest that stem cell-based therapies aimed to engineer tissue in vivo may be effective to treat acute and chronic VML.

  View details for PubMedID 28631758

• Aging of the skeletal muscle extracellular matrix drives a stem cell fibrogenic conversion AGING CELL Stearns-Reider, K. M., D'Amore, A., Beezhold, K., Rothrauff, B., Cavalli, L., Wagner, W. R., Vorp, D. A., Tsamis, A., Shinde, S., Zhang, C., Barchowsky, A., Rando, T. A., Tuan, R. S., Ambrosio, F. 2017; 16 (3): 518–28

  Abstract

  Age-related declines in skeletal muscle regeneration have been attributed to muscle stem cell (MuSC) dysfunction. Aged MuSCs display a fibrogenic conversion, leading to fibrosis and impaired recovery after injury. Although studies have demonstrated the influence of in vitro substrate characteristics on stem cell fate, whether and how aging of the extracellular matrix (ECM) affects stem cell behavior has not been investigated. Here, we investigated the direct effect of the aged muscle ECM on MuSC lineage specification. Quantification of ECM topology and muscle mechanical properties reveals decreased collagen tortuosity and muscle stiffening with increasing age. Age-related ECM alterations directly disrupt MuSC responses, and MuSCs seeded ex vivo onto decellularized ECM constructs derived from aged muscle display increased expression of fibrogenic markers and decreased myogenicity, compared to MuSCs seeded onto young ECM. This fibrogenic conversion is recapitulated in vitro when MuSCs are seeded directly onto matrices elaborated by aged fibroblasts. When compared to young fibroblasts, fibroblasts isolated from aged muscle display increased nuclear levels of the mechanosensors, Yes-associated protein (YAP)/transcriptional coactivator with PDZ-binding motif (TAZ), consistent with exposure to a stiff microenvironment in vivo. Accordingly, preconditioning of young fibroblasts by seeding them onto a substrate engineered to mimic the stiffness of aged muscle increases YAP/TAZ nuclear translocation and promotes secretion of a matrix that favors MuSC fibrogenesis. The findings here suggest that an age-related increase in muscle stiffness drives YAP/TAZ-mediated pathogenic expression of matricellular proteins by fibroblasts, ultimately disrupting MuSC fate.

  View details for PubMedID 28371268

• Evolved AAV Capsids for Intramuscular Passive Vaccine Administration to Human Skeletal Muscle Paulk, N. K., Pekrun, K., Charville, G., Maguire-Nguyen, K., Xu, J., Wosczyna, M., Lisowski, L., Lee, G., Shrager, J., Rando, T., Kay, M. A. CELL PRESS. 2017: 96

  View details for Web of Science ID 000401083600200

• HGFA Is an Injury-Regulated Systemic Factor that Induces the Transition of Stem Cells into G(Alert) CELL REPORTS Rodgers, J. T., Schroeder, M. D., Ma, C., Rando, T. A. 2017; 19 (3): 479-486

  Abstract

  The activation of quiescent stem cells into the cell cycle is a key step in initiating the process of tissue repair. We recently reported that quiescent stem cells can transition into GAlert, a cellular state in which they have an increased functional ability to activate and participate in tissue repair. However, the precise molecular signals that induce GAlert in stem cells have remained elusive. Here, we show that the injury-induced regulation of hepatocyte growth factor (HGF) proteolytic processing via the systemic protease, hepatocyte growth factor activator (HGFA), stimulates GAlert in skeletal muscle stem cells (MuSCs) and fibro-adipogenic progenitors (FAPs). We demonstrate that administering active HGFA to animals is sufficient to induce GAlert in stem cells throughout the body and to significantly accelerate the processes of stem cell activation and tissue repair. Our data suggest that factors that induce GAlert will have broad therapeutic applications for regenerative medicine and wound healing.

  View details for DOI 10.1016/j.celrep.2017.03.066

  View details for Web of Science ID 000401133100005

  View details for PubMedCentralID PMC5468096

• HGFA Is an Injury-Regulated Systemic Factor that Induces the Transition of Stem Cells into GAlert. Cell reports Rodgers, J. T., Schroeder, M. D., Ma, C., Rando, T. A. 2017; 19 (3): 479-486

  Abstract

  The activation of quiescent stem cells into the cell cycle is a key step in initiating the process of tissue repair. We recently reported that quiescent stem cells can transition into GAlert, a cellular state in which they have an increased functional ability to activate and participate in tissue repair. However, the precise molecular signals that induce GAlert in stem cells have remained elusive. Here, we show that the injury-induced regulation of hepatocyte growth factor (HGF) proteolytic processing via the systemic protease, hepatocyte growth factor activator (HGFA), stimulates GAlert in skeletal muscle stem cells (MuSCs) and fibro-adipogenic progenitors (FAPs). We demonstrate that administering active HGFA to animals is sufficient to induce GAlert in stem cells throughout the body and to significantly accelerate the processes of stem cell activation and tissue repair. Our data suggest that factors that induce GAlert will have broad therapeutic applications for regenerative medicine and wound healing.

  View details for DOI 10.1016/j.celrep.2017.03.066

  View details for PubMedID 28423312

  View details for PubMedCentralID PMC5468096

• Deltex2 represses MyoD expression and inhibits myogenic differentiation by acting as a negative regulator of Jmjd1c PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Luo, D., de Morree, A., Boutet, S., Quach, N., Natu, V., Rustagi, A., Rando, T. A. 2017; 114 (15): E3071-E3080

  Abstract

  The myogenic regulatory factor MyoD has been implicated as a key regulator of myogenesis, and yet there is little information regarding its upstream regulators. We found that Deltex2 inhibits myogenic differentiation in vitro, and that skeletal muscle stem cells from Deltex2 knockout mice exhibit precocious myogenic differentiation and accelerated regeneration in response to injury. Intriguingly, Deltex2 inhibits myogenesis by suppressing MyoD transcription, and the Deltex2 knockout phenotype can be rescued by a loss-of-function allele for MyoD In addition, we obtained evidence that Deltex2 regulates MyoD expression by promoting the enrichment of histone 3 modified by dimethylation at lysine 9 at a key regulatory region of the MyoD locus. The enrichment is attributed to a Deltex2 interacting protein, Jmjd1c, whose activity is directly inhibited by Deltex2 and whose expression is required for MyoD expression in vivo and in vitro. Finally, we find that Deltex2 causes Jmjd1c monoubiquitination and inhibits its demethylase activity. Mutation of the monoubiquitination site in Jmjd1c abolishes the inhibitory effect of Deltex2 on Jmjd1c demethylase activity. These results reveal a mechanism by which a member of the Deltex family of proteins can inhibit cellular differentiation, and demonstrate a role of Deltex in the epigenetic regulation of myogenesis.

  View details for DOI 10.1073/pnas.1613592114

  View details for Web of Science ID 000398789800012

  View details for PubMedID 28351977

• The protein tyrosine phosphatase 1B inhibitor MSI-1436 stimulates regeneration of heart and multiple other tissues NPJ REGENERATIVE MEDICINE Smith, A. M., Maguire-Nguyen, K. K., Rando, T. A., Zasloff, M. A., Strange, K. B., Yin, V. P. 2017; 2: 4

  Abstract

  Regenerative medicine holds substantial promise for repairing or replacing tissues and organs damaged by disease, injury, and degeneration. Much of the field has focused on development of cell-based therapeutics, gene-based therapeutics, and tissue engineering-based therapeutics. In contrast, development of small molecule regenerative medicine therapies is an emerging area. Using the adult zebrafish as a novel screening platform, we identified MSI-1436 as a first-in-class regenerative medicine drug candidate. MSI-1436 is a naturally occurring aminosterol that inhibits protein tyrosine phosphatase 1B. Treatment of adult zebrafish by intraperitoneal injection of MSI-1436 increased the rate of regeneration of the amputated caudal fin, which is comprised of bone, connective, skin, vascular and nervous tissues and also increased the rate of adult zebrafish heart regeneration. Intraperitoneal administration of MSI-1436 to adult mice for 4 weeks after induction of myocardial infarction increased survival, improved heart function, reduced infarct size, reduced ventricular wall thinning and increased cardiomyocyte proliferation. Satellite cell activation in injured mouse skeletal muscle was stimulated by MSI-1436. MSI-1436 was well tolerated by patients in Phase 1 and 1b obesity and type 2 diabetes clinical trials. Doses effective at stimulating regeneration are 5-50-times lower than the maximum well tolerated human dose. The demonstrated safety and well established pharmacological properties of MSI-1436 underscore the potential of this molecule as a novel treatment for heart attack and multiple other degenerative diseases.

  View details for PubMedID 29302341

• Fleeting factors, turning back time NATURE BIOTECHNOLOGY Rando, T. A. 2017; 35 (3): 218–20

  View details for PubMedID 28267729

• Interaction between epigenetic and metabolism in aging stem cells. Current opinion in cell biology Brunet, A., Rando, T. A. 2017; 45: 1-7

  Abstract

  Aging is accompanied by a decline in tissue function, regeneration, and repair. A large part of this decline is caused by the deterioration of tissue stem cell function. Understanding the mechanisms that drive stem cell aging and how to counteract them is a critical step for enhancing tissue repair and maintenance during aging. Emerging evidence indicates that epigenetic modifiers and metabolism regulators interact to impact lifespan, suggesting that this mechanism may also affect stem cell function with age. This review focuses on the interaction between chromatin and metabolism in the regulation of tissue stem cells during aging. We also discuss how these mechanisms integrate environmental stimuli such as nutrient stress to regulate stem cell function. Finally, this review examines new perspectives for regeneration, rejuvenation, and treatment of age-related decline of stem cell function.

  View details for DOI 10.1016/j.ceb.2016.12.009

  View details for PubMedID 28129586

• Macrophage-released ADAMTS1 promotes muscle stem cell activation. Nature communications Du, H. n., Shih, C. H., Wosczyna, M. N., Mueller, A. A., Cho, J. n., Aggarwal, A. n., Rando, T. A., Feldman, B. J. 2017; 8 (1): 669

  Abstract

  Coordinated activation of muscle stem cells (known as satellite cells) is critical for postnatal muscle growth and regeneration. The muscle stem cell niche is central for regulating the activation state of satellite cells, but the specific extracellular signals that coordinate this regulation are poorly understood. Here we show that macrophages at sites of muscle injury induce activation of satellite cells via expression of Adamts1. Overexpression of Adamts1 in macrophages in vivo is sufficient to increase satellite cell activation and improve muscle regeneration in young mice. We demonstrate that NOTCH1 is a target of ADAMTS1 metalloproteinase activity, which reduces Notch signaling, leading to increased satellite cell activation. These results identify Adamts1 as a potent extracellular regulator of satellite cell activation and have significant implications for understanding the regulation of satellite cell activity and regeneration after muscle injury.Satellite cells are crucial for growth and regeneration of skeletal muscle. Here the authors show that in response to muscle injury, macrophages secrete Adamts1, which induces satellite cell activation by modulating Notch1 signaling.

  View details for PubMedID 28939843

• Intronic polyadenylation of PDGFR alpha in resident stem cells attenuates muscle fibrosis NATURE Mueller, A. A., van Velthoven, C. T., Fukumoto, K. D., Cheung, T. H., Rando, T. A. 2016; 540 (7632): 276-?

  Abstract

  Platelet-derived growth factor receptor α (PDGFRα) exhibits divergent effects in skeletal muscle. At physiological levels, signalling through this receptor promotes muscle development in growing embryos and angiogenesis in regenerating adult muscle. However, both increased PDGF ligand abundance and enhanced PDGFRα pathway activity cause pathological fibrosis. This excessive collagen deposition, which is seen in aged and diseased muscle, interferes with muscle function and limits the effectiveness of gene- and cell-based therapies for muscle disorders. Although compelling evidence exists for the role of PDGFRα in fibrosis, little is known about the cells through which this pathway acts. Here we show in mice that PDGFRα signalling regulates a population of muscle-resident fibro/adipogenic progenitors (FAPs) that play a supportive role in muscle regeneration but may also cause fibrosis when aberrantly regulated. We found that FAPs produce multiple transcriptional variants of Pdgfra with different polyadenylation sites, including an intronic variant that codes for a protein isoform containing a truncated kinase domain. This variant, upregulated during regeneration, acts as a decoy to inhibit PDGF signalling and to prevent FAP over-activation. Moreover, increasing the expression of this isoform limits fibrosis in vivo in mice, suggesting both biological relevance and therapeutic potential of modulating polyadenylation patterns in stem-cell populations.

  View details for DOI 10.1038/nature20160

  View details for Web of Science ID 000389548700058

  View details for PubMedID 27894125

  View details for PubMedCentralID PMC5384334

• Engineering Pre-Vascularized Skeletal Muscle for Treatment of Volumetric Muscle Loss Nakayama, K., Quarta, M., Garcia, V., Strassberg, Z., Akimenko, I., Abilez, O., Rando, T., Huang, N. MARY ANN LIEBERT, INC. 2016: S22

  View details for Web of Science ID 000390569200079

• An artificial niche preserves the quiescence of muscle stem cells and enhances their therapeutic efficacy. Nature biotechnology Quarta, M., Brett, J. O., DiMarco, R., de Morree, A., Boutet, S. C., Chacon, R., Gibbons, M. C., Garcia, V. A., Su, J., Shrager, J. B., Heilshorn, S., Rando, T. A. 2016; 34 (7): 752-759

  Abstract

  A promising therapeutic strategy for diverse genetic disorders involves transplantation of autologous stem cells that have been genetically corrected ex vivo. A major challenge in such approaches is a loss of stem cell potency during culture. Here we describe an artificial niche for maintaining muscle stem cells (MuSCs) in vitro in a potent, quiescent state. Using a machine learning method, we identified a molecular signature of quiescence and used it to screen for factors that could maintain mouse MuSC quiescence, thus defining a quiescence medium (QM). We also engineered muscle fibers that mimic the native myofiber of the MuSC niche. Mouse MuSCs maintained in QM on engineered fibers showed enhanced potential for engraftment, tissue regeneration and self-renewal after transplantation in mice. An artificial niche adapted to human cells similarly extended the quiescence of human MuSCs in vitro and enhanced their potency in vivo. Our approach for maintaining quiescence may be applicable to stem cells isolated from other tissues.

  View details for DOI 10.1038/nbt.3576

  View details for PubMedID 27240197

• UTX in muscle regeneration - the right dose and the right time JOURNAL OF CLINICAL INVESTIGATION Liu, L., Rando, T. A. 2016; 126 (4): 1233–35

  Abstract

  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.

  View details for DOI 10.1172/JCI86798

  View details for Web of Science ID 000373522300013

  View details for PubMedID 26999609

  View details for PubMedCentralID PMC4811162

• Stem cells and healthy aging SCIENCE Goodell, M. A., Rando, T. A. 2015; 350 (6265): 1199-1204

  Abstract

  Research into stem cells and aging aims to understand how stem cells maintain tissue health, what mechanisms ultimately lead to decline in stem cell function with age, and how the regenerative capacity of somatic stem cells can be enhanced to promote healthy aging. Here, we explore the effects of aging on stem cells in different tissues. Recent research has focused on the ways that genetic mutations, epigenetic changes, and the extrinsic environmental milieu influence stem cell functionality over time. We describe each of these three factors, the ways in which they interact, and how these interactions decrease stem cell health over time. We are optimistic that a better understanding of these changes will uncover potential strategies to enhance stem cell function and increase tissue resiliency into old age.

  View details for DOI 10.1126/science.aab3388

  View details for Web of Science ID 000365700500063

• Stem cells and healthy aging. Science (New York, N.Y.) Goodell, M. A., Rando, T. A. 2015; 350 (6265): 1199-204

  Abstract

  Research into stem cells and aging aims to understand how stem cells maintain tissue health, what mechanisms ultimately lead to decline in stem cell function with age, and how the regenerative capacity of somatic stem cells can be enhanced to promote healthy aging. Here, we explore the effects of aging on stem cells in different tissues. Recent research has focused on the ways that genetic mutations, epigenetic changes, and the extrinsic environmental milieu influence stem cell functionality over time. We describe each of these three factors, the ways in which they interact, and how these interactions decrease stem cell health over time. We are optimistic that a better understanding of these changes will uncover potential strategies to enhance stem cell function and increase tissue resiliency into old age.

  View details for DOI 10.1126/science.aab3388

  View details for PubMedID 26785478

• Ex Vivo Expansion and In Vivo Self-Renewal of Human Muscle Stem Cells STEM CELL REPORTS Charville, G. W., Cheung, T. H., Yoo, B., Santos, P. J., Lee, G. K., Shrager, J. B., Rando, T. A. 2015; 5 (4): 621-632

  Abstract

  Adult skeletal muscle stem cells, or satellite cells (SCs), regenerate functional muscle following transplantation into injured or diseased tissue. To gain insight into human SC (huSC) biology, we analyzed transcriptome dynamics by RNA sequencing of prospectively isolated quiescent and activated huSCs. This analysis indicated that huSCs differentiate and lose proliferative potential when maintained in high-mitogen conditions ex vivo. Further analysis of gene expression revealed that p38 MAPK acts in a transcriptional network underlying huSC self-renewal. Activation of p38 signaling correlated with huSC differentiation, while inhibition of p38 reversibly prevented differentiation, enabling expansion of huSCs. When transplanted, expanded huSCs differentiated to generate chimeric muscle and engrafted as SCs in the sublaminar niche with a greater frequency than freshly isolated cells or cells cultured without p38 inhibition. These studies indicate characteristics of the huSC transcriptome that promote expansion ex vivo to allow enhanced functional engraftment of a defined population of self-renewing huSCs.

  View details for DOI 10.1016/j.stemcr.2015.08.004

  View details for PubMedID 26344908

• Isolation of skeletal muscle stem cells by fluorescence-activated cell sorting NATURE PROTOCOLS Liu, L., Cheung, T. H., Charville, G. W., Rando, T. A. 2015; 10 (10): 1612-1624

  Abstract

  The prospective isolation of purified stem cell populations has dramatically altered the field of stem cell biology, and it has been a major focus of research across tissues in different organisms. Muscle stem cells (MuSCs) are now among the most intensely studied stem cell populations in mammalian systems, and the prospective isolation of these cells has allowed cellular and molecular characterizations that were not dreamed of a decade ago. In this protocol, we describe how to isolate MuSCs from limb muscles of adult mice by fluorescence-activated cell sorting (FACS). We provide a detailed description of the physical and enzymatic dissociation of mononucleated cells from limb muscles, a procedure that is essential in order to maximize cell yield. We also describe a FACS-based method that is used subsequently to obtain highly pure populations of either quiescent or activated MuSCs (VCAM(+)CD31(-)CD45(-)Sca1(-)). The isolation process takes ∼5-6 h to complete. The protocol also allows for the isolation of endothelial cells, hematopoietic cells and mesenchymal stem cells from muscle tissue.

  View details for DOI 10.1038/nprot.2015.110

  View details for PubMedID 26401916

• Mimicking the niche: cytokines expand muscle stem cells CELL RESEARCH Quarta, M., Rando, T. A. 2015; 25 (7): 761-762

  Abstract

  Muscle stem cells (MuSCs) have long been considered to be potential therapeutic vehicles for diseases of muscle such as muscular dystrophies. A recent study published in Cell Research by Fu et al. reveals that recapitulating in vitro the in vivo microenvironment of MuSCs that occurs during muscle regeneration might be a major step towards translation.

  View details for DOI 10.1038/cr.2015.78

  View details for Web of Science ID 000357516000001

  View details for PubMedID 26113258

  View details for PubMedCentralID PMC4493285

• Synergizing Engineering and Biology to Treat and Model Skeletal Muscle Injury and Disease ANNUAL REVIEW OF BIOMEDICAL ENGINEERING, VOL 17 Bursac, N., Juhas, M., Rando, T. A., Yarmush, M. L. 2015; 17: 217–42

  Abstract

  Although skeletal muscle is one of the most regenerative organs in our body, various genetic defects, alterations in extrinsic signaling, or substantial tissue damage can impair muscle function and the capacity for self-repair. The diversity and complexity of muscle disorders have attracted much interest from both cell biologists and, more recently, bioengineers, leading to concentrated efforts to better understand muscle pathology and develop more efficient therapies. This review describes the biological underpinnings of muscle development, repair, and disease, and discusses recent bioengineering efforts to design and control myomimetic environments, both to study muscle biology and function and to aid in the development of new drug, cell, and gene therapies for muscle disorders. The synergy between engineering-aided biological discovery and biology-inspired engineering solutions will be the path forward for translating laboratory results into clinical practice.

  View details for PubMedID 26643021

• The JAK-STAT Pathway Is Critical in Ventilator-Induced Diaphragm Dysfunction. Molecular medicine Tang, H., Smith, I. J., Hussain, S. N., Goldberg, P., Lee, M., Sugiarto, S., Godinez, G. L., Singh, B. K., Payan, D. G., Rando, T. A., Kinsella, T. M., Shrager, J. B. 2015; 20 (1): 579-589

  Abstract

  Mechanical ventilation (MV) is one of the lynchpins of modern intensive-care medicine and is life saving in many critically ill patients. Continuous ventilator support, however, results in ventilation-induced diaphragm dysfunction (VIDD) that likely prolongs patients' need for MV and thereby leads to major associated complications and avoidable intensive care unit (ICU) deaths. Oxidative stress is a key pathogenic event in the development of VIDD, but its regulation remains largely undefined. We report here that the JAK-STAT pathway is activated in MV in the human diaphragm, as evidenced by significantly increased phosphorylation of JAK and STAT. Blockage of the JAK-STAT pathway by a JAK inhibitor in a rat MV model prevents diaphragm muscle contractile dysfunction (by ~85%, p < 0.01). We further demonstrate that activated STAT3 compromises mitochondrial function and induces oxidative stress in vivo, and, interestingly, that oxidative stress also activates JAK-STAT. Inhibition of JAK-STAT prevents oxidative stress-induced protein oxidation and polyubiquitination and recovers mitochondrial function in cultured muscle cells. Therefore, in ventilated diaphragm muscle, activation of JAK-STAT is critical in regulating oxidative stress and is thereby central to the downstream pathogenesis of clinical VIDD. These findings establish the molecular basis for the therapeutic promise of JAK-STAT inhibitors in ventilated ICU patients.

  View details for DOI 10.2119/molmed.2014.00049

  View details for PubMedID 25286450

• A Wnt-TGF beta 2 axis induces a fibrogenic program in muscle stem cells from dystrophic mice SCIENCE TRANSLATIONAL MEDICINE Biressi, S., Miyabara, E. H., Gopinath, S. D., Carlig, P. M., Rando, T. A. 2014; 6 (267)

  Abstract

  We have previously observed that Wnt signaling activates a fibrogenic program in adult muscle stem cells, called satellite cells, during aging. We genetically labeled satellite cells in a mouse model of Duchenne muscular dystrophy to follow their fate during the progression of the disease. We observed that a fraction of satellite cells had a reduced myogenic potential and showed enhanced expression of profibrotic genes compared to age-matched controls. By combining in vitro and in vivo results, we found that expression of transforming growth factor-β2 (TGFβ2) was induced in response to elevated canonical Wnt signaling in dystrophic muscles and that the resulting increase in TGFβ activity affected the behavior of satellite cells in an autocrine or paracrine fashion. Indeed, pharmacological inhibition of the TGFβ pathway in vivo reduced the fibrogenic characteristics of satellite cells. These studies shed new light on the cellular and molecular mechanisms responsible for stem cell dysfunction in dystrophic muscle and may contribute to the development of more effective and specific therapeutic approaches for the prevention of muscle fibrosis.

  View details for DOI 10.1126/scitranslmed.3008411

  View details for Web of Science ID 000346463000001

  View details for PubMedCentralID PMC4350665

• A Wnt-TGFβ2 axis induces a fibrogenic program in muscle stem cells from dystrophic mice. Science translational medicine Biressi, S., Miyabara, E. H., Gopinath, S. D., Carlig, P. M., Rando, T. A. 2014; 6 (267): 267ra176

  Abstract

  We have previously observed that Wnt signaling activates a fibrogenic program in adult muscle stem cells, called satellite cells, during aging. We genetically labeled satellite cells in a mouse model of Duchenne muscular dystrophy to follow their fate during the progression of the disease. We observed that a fraction of satellite cells had a reduced myogenic potential and showed enhanced expression of profibrotic genes compared to age-matched controls. By combining in vitro and in vivo results, we found that expression of transforming growth factor-β2 (TGFβ2) was induced in response to elevated canonical Wnt signaling in dystrophic muscles and that the resulting increase in TGFβ activity affected the behavior of satellite cells in an autocrine or paracrine fashion. Indeed, pharmacological inhibition of the TGFβ pathway in vivo reduced the fibrogenic characteristics of satellite cells. These studies shed new light on the cellular and molecular mechanisms responsible for stem cell dysfunction in dystrophic muscle and may contribute to the development of more effective and specific therapeutic approaches for the prevention of muscle fibrosis.

  View details for DOI 10.1126/scitranslmed.3008411

  View details for PubMedID 25520397

  View details for PubMedCentralID PMC4350665

• Epigenetic Mechanisms of Stem Cell Aging and Rejuvenation Rando, T. A. AMER SOC HEMATOLOGY. 2014

  View details for Web of Science ID 000349233806161

• Stem Cell Energetics CELL STEM CELL Passegue, E., Teitell, M., Ding, S., Rando, T., Belmonte, J., Ramalho-Santos, M., Terzic, A., Brunet, A., Chandel, N. S., Bonnet, D., Jasper, H. 2014; 15 (6): 679–82

  View details for Web of Science ID 000347174300006

• Induction of autophagy supports the bioenergetic demands of quiescent muscle stem cell activation EMBO JOURNAL Tang, A. H., Rando, T. A. 2014; 33 (23): 2782-2797

  Abstract

  The exit of a stem cell out of quiescence into an activated state is characterized by major metabolic changes associated with increased biosynthesis of proteins and macromolecules. The regulation of this transition is poorly understood. Using muscle stem cells, or satellite cells (SCs), we found that autophagy, which catabolizes intracellular contents to maintain proteostasis and to produce energy during nutrient deprivation, was induced during SC activation. Inhibition of autophagy suppressed the increase in ATP levels and delayed SC activation, both of which could be partially rescued by exogenous pyruvate as an energy source, suggesting that autophagy may provide nutrients necessary to meet bioenergetic demands during this critical transition from quiescence to activation. We found that SIRT1, a known nutrient sensor, regulates autophagic flux in SC progeny. A deficiency of SIRT1 led to a delay in SC activation that could also be partially rescued by exogenous pyruvate. These studies suggest that autophagy, regulated by SIRT1, may play an important role during SC activation to meet the high bioenergetic demands of the activation process.

  View details for Web of Science ID 000345770000006

  View details for PubMedID 25316028

• Geroscience: Linking Aging to Chronic Disease CELL Kennedy, B. K., Berger, S. L., Brunet, A., Campisi, J., Cuervo, A., Epel, E. S., Franceschi, C., Lithgow, G. J., Morimoto, R. I., Pessin, J. E., Rando, T. A., Richardson, A., Schadt, E. E., Wyss-Coray, T., Sierra, F. 2014; 159 (4): 708–12

  Abstract

  Mammalian aging can be delayed with genetic, dietary, and pharmacologic approaches. Given that the elderly population is dramatically increasing and that aging is the greatest risk factor for a majority of chronic diseases driving both morbidity and mortality, it is critical to expand geroscience research directed at extending human healthspan.

  View details for PubMedID 25417146

  View details for PubMedCentralID PMC4852871

• Translational strategies and challenges in regenerative medicine NATURE MEDICINE Dimmeler, S., Ding, S., Rando, T. A., Trounson, A. 2014; 20 (8): 814-821

  Abstract

  The scientific community is currently witnessing substantial strides in understanding stem cell biology in humans; however, major disappointments in translating this knowledge into medical therapies are flooding the field as well. Despite these setbacks, investigators are determined to better understand the caveats of regeneration, so that major pathways of repair and regrowth can be exploited in treating aged and diseased tissues. Last year, in an effort to contribute to this burgeoning field, Nature Medicine, in collaboration with the Volkswagen Foundation, organized a meeting with a panel of experts in regenerative medicine to identify the most pressing challenges, as well as the crucial strategies and stem cell concepts that can best help advance the translational regenerative field. Here some experts who participated in the meeting provide an outlook at some of those key issues and concepts.

  View details for DOI 10.1038/nm.3627

  View details for Web of Science ID 000340074600013

  View details for PubMedID 25100527

• H3K4me3 Breadth Is Linked to Cell Identity and Transcriptional Consistency. Cell Benayoun, B. A., Pollina, E. A., Ucar, D., Mahmoudi, S., Karra, K., Wong, E. D., Devarajan, K., Daugherty, A. C., Kundaje, A. B., Mancini, E., Hitz, B. C., Gupta, R., Rando, T. A., Baker, J. C., Snyder, M. P., Cherry, J. M., Brunet, A. 2014; 158 (3): 673-688

  Abstract

  Trimethylation of histone H3 at lysine 4 (H3K4me3) is a chromatin modification known to mark the transcription start sites of active genes. Here, we show that H3K4me3 domains that spread more broadly over genes in a given cell type preferentially mark genes that are essential for the identity and function of that cell type. Using the broadest H3K4me3 domains as a discovery tool in neural progenitor cells, we identify novel regulators of these cells. Machine learning models reveal that the broadest H3K4me3 domains represent a distinct entity, characterized by increased marks of elongation. The broadest H3K4me3 domains also have more paused polymerase at their promoters, suggesting a unique transcriptional output. Indeed, genes marked by the broadest H3K4me3 domains exhibit enhanced transcriptional consistency and [corrected] increased transcriptional levels, and perturbation of H3K4me3 breadth leads to changes in transcriptional consistency. Thus, H3K4me3 breadth contains information that could ensure transcriptional precision at key cell identity/function genes.

  View details for DOI 10.1016/j.cell.2014.06.027

  View details for PubMedID 25083876

• Lineage of origin in rhabdomyosarcoma informs pharmacological response. Genes & development Abraham, J., Nuñez-Álvarez, Y., Hettmer, S., Carrió, E., Chen, H. H., Nishijo, K., Huang, E. T., Prajapati, S. I., Walker, R. L., Davis, S., Rebeles, J., Wiebush, H., McCleish, A. T., Hampton, S. T., Bjornson, C. R., Brack, A. S., Wagers, A. J., Rando, T. A., Capecchi, M. R., Marini, F. C., Ehler, B. R., Zarzabal, L. A., Goros, M. W., Michalek, J. E., Meltzer, P. S., Langenau, D. M., Legallo, R. D., Mansoor, A., Chen, Y., Suelves, M., Rubin, B. P., Keller, C. 2014; 28 (14): 1578-1591

  Abstract

  Lineage or cell of origin of cancers is often unknown and thus is not a consideration in therapeutic approaches. Alveolar rhabdomyosarcoma (aRMS) is an aggressive childhood cancer for which the cell of origin remains debated. We used conditional genetic mouse models of aRMS to activate the pathognomonic Pax3:Foxo1 fusion oncogene and inactivate p53 in several stages of prenatal and postnatal muscle development. We reveal that lineage of origin significantly influences tumor histomorphology and sensitivity to targeted therapeutics. Furthermore, we uncovered differential transcriptional regulation of the Pax3:Foxo1 locus by tumor lineage of origin, which led us to identify the histone deacetylase inhibitor entinostat as a pharmacological agent for the potential conversion of Pax3:Foxo1-positive aRMS to a state akin to fusion-negative RMS through direct transcriptional suppression of Pax3:Foxo1.

  View details for DOI 10.1101/gad.238733.114

  View details for PubMedID 25030697

• The JAK-STAT Pathway Is Critical in Ventilator-Induced Diaphragm Dysfunction MOLECULAR MEDICINE Tang, H., Smith, I. J., Hussain, S. N., Goldberg, P., Lee, M., Sugiarto, S., Godinez, G. L., Singh, B. K., Payan, D. G., Rando, T. A., Kinsella, T. M., Shrager, J. B. 2014; 20: 579-589

  Abstract

  Mechanical ventilation (MV) is one of the lynchpins of modern intensive-care medicine and is life saving in many critically ill patients. Continuous ventilator support, however, results in ventilation-induced diaphragm dysfunction (VIDD) that likely prolongs patients' need for MV and thereby leads to major associated complications and avoidable intensive care unit (ICU) deaths. Oxidative stress is a key pathogenic event in the development of VIDD, but its regulation remains largely undefined. We report here that the JAK-STAT pathway is activated in MV in the human diaphragm, as evidenced by significantly increased phosphorylation of JAK and STAT. Blockage of the JAK-STAT pathway by a JAK inhibitor in a rat MV model prevents diaphragm muscle contractile dysfunction (by ~85%, p < 0.01). We further demonstrate that activated STAT3 compromises mitochondrial function and induces oxidative stress in vivo, and, interestingly, that oxidative stress also activates JAK-STAT. Inhibition of JAK-STAT prevents oxidative stress-induced protein oxidation and polyubiquitination and recovers mitochondrial function in cultured muscle cells. Therefore, in ventilated diaphragm muscle, activation of JAK-STAT is critical in regulating oxidative stress and is thereby central to the downstream pathogenesis of clinical VIDD. These findings establish the molecular basis for the therapeutic promise of JAK-STAT inhibitors in ventilated ICU patients.

  View details for DOI 10.2119/molmed.2014.00049

  View details for Web of Science ID 000349988800001

  View details for PubMedCentralID PMC4365068

• mTORC1 controls the adaptive transition of quiescent stem cells from G(0) to G(Alert) NATURE Rodgers, J. T., King, K. Y., Brett, J. O., Cromie, M. J., Charville, G. W., Maguire, K. K., Brunson, C., Mastey, N., Liu, L., Tsai, C., Goodell, M. A., Rando, T. A. 2014; 510 (7505): 393-?

  Abstract

  A unique property of many adult stem cells is their ability to exist in a non-cycling, quiescent state. Although quiescence serves an essential role in preserving stem cell function until the stem cell is needed in tissue homeostasis or repair, defects in quiescence can lead to an impairment in tissue function. The extent to which stem cells can regulate quiescence is unknown. Here we show that the stem cell quiescent state is composed of two distinct functional phases, G0 and an 'alert' phase we term G(Alert). Stem cells actively and reversibly transition between these phases in response to injury-induced systemic signals. Using genetic mouse models specific to muscle stem cells (or satellite cells), we show that mTORC1 activity is necessary and sufficient for the transition of satellite cells from G0 into G(Alert) and that signalling through the HGF receptor cMet is also necessary. We also identify G0-to-G(Alert) transitions in several populations of quiescent stem cells. Quiescent stem cells that transition into G(Alert) possess enhanced tissue regenerative function. We propose that the transition of quiescent stem cells into G(Alert) functions as an 'alerting' mechanism, an adaptive response that positions stem cells to respond rapidly under conditions of injury and stress, priming them for cell cycle entry.

  View details for DOI 10.1038/nature13255

  View details for Web of Science ID 000337350200034

  View details for PubMedCentralID PMC4065227

• mTORC1 controls the adaptive transition of quiescent stem cells from G0 to G(Alert). Nature Rodgers, J. T., King, K. Y., Brett, J. O., Cromie, M. J., Charville, G. W., Maguire, K. K., Brunson, C., Mastey, N., Liu, L., Tsai, C., Goodell, M. A., Rando, T. A. 2014; 510 (7505): 393-396

  Abstract

  A unique property of many adult stem cells is their ability to exist in a non-cycling, quiescent state. Although quiescence serves an essential role in preserving stem cell function until the stem cell is needed in tissue homeostasis or repair, defects in quiescence can lead to an impairment in tissue function. The extent to which stem cells can regulate quiescence is unknown. Here we show that the stem cell quiescent state is composed of two distinct functional phases, G0 and an 'alert' phase we term G(Alert). Stem cells actively and reversibly transition between these phases in response to injury-induced systemic signals. Using genetic mouse models specific to muscle stem cells (or satellite cells), we show that mTORC1 activity is necessary and sufficient for the transition of satellite cells from G0 into G(Alert) and that signalling through the HGF receptor cMet is also necessary. We also identify G0-to-G(Alert) transitions in several populations of quiescent stem cells. Quiescent stem cells that transition into G(Alert) possess enhanced tissue regenerative function. We propose that the transition of quiescent stem cells into G(Alert) functions as an 'alerting' mechanism, an adaptive response that positions stem cells to respond rapidly under conditions of injury and stress, priming them for cell cycle entry.

  View details for DOI 10.1038/nature13255

  View details for PubMedID 24870234

• Stem cells as vehicles for youthful regeneration of aged tissues. journals of gerontology. Series A, Biological sciences and medical sciences Rando, T. A., Wyss-Coray, T. 2014; 69: S39-42

  Abstract

  Stem cells hold great promise for regenerative therapies for a wide spectrum of diseases and disorders of aging by virtue of their ability to regenerate tissues and contribute to their homeostasis. Aging is associated with a marked decline in these functionalities of adult stem cells. As such, regeneration of aged tissues is both less efficient and less effective than that of young tissues. Recent studies have revealed the remarkably dynamic responses of stem cells to systemic signals, including the ability of "youthful" factors in the blood of young animals to enhance the functionality of aged stem cells. Thus, there is much hope that even aged stem cells retain a remarkable regenerative potential if provided with the correct cues and environment to engage in tissue repair. The overall focus of the presentations of this session is to address the determinants of changes in stem cell functionality with age, the key characteristics of stem cells in aged tissues, the extent to which those characteristics are capable of being rejuvenated and by what signals, and the potential for stem cell therapeutics for chronic diseases and acute injuries in aged individuals.

  View details for DOI 10.1093/gerona/glu043

  View details for PubMedID 24833585

  View details for PubMedCentralID PMC4022127

• Alive and well? Exploring disease by studying lifespan. Current opinion in genetics & development Brett, J. O., Rando, T. A. 2014; 26: 33-40

  Abstract

  A common concept in aging research is that chronological age is the most important risk factor for the development of diverse diseases, including degenerative diseases and cancers. The mechanistic link between the aging process and disease pathogenesis, however, is still enigmatic. Nevertheless, measurement of lifespan, as a surrogate for biological aging, remains among the most frequently used assays in aging research. In this review, we examine the connection between 'normal aging' and age-related disease from the point of view that they form a continuum of aging phenotypes. This notion of common mechanisms gives rise to the converse postulate that diseases may be risk factors for accelerated aging. We explore the advantages and caveats associated with using lifespan as a metric to understand cell and tissue aging, focusing on the elucidation of molecular mechanisms and potential therapies for age-related diseases.

  View details for DOI 10.1016/j.gde.2014.05.004

  View details for PubMedID 25005743

  View details for PubMedCentralID PMC4253307

• Alive and well? Exploring disease by studying lifespan CURRENT OPINION IN GENETICS & DEVELOPMENT Brett, J. O., Rando, T. A. 2014; 26: 33-40

  Abstract

  A common concept in aging research is that chronological age is the most important risk factor for the development of diverse diseases, including degenerative diseases and cancers. The mechanistic link between the aging process and disease pathogenesis, however, is still enigmatic. Nevertheless, measurement of lifespan, as a surrogate for biological aging, remains among the most frequently used assays in aging research. In this review, we examine the connection between 'normal aging' and age-related disease from the point of view that they form a continuum of aging phenotypes. This notion of common mechanisms gives rise to the converse postulate that diseases may be risk factors for accelerated aging. We explore the advantages and caveats associated with using lifespan as a metric to understand cell and tissue aging, focusing on the elucidation of molecular mechanisms and potential therapies for age-related diseases.

  View details for DOI 10.1016/j.gde.2014.05.004

  View details for Web of Science ID 000345060800006

  View details for PubMedCentralID PMC4253307

• Stem cells as vehicles for youthful regeneration of aged tissues. journals of gerontology. Series A, Biological sciences and medical sciences Rando, T. A., Wyss-Coray, T. 2014; 69: S39-42

  View details for DOI 10.1093/gerona/glu043

  View details for PubMedID 24833585

• FOXO3 Promotes Quiescence in Adult Muscle Stem Cells during the Process of Self-Renewal. Stem cell reports Gopinath, S. D., Webb, A. E., Brunet, A., Rando, T. A. 2014; 2 (4): 414-426

  Abstract

  Skeletal muscle stem cells, or "satellite cells" (SCs), are required for the regeneration of damaged muscle tissue. Although SCs self-renew during regeneration, the mechanisms that govern SC re-entry into quiescence remain elusive. We show that FOXO3, a member of the forkhead family of transcription factors, is expressed in quiescent SCs (QSCs). Conditional deletion of Foxo3 in QSCs impairs self-renewal and increases the propensity of SCs to adopt a differentiated fate. Transcriptional analysis of SCs lacking FOXO3 revealed a downregulation of Notch signaling, a key regulator of SC quiescence. Conversely, overexpression of Notch intracellular domain (NICD) rescued the self-renewal deficit of FOXO3-deficient SCs. We show that FOXO3 regulates NOTCH1 and NOTCH3 receptor expression and that decreasing expression of NOTCH1 and NOTCH3 receptors phenocopies the effect of FOXO3 deficiency in SCs. We demonstrate that FOXO3, perhaps by activating Notch signaling, promotes the quiescent state during SC self-renewal in adult muscle regeneration.

  View details for DOI 10.1016/j.stemcr.2014.02.002

  View details for PubMedID 24749067

  View details for PubMedCentralID PMC3986584

• Of fish aid men NATURE CHEMICAL BIOLOGY Rando, T. A. 2014; 10 (2): 91–92

  View details for PubMedID 24441638

• Numb-deficient satellite cells have regeneration and proliferation defects PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA George, R. M., Biressi, S., Beres, B. J., Rogers, E., Mulia, A. K., Allen, R. E., Rawls, A., Rando, T. A., Wilson-Rawls, J. 2013; 110 (46): 18549-18554

  Abstract

  The adaptor protein Numb has been implicated in the switch between cell proliferation and differentiation made by satellite cells during muscle repair. Using two genetic approaches to ablate Numb, we determined that, in its absence, muscle regeneration in response to injury was impaired. Single myofiber cultures demonstrated a lack of satellite cell proliferation in the absence of Numb, and the proliferation defect was confirmed in satellite cell cultures. Quantitative RT-PCR from Numb-deficient satellite cells demonstrated highly up-regulated expression of p21 and Myostatin, both inhibitors of myoblast proliferation. Transfection with Myostatin-specific siRNA rescued the proliferation defect of Numb-deficient satellite cells. Furthermore, overexpression of Numb in satellite cells inhibited Myostatin expression. These data indicate a unique function for Numb during the initial activation and proliferation of satellite cells in response to muscle injury.

  View details for DOI 10.1073/pnas.1311628110

  View details for Web of Science ID 000326830900059

  View details for PubMedID 24170859

  View details for PubMedCentralID PMC3831958

• The CHC22 Clathrin-GLUT4 Transport Pathway Contributes to Skeletal Muscle Regeneration PLOS ONE Hoshino, S., Sakamoto, K., Vassilopoulos, S., Camus, S. M., Griffin, C. A., Esk, C., Torres, J. A., Ohkoshi, N., Ishii, A., Tamaoka, A., Funke, B. H., Kucherlapati, R., Margeta, M., Rando, T. A., Brodsky, F. M. 2013; 8 (10)

  Abstract

  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 these findings, CHC22 and GLUT4 can be considered markers of muscle regeneration in humans.

  View details for DOI 10.1371/journal.pone.0077787

  View details for Web of Science ID 000326334500060

  View details for PubMedID 24204966

  View details for PubMedCentralID PMC3813726

• FOXO3 Shares Common Targets with ASCL1 Genome-wide and Inhibits ASCL1-Dependent Neurogenesis. Cell reports Webb, A. E., Pollina, E. A., Vierbuchen, T., Urbán, N., Ucar, D., Leeman, D. S., Martynoga, B., Sewak, M., Rando, T. A., Guillemot, F., Wernig, M., Brunet, A. 2013; 4 (3): 477-491

  Abstract

  FOXO transcription factors are central regulators of longevity from worms to humans. FOXO3, the FOXO isoform associated with exceptional human longevity, preserves adult neural stem cell pools. Here, we identify FOXO3 direct targets genome-wide in primary cultures of adult neural progenitor cells (NPCs). Interestingly, FOXO3-bound sites are enriched for motifs for bHLH transcription factors, and FOXO3 shares common targets with the proneuronal bHLH transcription factor ASCL1/MASH1 in NPCs. Analysis of the chromatin landscape reveals that FOXO3 and ASCL1 are particularly enriched at the enhancers of genes involved in neurogenic pathways. Intriguingly, FOXO3 inhibits ASCL1-dependent neurogenesis in NPCs and direct neuronal conversion in fibroblasts. FOXO3 also restrains neurogenesis in vivo. Our study identifies a genome-wide interaction between the prolongevity transcription factor FOXO3 and the cell-fate determinant ASCL1 and raises the possibility that FOXO3's ability to restrain ASCL1-dependent neurogenesis may help preserve the neural stem cell pool.

  View details for DOI 10.1016/j.celrep.2013.06.035

  View details for PubMedID 23891001

• Cardiac Aging and Rejuvenation A Sense of Humors? NEW ENGLAND JOURNAL OF MEDICINE Rando, T. A., Finkel, T. 2013; 369 (6): 575–76

  View details for PubMedID 23924010

• The mortal strand hypothesis: Non-random chromosome inheritance and the biased segregation of damaged DNA. Seminars in cell & developmental biology Charville, G. W., Rando, T. A. 2013; 24 (8-9): 653-660

  Abstract

  If a eukaryotic cell is to reproduce, it must duplicate its genetic information in the form of DNA, and faithfully segregate that information during a complex process of cell division. During this division process, the resulting cells inherit one, and only one, copy of each chromosome. Over thirty years ago, it was predicted that the segregation of sister chromosomes could occur non-randomly, such that a daughter cell would preferentially inherit one of the two sister chromosomes according to some characteristic of that chromosome's template DNA strand. Although this prediction has been confirmed in studies of various cell-types, we know little of both the mechanism by which the asymmetric inheritance occurs and the significance it has to cells. In this essay, we propose a new model of non-random chromosome segregation-the mortal strand hypothesis-and discuss tests of the model that will provide insight into the molecular choreography of this intriguing phenomenon.

  View details for DOI 10.1016/j.semcdb.2013.05.006

  View details for PubMedID 23701893

• Myf5 expression during fetal myogenesis defines the developmental progenitors of adult satellite cells DEVELOPMENTAL BIOLOGY Biressi, S., Bjornson, C. R., Carlig, P. M., Nishijo, K., Keller, C., Rando, T. A. 2013; 379 (2): 195-207

  Abstract

  Myf5 is a member of the muscle-specific determination genes and plays a critical role in skeletal muscle development. Whereas the expression of Myf5 during embryonic and fetal myogenesis has been extensively studied, its expression in progenitors that will ultimately give rise to adult satellite cells, the stem cells responsible for muscle repair, is still largely unexplored. To investigate this aspect, we have generated a mouse strain carrying a CreER coding sequence in the Myf5 locus. In this strain, Tamoxifen-inducible Cre activity parallels endogenous Myf5 expression. Combining Myf5(CreER) and Cre reporter alleles, we were able to evaluate the contribution of cells expressing Myf5 at distinct developmental stages to the pool of satellite cells in adult hindlimb muscles. Although it was possible to trace back the origin of some rare satellite cells to a subpopulation of Myf5(+ve) progenitors in the limb buds at the late embryonic stage (∼E12), a significant number of satellite cells arise from cells which expressed Myf5 for the first time at the fetal stage (∼E15). These studies provide direct evidence that adult satellite cells derive from progenitors that first express the myogenic determination gene Myf5 during fetal stages of myogenesis.

  View details for DOI 10.1016/j.ydbio.2013.04.021

  View details for Web of Science ID 000320412300006

  View details for PubMedID 23639729

  View details for PubMedCentralID PMC3679295

• Chromatin Modifications as Determinants of Muscle Stem Cell Quiescence and Chronological Aging CELL REPORTS Liu, L., Cheung, T. H., Charville, G. W., Hurgo, B. M., Leavitt, T., Shih, J., Brunet, A., Rando, T. A. 2013; 4 (1): 189-204

  Abstract

  The ability to maintain quiescence is critical for the long-term maintenance of a functional stem cell pool. To date, the epigenetic and transcriptional characteristics of quiescent stem cells and how they change with age remain largely unknown. In this study, we explore the chromatin features of adult skeletal muscle stem cells, or satellite cells (SCs), which reside predominantly in a quiescent state in fully developed limb muscles of both young and aged mice. Using a ChIP-seq approach to obtain global epigenetic profiles of quiescent SCs (QSCs), we show that QSCs possess a permissive chromatin state in which few genes are epigenetically repressed by Polycomb group (PcG)-mediated histone 3 lysine 27 trimethylation (H3K27me3), and a large number of genes encoding regulators that specify nonmyogenic lineages are demarcated by bivalent domains at their transcription start sites (TSSs). By comparing epigenetic profiles of QSCs from young and old mice, we also provide direct evidence that, with age, epigenetic changes accumulate and may lead to a functional decline in quiescent stem cells. These findings highlight the importance of chromatin mapping in understanding unique features of stem cell identity and stem cell aging.

  View details for DOI 10.1016/j.celrep.2013.05.043

  View details for PubMedID 23810552

• A sexy spin on nonrandom chromosome segregation. Cell stem cell Charville, G. W., Rando, T. A. 2013; 12 (6): 641-643

  Abstract

  Nonrandom chromosome segregation is an intriguing phenomenon linked to certain asymmetric stem cell divisions. In a recent report in Nature, Yadlapalli and Yamashita (2013) observe nonrandom segregation of X and Y chromosomes in Drosophila germline stem cells and shed light on the complex mechanisms of this fascinating process.

  View details for DOI 10.1016/j.stem.2013.05.013

  View details for PubMedID 23746972

• Molecular regulation of stem cell quiescence. Nature reviews. Molecular cell biology Cheung, T. H., Rando, T. A. 2013; 14 (6): 329-340

  Abstract

  Subsets of mammalian adult stem cells reside in the quiescent state for prolonged periods of time. This state, which is reversible, has long been viewed as dormant and with minimal basal activity. Recent advances in adult stem cell isolation have provided insights into the epigenetic, transcriptional and post-transcriptional control of quiescence and suggest that quiescence is an actively maintained state in which signalling pathways are involved in maintaining a poised state that allows rapid activation. Deciphering the molecular mechanisms regulating adult stem cell quiescence will increase our understanding of tissue regeneration mechanisms and how they are dysregulated in pathological conditions and in ageing.

  View details for DOI 10.1038/nrm3591

  View details for PubMedID 23698583

• Collagen VI regulates satellite cell self-renewal and muscle regeneration NATURE COMMUNICATIONS Urciuolo, A., Quarta, M., Morbidoni, V., Gattazzo, F., Molon, S., Grumati, P., Montemurro, F., Tedesco, F. S., Blaauw, B., Cossu, G., Vozzi, G., Rando, T. A., Bonaldo, P. 2013; 4

  Abstract

  Adult muscle stem cells, or satellite cells have essential roles in homeostasis and regeneration of skeletal muscles. Satellite cells are located within a niche that includes myofibers and extracellular matrix. The function of specific extracellular matrix molecules in regulating SCs is poorly understood. Here, we show that the extracellular matrix protein collagen VI is a key component of the satellite cell niche. Lack of collagen VI in Col6a1(-/-) mice causes impaired muscle regeneration and reduced satellite cell self-renewal capability after injury. Collagen VI null muscles display significant decrease of stiffness, which is able to compromise the in vitro and in vivo activity of wild-type satellite cells. When collagen VI is reinstated in vivo by grafting wild-type fibroblasts, the biomechanical properties of Col6a1(-/-) muscles are ameliorated and satellite cell defects rescued. Our findings establish a critical role for an extracellular matrix molecule in satellite cell self-renewal and open new venues for therapies of collagen VI-related muscle diseases.

  View details for DOI 10.1038/ncomms2964

  View details for Web of Science ID 000323624100032

  View details for PubMedID 23743995

  View details for PubMedCentralID PMC3682802

• Heterochronic parabiosis: historical perspective and methodological considerations for studies of aging and longevity. Aging cell Conboy, M. J., Conboy, I. M., Rando, T. A. 2013; 12 (3): 525-530

  Abstract

  Pairing two animals in parabiosis to test for systemic or circulatory factors from one animal affecting the other animal has been used in scientific studies for at least 150 years. These studies have led to advances in fields as diverse as endocrinology, immunology, and oncology. A variation on the technique, heterochronic parabiosis, whereby two animals of different ages are joined to test for systemic regulators of aspects of aging or age-related diseases also has almost a century-long scientific history. In this review, we focus on the history of heterochronic parabiosis, methodological considerations and caveats, and the major advances that have emerged from those studies, including recent advances in our understanding of stem cell aging.

  View details for DOI 10.1111/acel.12065

  View details for PubMedID 23489470

• Assessment of disease activity in muscular dystrophies by noninvasive imaging. journal of clinical investigation Maguire, K. K., Lim, L., Speedy, S., Rando, T. A. 2013; 123 (5): 2298-2305

  Abstract

  Muscular dystrophies are a class of disorders that cause progressive muscle wasting. A major hurdle for discovering treatments for the muscular dystrophies is a lack of reliable assays to monitor disease progression in animal models. We have developed a novel mouse model to assess disease activity noninvasively in mice with muscular dystrophies. These mice express an inducible luciferase reporter gene in muscle stem cells. In dystrophic mice, muscle stem cells activate and proliferate in response to muscle degeneration, resulting in an increase in the level of luciferase expression, which can be monitored by noninvasive, bioluminescence imaging. We applied this noninvasive imaging to assess disease activity in a mouse model of the human disease limb girdle muscular dystrophy 2B (LGMD2B), caused by a mutation in the dysferlin gene. We monitored the natural history and disease progression in these dysferlin-deficient mice up to 18 months of age and were able to detect disease activity prior to the appearance of any overt disease manifestation by histopathological analyses. Disease activity was reflected by changes in luciferase activity over time, and disease burden was reflected by cumulative luciferase activity, which paralleled disease progression as determined by histopathological analysis. The ability to monitor disease activity noninvasively in mouse models of muscular dystrophy will be invaluable for the assessment of disease progression and the effectiveness of therapeutic interventions.

  View details for DOI 10.1172/JCI68458

  View details for PubMedID 23619364

  View details for PubMedCentralID PMC3638910

• Type 2 Innate Signals Stimulate Fibro/Adipogenic Progenitors to Facilitate Muscle Regeneration CELL Heredia, J. E., Mukundan, L., Chen, F. M., Mueller, A. A., Deo, R. C., Locksley, R. M., Rando, T. A., Chawla, A. 2013; 153 (2): 376-388

  Abstract

  In vertebrates, activation of innate immunity is an early response to injury, implicating it in the regenerative process. However, the mechanisms by which innate signals might regulate stem cell functionality are unknown. Here, we demonstrate that type 2 innate immunity is required for regeneration of skeletal muscle after injury. Muscle damage results in rapid recruitment of eosinophils, which secrete IL-4 to activate the regenerative actions of muscle resident fibro/adipocyte progenitors (FAPs). In FAPs, IL-4/IL-13 signaling serves as a key switch to control their fate and functions. Activation of IL-4/IL-13 signaling promotes proliferation of FAPs to support myogenesis while inhibiting their differentiation into adipocytes. Surprisingly, type 2 cytokine signaling is also required in FAPs, but not in myeloid cells, for rapid clearance of necrotic debris, a process that is necessary for timely and complete regeneration of tissues.

  View details for DOI 10.1016/j.cell.2013.02.053

  View details for Web of Science ID 000317349700016

  View details for PubMedID 23582327

  View details for PubMedCentralID PMC3663598

• All's well that ends well: alternative polyadenylation and its implications for stem cell biology CURRENT OPINION IN CELL BIOLOGY Mueller, A. A., Cheung, T. H., Rando, T. A. 2013; 25 (2): 222-232

  Abstract

  Stem cell quiescence, activation, and differentiation are governed by a complex network of molecular pathways. There has been a growing recognition that posttranscriptional modifications, such as alternative polyadenylation (APA) of transcripts, play an important role in regulating gene expression and function. Recent analyses of stem cell populations have suggested that APA controls stem cell fate and behavior. Here, we review recent developments that have shaped our understanding of the control of stem cell behavior by APA and we highlight promising areas for future investigation.

  View details for DOI 10.1016/j.ceb.2012.12.008

  View details for Web of Science ID 000317886100012

  View details for PubMedID 23357469

  View details for PubMedCentralID PMC3615088

• The Ins and Outs of Aging and Longevity ANNUAL REVIEW OF PHYSIOLOGY, VOL 75 Rando, T. A. 2013; 75: 617-619

  Abstract

  As a nod to the oft-quoted evolutionary theorist George Williams, "It is remarkable that after a seemingly miraculous feat of morphogenesis, a complex metazoan should be unable to perform the much simpler task of merely maintaining what is already formed". How and why we age are mysteries of the ages. The "how" of this mystery is the purview of experimental biologists who try to understand the basic processes that lead to system maintenance failure-from the level of molecules to that of entire organisms-that we term "aging". The "why" of this mystery is the purview of evolutionary theorists whose ideas shape the questions that biogerontologists pose, on the basis of the premise put forth by another preeminent geneticist and evolutionary biologist, Theodosius Dobzhansky, that "[n]othing in biology makes sense except in the light of evolution". These experimental and evolutionary perspectives converge in the modern science of aging, and its curious cousin "longevity", in an attempt to unify extensive findings from diverse areas of biology.

  View details for DOI 10.1146/annurev-physiol-092712-103439

  View details for Web of Science ID 000316381400027

  View details for PubMedID 23398156

• Guest Editorial: Emergent themes from Second Annual Symposium on Regenerative Rehabilitation, Pittsburgh, Pennsylvania. Journal of rehabilitation research and development Ambrosio, F. n., Boninger, M. L., Brubaker, C. E., Delitto, A. n., Wagner, W. R., Shields, R. K., Wolf, S. L., Rando, T. A. 2013; 50 (3): vii-xiv

  View details for PubMedID 23881770

• Sprouting a new take on stem cell aging EMBO JOURNAL Rodgers, J. T., Rando, T. A. 2012; 31 (21): 4103-4105

  View details for DOI 10.1038/emboj.2012.281

  View details for Web of Science ID 000310724300004

  View details for PubMedID 23042560

  View details for PubMedCentralID PMC3492738

• Recent advances in the pathogenesis and treatment of neuromuscular diseases CURRENT OPINION IN NEUROLOGY Rando, T. A. 2012; 25 (5): 586-587

  View details for DOI 10.1097/WCO.0b013e328357f2a1

  View details for Web of Science ID 000308728000010

  View details for PubMedID 22892955

• The place of genetics in ageing research NATURE REVIEWS GENETICS Barzilai, N., Guarente, L., Kirkwood, T. B., Partridge, L., Rando, T. A., Slagboom, P. E. 2012; 13 (8): 589-594

  Abstract

  Rapidly increasing numbers of older people present many countries with growing social and economic challenges. Yet despite the far-reaching implications of ageing, its biological basis remains a topic of much debate. Recent advances in genomics have spurred research on ageing and lifespan in human populations, adding to extensive genetic studies being carried out in model organisms. But how far is ageing controlled by our genes? In this Viewpoint, six experts present their opinions and comment on future directions in ageing research.

  View details for DOI 10.1038/nrg3290

  View details for Web of Science ID 000306524400013

  View details for PubMedID 22777128

• Heterochronic parabiosis for the study of the effects of aging on stem cells and their niches CELL CYCLE Conboy, I. M., Rando, T. A. 2012; 11 (12): 2260-2267

  Abstract

  Aging is unmistakable and undeniable in mammals. Interestingly, mice develop cataracts, muscle atrophy, osteoporosis, obesity, diabetes and cognitive deficits after just 2-3 postnatal years, while it takes seven or more decades for the same age-specific phenotypes to develop in humans. Thus, chronological age corresponds differently with biological age in metazoan species and although many theories exist, we do not understand what controls the rate of mammalian aging. One interesting idea is that species-specific rate of aging represents a ratio of tissue attrition to tissue regeneration. Furthermore, current findings suggest that the age-imposed biochemical changes in the niches of tissue stem cells inhibit performance of this regenerative pool, which leads to the decline of tissue maintenance and repair. If true, slowing down stem cell and niche aging, thereby promoting tissue regeneration, could slow down the process of tissue and organismal aging. In this regard, recent studies of heterochronic parabiosis provide important clues as to the mechanisms of stem cell aging and suggest novel strategies for enhancing tissue repair in the old. Here we review current literature on the relationship between the vigor of tissue stem cells and the process of aging, with an emphasis on the rejuvenation of old tissues by the extrinsic modifications of stem cell niches.

  View details for DOI 10.4161/cc.20437

  View details for Web of Science ID 000305353000015

  View details for PubMedID 22617385

  View details for PubMedCentralID PMC3383588

• Tissue-Specific Stem Cells: Lessons from the Skeletal Muscle Satellite Cell CELL STEM CELL Brack, A. S., Rando, T. A. 2012; 10 (5): 504-514

  Abstract

  In 1961, the satellite cell was first identified when electron microscopic examination of skeletal muscle demonstrated a cell wedged between the plasma membrane of the muscle fiber and the basement membrane. In recent years it has been conclusively demonstrated that the satellite cell is the primary cellular source for muscle regeneration and is equipped with the potential to self renew, thus functioning as a bona fide skeletal muscle stem cell (MuSC). As we move past the 50(th) anniversary of the satellite cell, we take this opportunity to discuss the current state of the art and dissect the unknowns in the MuSC field.

  View details for DOI 10.1016/j.stem.2012.04.001

  View details for Web of Science ID 000304234600009

  View details for PubMedID 22560074

  View details for PubMedCentralID PMC3348769

• Alternative Polyadenylation Mediates MicroRNA Regulation of Muscle Stem Cell Function CELL STEM CELL Boutet, S. C., Cheung, T. H., Quach, N. L., Liu, L., Prescott, S. L., Edalati, A., Iori, K., Rando, T. A. 2012; 10 (3): 327-336

  Abstract

  Pax3, a key myogenic regulator, is transiently expressed during activation of adult muscle stem cells, or satellite cells (SCs), and is also expressed in a subset of quiescent SCs (QSCs), but only in specific muscles. The mechanisms regulating these variations in expression are not well understood. Here we show that Pax3 levels are regulated by miR-206, a miRNA with a previously demonstrated role in myogenic differentiation. In most QSCs and activated SCs, miR-206 expression suppresses Pax3 expression. Paradoxically, QSCs that express high levels of Pax3 also express high levels of miR-206. In these QSCs, Pax3 transcripts are subject to alternative polyadenylation, resulting in transcripts with shorter 3' untranslated regions (3'UTRs) that render them resistant to regulation by miR-206. Similar alternate polyadenylation of the Pax3 transcript also occurs in myogenic progenitors during development. Our findings may reflect a general role of alternative polyadenylation in circumventing miRNA-mediated regulation of stem cell function.

  View details for DOI 10.1016/j.stem.2012.01.017

  View details for Web of Science ID 000301466500013

  View details for PubMedID 22385659

  View details for PubMedCentralID PMC3306803

• Maintenance of muscle stem-cell quiescence by microRNA-489 NATURE Cheung, T. H., Quach, N. L., Charville, G. W., Liu, L., Park, L., Edalati, A., Yoo, B., Hoang, P., Rando, T. A. 2012; 482 (7386): 524-U247

  Abstract

  Among the key properties that distinguish adult mammalian stem cells from their more differentiated progeny is the ability of stem cells to remain in a quiescent state for prolonged periods of time. However, the molecular pathways for the maintenance of stem-cell quiescence remain elusive. Here we use adult mouse muscle stem cells (satellite cells) as a model system and show that the microRNA (miRNA) pathway is essential for the maintenance of the quiescent state. Satellite cells that lack a functional miRNA pathway spontaneously exit quiescence and enter the cell cycle. We identified quiescence-specific miRNAs in the satellite-cell lineage by microarray analysis. Among these, miRNA-489 (miR-489) is highly expressed in quiescent satellite cells and is quickly downregulated during satellite-cell activation. Further analysis revealed that miR-489 functions as a regulator of satellite-cell quiescence, as it post-transcriptionally suppresses the oncogene Dek, the protein product of which localizes to the more differentiated daughter cell during asymmetric division of satellite cells and promotes the transient proliferative expansion of myogenic progenitors. Our results provide evidence of the miRNA pathway in general, and of a specific miRNA, miR-489, in actively maintaining the quiescent state of an adult stem-cell population.

  View details for DOI 10.1038/nature10834

  View details for PubMedID 22358842

• Aging, Rejuvenation, and Epigenetic Reprogramming: Resetting the Aging Clock CELL Rando, T. A., Chang, H. Y. 2012; 148 (1-2): 46-57

  Abstract

  The underlying cause of aging remains one of the central mysteries of biology. Recent studies in several different systems suggest that not only may the rate of aging be modified by environmental and genetic factors, but also that the aging clock can be reversed, restoring characteristics of youthfulness to aged cells and tissues. This Review focuses on the emerging biology of rejuvenation through the lens of epigenetic reprogramming. By defining youthfulness and senescence as epigenetic states, a framework for asking new questions about the aging process emerges.

  View details for DOI 10.1016/j.cell.2012.01.003

  View details for PubMedID 22265401

• Losartan Improves Adipose Tissue-Derived Stem Cell Niche by Inhibiting Transforming Growth Factor-beta and Fibrosis in Skeletal Muscle Injury CELL TRANSPLANTATION Park, J., Ki, M., Lee, E., Kim, A., You, S., Han, S., Lee, E., Hong, I., Kwon, S., Kim, S., Rando, T. A., Jeong, K. 2012; 21 (11): 2407-2424

  Abstract

  Recently, adipose tissue-derived stem cells (ASCs) were emerged as an alternative, abundant, and easily accessible source of stem cell therapy. Previous studies revealed losartan (an angiotensin II type I receptor blocker) treatment promoted the healing of skeletal muscle by attenuation of the TGF-β signaling pathway, which inhibits muscle differentiation. Therefore, we hypothesized that a combined therapy using ASCs and losartan might dramatically improve the muscle remodeling after muscle injury. To determine the combined effect of losartan with ASC transplantation, we created a muscle laceration mouse model. EGFP-labeled ASCs were locally transplanted to the injured gastrocnemius muscle after muscle laceration. The dramatic muscle regeneration and the remarkably inhibited muscular fibrosis were observed by combined treatment. Transplanted ASCs fused with the injured or differentiating myofibers. Myotube formation was also enhanced by ASC(+) satellite coculture and losartan treatment. Thus, the present study indicated that ASC transplantation effect for skeletal muscle injury can be dramatically improved by losartan treatment inducing better niche.

  View details for DOI 10.3727/096368912X637055

  View details for Web of Science ID 000313143600007

  View details for PubMedID 22507443

• The ageing systemic milieu negatively regulates neurogenesis and cognitive function NATURE Villeda, S. A., Luo, J., Mosher, K. I., Zou, B., Britschgi, M., Bieri, G., Stan, T. M., Fainberg, N., Ding, Z., Eggel, A., Lucin, K. M., Czirr, E., Park, J., Couillard-Despres, S., Aigner, L., Li, G., Peskind, E. R., Kaye, J. A., Quinn, J. F., Galasko, D. R., Xie, X. S., Rando, T. A., Wyss-Coray, T. 2011; 477 (7362): 90-U157

  Abstract

  In the central nervous system, ageing results in a precipitous decline in adult neural stem/progenitor cells and neurogenesis, with concomitant impairments in cognitive functions. Interestingly, such impairments can be ameliorated through systemic perturbations such as exercise. Here, using heterochronic parabiosis we show that blood-borne factors present in the systemic milieu can inhibit or promote adult neurogenesis in an age-dependent fashion in mice. Accordingly, exposing a young mouse to an old systemic environment or to plasma from old mice decreased synaptic plasticity, and impaired contextual fear conditioning and spatial learning and memory. We identify chemokines--including CCL11 (also known as eotaxin)--the plasma levels of which correlate with reduced neurogenesis in heterochronic parabionts and aged mice, and the levels of which are increased in the plasma and cerebrospinal fluid of healthy ageing humans. Lastly, increasing peripheral CCL11 chemokine levels in vivo in young mice decreased adult neurogenesis and impaired learning and memory. Together our data indicate that the decline in neurogenesis and cognitive impairments observed during ageing can be in part attributed to changes in blood-borne factors.

  View details for DOI 10.1038/nature10357

  View details for Web of Science ID 000294404300037

  View details for PubMedID 21886162

  View details for PubMedCentralID PMC3170097

• Emerging models and paradigms for stem cell ageing NATURE CELL BIOLOGY Jones, D. L., Rando, T. A. 2011; 13 (5): 506-512

  Abstract

  Ageing is accompanied by a progressive decline in stem cell function, resulting in less effective tissue homeostasis and repair. Here we discuss emerging invertebrate models that provide insights into molecular pathways of age-related stem cell dysfunction in mammals, and we present various paradigms of how stem cell functionality changes with age, including impaired self-renewal and aberrant differentiation potential.

  View details for DOI 10.1038/ncb0511-506

  View details for Web of Science ID 000290148700005

  View details for PubMedID 21540846

  View details for PubMedCentralID PMC3257978

• Manifestations and mechanisms of stem cell aging JOURNAL OF CELL BIOLOGY Liu, L., Rando, T. A. 2011; 193 (2): 257-266

  Abstract

  Adult stem cells exist in most mammalian organs and tissues and are indispensable for normal tissue homeostasis and repair. In most tissues, there is an age-related decline in stem cell functionality but not a depletion of stem cells. Such functional changes reflect deleterious effects of age on the genome, epigenome, and proteome, some of which arise cell autonomously and others of which are imposed by an age-related change in the local milieu or systemic environment. Notably, some of the changes, particularly epigenomic and proteomic, are potentially reversible, and both environmental and genetic interventions can result in the rejuvenation of aged stem cells. Such findings have profound implications for the stem cell-based therapy of age-related diseases.

  View details for DOI 10.1083/jcb.201010131

  View details for Web of Science ID 000289673000003

  View details for PubMedID 21502357

  View details for PubMedCentralID PMC3080271

• Stem cell ageing and non-random chromosome segregation PHILOSOPHICAL TRANSACTIONS OF THE ROYAL SOCIETY B-BIOLOGICAL SCIENCES Charville, G. W., Rando, T. A. 2011; 366 (1561): 85-93

  Abstract

  Adult stem cells maintain the mature tissues of metazoans. They do so by reproducing in such a way that their progeny either differentiate, and thus contribute functionally to a tissue, or remain uncommitted and replenish the stem cell pool. Because ageing manifests as a general decline in tissue function, diminished stem cell-mediated tissue maintenance may contribute to age-related pathologies. Accordingly, the mechanisms by which stem cell regenerative potential is sustained, and the extent to which these mechanisms fail with age, are fundamental determinants of tissue ageing. Here, we explore the mechanisms of asymmetric division that account for the sustained fitness of adult stem cells and the tissues that comprise them. In particular, we summarize the theory and experimental evidence underlying non-random chromosome segregation-a mitotic asymmetry arising from the unequal partitioning of chromosomes according to the age of their template DNA strands. Additionally, we consider the possible consequences of non-random chromosome segregation, especially as they relate to both replicative and chronological ageing in stem cells. While biased segregation of chromosomes may sustain stem cell replicative potential by compartmentalizing the errors derived from DNA synthesis, it might also contribute to the accrual of replication-independent DNA damage in stem cells and thus hasten chronological ageing.

  View details for DOI 10.1098/rstb.2010.0279

  View details for PubMedID 21115534

• Taf1 Regulates Pax3 Protein by Monoubiquitination in Skeletal Muscle Progenitors MOLECULAR CELL Boutet, S. C., Biressi, S., Iori, K., Natu, V., Rando, T. A. 2010; 40 (5): 749-761

  Abstract

  Pax3 plays critical roles during developmental and postnatal myogenesis. We have previously shown that levels of Pax3 protein are regulated by monoubiquitination and proteasomal degradation during postnatal myogenesis, but none of the key regulators of the monoubiquitination process were known. Here we show that Pax3 monoubiquitination is mediated by the ubiquitin-activating/conjugating activity of Taf1, a component of the core transcriptional machinery that was recently reported to be downregulated during myogenic differentiation. We show that Taf1 binds directly to Pax3 and overexpression of Taf1 increases the level of monoubiquitinated Pax3 and its degradation by the proteasome. A decrease of Taf1 results in a decrease in Pax3 monoubiquitination, an increase in the levels of Pax3 protein, and a concomitant increase in Pax3-mediated inhibition of myogenic differentiation and myoblast migration. These results suggest that Taf1 regulates Pax3 protein levels through its ability to mediate monoubiquitination, revealing a critical interaction between two proteins that are involved in distinct aspects of myogenic differentiation. Finally, these results suggest that the components of the core transcriptional are integrally involved in the process of myogenic differentiation, acting as nodal regulators of the differentiation program.

  View details for DOI 10.1016/j.molcel.2010.09.029

  View details for Web of Science ID 000285405800009

  View details for PubMedID 21145483

  View details for PubMedCentralID PMC3023311

• Heterogeneity in the muscle satellite cell population SEMINARS IN CELL & DEVELOPMENTAL BIOLOGY Biressi, S., Rando, T. A. 2010; 21 (8): 845-854

  Abstract

  Satellite cells, the adult stem cells responsible for skeletal muscle regeneration, are defined by their location between the basal lamina and the fiber sarcolemma. Increasing evidence suggests that satellite cells represent a heterogeneous population of cells with distinct embryological origin and multiple levels of biochemical and functional diversity. This review focuses on the rich diversity of the satellite cell population based on studies across species. Ultimately, a more complete characterization of the heterogeneity of satellite cells will be essential to understand the functional significance in terms of muscle growth, homeostasis, tissue repair, and aging.

  View details for DOI 10.1016/j.semcdb.2010.09.003

  View details for Web of Science ID 000283598400011

  View details for PubMedID 20849971

  View details for PubMedCentralID PMC2967620

• Tumour stem cells in oncogenic RAS-dependent rhabdomyosarcoma Schaaf, G., Sage, F., Chan, A., Chen, F., Hamdi, M., Zwijnenburg, D., Boutet, S., Kool, M., Rando, T. PERGAMON-ELSEVIER SCIENCE LTD. 2010: 138

  View details for DOI 10.1016/S1359-6349(10)71343-4

  View details for Web of Science ID 000288603100525

• Epigenetics and aging EXPERIMENTAL GERONTOLOGY Rando, T. A. 2010; 45 (4): 253-254

  View details for DOI 10.1016/j.exger.2009.12.007

  View details for Web of Science ID 000276428300001

  View details for PubMedID 20034555

  View details for PubMedCentralID PMC2838984

• Impact papers on aging in 2009 AGING-US Blagosklonny, M. V., Campisi, J., Sinclair, D. A., Bartke, A., Blasco, M. A., Bonner, W. M., Bohr, V. A., Brosh, R. M., Brunet, A., DePinho, R. A., Donehower, L. A., Finch, C. E., Finkel, T., Gorospe, M., Gudkov, A. V., Hall, M. N., Hekimi, S., Helfand, S. L., Karlseder, J., Kenyon, C., Kroemer, G., Longo, V., Nussenzweig, A., Osiewacz, H. D., Peeper, D. S., Rando, T. A., Rudolph, K. L., Sassone-Corsi, P., Serrano, M., Sharpless, N. E., Skulachev, V. P., Tilly, J. L., Tower, J., Verdin, E., Vijg, J. 2010; 2 (3): 111-121

  Abstract

  The Editorial Board of Aging reviews research papers published in 2009, which they believe have or will have significant impact on aging research. Among many others, the topics include genes that accelerate aging or in contrast promote longevity in model organisms, DNA damage responses and telomeres, molecular mechanisms of life span extension by calorie restriction and pharmacological interventions into aging. The emerging message in 2009 is that aging is not random but determined by a genetically-regulated longevity network and can be decelerated both genetically and pharmacologically.

  View details for Web of Science ID 000277115300001

  View details for PubMedID 20351400

  View details for PubMedCentralID PMC2871240

• Enhanced gene repair mediated by methyl-CpG-modified single-stranded oligonucleotides NUCLEIC ACIDS RESEARCH Bertoni, C., Rustagi, A., Rando, T. A. 2009; 37 (22): 7468-7482

  Abstract

  Gene editing mediated by oligonucleotides has been shown to induce stable single base alterations in genomic DNA in both prokaryotic and eukaryotic organisms. However, the low frequencies of gene repair have limited its applicability for both basic manipulation of genomic sequences and for the development of therapeutic approaches for genetic disorders. Here, we show that single-stranded oligodeoxynucleotides (ssODNs) containing a methyl-CpG modification and capable of binding to the methyl-CpG binding domain protein 4 (MBD4) are able to induce >10-fold higher levels of gene correction than ssODNs lacking the specific modification. Correction was stably inherited through cell division and was confirmed at the protein, transcript and genomic levels. Downregulation of MBD4 expression using RNAi prevented the enhancement of gene correction efficacy obtained using the methyl-CpG-modified ssODN, demonstrating the specificity of the repair mechanism being recruited. Our data demonstrate that efficient manipulation of genomic targets can be achieved and controlled by the type of ssODN used and by modulation of the repair mechanism involved in the correction process. This new generation of ssODNs represents an important technological advance that is likely to have an impact on multiple applications, especially for gene therapy where permanent correction of the genetic defect has clear advantages over viral and other nonviral approaches currently being tested.

  View details for DOI 10.1093/nar/gkp757

  View details for Web of Science ID 000272935000020

  View details for PubMedID 19854937

  View details for PubMedCentralID PMC2794159

• BCL9 is an essential component of canonical Wnt signaling that mediates the differentiation of myogenic progenitors during muscle regeneration DEVELOPMENTAL BIOLOGY Brack, A. S., Murphy-Seiler, F., Hanifi, J., Deka, J., Eyckerman, S., Keller, C., Aguet, M., Rando, T. A. 2009; 335 (1): 93-105

  Abstract

  Muscle stem cells and their progeny play a fundamental role in the regeneration of adult skeletal muscle. We have previously shown that activation of the canonical Wnt/beta-catenin signaling pathway in adult myogenic progenitors is required for their transition from rapidly dividing transient amplifying cells to more differentiated progenitors. Whereas Wnt signaling in Drosophila is dependent on the presence of the co-regulator Legless, previous studies of the mammalian ortholog of Legless, BCL9 (and its homolog, BCL9-2), have not revealed an essential role of these proteins in Wnt signaling in specific tissues during development. Using Cre-lox technology to delete BCL9 and BCL9-2 in the myogenic lineage in vivo and RNAi technology to knockdown the protein levels in vitro, we show that BCL9 is required for activation of the Wnt/beta-catenin cascade in adult mammalian myogenic progenitors. We observed that the nuclear localization of beta-catenin and downstream TCF/LEF-mediated transcription, which are normally observed in myogenic progenitors upon addition of exogenous Wnt and during muscle regeneration, were abrogated when BCL9/9-2 levels were reduced. Furthermore, reductions of BCL9/9-2 inhibited the promotion of myogenic differentiation by Wnt and the normal regenerative response of skeletal muscle. These results suggest a critical role of BCL9/9-2 in the Wnt-mediated regulation of adult, as opposed to embryonic, myogenic progenitors.

  View details for DOI 10.1016/j.ydbio.2009.08.014

  View details for Web of Science ID 000271080300008

  View details for PubMedID 19699733

  View details for PubMedCentralID PMC3259687

• Biomarker system for studying muscle, stem cells, and cancer in vivo FASEB JOURNAL Nishijo, K., Hosoyama, T., Bjornson, C. R., Schaffer, B. S., Prajapati, S. I., Bahadur, A. N., Hansen, M. S., Blandford, M. C., McCleish, A. T., Rubin, B. P., Epstein, J. A., Rando, T. A., Capecchi, M. R., Keller, C. 2009; 23 (8): 2681-2690

  Abstract

  Bioluminescent reporter genes are sensitive in situ tools for following disease progression in preclinical models, albeit they are subject to scattering and absorption in deep tissues. We have generated a bicistronic Cre/LoxP reporter mouse line that pairs the expression of firefly luciferase with quantifiable expression of a human placental alkaline phosphatase that is secreted into the serum (SeAP). With the use of this dual-modality bioreporter with a novel, inducible Pax7-CreER line for tracking muscle satellite cells, we demonstrate the longitudinal kinetics of muscle stem cell turnover, accounting for a doubling of the signal from satellite cell and progeny every 3.93 wk in the transition from adolescence to early adulthood. We also show that this dual-modality bioreporter can be incorporated in preclinical cancer models, whereby SeAP activity is reflective of tumor burden. Thus, this dual bioreporter permits both spatial localization and accurate quantification of biological processes in vivo even when the tissue of interest is deep within the animal.

  View details for DOI 10.1096/fj.08-128116

  View details for Web of Science ID 000268836700036

  View details for PubMedID 19332644

  View details for PubMedCentralID PMC2717773

• Focal Adhesion Kinase Signaling Regulates the Expression of Caveolin 3 and beta 1 Integrin, Genes Essential for Normal Myoblast Fusion MOLECULAR BIOLOGY OF THE CELL Quach, N. L., Biressi, S., Reichardt, L. F., Keller, C., Rando, T. A. 2009; 20 (14): 3422-3435

  Abstract

  An essential phase of skeletal myogenesis is the fusion of mononucleated myoblasts to form multinucleated myotubes. Many cell adhesion proteins, including integrins, have been shown to be important for myoblast fusion in vertebrates, but the mechanisms by which these proteins regulate cell fusion remain mostly unknown. Here, we focused on the role of focal adhesion kinase (FAK), an important nonreceptor protein tyrosine kinase involved in integrin signaling, as a potential mediator by which integrins may regulate myoblast fusion. To test this hypothesis in vivo, we generated mice in which the Fak gene was disrupted specifically in muscle stem cells ("satellite cells") and we found that this resulted in impaired myotube formation during muscle regeneration after injury. To examine the role of FAK in the fusion of myogenic cells, we examined the expression of FAK and the effects of FAK deletion on the differentiation of myoblasts in vitro. Differentiation of mouse primary myoblasts was accompanied by a rapid and transient increase of phosphorylated FAK. To investigate the requirement of FAK in myoblast fusion, we used two loss-of-function approaches (a dominant-negative inhibitor of FAK and FAK small interfering RNA [siRNA]). Inhibition of FAK resulted in markedly impaired fusion but did not inhibit other biochemical measures of myogenic differentiation, suggesting a specific role of FAK in the morphological changes of cell fusion as part of the differentiation program. To examine the mechanisms by which FAK may be regulating fusion, we used microarray analysis to identify the genes that failed to be normally regulated in cells that were fusion defective due to FAK inhibition. Several genes that have been implicated in myoblast fusion were aberrantly regulated during differentiation when FAK was inhibited. Intriguingly, the normal increases in the transcript of caveolin 3 as well as an integrin subunit, the beta1D isoform, were suppressed by FAK inhibition. We confirmed this also at the protein level and show that direct inhibition of beta1D subunit expression by siRNA inhibited myotube formation with a prominent effect on secondary fusion. These data suggest that FAK regulation of profusion genes, including caveolin 3 and the beta1D integrin subunit, is essential for morphological muscle differentiation.

  View details for DOI 10.1091/mbc.E09-02-0175

  View details for Web of Science ID 000267981600023

  View details for PubMedID 19458188

  View details for PubMedCentralID PMC2710835

• Preventing oxidative stress: a new role for XBP1 CELL DEATH AND DIFFERENTIATION Liu, Y., Adachi, M., Zhao, S., Hareyama, M., Koong, A. C., Luo, D., Rando, T. A., Imai, K., Shinomura, Y. 2009; 16 (6): 847-857

  Abstract

  Antioxidant molecules reduce oxidative stress and protect cells from reactive oxygen species (ROS)-mediated cellular damage and probably the development of cancer. We have investigated the contribution of X-box-binding protein (XBP1), a major endoplasmic reticulum stress-linked transcriptional factor, to cellular resistance to oxidative stress. After exposure to hydrogen peroxide (H(2)O(2)) or a strong ROS inducer parthenolide, loss of mitochondrial membrane potential (MMP) and subsequent cell death occurred more extensively in XBP1-deficient cells than wild-type mouse embryonic fibroblast cells, whereas two other anticancer agents induced death similarly in both cells. In XBP1-deficient cells, H(2)O(2) exposure induced more extensive ROS generation and prolonged p38 phosphorylation, and expression of several antioxidant molecules including catalase was lower. Knockdown of XBP1 decreased catalase expression, enhanced ROS generation and MMP loss after H(2)O(2) exposure, but extrinsic catalase supply rescued them. Overexpression of XBP1 recovered catalase expression in XBP1-deficient cells and diminished ROS generation after H(2)O(2) exposure. Mutation analysis of the catalase promoter region suggests a pivotal role of CCAAT boxes, NF-Y-binding sites, for the XBP1-mediated enhancing effect. Taken together, these results indicate a protective role of XBP1 against oxidative stress, and its positive regulation of catalase expression may at least in part account for this function.

  View details for DOI 10.1038/cdd.2009.14

  View details for Web of Science ID 000266412400005

  View details for PubMedID 19247368

  View details for PubMedCentralID PMC2826168

• Turning back time: Reversing tissue pathology to enhance stem cell engraftment CELL STEM CELL Rando, T. A. 2008; 3 (3): 232-234

  View details for DOI 10.1016/j.stem.2008.08.006

  View details for Web of Science ID 000259665000003

  View details for PubMedID 18786408

• Stem cell review series: Aging of the skeletal muscle stem cell niche AGING CELL Gopinath, S. D., Rando, T. A. 2008; 7 (4): 590-598

  Abstract

  Declining stem cell function during aging contributes to impaired tissue function. Muscle-specific stem cells ('satellite cells') are responsible for generating new muscle in response to injury in the adult. However, aged muscle displays a significant reduction in regenerative abilities and an increased susceptibility to age-related pathologies. This review describes components of the satellite cell niche and addresses how age-related changes in these components impinge on satellite cell function. In particular, we review changes in the key niche elements, the myofiber and the basal lamina that are in intimate contact with satellite cells. We address how these elements are influenced by factors secreted by interstitial cells, cells of the immune system, and cells associated with the vasculature, all of which change with age. In addition, we consider more distant sources of influence on the satellite cell niche that change with age, such as neural-mediated trophic factors and electrical activity and systemic factors present in the circulation. A better understanding of the niche elements and their influence on the satellite cell will facilitate the development of therapeutic interventions aimed at improving satellite cell activity and ultimately tissue response to injury in aged individuals.

  View details for DOI 10.1111/j.1474-9726.2008.00399.x

  View details for Web of Science ID 000257513500015

  View details for PubMedID 18462272

• Tissue ageing: Do insights into molecular mechanisms of ageing lead to new therapeutic strategies? EXPERIMENTAL GERONTOLOGY Simm, A., Rando, T. A. 2008; 43 (7): 603-604

  View details for DOI 10.1016/j.exger.2008.05.009

  View details for Web of Science ID 000258310600001

  View details for PubMedID 18599230

  View details for PubMedCentralID PMC2536625

• Get personal with gene therapy for muscular dystrophy LANCET NEUROLOGY Rando, T. A. 2008; 7 (3): 196-198

  View details for Web of Science ID 000253580000005

  View details for PubMedID 18275918

• Technology Insight: therapy for Duchenne muscular dystrophy-an opportunity for personalized medicine? NATURE CLINICAL PRACTICE NEUROLOGY Lim, L. E., Rando, T. A. 2008; 4 (3): 149-158

  Abstract

  Since the identification of dystrophin as the protein product of the Duchenne and Becker muscular dystrophy locus, many different mutations, encompassing the entire spectrum of gene mutations ranging from point mutations to large deletions, have been found. These discoveries have led to the investigation of a variety of methods aimed at the treatment of muscular dystrophy, including strategies for gene replacement, gene correction, and modification of the gene product. The preferred approach in each case depends on the nature of the gene defect. In this Review, we focus on methods that have been developed for gene correction and for the modification of gene products. This mutation-focused approach offers the opportunity for 'personalized' gene therapy for muscular dystrophy and might also be a logical strategy for the treatment of other genetic disorders.

  View details for DOI 10.1038/ncpneuro0737

  View details for Web of Science ID 000253634300010

  View details for PubMedID 18268530

• A temporal switch from Notch to Wnt signaling in muscle stem cells is necessary for normal adult myogenesis CELL STEM CELL Brack, A. S., Conboy, I. M., Conboy, M. J., Shen, J., Rando, T. A. 2008; 2 (1): 50-59

  Abstract

  The temporal switch from progenitor cell proliferation to differentiation is essential for effective adult tissue repair. We previously reported the critical role of Notch signaling in the proliferative expansion of myogenic progenitors in mammalian postnatal myogenesis. We now show that the onset of differentiation is due to a transition from Notch signaling to Wnt signaling in myogenic progenitors and is associated with an increased expression of Wnt in the tissue and an increased responsiveness of progenitors to Wnt. Crosstalk between these two pathways occurs via GSK3beta, which is maintained in an active form by Notch but is inhibited by Wnt in the canonical Wnt signaling cascade. These results demonstrate that the temporal balance between Notch and Wnt signaling orchestrates the precise progression of muscle precursor cells along the myogenic lineage pathway, through stages of proliferative expansion and then differentiation, during postnatal myogenesis.

  View details for DOI 10.1016/j.stem.2007.10.006

  View details for Web of Science ID 000252606400011

  View details for PubMedID 18371421

• Proteasomal degradation of Pax3 in skeletal muscle progenitors: one ubiquitin does the trick! M S-MEDECINE SCIENCES Boutet, S. C., Rondo, T. A. 2008; 24 (1): 31-33

  View details for Web of Science ID 000252834800013

  View details for PubMedID 18198106

• Ageing - From stem to stern NATURE Brunet, A., Rando, T. A. 2007; 449 (7160): 288-?

  View details for DOI 10.1038/449288a

  View details for Web of Science ID 000249557300030

  View details for PubMedID 17882206

• Increased Wnt signaling during aging alters muscle stem cell fate and increases fibrosis SCIENCE Brack, A. S., Conboy, M. J., Roy, S., Lee, M., Kuo, C. J., Keller, C., Rando, T. A. 2007; 317 (5839): 807-810

  Abstract

  The regenerative potential of skeletal muscle declines with age, and this impairment is associated with an increase in tissue fibrosis. We show that muscle stem cells (satellite cells) from aged mice tend to convert from a myogenic to a fibrogenic lineage as they begin to proliferate and that this conversion is mediated by factors in the systemic environment of the old animals. We also show that this lineage conversion is associated with an activation of the canonical Wnt signaling pathway in aged myogenic progenitors and can be suppressed by Wnt inhibitors. Furthermore, components of serum from aged mice that bind to the Frizzled family of proteins, which are Wnt receptors, may account for the elevated Wnt signaling in aged cells. These results indicate that the Wnt signaling pathway may play a critical role in tissue-specific stem cell aging and an increase in tissue fibrosis with age.

  View details for DOI 10.1126/science.1144090

  View details for Web of Science ID 000248624500041

  View details for PubMedID 17690295

• Regulation of Pax3 by proteasomal degradation of monoubiquitinated protein in skeletal muscle progenitors CELL Boutet, S. C., Disatnik, M., Chan, L. S., Iori, K., Rando, T. A. 2007; 130 (2): 349-362

  Abstract

  Pax3 and Pax7 play distinct but overlapping roles in developmental and postnatal myogenesis. The mechanisms involved in the differential regulation of these highly homologous proteins are unknown. We present evidence that Pax3, but not Pax7, is regulated by ubiquitination and proteasomal degradation during adult muscle stem cell activation. Intriguingly, only monoubiquitinated forms of Pax3 could be detected. Mutation of two specific lysine residues in the C-terminal region of Pax3 reduced the extent of its monoubiquitination and susceptibility to proteasomal degradation, whereas introduction of a key lysine into the C-terminal region of Pax7 rendered that protein susceptible to monoubiquitination and proteasomal degradation. Monoubiquitinated Pax3 was shuttled to the intrinsic proteasomal protein S5a by interacting specifically with the ubiquitin-binding protein Rad23B. Functionally, sustained expression of Pax3 proteins inhibited myogenic differentiation, demonstrating that Pax3 degradation is an essential step for the progression of the myogenic program. These results reveal an important mechanism of Pax3 regulation in muscle progenitors and an unrecognized role of protein monoubiquitination in mediating proteasomal degradation.

  View details for DOI 10.1016/j.cell.2007.05.044

  View details for Web of Science ID 000248588000023

  View details for PubMedID 17662948

• The immortal strand hypothesis: Segregation and reconstruction CELL Rando, T. A. 2007; 129 (7): 1239-1243

  Abstract

  The immortal strand hypothesis posits that the propensity of stem cell compartments to give rise to cancer in later life can be minimized if stem cells, during the process of self-renewal, retain those DNA strands with the fewest mutations acquired during DNA replication. In this Essay, I explore evidence in support of the hypothesis, the biological implications, and the key questions that remain to be answered experimentally to address the fundamental tenets of the hypothesis.

  View details for DOI 10.1016/j.cell.2007.06.019

  View details for Web of Science ID 000247911400006

  View details for PubMedID 17604710

• High incidence of non-random template strand segregation and asymmetric fate determination in dividing stem cells and their progeny PLOS BIOLOGY Conboy, M. J., Karasov, A. O., Rando, T. A. 2007; 5 (5): 1120-1126

  Abstract

  Decades ago, the "immortal strand hypothesis" was proposed as a means by which stem cells might limit acquiring mutations that could give rise to cancer, while continuing to proliferate for the life of an organism. Originally based on observations in embryonic cells, and later studied in terms of stem cell self-renewal, this hypothesis has remained largely unaccepted because of few additional reports, the rarity of the cells displaying template strand segregation, and alternative interpretations of experiments involving single labels or different types of labels to follow template strands. Using sequential pulses of halogenated thymidine analogs (bromodeoxyuridine [BrdU], chlorodeoxyuridine [CldU], and iododeoxyuridine [IdU]), and analyzing stem cell progeny during induced regeneration in vivo, we observed extraordinarily high frequencies of segregation of older and younger template strands during a period of proliferative expansion of muscle stem cells. Furthermore, template strand co-segregation was strongly associated with asymmetric cell divisions yielding daughters with divergent fates. Daughter cells inheriting the older templates retained the more immature phenotype, whereas daughters inheriting the newer templates acquired a more differentiated phenotype. These data provide compelling evidence of template strand co-segregation based on template age and associated with cell fate determination, suggest that template strand age is monitored during stem cell lineage progression, and raise important caveats for the interpretation of label-retaining cells.

  View details for DOI 10.1371/journal.pbio.0050102

  View details for Web of Science ID 000246716700019

  View details for PubMedID 17439301

  View details for PubMedCentralID PMC1852584

• Non-viral gene therapy for Duchenne muscular dystrophy: Progress and challenges BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR BASIS OF DISEASE Rando, T. A. 2007; 1772 (2): 263-271

  Abstract

  Duchenne muscular dystrophy (DMD) is one of the most common lethal, hereditary diseases of childhood. Since the identification of the genetic basis of this disorder, there has been the hope that a cure would be developed in the form of gene therapy. This has yet to be realized, but many different gene therapy approaches have seen dramatic advances in recent years. Although viral-mediated gene therapy has been at the forefront of the field, several non-viral gene therapy approaches have been applied to animal and cellular models of DMD. These include plasmid-mediated gene delivery, antisense-mediated exon skipping, and oligonucleotide-mediated gene editing. In the past several years, non-viral gene therapy has moved from the laboratory to the clinic. Advances in vector design, formulation, and delivery are likely to lead to even more rapid advances in the coming decade. Given the relative simplicity, safety, and cost-effectiveness of these methodologies, non-viral gene therapy continues to have great promise for future gene therapy approaches to the treatment of DMD.

  View details for DOI 10.1016/j.bbadis.2006.07.009

  View details for Web of Science ID 000244193800015

  View details for PubMedID 17005381

• Intrinsic changes and extrinsic influences of myogenic stem cell function during aging STEM CELL REVIEWS Brack, A. S., Rando, T. A. 2007; 3 (3): 226-237

  Abstract

  The myogenic stem cell (satellite cell) is almost solely responsible for the remarkable regeneration of adult skeletal muscle fibers after injury. The availability and the functionality of satellite cells are the determinants of efficient muscle regeneration. During aging, the efficiency of muscle regeneration declines, suggesting that the functionality of satellite cells and their progeny may be altered. Satellite cells do not sit in isolation but rather are surrounded by, and influenced by, many extrinsic factors within the muscle tissue that can alter their functionality. These factors likely change during aging and impart both reversible and irreversible changes to the satellite cells and on their proliferating progeny. In this review, we discuss the possible mechanisms of impaired muscle regeneration with respect to the biology of satellite cells. Future studies that enhance our understanding of the interactions between stem cells and the environment in which they reside will offer promise for therapeutic applications in age-related diseases.

  View details for DOI 10.1007/s12015-007-9000-2

  View details for Web of Science ID 000249929800006

  View details for PubMedID 17917136

• Muscular Dystrophies NEUROBIOLOGY OF DISEASE Lim, L. E., Thornton, C. A., Rando, T. A., Gilman, S. 2007: 925–34

  View details for Web of Science ID 000310853000085

• Long-term increase in mVEGF164 in mouse hindlimb muscle mediated by phage phi C31 integrase after nonviral DNA delivery HUMAN GENE THERAPY Portlock, J. L., Keravala, A., Bertoni, C., Lee, S., Rando, T. A., Calos, M. P. 2006; 17 (8): 871-876

  Abstract

  Peripheral vascular disease (PVD), characterized by insufficient blood supply to extremities, can be a devastating illness. Although many gene therapy strategies for PVD using vascular endothelial growth factor (VEGF) have resulted in increased blood vessel formation, the vessels are often impermanent and regress after therapy, probably because of the short-lived VEGF expression mediated by gene therapy vectors (14 days or less). phiC31 integrase is a recombinase originally isolated from a bacteriophage of Streptomyces. This integrase performs efficient chromosomal integration of plasmid DNA into mammalian genomes that results in long-term transgene expression. In this study, gene transfer was achieved by intramuscular injection of VEGF and integrase plasmid DNAs into the tibialis anterior muscle in the mouse hindlimb, followed by electroporation of the muscle with needle electrodes. We observed VEGF levels significantly above background 40 days after injection in animals that received phiC31 integrase and the VEGF plasmid. Site-specific integration of plasmid DNA in the chromosomes of muscle tissue was verified by polymerase chain reaction at a common integration site. These results suggest the possible utility of the phiC31 integrase system to treat ischemic disease.

  View details for PubMedID 16942446

• Prognostic value of telomere length: The long and short of it ANNALS OF NEUROLOGY Rando, T. A. 2006; 60 (2): 155-157

  View details for Web of Science ID 000239536600002

  View details for PubMedID 16862590

• Stem cells, ageing and the quest for immortality NATURE Rando, T. A. 2006; 441 (7097): 1080-1086

  Abstract

  Adult stem cells reside in most mammalian tissues, but the extent to which they contribute to normal homeostasis and repair varies widely. There is an overall decline in tissue regenerative potential with age, and the question arises as to whether this is due to the intrinsic ageing of stem cells or, rather, to the impairment of stem-cell function in the aged tissue environment. Unravelling these distinct contributions to the aged phenotype will be critical to the success of any therapeutic application of stem cells in the emerging field of regenerative medicine with respect to tissue injury, degenerative diseases or normal functional declines that accompany ageing.

  View details for DOI 10.1038/nature04958

  View details for Web of Science ID 000238615500035

  View details for PubMedID 16810243

• Enhanced Level of Gene Correction Mediated by Oligonucleotides Containing CpG Modification in the mdx Mouse Model for Duchenne Muscular Dystrophy Bertoni, C., Rustagi, A., Rando, T. A. NATURE PUBLISHING GROUP. 2006: S221

  View details for DOI 10.1016/j.ymthe.2006.08.646

  View details for Web of Science ID 000208933501497

• Phi C31 Integrase System Enhances Dystrophin Gene Expression in Skeletal Muscle of Mouse Models for Duchenne Muscular Dystrophy Bertoni, C., Jarrahian, S., Wheeler, T. M., Li, Y., Olivares, E. C., Calos, M. P., Rando, T. A. NATURE PUBLISHING GROUP. 2006: S14–S15

  View details for DOI 10.1016/j.ymthe.2006.08.047

  View details for Web of Science ID 000208933500035

• The regulation of Notch signaling controls satellite cell activation and cell fate determination in postnatal myogenesis (vol 3, pg 397, 2002) DEVELOPMENTAL CELL Conboy, I. M., Rando, T. A. 2006; 10 (2): 273

  View details for DOI 10.1016/j.devcel.2006.01.003

  View details for Web of Science ID 000235480500018

• Enhancement of plasmid-mediated gene therapy for muscular dystrophy by directed plasmid integration PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Bertoni, C., Jarrahian, S., Wheeler, T. M., Li, Y. N., Olivares, E. C., Calos, M. P., Rando, T. A. 2006; 103 (2): 419-424

  Abstract

  Plasmid-mediated gene therapy can restore dystrophin expression in skeletal muscle in the mdx mouse, a model of Duchenne muscular dystrophy. However, sufficient long-term expression and distribution of dystrophin remain a hurdle for translating this technology into a viable treatment for Duchenne muscular dystrophy. To improve plasmid-mediated gene therapy for muscle diseases, we studied the effects of targeted plasmid integration using a phage integrase (phiC31) that can mediate the integration of suitably modified plasmids into the mammalian genome. Using a luciferase expression plasmid, we monitored plasmid gene expression noninvasively in living mice by bioluminescence imaging. Coinjection of an integrase plasmid resulted in 5- to 10-fold higher levels of sustained luciferase expression. Likewise, plasmid-mediated dystrophin expression in mdx muscle was enhanced by integration. Using a combination of dystrophin and luciferase plasmids, we analyzed the functional benefit of dystrophin expression in the dystrophic muscle. The expression of dystrophin slowed the loss of luciferase expression associated with muscle degeneration, and that protection was enhanced by targeted integration of the dystrophin plasmid. In the presence of integrase, dystrophin expression was distributed along a much greater length of individual fibers, and this was associated with increased protection against degenerative changes. These data demonstrate the importance of both the level and distribution of dystrophin expression to achieve therapeutic efficacy, and that the efficacy can be enhanced by targeted plasmid integration.

  View details for Web of Science ID 000234624100031

  View details for PubMedID 16387861

• Stem cells in postnatal myogenesis: molecular mechanisms of satellite cell quiescence, activation and replenishment TRENDS IN CELL BIOLOGY Dhawan, J., Rando, T. A. 2005; 15 (12): 666-673

  Abstract

  Satellite cells are the primary stem cells in adult skeletal muscle, and are responsible for postnatal muscle growth, hypertrophy and regeneration. In mature muscle, most satellite cells are in a quiescent state, but they activate and begin proliferating in response to extrinsic signals. Following activation, a subset of satellite cell progeny returns to the quiescent state during the process of self-renewal. Here, we review recent studies of satellite cell biology and focus on the key transitions from the quiescent state to the state of proliferative activation and myogenic lineage progression and back to the quiescent state. The molecular mechanisms of these transitions are considered in the context of the biology of the satellite cell niche, changes with age, and interactions with established pathways of myogenic commitment and differentiation.

  View details for DOI 10.1016/j.tcb.2005.10.007

  View details for Web of Science ID 000234316700008

  View details for PubMedID 16243526

• The regulation of Notch signaling in muscle stem cell activation and postnatal myogenesis SEMINARS IN CELL & DEVELOPMENTAL BIOLOGY Luo, D., Renault, V. M., Rando, T. A. 2005; 16 (4-5): 612-622

  Abstract

  The Notch signaling pathway is an evolutionarily conserved pathway that is critical for tissue morphogenesis during development, but is also involved in tissue maintenance and repair in the adult. In skeletal muscle, regulation of Notch signaling is involved in somitogenesis, muscle development, and the proliferation and cell fate determination of muscle stems cells during regeneration. During each of these processes, the spatial and temporal control of Notch signaling is essential for proper tissue formation. That control is mediated by a series of regulatory proteins and protein complexes that enhance or inhibit Notch signaling by regulating protein processing, localization, activity, and stability. In this review, we focus on the regulation of Notch signaling during postnatal muscle regeneration when muscle stem cells ("satellite cells") must activate, proliferate, progress along a myogenic lineage pathway, and ultimately differentiate to form new muscle. We review the regulators of Notch signaling, such as Numb and Deltex, that have documented roles in myogenesis as well as other regulators that may play a role in modulating Notch signaling during satellite cell activation and postnatal myogenesis.

  View details for DOI 10.1016/j.semcdb.2005.07.002

  View details for PubMedID 16087370

• The adult muscle stem cell comes of age NATURE MEDICINE Rando, T. A. 2005; 11 (8): 829-831

  View details for DOI 10.1038/nm0805-829

  View details for Web of Science ID 000230964700019

  View details for PubMedID 16079875

• NF90 regulates cell cycle exit and terminal myogenic differentiation by direct binding to the 3 '-untranslated region of MyoD and p21(WAF1/CIP1) mRNAs JOURNAL OF BIOLOGICAL CHEMISTRY Shi, L. F., Zhao, G. H., Qiu, D. M., Godfrey, W. R., Vogel, H., Rando, T. A., Hu, H., Kao, P. N. 2005; 280 (19): 18981-18989

  Abstract

  NF90 and splice variant NF110/ILF3/NFAR are double-stranded RNA-binding proteins that regulate gene expression. Mice with targeted disruption of NF90 were engineered. NF90(-/-) mice were born small and weak and succumbed to perinatal death within 12 h because of neuromuscular respiratory failure. Lung inflation and morphology were normal in NF90(-/-) mice. The diaphragm and other skeletal muscles in NF90(-/-) mice demonstrated disorganized arrangement and paucity of myofibers, evidence of myocyte degeneration and increased apoptosis. The expression of myogenic regulators, MyoD, myogenin, and p21WAF1/CIP1, was severely decreased in NF90(-/-) mice. These myogenic transcription factors and cell cycle inhibitors are regulated in part through post-transcriptional mRNA stabilization. Northwestern blotting revealed that NF90 is the principal and specific p21WAF1/CIP1 and MyoD 3'-untranslated region RNA-binding protein in developing skeletal muscles. NF90 regulates transcription factors and a cell cycle inhibitor essential for skeletal muscle differentiation and for survival.

  View details for DOI 10.1074/jbc.M411034200

  View details for PubMedID 15746098

• Aging, stem cells and tissue regeneration - Lessons from muscle CELL CYCLE Conboy, I. M., Rando, T. A. 2005; 4 (3): 407-410

  Abstract

  With age, there is a gradual decline in the regenerative properties of most tissues due to a combination of age-dependent changes in tissue-specific stem cells and in the environmental cues that promote those cells to participate in tissue maintenance and repair. In adult skeletal muscle, where the resident dedicated stem cells ("satellite cells") are capable of rapid and highly effective regeneration in response to injury, there is just such a loss of regenerative potential with age. Satellite cell activation and cell fate determination are controlled by the Notch signaling pathway that is initiated by the rapid increase in expression of the Notch ligand, Delta, following injury. In old muscle, this upregulation of Delta is blunted and thus satellite cell activation is markedly diminished. However, by indirectly inducing Notch activity, the regenerative potential of aged satellite cells can be restored. Furthermore, exposure of aged satellite cells to serum from young mice, either in vivo by heterochronic parabiotic pairings or in vitro, rejuvenates the satellite cell response. This restorative potential suggests that tissue-specific stem cells do not lose their ability to participate in tissue maintenance and repair. Therefore, it may be that even very old stem cells may be capable of maintaining and repairing aged tissues if provided with optimal environmental cues.

  View details for Web of Science ID 000229538500016

  View details for PubMedID 15725724

• Rejuvenation of aged progenitor cells by exposure to a young systemic environment NATURE Conboy, I. M., Conboy, M. J., Wagers, A. J., Girma, E. R., Weissman, I. L., Rando, T. A. 2005; 433 (7027): 760-764

  Abstract

  The decline of tissue regenerative potential is a hallmark of ageing and may be due to age-related changes in tissue-specific stem cells. A decline in skeletal muscle stem cell (satellite cell) activity due to a loss of Notch signalling results in impaired regeneration of aged muscle. The decline in hepatic progenitor cell proliferation owing to the formation of a complex involving cEBP-alpha and the chromatin remodelling factor brahma (Brm) inhibits the regenerative capacity of aged liver. To examine the influence of systemic factors on aged progenitor cells from these tissues, we established parabiotic pairings (that is, a shared circulatory system) between young and old mice (heterochronic parabioses), exposing old mice to factors present in young serum. Notably, heterochronic parabiosis restored the activation of Notch signalling as well as the proliferation and regenerative capacity of aged satellite cells. The exposure of satellite cells from old mice to young serum enhanced the expression of the Notch ligand (Delta), increased Notch activation, and enhanced proliferation in vitro. Furthermore, heterochronic parabiosis increased aged hepatocyte proliferation and restored the cEBP-alpha complex to levels seen in young animals. These results suggest that the age-related decline of progenitor cell activity can be modulated by systemic factors that change with age.

  View details for DOI 10.1038/nature03260

  View details for Web of Science ID 000227039200043

  View details for PubMedID 15716955

• Strand bias in oligonucleotide-mediated dystrophin gene editing HUMAN MOLECULAR GENETICS Bertoni, C., Morris, G. E., Rando, T. A. 2005; 14 (2): 221-233

  Abstract

  Defects in the dystrophin gene cause the severe degenerative muscle disorder, Duchenne muscular dystrophy (DMD). Among the gene therapy approaches to DMD under investigation, a gene editing approach using oligonucleotide vectors has yielded encouraging results. Here, we extend our studies of gene editing with self-pairing, chimeric RNA/DNA oligonucleotides (RDOs) to the use of oligodeoxynucleotides (ODNs) to correct point mutations in the dystrophin gene. The ODN vectors offer many advantages over the RDO vectors, and we compare the targeting efficiencies in the mdx(5cv) mouse model of DMD. We found that ODNs targeted to either the transcribed or the non-transcribed strand of the dystrophin gene were capable of inducing gene repair, with efficiencies comparable to that seen with RDO vectors. Oligonucleotide-mediated repair was demonstrated at the genomic, mRNA and protein levels in muscle cells both in vitro and in vivo, and the correction was stable over time. Interestingly, there was a strand bias observed with the ODNs, with more efficient correction of the non-transcribed strand even though the dystrophin gene is not transcribed in proliferating myoblasts. This finding demonstrates that strand bias of ODN-mediated gene repair is likely to be due to the specific sequence of the target gene in addition to any effects of transcription. A better understanding of how the efficiency of gene editing relates to the target sequence will offer the opportunity for rational oligonucleotide design for further development of this elegant approach to gene therapy for DMD and other genetic diseases.

  View details for DOI 10.1093/hmg/ddi020

  View details for Web of Science ID 000226199400004

  View details for PubMedID 15563511

• Isolation of adult mouse myogenic progenitors: Functional heterogeneity of cells within and engrafting skeletal muscle CELL Sherwood, R. I., Christensen, J. L., Conboy, I. M., Conboy, M. J., Rando, T. A., Weissman, I. L., Wagers, A. J. 2004; 119 (4): 543-554

  Abstract

  Skeletal muscle regeneration in adults is thought to occur through the action of myogenic satellite cells located in close association with mature muscle fibers; however, these precursor cells have not been prospectively isolated, and recent studies have suggested that additional muscle progenitors, including cells of bone marrow or hematopoietic origin, may exist. To clarify the origin(s) of adult myogenic cells, we used phenotypic, morphological, and functional criteria to identify and prospectively isolate a subset of myofiber-associated cells capable at the single cell level of generating myogenic colonies at high frequency. Importantly, although muscle-engrafted cells from marrow and/or circulation localized to the same anatomic compartment as myogenic satellite cells and expressed some though not all satellite cell markers, they displayed no intrinsic myogenicity. Together, these studies describe the clonal isolation of functional adult myogenic progenitors and demonstrate that these cells do not arise from hematopoietic or other bone marrow or circulating precursors.

  View details for Web of Science ID 000225183200012

  View details for PubMedID 15537543

• Understanding and reversing stem cell aging 44th Annual Meeting of the American-Society-for-Cell-Biology Conboy, I. M., Conboy, M. J., Wagers, A. J., Weissman, I. L., Rando, T. A. AMER SOC CELL BIOLOGY. 2004: 361A–361A

  View details for Web of Science ID 000224648803066

• Focal adhesion kinase (FAK) localization at focal adhesions is essential for myoblast fusion and sarcomere assembly Quach, N., Disatnik, M., Rando, T. A. AMER SOC CELL BIOLOGY. 2004: 33A

  View details for Web of Science ID 000224648800179

• Artificial sweeteners - Enhancing glycosylation to treat muscular dystrophies NEW ENGLAND JOURNAL OF MEDICINE Rando, T. A. 2004; 351 (12): 1254-1256

  View details for Web of Science ID 000223861300017

  View details for PubMedID 15371584

• The bi-directional translocation of MARCKS between membrane and cytosol regulates integrin-mediated muscle cell spreading JOURNAL OF CELL SCIENCE Disatnik, M. H., Boutet, S. C., Pacio, W., Chan, A. Y., Ross, L. B., Lee, C. H., Rando, T. A. 2004; 117 (19): 4469-4479

  Abstract

  The regulation of the cytoskeleton is critical to normal cell function during tissue morphogenesis. Cell-matrix interactions mediated by integrins regulate cytoskeletal dynamics, but the signaling cascades that control these processes remain largely unknown. Here we show that myristoylated alanine-rich C-kinase substrate (MARCKS) a specific substrate of protein kinase C (PKC), is regulated by alpha5beta1 integrin-mediated activation of PKC and is critical to the regulation of actin stress fiber formation during muscle cell spreading. Using MARCKS mutants that are defective in membrane association or responsiveness to PKC-dependent phosphorylation, we demonstrate that the translocation of MARCKS from the membrane to the cytosol in a PKC-dependent manner permits the initial phases of cell adhesion. The dephosphorylation of MARCKS and its translocation back to the membrane permits the later stages of cell spreading during the polymerization and cross-linking of actin and the maturation of the cytoskeleton. All of these processes are directly dependent on the binding of alpha5beta1 integrin to its extracellular matrix receptor, fibronectin. These results demonstrate a direct biochemical pathway linking alpha5beta1 integrin signaling to cytoskeletal dynamics and involving bi-directional translocation of MARCKS during the dramatic changes in cellular morphology that occur during cell migration and tissue morphogenesis.

  View details for DOI 10.1242/jcs.01309

  View details for PubMedID 15316066

• Site-specific integration enhances expression of DNA introduced into skeletal muscle 7th Annual Meeting of the American-Society-of-Gene-Therapy Wheeler, T. M., Jarrahian, S., Bertoni, C., Olivares, E. C., Doyle, T., Contag, C. H., Calos, M. P., Rando, T. A. NATURE PUBLISHING GROUP. 2004: S307–S307

  View details for Web of Science ID 000222316600811

• A caveolin-3 mutant that causes limb girdle muscular dystrophy type 1C disrupts Src localization and activity and induces apoptosis in skeletal myotubes JOURNAL OF CELL SCIENCE Smythe, G. M., Eby, J. C., Disatnik, M. H., Rando, T. A. 2003; 116 (23): 4739-4749

  Abstract

  Caveolins are membrane proteins that are the major coat proteins of caveolae, specialized lipid rafts in the plasma membrane that serve as scaffolding sites for many signaling complexes. Among the many signaling molecules associated with caveolins are the Src tyrosine kinases, whose activation regulates numerous cellular functions including the balance between cell survival and cell death. Several mutations in the muscle-specific caveolin, caveolin-3, lead to a form of autosomal dominant muscular dystrophy referred to as limb girdle muscular dystrophy type 1C (LGMD-1C). One of these mutations (here termed the 'TFT mutation') results in a deletion of a tripeptide (DeltaTFT(63-65)) that affects the scaffolding and oligomerization domains of caveolin-3. This mutation causes a 90-95% loss of caveolin-3 protein levels and reduced formation of caveolae in skeletal muscle fibers. However, the effects of this mutation on the specific biochemical processes and cellular functions associated with caveolae have not been elucidated. We demonstrate that the TFT caveolin-3 mutation in post-mitotic skeletal myotubes causes severely reduced localization of caveolin-3 to the plasma membrane and to lipid rafts, and significantly inhibits caveolar function. The TFT mutation reduced the binding of Src to caveolin-3, diminished targeting of Src to lipid rafts, and caused abnormal perinuclear accumulation of Src. Along with these alterations of Src localization and targeting, there was elevated Src activation in myotubes expressing the TFT mutation and an increased incidence of apoptosis in those cells compared with control myotubes. The results of this study demonstrate that caveolin-3 mutations associated with LGMD-1C disrupt normal cellular signal transduction pathways associated with caveolae and cause apoptosis in muscle cells, all of which may reflect pathogenetic pathways that lead to muscle degeneration in these disorders.

  View details for DOI 10.1242/jcs.00806

  View details for Web of Science ID 000187395600006

  View details for PubMedID 14600260

• Notch-mediated restoration of regenerative potential to aged muscle SCIENCE Conboy, I. M., Conboy, M. J., Smythe, G. M., Rando, T. A. 2003; 302 (5650): 1575-1577

  Abstract

  A hallmark of aging is diminished regenerative potential of tissues, but the mechanism of this decline is unknown. Analysis of injured muscle revealed that, with age, resident precursor cells (satellite cells) had a markedly impaired propensity to proliferate and to produce myoblasts necessary for muscle regeneration. This was due to insufficient up-regulation of the Notch ligand Delta and, thus, diminished activation of Notch in aged, regenerating muscle. Inhibition of Notch impaired regeneration of young muscle, whereas forced activation of Notch restored regenerative potential to old muscle. Thus, Notch signaling is a key determinant of muscle regenerative potential that declines with age.

  View details for Web of Science ID 000186802200047

  View details for PubMedID 14645852

• Remarkable preservation of undigested muscle-tissue within a late Cretaceous tyrannosaurid coprolite from Alberta, Canada PALAIOS Chin, K., Eberth, D. A., Schweitzer, M. H., Rando, T. A., Sloboda, W. J., Horner, J. R. 2003; 18 (3): 286-294

  Abstract

  Exceptionally detailed soft tissues have been identified within the fossilized feces of a large Cretaceous tyrannosaurid. Microscopic cord-like structures in the coprolitic ground mass are visible in thin section and with scanning electron microscopy. The morphology, organization, and context of these structures indicate that they are the fossilized remains of undigested muscle tissue. This unusual discovery indicates specific digestive and taphonomic conditions, including a relatively short gut-residence time, rapid lithification, and minimal diagenetic recrystallization. Rapid burial of the feces probably was facilitated by a flood event on the ancient coastal lowland plain on which the fecal mass was deposited.

  View details for Web of Science ID 000183849200009

  View details for PubMedID 12866547

• Restoration of dystrophin expression in mdx muscle cells by chimeraplast-mediated exon skipping HUMAN MOLECULAR GENETICS Bertoni, C., Lau, C., Rando, T. A. 2003; 12 (10): 1087-1099

  Abstract

  The most common types of dystrophin gene mutations that cause Duchenne muscular dystrophy (DMD) are large deletions that result in a shift of the translational reading frame. Such mutations generally lead to a complete absence of dystrophin protein in the muscle cells of affected individuals. Any therapeutic modality that could restore the reading frame would have the potential to substantially reduce the severity of the disease by allowing the production of an internally deleted, but partially functional, dystrophin protein as occurs in Becker muscular dystrophy (BMD). One approach to restoring the reading frame would be to alter the splicing of the pre-mRNA to produce an in-frame transcript. We have tested the ability of chimeric RNA/DNA oligonucleotides (chimeraplasts) to alter key bases in specific splice sequences in the dystrophin gene to induce exon skipping. Using cells from the mdx mouse as a model system, we show that chimeraplast-mediated base conversion in the intron 22/exon 23 splice junction induces alternative splicing and the production of in-frame transcripts. Interestingly, multiple alternative transcripts were induced by this targeted splice site mutation. Direct sequencing indicated that several of these were predicted to produce in-frame dystrophin transcripts with internal deletions. Indeed, multiple forms of dystrophin protein were observed by western blot analysis, and the functionality of the products was demonstrated by the restoration of expression and localization of a dystrophin-associated protein, alpha-dystroglycan, in differentiated cells. These data demonstrate that chimeraplasts can induce exon skipping by altering splice site sequences at the genomic level. As such, chimeraplast-mediated exon skipping has the potential to be used to transform a severe DMD phenotype into a much milder BMD phenotype.

  View details for DOI 10.1093/hmg/ddg133

  View details for Web of Science ID 000182950200002

  View details for PubMedID 12719373

• The regulation of catalase gene expression in mouse muscle cells is dependent on the CCAAT-binding factor NF-Y BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Luo, D., Rando, T. A. 2003; 303 (2): 609-618

  Abstract

  Catalase is an antioxidant enzyme whose expression is transcriptionally regulated and tissue-specific. The level of expression determines, in part, the susceptibility of a cell to oxidative stress. Skeletal muscle is a tissue that experiences high levels of oxidative stress during normal metabolic activity, so the expression of antioxidant enzymes is critical to preventing cellular damage. To study the transcriptional regulation of the catalase gene in mouse muscle cells, the 5'-flanking region of the mouse catalase gene was isolated from genomic DNA. The transcriptional activity of the 5'-flanking region was investigated in transiently transfected murine myoblasts using a promoter-less luciferase reporter vector and site-directed mutagenesis. Strikingly, we found that nearly all of the transcriptional activity was restricted to the final 191 bp of the greater than 2.5 kb of the 5'-flanking region examined. Of the potential consensus binding sites for transcriptional regulators within this 191-bp region, we identified two CCAAT boxes and no other consensus sites that were important for the transcriptional activity of this promoter. Gel shift and super shift assays indicated that the transcription factor NF-Y bound to both CCAAT boxes. Furthermore, co-transfection of reporter constructs with NF-Y expression vectors into Drosophila SL2 cells demonstrated NF-Y-mediated transcriptional activation of the catalase gene. Interestingly, there were no nearby sites that appeared to interact with either NF-Y binding sites, and thus it appears that NF-Y acts as a bona fide transcription factor for catalase gene expression in mouse muscle cells. These data provide the first examination of the regulation of the mouse catalase gene and indicate unique aspects of its regulation that may pertain to the tissue-specific patterns of expression.

  View details for DOI 10.1016/S0006-291X(03)00397-8

  View details for Web of Science ID 000182177800035

  View details for PubMedID 12659863

• Oxidative stress and the pathogenesis of muscular dystrophies Conference on Role of Physical Activity and Exercise Training in Neuromuscular Diseases Rando, T. A. LIPPINCOTT WILLIAMS & WILKINS. 2002: S175–S186

  Abstract

  The muscular dystrophies represent a diverse group of diseases differing in underlying genetic basis, age of onset, mode of inheritance, and severity of progression, but they share certain common pathologic features. Most prominent among these features is the necrotic degeneration of muscle fibers. Although the genetic basis of many of the dystrophies has been known for over a decade and new disease genes continue to be discovered, the pathogenetic mechanisms leading to muscle cell death in the dystrophies remain a mystery. This review focuses on the oxidative stress theory, which states that the final common pathway of muscle cell death in these diseases involves oxidative damage.

  View details for DOI 10.1097/01.PHM.0000029774.56528.A6

  View details for Web of Science ID 000178883700018

  View details for PubMedID 12409822

• The regulation of notch signaling controls satellite cell activation and cell fate determination in postnatal myogenesis DEVELOPMENTAL CELL Conboy, I. M., Rando, T. A. 2002; 3 (3): 397-409

  Abstract

  We have studied the role of Notch-1 and its antagonist Numb in the activation of satellite cells during postnatal myogenesis. Activation of Notch-1 promoted the proliferation of myogenic precursor cells expressing the premyoblast marker Pax3. Attenuation of Notch signaling by increases in Numb expression led to the commitment of progenitor cells to the myoblast cell fate and the expression of myogenic regulatory factors, desmin, and Pax7. In many intermediate progenitor cells, Numb was localized asymmetrically in actively dividing cells, suggesting an asymmetric cell division and divergent cell fates of daughter cells. The results indicate that satellite cell activation results in a heterogeneous population of precursor cells with respect to Notch-1 activity and that the balance between Notch-1 and Numb controls cellular homeostasis and cell fate determination.

  View details for Web of Science ID 000178132800013

  View details for PubMedID 12361602

• Activation of an adipogenic program in adult myoblasts with age MECHANISMS OF AGEING AND DEVELOPMENT Taylor-Jones, J. M., McGehee, R. E., Rando, T. A., Lecka-Czernik, B., Lipschitz, D. A., Peterson, C. A. 2002; 123 (6): 649-661

  Abstract

  Myoblasts isolated from mouse hindlimb skeletal muscle demonstrated increased adipogenic potential as a function of age. Whereas myoblasts from 8-month-old adult mice did not significantly accumulate terminal markers of adipogenesis regardless of culture conditions, myoblasts from 23-month-old mice accumulated fat and expressed genes characteristic of differentiated adipocytes, such as the fatty acid binding protein aP2. This change in differentiation potential was associated with a change in the abundance of the mRNA encoding the transcription factor C/EBPalpha, and in the relative abundance of PPARgamma2 to PPARgamma1 mRNAs. Furthermore, PPARgamma activity appeared to be regulated at the level of phosphorylation, being more highly phosphorylated in myoblasts isolated from younger animals. Although adipogenic gene expression in myoblasts from aged animals was activated, presumably in response to PPARgamma and C/EBPalpha, unexpectedly, myogenic gene expression was not effectively repressed. The Wnt signaling pathway may also alter differentiation potential in muscle with age. Wnt-10b mRNA was more abundantly expressed in muscle tissue and cultured myoblasts from adult compared with aged mice, resulting in stabilization of cytosolic beta-catenin, that may potentially contribute to inhibition of adipogenic gene expression in adult myoblasts. The changes reported here, together with those reported in bone marrow stroma with age, suggest that a default program may be activated in mesenchymal cells with increasing age resulting in a more adipogenic-like phenotype. Whether this change in differentiation potential contributes to the increased adiposity in muscle with age remains to be determined.

  View details for Web of Science ID 000177238100010

  View details for PubMedID 11850028

• Myonuclear apoptosis and skeletal muscle atrophy Xing, J., Yu, Y., Rando, T. A. AMER SOC CELL BIOLOGY. 2001: 516A–516A

  View details for Web of Science ID 000172372502838

• Copper/zinc superoxide dismutase: More is not necessarily better! ANNALS OF NEUROLOGY Rando, T. A., Epstein, C. J. 1999; 46 (1): 135-136

  View details for Web of Science ID 000081200900023

  View details for PubMedID 10401795

• Evidence of oxidative stress in mdx mouse muscle: Studies of the pre-necrotic state JOURNAL OF THE NEUROLOGICAL SCIENCES Disatnik, M. H., Dhawan, J., Yu, Y., Beal, M. F., Whirl, M. M., Franco, A. A., Rando, T. A. 1998; 161 (1): 77-84

  Abstract

  Considerable evidence indicates that free radical injury may underlie the pathologic changes in muscular dystrophies from mammalian and avian species. We have investigated the role of oxidative injury in muscle necrosis in mice with a muscular dystrophy due to a defect in the dystrophin gene (the mdx strain). In order to avoid secondary consequences of muscle necrosis, all experiments were done on muscle prior to the onset of the degenerative process (i.e. during the 'pre-necrotic' phase) which lasted up to 20 days of age in the muscles examined. In pre-necrotic mdx muscle, there was an induction of expression of genes encoding antioxidant enzymes, indicative of a cellular response to oxidative stress. In addition, the levels of lipid peroxidation were greater in mdx muscle than in the control. Since the free radical nitric oxide (NO*) has been shown to mediate oxidative injury in various disease states, and because dystrophin has been shown to form a complex with the enzyme nitric oxide synthase, we examined pre-necrotic mdx muscle for evidence of NO*-mediated injury by measuring cellular nitrotyrosine formation. By both immunohistochemical and electrochemical analyses, no evidence of increased nitrotyrosine levels in mdx muscle was detected. Therefore, although no relationship with NO*-mediated toxicity was found, we found evidence of increased oxidative stress preceding the onset of muscle cell death in dystrophin-deficient mice. These results lend support to the hypothesis that free radical-mediated injury may contribute to the pathogenesis of muscular dystrophies.

  View details for Web of Science ID 000077605200013

  View details for PubMedID 9879685

• Dystrophic muscle in mice chimeric for expression of alpha 5 integrin JOURNAL OF CELL BIOLOGY Taverna, D., Disatnik, M. H., Rayburn, H., Bronson, R. T., Yang, J., Rando, T. A., Hynes, R. O. 1998; 143 (3): 849-859

  Abstract

  alpha5-deficient mice die early in embryogenesis (). To study the functions of alpha5 integrin later in mouse embryogenesis and during adult life we generated alpha5 -/-;+/+ chimeric mice. These animals contain alpha5-negative and positive cells randomly distributed. Analysis of the chimerism by glucose- 6-phosphate isomerase (GPI) assay revealed that alpha5 -/- cells contributed to all the tissues analyzed. High contributions were observed in the skeletal muscle. The perinatal survival of the mutant chimeras was lower than for the controls, however the subsequent life span of the survivors was only slightly reduced compared with controls (). Histological analysis of alpha5 -/-;+/+ mice from late embryogenesis to adult life revealed an alteration in the skeletal muscle structure resembling a typical muscle dystrophy. Giant fibers, increased numbers of nuclei per fiber with altered position and size, vacuoli and signs of muscle degeneration-regeneration were observed in head, thorax and limb muscles. Electron microscopy showed an increase in the number of mitochondria in some muscle fibers of the mutant mice. Increased apoptosis and immunoreactivity for tenascin-C were observed in mutant muscle fibers. All the alterations were already visible at late stages of embryogenesis. The number of altered muscle fibers varied in different animals and muscles and was often increased in high percentage chimeric animals. Differentiation of alpha5 -/- ES cells or myoblasts showed that in vitro differentiation into myotubes was achieved normally. However proper adhesion and survival of myoblasts on fibronectin was impaired. Our data suggest that a novel form of muscle dystrophy in mice is alpha5-integrin-dependent.

  View details for Web of Science ID 000076894300023

  View details for PubMedID 9813102

  View details for PubMedCentralID PMC2148145

• Overexpression of copper/zinc superoxide dismutase: A novel cause of murine muscular dystrophy ANNALS OF NEUROLOGY Rando, T. A., Crowley, R. S., Carlson, E. J., Epstein, C. J., Mohapatra, P. K. 1998; 44 (3): 381-386

  Abstract

  Oxidative injury underlies the cellular injury and cell death in a variety of disease states. In muscular dystrophies, evidence from in vivo and in vitro studies suggests that muscle degeneration may be secondary to an increased susceptibility to oxidative stress. To address the role of free radical metabolism in the pathogenetic process of muscular dystrophies, we examined the muscle of transgenic mice that overexpress copper/zinc (Cu/Zn) superoxide dismutase. Overexpression of this enzyme can sensitize cells to oxidative injury, and Cu/Zn superoxide dismutase activity was elevated approximately fourfold above control levels in skeletal muscle of the transgenic strain. Examination of serum creatine phosphokinase levels in these mice revealed significant elevations after 2 months of age, indicative of active muscle breakdown. By 8 months of age, there was gross atrophy of the quadriceps muscle, and other hindlimb muscles were variably affected. Histologically, there was evidence of widespread muscle necrosis and regeneration, fiber splitting, and replacement of muscle with adipose and fibrous connective tissue, typical of a muscular dystrophy. Associated with the development of this degeneration was an increase in the levels of lipid peroxidation in the muscle of Cu/Zn superoxide dismutase transgenic mice, highlighting the central role of oxidative injury in this pathogenetic process. These results demonstrate that oxidative damage can be the primary pathogenetic process underlying a muscular dystrophy.

  View details for Web of Science ID 000075744700014

  View details for PubMedID 9749606

• Heterogeneity among muscle precursor cells in adult skeletal muscles with differing regenerative capacities DEVELOPMENTAL DYNAMICS PAVLATH, G. K., Thaloor, D., Rando, T. A., Cheong, M., English, A. W., Zheng, B. 1998; 212 (4): 495-508

  Abstract

  Skeletal muscle has a remarkable capacity to regenerate after injury, although studies of muscle regeneration have heretofore been limited almost exclusively to limb musculature. Muscle precursor cells in skeletal muscle are responsible for the repair of damaged muscle. Heterogeneity exists in the growth and differentiation properties of muscle precursor cell (myoblast) populations throughout limb development but whether the muscle precursor cells differ among adult skeletal muscles is unknown. Such heterogeneity among myoblasts in the adult may give rise to skeletal muscles with different regenerative capacities. Here we compare the regenerative response of a masticatory muscle, the masseter, to that of limb muscles. After exogenous trauma (freeze or crush injuries), masseter muscle regenerated much less effectively than limb muscle. In limb muscle, normal architecture was restored 12 days after injury, whereas in masseter muscle, minimal regeneration occurred during the same time period. Indeed, at late time points, masseter muscles exhibited increased fibrous connective tissue in the region of damage, evidence of ineffective muscle regeneration. Similarly, in response to endogenous muscle injury due to a muscular dystrophy, widespread evidence of impaired regeneration was present in masseter muscle but not in limb muscle. To explore the cellular basis of these different regenerative capacities, we analyzed the myoblast populations of limb and masseter muscles both in vivo and in vitro. From in vivo analyses, the number of myoblasts in regenerating muscle was less in masseter compared with limb muscle. Assessment of population growth in vitro indicated that masseter myoblasts grow more slowly than limb myoblasts under identical conditions. We conclude that the impaired regeneration in masseter muscles is due to differences in the intrinsic myoblast populations compared to limb muscles.

  View details for Web of Science ID 000075132500003

  View details for PubMedID 9707323

• Two regions of the ryanodine receptor involved in coupling with L-type Ca2+ channels JOURNAL OF BIOLOGICAL CHEMISTRY Nakai, J., Sekiguchi, N., Rando, T. A., Allen, P. D., Beam, K. G. 1998; 273 (22): 13403-13406

  Abstract

  Ryanodine receptors (RyRs) are present in the endoplasmic reticulum of virtually every cell type and serve critical roles, including excitation-contraction (EC) coupling in muscle cells. In skeletal muscle the primary control of RyR-1 (the predominant skeletal RyR isoform) occurs via an interaction with plasmalemmal dihydropyridine receptors (DHPRs), which function as both voltage sensors for EC coupling and as L-type Ca2+ channels (Rios, E., and Brum, G. (1987) Nature 325, 717-720). In addition to "receiving" the EC coupling signal from the DHPR, RyR-1 also "transmits" a retrograde signal that enhances the Ca2+ channel activity of the DHPR (Nakai, J., Dirksen, R. T., Nguyen, H. T., Pessah, I. N., Beam, K. G., and Allen, P. D. (1996) Nature 380, 72-76). A similar kind of retrograde signaling (from RyRs to L-type Ca2+ channels) has also been reported in neurons (Chavis, P., Fagni, L., Lansman, J. B., and Bockaert, J. (1996) Nature 382, 719-722). To investigate the molecular mechanism of reciprocal signaling, we constructed cDNAs encoding chimeras of RyR-1 and RyR-2 (the predominant cardiac RyR isoform) and expressed them in dyspedic myotubes, which lack an endogenous RyR-1. We found that a chimera that contained residues 1,635-2,636 of RyR-1 both mediated skeletal-type EC coupling and enhanced Ca2+ channel function, whereas a chimera containing adjacent RyR-1 residues (2, 659-3,720) was only able to enhance Ca2+ channel function. These results demonstrate that two distinct regions are involved in the reciprocal interactions of RyR-1 with the skeletal DHPR.

  View details for Web of Science ID 000073919100010

  View details for PubMedID 9593671

• Muscle cells from mdx mice have an increased susceptibility to oxidative stress NEUROMUSCULAR DISORDERS Rando, T. A., Disatnik, M. H., Yu, Y., Franco, A. 1998; 8 (1): 14-21

  Abstract

  Several lines of evidence suggest that free radical mediated injury and oxidative stress may lead to muscle necrosis in the muscular dystrophies, including those related to defects in the dystrophin gene. We have examined muscle cell death using an in vitro assay in which the processes that lead to myofiber necrosis in vivo may be amenable to investigation in a simplified cell culture system. Using myotube cultures from normal and dystrophin-deficient (mdx) mice, we have examined the susceptibilities of the cells to different metabolic stresses. Dystrophin-deficient cells were more susceptible to free radical induced injury when compared to normal cells, but the two populations were equally susceptible to other forms of metabolic stress. The differential response appeared to be specifically related to dystrophin expression since undifferentiated myoblasts (which do not express dystrophin) from normal and mdx mice were equally sensitive to oxidative stress. Thus, the absence of dystrophin appears to render muscle specifically more susceptible to free radical induced injury. These results support the hypothesis that oxidative stress may lead to myofiber necrosis in these disorders. Elucidating the mechanisms leading to cell death may help to explain the variabilities in disease expression that are seen as a function of age, among different muscles, and across species in animals with muscular dystrophy due to dystrophin deficiency.

  View details for Web of Science ID 000073054000003

  View details for PubMedID 9565986

• Genetic analysis of alpha(4) integrin functions in the development of mouse skeletal muscle JOURNAL OF CELL BIOLOGY Yang, J. T., Rando, T. A., Mohler, W. A., Rayburn, H., Blau, H. M., HYNES, R. O. 1996; 135 (3): 829-835

  Abstract

  It has been suggested, on the basis of immunolocalization studies in vivo and antibody blocking experiments in vitro, that alpha 4 integrins interacting with vascular cell adhesion molecule 1 (VCAM-1) are involved in myogenesis and skeletal muscle development. To test this proposal, we generated embryonic stem (ES) cells homozygous null for the gene encoding the alpha 4 subunit and used them to generate chimeric mice. These chimeric mice showed high contributions of alpha 4-null cells in many tissues, including skeletal muscle, and muscles lacking any detectable (< 2%) alpha 4-positive cells did not reveal any gross morphological abnormalities. Furthermore, assays for in vitro myogenesis using either pure cultures of alpha 4-null myoblasts derived from the chimeras or alpha 4-null ES cells showed conclusively that alpha 4 integrins are not essential for muscle cell fusion and differentiation. Taking these results together, we conclude that alpha 4 integrins appear not to play essential roles in normal skeletal muscle development.

  View details for Web of Science ID A1996VR26800023

  View details for PubMedID 8909554

  View details for PubMedCentralID PMC2121061

• Myoblast-mediated expression of colony stimulating factor-1 (CSF-1) in the cytokine-deficient op/op mouse SOMATIC CELL AND MOLECULAR GENETICS Dhawan, J., Rando, T. A., Elson, S. E., Lee, F., Stanley, E. R., Blau, H. M. 1996; 22 (5): 363-381

  Abstract

  The osteopetrotic (op/op) mouse lacks colony stimulating factor-1 (CSF-1) due to an inactivating mutation in the CSF-1 gene. Intramuscular transplantation of engineered myoblasts was used to introduce CSF-1 into the circulation of op/op mice. The CSF-1 cDNA was introduced into C2C12 mouse myoblasts in culture using retroviral mediated gene transfer. Upon transplantation into the skeletal muscle of mutant mice, physiological levels of the cytokine were achieved systemically and elicited a biological response: circulating monocytes were induced. Howvever, both circulating CSF-1 levels and the induction of monocytes were transient. Analysis of the site of cell transplantation revealed local changes that may account for the transience of serum cytokine levels. Macrophage markers were induced in muscle tissue implanted with CSF-1 expressing myoblasts: c-fms, the CSF-1 receptor as well as the lineage-restricted antigen F4/80. We propose that in addition to CSF-1 clearance by Kupffer cells of the liver, macrophages that accumulated at the site of cell transplantation bound the CSF-1 produced by the muscle cell transplants, precluding the sustained release of this cytokine into the systemic circulation. Our studies also revealed that damage to muscle caused during cell transplantation or by freeze injury resulted in the accumulation of macrophages in op/op mouse muscle tissue. Indeed, op/op mice were fully capable of regenerating injured muscle suggesting the presence of as yet unidentified CSF-1-independent factors capable of generating macrophages that presumably participate in tissue remodeling in this cytokine-deficient mouse.

  View details for Web of Science ID A1996WK59300002

  View details for PubMedID 9039846

• THE FATE OF MYOBLASTS FOLLOWING TRANSPLANTATION INTO MATURE MUSCLE EXPERIMENTAL CELL RESEARCH Rando, T. A., PAVLATH, G. K., Blau, H. M. 1995; 220 (2): 383-389

  Abstract

  Cell transplantation has potential benefits for tissue replacement in the the enhancement of tissue regeneration and as cell-mediated gene therapy for systemic diseases. The transplantation of myoblasts into skeletal muscle also allows gene transfer into cells of the host since myoblasts fuse with host fibers thereby forming hybrid myofibers. The success of myoblast transplantation can be determined by a variety of measures, such as the percentage of myoblasts that fuse, the number of hybrid myofibers formed, or the level of transgene expression. Each measure is a reflection of the fate of the transplanted cells. In order to compare different measures of transplantation efficacy, we followed the fate of transplanted myoblasts expressing the marker enzyme beta-galactosidase (beta-gal) in two different assays. Two weeks after transplantation, the number of hybrid myofibers was determined histochemically, whereas transgene (beta-gal) expression was measured biochemically. To control for variabilities of transplantation among different animals, we obtained both measurements from each muscle by using alternate cryosections in the two assays. Within each individual muscle, both hybrid fiber number and beta-gal expression were maximal at the site of implantation and diminished in parallel with distance from the site. However, for determining the success of transplantation among groups of muscles, these two measures of efficacy yielded discordant results: the transplants with the highest number of hybrid fibers were not the transplants with the greatest beta-gal activity. Such discrepancies are likely due to regional variations at the transplantation site that arise when cells are introduced into a solid tissue. These results demonstrate the importance of multiple measures of cell fate and transplantation efficacy for studies of cell transplantation and for the application of such studies to cell therapy and cell-mediated gene therapy.

  View details for Web of Science ID A1995RX76400017

  View details for PubMedID 7556447

• TETRACYCLINE-REGULATED GENE-EXPRESSION FOLLOWING DIRECT GENE-TRANSFER INTO MOUSE SKELETAL-MUSCLE SOMATIC CELL AND MOLECULAR GENETICS Dhawan, J., Rando, T. A., ELSON, S. L., Bujard, H., Blau, H. M. 1995; 21 (4): 233-240

  Abstract

  For most experimental and therapeutic applications of gene transfer, regulation of the timing and level of gene expression is preferable to constitutive gene expression. Among the systems that have been developed for pharmacologically controlled gene expression in mammalian cells, the bacterial tetracycline (tet)-responsive system has the advantage that it is dependent on a drug (tet) that is both highly specific and non-toxic. The tet-responsive system has been previously used to modulate expression of cell cycle regulatory proteins in cultured cells, reporter genes in plants and transgenic mice and reporter genes directly injected into the heart. Here we show that orally or parenterally administered tet regulates expression of tet-responsive plasmids injected directly into mouse skeletal muscle. Reporter gene expression was suppressed by two orders of magnitude in the presence of tet, and that suppression was reversed when tet was withdrawn. These data show that skeletal muscle offers an accessible and well characterized target tissue for tet-controlled expression of genes in vivo, suggesting applications to developmental studies and gene therapy.

  View details for Web of Science ID A1995TK30000002

  View details for PubMedID 8525429

• TRANSIENT IMMUNOSUPPRESSIVE TREATMENT LEADS TO LONG-TERM RETENTION OF ALLOGENEIC MYOBLASTS IN HYBRID MYOFIBERS JOURNAL OF CELL BIOLOGY PAVLATH, G. K., Rando, T. A., Blau, H. M. 1994; 127 (6): 1923-1932

  Abstract

  Normal and genetically engineered skeletal muscle cells (myoblasts) show promise as drug delivery vehicles and as therapeutic agents for treating muscle degeneration in muscular dystrophies. A limitation is the immune response of the host to the transplanted cells. Allogeneic myoblasts are rapidly rejected unless immunosuppressants are administered. However, continuous immunosuppression is associated with significant toxic side effects. Here we test whether immunosuppressive treatment, administered only transiently after allogeneic myoblast transplantation, allows the long-term survival of the transplanted cells in mice. Two immunosuppressive treatments with different modes of action were used: (a) cyclosporine A (CSA); and (b) monoclonal antibodies to intracellular adhesion molecule-1 and leukocyte function-associated molecule-1. The use of myoblasts genetically engineered to express beta-galactosidase allowed quantitation of the survival of allogeneic myoblasts at different times after cessation of the immunosuppressive treatments. Without host immunosuppression, allogeneic myoblasts were rejected from all host strains tested, although the relative time course differed as expected for low and high responder strains. The allogeneic myoblasts initially fused with host myofibers, but these hybrid cells were later destroyed by the massive immunological response of the host. However, transient immunosuppressive treatment prevented the hybrid myofiber destruction and led to their long-term retention. Even four months after the cessation of treatment, the hybrid myofibers persisted and no inflammatory infiltrate was present in the tissue. Such long-term survival indicates that transient immunosuppression may greatly increase the utility of myoblast transplantation as a therapeutic approach to the treatment of muscle and nonmuscle disease.

  View details for Web of Science ID A1994PZ27200014

  View details for PubMedID 7806570

• PSEUDOCHOREOATHETOSIS - MOVEMENTS ASSOCIATED WITH LOSS OF PROPRIOCEPTION ARCHIVES OF NEUROLOGY Sharp, F. R., Rando, T. A., Greenberg, S. A., Brown, L., Sagar, S. M. 1994; 51 (11): 1103-1109

  Abstract

  To describe seven patients with proprioceptive sensory loss and choreoathetoid movements.Case series.Outpatient and inpatient university referral.Patients with sensory loss and abnormal movements.None.None.One patient had a parietal cortex injury, one had a thalamic infarction, two had spinal cord lesions, two had dorsal root ganglion neuronopathies, and one had an ulnar neuropathy. In each case, the duration of abnormal movements correlated with the duration of proprioceptive sensory loss, and the abnormal movements were restricted to body parts with proprioceptive sensory loss. The movements varied from chorea and athetosis to dystonia.These cases suggest that proprioceptive sensory loss can lead to a movement disorder, termed pseudochoreoathetosis, which occurs following the appearance of lesions anywhere along proprioceptive sensory pathways, from peripheral nerves to the cerebral cortex. It is hypothesized that pseudochoreoathetosis occurs because of the failure to process limb proprioceptive information in the striatum. Therefore, both choreoathetosis and pseudochoreoathetosis may be manifestations of the failure of the striatum to properly integrate cortical motor and sensory inputs.

  View details for Web of Science ID A1994PQ07500005

  View details for PubMedID 7980104

• PRIMARY MOUSE MYOBLAST PURIFICATION, CHARACTERIZATION, AND TRANSPLANTATION FOR CELL-MEDIATED GENE-THERAPY JOURNAL OF CELL BIOLOGY Rando, T. A., Blau, H. M. 1994; 125 (6): 1275-1287

  Abstract

  The transplantation of cultured myoblasts into mature skeletal muscle is the basis for a new therapeutic approach to muscle and non-muscle diseases: myoblast-mediated gene therapy. The success of myoblast transplantation for correction of intrinsic muscle defects depends on the fusion of implanted cells with host myofibers. Previous studies in mice have been problematic because they have involved transplantation of established myogenic cell lines or primary muscle cultures. Both of these cell populations have disadvantages: myogenic cell lines are tumorigenic, and primary cultures contain a substantial percentage of non-myogenic cells which will not fuse to host fibers. Furthermore, for both cell populations, immune suppression of the host has been necessary for long-term retention of transplanted cells. To overcome these difficulties, we developed novel culture conditions that permit the purification of mouse myoblasts from primary cultures. Both enriched and clonal populations of primary myoblasts were characterized in assays of cell proliferation and differentiation. Primary myoblasts were dependent on added bFGF for growth and retained the ability to differentiate even after 30 population doublings. The fate of the pure myoblast populations after transplantation was monitored by labeling the cells with the marker enzyme beta-galactosidase (beta-gal) using retroviral mediated gene transfer. Within five days of transplantation into muscle of mature mice, primary myoblasts had fused with host muscle cells to form hybrid myofibers. To examine the immunobiology of primary myoblasts, we compared transplanted cells in syngeneic and allogeneic hosts. Even without immune suppression, the hybrid fibers persisted with continued beta-gal expression up to six months after myoblast transplantation in syngeneic hosts. In allogeneic hosts, the implanted cells were completely eliminated within three weeks. To assess tumorigenicity, primary myoblasts and myoblasts from the C2 myogenic cell line were transplanted into immunodeficient mice. Only C2 myoblasts formed tumors. The ease of isolation, growth, and transfection of primary mouse myoblasts under the conditions described here expand the opportunities to study muscle cell growth and differentiation using myoblasts from normal as well as mutant strains of mice. The properties of these cells after transplantation--the stability of resulting hybrid myofibers without immune suppression, the persistence of transgene expression, and the lack of tumorigenicity--suggest that studies of cell-mediated gene therapy using primary myoblasts can now be broadly applied to mouse models of human muscle and non-muscle diseases.

  View details for Web of Science ID A1994NT42000008

  View details for PubMedID 8207057

• TUMOR SUPPRESSION BY RNA FROM THE 3' UNTRANSLATED REGION OF ALPHA-TROPOMYOSIN CELL Rastinejad, F., Conboy, M. J., Rando, T. A., Blau, H. M. 1993; 75 (6): 1107-1117

  Abstract

  NMU2, a nondifferentiating mutant myogenic cell line, gives rise to rhabdomyosarcomas in mice. We show that constitutive expression of RNA from 0.2 kb of the alpha-tropomyosin (Tm) 3' untranslated region (UTR), but not control 3'UTRs, suppresses anchorage-independent growth and tumor formation by NMU2 cells. When beta-galactosidase (beta-gal)-labeled cells were implanted into muscles of adult mouse hindlimbs, Tm 3'UTR expression suppressed the proliferation, invasion, and destruction of muscle tissues characteristic of NMU2. In the rare tumors that developed from Tm 3'UTR transfectants, RNA expression was extinguished. These results suggest that suppression of tumorigenicity is dependent on the continued expression of Tm transcripts lacking a coding region. We conclude that untranslated RNAs can function as regulators (riboregulators) that suppress tumor formation.

  View details for Web of Science ID A1993MM89300009

  View details for PubMedID 7505203

• WOLFRAM SYNDROME - EVIDENCE OF A DIFFUSE NEURODEGENERATIVE DISEASE BY MAGNETIC-RESONANCE-IMAGING 43RD ANNUAL MEETING OF THE AMERICAN ACADEMY OF NEUROLOGY Rando, T. A., Horton, J. C., LAYZER, R. B. LIPPINCOTT WILLIAMS & WILKINS. 1992: 1220–24

  Abstract

  Wolfram syndrome is an autosomal recessive disorder beginning in childhood that consists of four cardinal features: optic atrophy, diabetes mellitus, diabetes insipidus, and neurosensory hearing loss. Aside from these features, the clinical picture is highly variable and may include other neurologic abnormalities such as ataxia, nystagmus, mental retardation, and seizures. We present two unrelated patients with Wolfram syndrome, both of whom had the four cardinal features and several other neurologic abnormalities. MRIs showed widespread atrophic changes throughout the brain, some of which correlated with the major neurologic features of the syndrome.

  View details for Web of Science ID A1992HX95900017

  View details for PubMedID 1603350

• SPONTANEOUS INTRACRANIAL HYPOTENSION - REPORT OF 2 CASES AND REVIEW OF THE LITERATURE NEUROLOGY Rando, T. A., Fishman, R. A. 1992; 42 (3): 481-487

  Abstract

  We report two patients with spontaneous intracranial hypotension. In addition to the cardinal features of a postural headache and a low CSF pressure, the patients also had subdural fluid collections demonstrated by head MRI. In both patients, radionuclide cisternography revealed a CSF leak along the spinal axis and rapid accumulation of radioisotope in the bladder. CSF leakage from spinal meningeal defects may be the most common cause of this syndrome. The headache is a consequence of the low CSF pressure producing displacement of pain-sensitive structures. Associated symptoms, including tinnitus and vertigo, and subdural fluid collections are presumably from hydrostatic changes among intracranial fluid compartments that occur at low CSF pressures. Methods of treatment are identical to those for post-dural puncture headaches. Epidural blood patches and epidural saline infusions have rapidly ameliorated the symptoms of spontaneous intracranial hypotension.

  View details for Web of Science ID A1992HJ99200003

  View details for PubMedID 1549206

• RAPID AND SLOW GATING OF VERATRIDINE-MODIFIED SODIUM-CHANNELS IN FROG MYELINATED NERVE JOURNAL OF GENERAL PHYSIOLOGY Rando, T. A. 1989; 93 (1): 43-65

  Abstract

  The properties of voltage-dependent Na channels modified by veratridine (VTD) were studied in voltage-clamped nodes of Ranvier of the frog Rana pipiens. Two modes of gating of VTD-modified channels are described. The first, occurring on a time scale of milliseconds, is shown to be the transition of channels between a modified resting state and a modified open state. There are important qualitative and quantitative differences of this gating process in nerve compared with that in muscle (Leibowitz et al., 1986). A second gating process occurring on a time scale of seconds, was originally described as a modified activation process (Ulbricht, 1969). This process is further analyzed here, and a model is presented in which the slow process represents the gating of VTD-modified channels between open and inactivated states. An expanded model is a step in the direction of unifying the known rapid and slow physiologic processes of Na channels modified by VTD and related alkaloid neurotoxins.

  View details for Web of Science ID A1989R845800003

  View details for PubMedID 2536798

• THE INTERACTION BETWEEN THE ACTIVATOR AGENTS BATRACHOTOXIN AND VERATRIDINE AND THE GATING PROCESSES OF NEURONAL SODIUM-CHANNELS MOLECULAR PHARMACOLOGY Rando, T. A., Wang, G. K., Strichartz, G. R. 1986; 29 (5): 467-477

  Abstract

  The depolarization of frog sciatic nerves by the Na channel-activating toxins, batrachotoxin and veratridine, was studied using the sucrose-gap technique. To study the interaction between the activators and the gating processes of Na channels, we measured the depolarizations of unstimulated nerves, of nerves during repetitive stimulation, and of nerves whose Na channel inactivation process had been pharmacologically modified. Stimulation enhanced the rates of depolarization by the activators but did not effect the steady state depolarization values. Of the three inhibitors of Na channel inactivation that were tested (Leiurus alpha-scorpion toxin, chloramine T, and Ni2+), only Leiurus toxin enhanced the potencies of the activators. Neither chloramine T nor Ni2+ had any effect on the steady state level of depolarization produced by either activator. Both chloramine T and Ni2+, however, enhanced the rate of batrachotoxin action, although neither affected the rate of veratridine action. Leiurus toxin also potentiated the effects of the activators in chloramine T-treated nerves. We tested the interaction between the Na channel activators and a class of agents, local anesthetics, that stabilize a non-conducting state of the Na channel. The presence of lidocaine inhibited the depolarization produced by addition of either activator, although the addition of lidocaine subsequent to the development of batrachotoxin-induced depolarization produced repolarization very weakly and slowly. We also found that the lidocaine homologue, RAC 109I, was about 3 times as potent as its stereoisomer, RAC 109II, in its ability both to reduce the compound action potential amplitude and to inhibit the veratridine-induced depolarization.

  View details for Web of Science ID A1986C418900004

  View details for PubMedID 2422536

• SAXITOXIN BLOCKS BATRACHOTOXIN-MODIFIED SODIUM-CHANNELS IN THE NODE OF RANVIER IN A VOLTAGE-DEPENDENT MANNER BIOPHYSICAL JOURNAL Rando, T. A., Strichartz, G. R. 1986; 49 (3): 785-794

  Abstract

  The inhibition by saxitoxin (STX) of single Na channels incorporated into planar lipid bilayers and modified by batrachotoxin (BTX) previously has been shown to be voltage dependent (Krueger, B.K.,J.F. Worley, and R. J. French, 1983, Nature [Lond.], 303:172-175; Moczydlowski, E., S. Hall, S. S. Garber, G. S. Strichartz, and C. Miller, 1984, J. Gen. Physiol., 84:687-704). We tested for such a voltage dependence of STX block of the Na current in voltage-clamped frog nodes of Ranvier. The block by STX of normal Na channels showed no modulation in response to maintained (20 s) changes of the membrane potential or to a train of brief pulses to potentials more positive than the holding potential. However, when the nodal channels were modified by BTX, the train of pulses produced a modulation of the block of the Na current by STX. The modulation of STX block depended on the voltage of the conditioning pulses and this voltage dependence agreed well with that predicted from the single channel studies over the membrane potential range used in those studies. In addition, we found that the voltage dependence of STX block was manifest only at potentials equal to or more positive than required to activate the channels. Most of the apparent differences among data from single channels in bilayers, equilibrium binding studies of STX, and the experiments described here are resolved by the hypotheses that (a) STX binding to open channels is voltage dependent, and (b) the affinities of STX for closed and inactivated channels are independent of voltage, equal, and less than the open channel affinity at potentials less than 0 mV. Whether these hypotheses apply to the STX block of all Na channels or just of BTX-modified channels remains to be determined.

  View details for Web of Science ID A1986A473600021

  View details for PubMedID 2421797

• LOCALIZATION OF NEURONS IN THE RAT SPINAL-CORD WHICH PROJECT TO THE SUPERIOR CERVICAL-GANGLION JOURNAL OF COMPARATIVE NEUROLOGY Rando, T. A., Bowers, C. W., Zigmond, R. E. 1981; 196 (1): 73-83

  Abstract

  Horseradish peroxidase (HRP) was used to determine the location in the spinal cord of neurons projecting to the superior cervical ganglion of the rat. HRP was applied to the proximal cut end of the cervical sympathetic trunk, close to its entry into the superior cervical ganglion. After survival times of 3, 6, or 9 days, the animals were sacrificed and their spinal cords were processed to visualize HRP using diaminobenzidine, benzidine dihydrochloride, or tetramethylbenzidine. Labeled neurons were found only ipsilateral to the site of HRP application and were restricted to spinal segments C8-T5. Ninety percent of these neurons were located in segments T1-T3. Similar numbers of labeled neurons were found at survival times of 3 and 6 days and the mean number +/- S.E.M. for 11 experiments at these two survival times was 1575 +/- 89. Nine days after application of HRP the mean number of labeled cells and the density of label per cell were reduced. Labeled neurons were found in four regions of the spinal cord: the intermediolateral nucleus (75%), the lateral funiculus (23%), the central autonomic area (1%), and the intercalated region (1%). The cells of the intermediolateral nucleus did not form a continuous column along the rostrocaudal axis of the spinal cord, but instead were often found in clusters, several clusters being present per spinal segment.

  View details for Web of Science ID A1981LC36200006

  View details for PubMedID 7204667