The myogenic precursor cells in postnatal and adult skeletal muscle (satellite cells) are situated adjacent to the myofiber. In growing animals some satellite cells proliferate, adding nuclei to myofibers. In adults these precursors are mitotically quiescent, but can reinitiate proliferative activity following a broad range of muscular stresses ranging from exercise to overt injury. Cell culture studies have identified several peptide growth factors that can regulate proliferation and differentiation of satellite cells; however, the in vivo association of satellite cells with their neighboring myofibers, which might be important in regulating myogenesis of satellite cells, is not maintained under such conditions. The present application focuses on the role of fibroblast growth factors (FGFs) in regulating activation and proliferation of satellite cells situated in their native position. Cultures of isolated rat muscle fibers are employed in many of the studies. The isolated fibers retain their basement membrane and satellite cells, providing a model for evaluating 'in situ' myogenesis of satellite cells. The long term goal of the proposal is the understanding of mechanisms controlling growth and regeneration of skeletal muscle. The overall hypothesis of the proposal is that specific FGFs regulate specific phases of satellite cell activation and proliferation, and that this program is linked to a specific pattern of expression of FGF receptors; FGFs are produced locally in growing and adult muscle, supporting satellite cell myogeneSis, but their levels are reduced in aging muscle.
The specific aims of the proposal are: *To determine the roles of aFGF (FGF-1) and bFGF (FGF-2) during activation and proliferation of satellite cells on adult fibers. This will include analysis of FGF effects on the activation of mitogen- activated protein kinase and investigations of the nature of FGF contributing by the fiber itself. *To analyze the expression of the high affinity FGF receptors 1 and 4 (FGFR-1 and FGFR-4) by satellite cells during their activation and proliferation on adult fibers. Analysis will focus on mRNA expression. *To compare the role of FGF during myogenesis of satellite cells on fibers from growing, adult and old rats. Analysis will focus on the effects of aFGF and bFGF, expression of FGFR-1 and FGFR-4, and the contribution of endogenous FGF by the fibers. Methods to be used include fiber isolation and culture, monitoring the expression of specific proteins via immunofluorescence and Western blotting, and analysis of mRNA expression by Northern blotting, in situ hybridization and polymerase chain reaction. The proposed studies are likely to contribute to better understanding of muscle growth and aging, and can prove important for improving muscle rehabilitation following muscle disuse and injury.

Agency
National Institute of Health (NIH)
Institute
National Institute on Aging (NIA)
Type
Research Project (R01)
Project #
5R01AG013798-08
Application #
2517047
Study Section
Molecular Cytology Study Section (CTY)
Program Officer
Mccormick, Anna M
Project Start
1989-01-01
Project End
1999-08-31
Budget Start
1997-09-01
Budget End
1998-08-31
Support Year
8
Fiscal Year
1997
Total Cost
Indirect Cost
Name
University of Washington
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195
Keire, Paul; Shearer, Andrew; Shefer, Gabi et al. (2013) Isolation and culture of skeletal muscle myofibers as a means to analyze satellite cells. Methods Mol Biol 946:431-68
Danoviz, Maria Elena; Yablonka-Reuveni, Zipora (2012) Skeletal muscle satellite cells: background and methods for isolation and analysis in a primary culture system. Methods Mol Biol 798:21-52
Day, Kenneth; Shefer, Gabi; Shearer, Andrew et al. (2010) The depletion of skeletal muscle satellite cells with age is concomitant with reduced capacity of single progenitors to produce reserve progeny. Dev Biol 340:330-43
Kirkpatrick, Lisa J; Yablonka-Reuveni, Zipora; Rosser, Benjamin W C (2010) Retention of Pax3 expression in satellite cells of muscle spindles. J Histochem Cytochem 58:317-27
Ieronimakis, Nicholas; Balasundaram, Gayathri; Rainey, Sabrina et al. (2010) Absence of CD34 on murine skeletal muscle satellite cells marks a reversible state of activation during acute injury. PLoS One 5:e10920
Day, Kenneth; Paterson, Bruce; Yablonka-Reuveni, Zipora (2009) A distinct profile of myogenic regulatory factor detection within Pax7+ cells at S phase supports a unique role of Myf5 during posthatch chicken myogenesis. Dev Dyn 238:1001-9
Kirkpatrick, Lisa J; Allouh, Mohammed Z; Nightingale, Chantale N et al. (2008) Pax7 shows higher satellite cell frequencies and concentrations within intrafusal fibers of muscle spindles. J Histochem Cytochem 56:831-40
Yablonka-Reuveni, Z; Day, K; Vine, A et al. (2008) Defining the transcriptional signature of skeletal muscle stem cells. J Anim Sci 86:E207-16
Kwiatkowski, Boguslaw A; Kirillova, Irina; Richard, Robert E et al. (2008) FGFR4 and its novel splice form in myogenic cells: Interplay of glycosylation and tyrosine phosphorylation. J Cell Physiol 215:803-17
Allouh, Mohammed Z; Yablonka-Reuveni, Zipora; Rosser, Benjamin W C (2008) Pax7 reveals a greater frequency and concentration of satellite cells at the ends of growing skeletal muscle fibers. J Histochem Cytochem 56:77-87

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