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Description
Postnatal skeletal muscle repair is dependent on the tight regulation of an adult stem cell population known as satellite cells. In response to injury, these quiescent cells are activated, proliferate and express skeletal muscle-specific genes. The majority of satellite cells will fuse to damaged fibers or form new muscle fibers, while a subset will return to a quiescent state, where they are available for future rounds of repair. Robust muscle repair is dependent on the signals that regulate the mutually exclusive decisions of differentiation and self-renewal. A likely candidate for regulating this process is NUMB, an inhibitor of Notch signaling pathway that has been shown to asymmetrically localize in daughter cells undergoing cell fate decisions. In order to study the role of this protein in muscle repair, an inducible knockout of Numb was made in mice. Numb deficient muscle had a defective repair response to acute induced damage as characterized by smaller myofibers, increased collagen deposition and infiltration of fibrotic cells. Satellite cells isolated from Numb-deficient mice show decreased proliferation rates. Subsequent analyses of gene expression demonstrated that these cells had an aberrantly up-regulated Myostatin (Mstn), an inhibitor of myoblast proliferation. Further, this defect could be rescued with Mstn specific siRNAs. These data indicate that NUMB is necessary for postnatal muscle repair and early proliferative expansion of satellite cells. We used an evolutionary compatible to examine processes controlling satellite cell fate decisions, primary satellite cell lines were generated from Anolis carolinensis. This green anole lizard is evolutionarily the closet animal to mammals that forms de novo muscle tissue while undergoing tail regeneration. The mechanism of regeneration in anoles and the sources of stem cells for skeletal muscle, cartilage and nerves are poorly understood. Thus, satellite cells were isolated from A. carolinensis and analyzed for their plasticity. Anole satellite cells show increased plasticity as compared to mouse as determined by expression of key markers specific for bone and cartilage without administration of exogenous morphogens. These novel data suggest that satellite cells might contribute to more than muscle in tail regeneration of A. carolinensis.
ContributorsGeorge, Rajani M (Author) / Wilson-Rawls, Jeanne (Thesis advisor) / Rawls, Alan (Committee member) / Whitfield, Kerr (Committee member) / Kusumi, Kenro (Committee member) / Arizona State University (Publisher)
Created2012
Description
Satellite cells are adult muscle stem cells that activate, proliferate, and differentiate into myofibers upon muscle damage. Satellite cells can be cultured and manipulated in vitro, and thus represent an accessible model for studying skeletal muscle biology, and a potential source of autologous stem cells for regenerative medicine. This work summarizes efforts to further understanding of satellite cell biology, using novel model organisms, bioengineering, and molecular and cellular approaches. Lizards are evolutionarily the closest vertebrates to humans that regenerate entire appendages. An analysis of lizard myoprogenitor cell transcriptome determined they were most transcriptionally similar to mammalian satellite cells. Further examination showed that among genes with the highest level of expression in lizard satellite cells were an increased number of regulators of chondrogenesis. In micromass culture, lizard satellite cells formed nodules that expressed chondrogenic regulatory genes, thus demonstrating increased musculoskeletal plasticity. However, to exploit satellite cells for therapeutics, development of an ex vivo culture is necessary. This work investigates whether substrates composed of extracellular matrix (ECM) proteins, as either coatings or hydrogels, can support expansion of this population whilst maintaining their myogenic potency. Stiffer substrates are necessary for in vitro proliferation and differentiation of satellite cells, while the ECM composition was not significantly important. Additionally, satellite cells on hydrogels entered a quiescent state that could be reversed when the cells were subsequently cultured on Matrigel. Proliferation and gene expression data further indicated that C2C12 cells are not a good proxy for satellite cells. To further understand how different signaling pathways control satellite cell behavior, an investigation of the Notch inhibitor protein Numb was carried out. Numb deficient satellite cells fail to activate, proliferate and participate in muscle repair. Examination of Numb isoform expression in satellite cells and embryonic tissues revealed that while developing limb bud, neural tube, and heart express the long and short isoforms of NUMB, satellite cells predominantly express the short isoforms. A preliminary immunoprecipitation- proteomics experiment suggested that the roles of NUMB in satellite cells are related to cell cycle modulation, cytoskeleton dynamics, and regulation of transcription factors necessary for satellite cell function.
ContributorsPalade, Joanna (Author) / Wilson-Rawls, Norma (Thesis advisor) / Rawls, Jeffrey (Committee member) / Kusumi, Kenro (Committee member) / Newbern, Jason (Committee member) / Stabenfeldt, Sarah (Committee member) / Arizona State University (Publisher)
Created2020