Progress 01/01/05 to 12/31/08
Outputs
OUTPUTS: OUTPUTS: We have been studying the activation of the resident stem cells in skeletal muscle, satellite cells. These cells are important in normal muscle growth and in repair of damaged muscle, a very common occurrence in most animals. In the last objective of the proposal, we used microarrays to identify genes that are either turned on or off during activation. These experiments have led to discovery of new and potentially important aspects of satellite cell function. The most recent gene being explored encodes a cell surface receptor that may mediate satellite cell motility and movement to a site of injury where it will proliferate, differentiate and repair the damaged muscle. This receptor is CXCR4 and is activated by stromal-derived factor. The expression of the receptor and ligand in growing and damaged muscle is being evaluated. This knowledge contributes to our basic understanding of muscle growth in meat producing animals. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project. PARTICIPANTS: Dr. Robert Rhoads worked on this project and is now an assistant professor in the Department of Animal Sciences. He is currently PI of his own independent NRI grant. Dr. Xiaosong Liu was funded on this grant and continues to be a Research Associate in the Department of Animal Sciences. TARGET AUDIENCES: The target audience is composed of other scientists in the animal sciences that are concerned with the growth of skeletal muscle. These include people interested in the basic biology of muscle growth as well as folks studying applied nutrition and health of growing meat animals. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
The primary outcome is a change in knowledge about a new molecular system that may play a key role in satellite cell function. We found that a variety of interesting events occur as satellite cells are becoming activated; the most novel came from analyses of microarrays of messages that change expression during activation. Interestingly we found that a pro-angiogenic program is initiated and that activated satellite cells can stimulate angiogenesis. Through this mechanism, the regeneration of new fibers is coordinated with the regeneration of vasculature to support new fibers. We also found that a ligand/receptor system not previously known to be expressed in satellite cells (SDF-1/CXCR4)is turned on during activation. CXCR4 is not expressed in quiescent satellite cells, but is rapidly expressed following activation. In dividing satellite cells, the receptor is present and presumably directs migration of cells; receptor expression diminishes with differentiation and is not found in muscle fibers. The receptor is not found in muscle-derived fibroblasts. The ligand, SDF1 is found in muscle fibroblasts and is especially evident in new myotubes in culture. In undamaged muscle, very little CXCR4 is found, and only modest amounts of SDF1 can be seen around muscle fibers. In damaged tissue, however, satellite cells express CXCR4, and SDF1 is plentiful in the damage region. These are but two aspects of satellite cell biology that have been discovered in this project that are important to meat animal muscle growth and repair.
Publications
- Mendias, C.L., Tatsumi, R., and Allen, R.E. (2004). The role of cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) in skeletal muscle satellite cell proliferation, differentiation and fusion. Muscle and Nerve 30:497-500.
- Tatsumi, R and Allen, RE. (2008). Mechano-biology of resident myogenic stem cells: Molecular mechanism of stretch-induced activation of satellite cells. Animal Science J. 79:279-290.
- Wagner, K.R., Liu, X., Chang, X., and Allen, R.E. (2005). Muscle regeneration in the prolonged absence of myostatin. Proc. Nat. Acad. Sci. USA 102:2519-2524.
- Tatsumi, R., Liu, X., Pulido, A., Morales, M., Sakata, T., Dial, S., Hattori, A., Ikeuchi, Y., and Allen, R.E.(2006). Satellite cell activation in stretched skeletal muscle and the role of nitric oxide and hepatocyte growth factor. Am. J. Physiol. Cell Physiol. 290:C1487-94.
- Yamada, M., Tatsumi, R., Kikuri, T., Okamoto S., Nonoshita, S., Mizunoya, W., Ikeuchi, Y., Shimokawa, H., Sunagawa, K., and Allen, R.E. (2006). Matrix metalloproteinases are involved in mechanical stretch-induced activation of skeletal muscle satellite cells. Muscle Nerve 34:313-319.
- Katsuki, Y., Tatsumi, R., Yamada, M., Ishizaki, J., Mizunoya, W., Ikeuchi, Y., Hattori, A., Shimokawa, H., Sungawa, K., and Allen, R. E. (2006). Low-pH preparation of skeletal muscle satellite cells can be used to study activation in vitro. Int. J. Biochem. Cell Biol. 38:1678-1685.
- Yamada, M, Sankoda, Y, Tatsumi, R Mizunoya, W, Ikeuchi, Y, Sunagawa, K and Allen, RE. (2008). Matrix metalloproteinase-2 mediates stretch-induced activation of skeletal muscle satellite cells in a nitric oxide-depeandent manner. Int. J. Biochem. Cell Biol. 40:2183-2191.
- Rhoads RP, Johnson, RM, Rathbone, CR, X Liu, Temm-Grove, C, Sheehan, SM, Hoying, JB and Allen, RE. (2009). Satellite cell mediated angiogenesis coincides with a functional hypoxia inducible factor (HIF) pathway. (in revision).
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Progress 01/01/07 to 12/31/07
Outputs
OUTPUTS: We have been studying the activation of the resident stem cells in skeletal muscle, satellite cells. These cells are important in normal muscle growth and in repair of damaged muscle, a very common occurrence in most animals. In the last objective of the proposal, we used microarrays to identify genes that are either turned on or off during activation. These experiments have led to discovery of new and potentially important aspects of satellite cell function. The most recent gene being explored encodes a cell surface receptor that may mediate satellite cell motility and movement to a site of injury where it will proliferate, differentiate and repair the damaged muscle. This receptor is CXCR4 and is activated by stromal-derived factor. The expression of the receptor and ligand in growing and damaged muscle is being evaluated. This knowledge contributes to our basic understanding of muscle growth in meat producing animals. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
The primary outcome is a change in knowledge about a new molecular system that may play a key role in satellite cell function. We are finding that CXCR4 is not expressed in quiescent satellite cells, but is rapidly expressed following activation. In dividing satellite cells, the receptor is present and presumably directs migration of cells; receptor expression diminishes with differentiation and is not found in muscle fibers. The receptor is not found in muscle-derived fibroblasts. The ligand, SDF1 is found in muscle fibroblasts and is especially evident in new myotubes in culture. In undamaged muscle, very little CXCR4 is found, and only modest amounts of SDF1 can be seen around muscle fibers. In damaged tissue, however, satellite cells express CXCR4, and SDF1 is plentiful in the damage region. Future studies will examine the extent to which CXCR4 and SDF1 mediate satellite cell movement in muscle.
Publications
- Meeson AP, Shi X, Alexander MS, Williams RS, Allen RE, Jiang N, Adham IM, Goetsch SC, Hammer RE, Garry DJ. (2007). Sox15 and Fhl3 transcriptionally coactivate Foxk1 and regulate myogenic progenitor cells. EMBO J. 6:1902-12.
- Reed SA, Ouellette SE, Liu X, Allen RE, Johnson SE. (2007). E2F5 and LEK1 translocation to the nucleus is an early event demarcating myoblast quiescence. J Cell Biochem. 101:1394-408.
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Progress 01/01/06 to 01/01/07
Outputs
When skeletal muscle is stretched or injured, myogenic satellite cells are activated to enter the cell cycle. This process depends on nitric oxide (NO) production, release of hepatocyte growth factor (HGF) from the extracellular matrix, and presentation of HGF to the c-met receptor. Experiments reported herein provide new evidence that matrix metalloproteinases (MMPs) are involved in the NO-dependent release of HGF in vitro. When rat satellite cells were treated with 10 ng/ml recombinant tissue inhibitor-1 of MMPs (TIMP-1) and subjected to treatments that induce activation in vitro, sodium nitroprusside (SNP) of a NO donor or mechanical cyclic stretch, the activation response was inhibited. In addition, conditioned medium generated by cultures treated with TIMP-1 plus SNP or mechanical stretch failed to activate cultured satellite cells and did not contain HGF. Moreover, NOx assay demonstrated that TIMP-1 does not impair NO synthase activity of stretched satellite cell
cultures. Therefore, results from these experiments provide strong evidence that MMP(s) mediates HGF release from the matrix and that this step in the pathway is downstream from NO synthesis.
Impacts
The work reported this year provides part of the mechanism by which mechanical changes in muscle, such as with damage, growth or exercise, stimulates the resident muscle stem cells to activate and mediate the repair of hypertrophy of muscle.
Publications
- 1. Yamada, M., Tatsumi, R., Kikuri, T., Okamoto S., Nonoshita, S., Mizunoya, W., Ikeuchi, Y., Shimokawa, H., Sunagawa, K., and Allen, R.E. (2006). Matrix metalloproteinases are involved in mechanical stretch-induced activation of skeletal muscle satellite cells. Muscle Nerve 34:313-319.
- 2. Katsuki, Y., Tatsumi, R., Yamada, M., Ishizaki, J., Mizunoya, W., Ikeuchi, Y., Hattori, A., Shimokawa, H., Sungawa, K., and Allen, R. E. (2006). Low-pH preparation of skeletal muscle satellite cells can be used to study activation in vitro. Int. J. Biochem. Cell Biol. 38:1678-1685.
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Progress 01/01/05 to 12/31/05
Outputs
The roles of hepatocyte growth factor (HGF) and nitric oxide (NO) in activation of satellite cells in passively stretched rat skeletal muscle were examined. A hind limb suspension model was developed in which the vastus, adductor and gracilis muscles were subjected to stretch for 1 hr. Satellite cells were activated by stretch, as determined by BrdU incorporation in vivo. Extracts from stretched muscles stimulated BrdU incorporation in freshly isolated control rat satellite cells in a concentration-dependent manner. Extracts from stretched muscles contained the active form of HGF, and the satellite cell activating activity could be neutralized by incubation with anti-HGF antibody. The involvement of NO was investigated by administering L-NAME or the inactive enantiomer D-NAME, prior to stretch treatment. In vivo activation of satellite cells in stretched muscle was not inhibited by D-NAME but was inhibited by the presence of L-NAME. The activity of stretched muscle
extract was abolished by L-NAME treatment, but could be restored by addition of HGF, indicating that the extract was not inhibitory. Finally, nitric oxide synthase activity in stretched and un-stretched muscle was assayed in muscle extracts immediately following 2 hr stretch treatment and was found to be elevated in stretched muscle but not in stretched muscle from L-NAME treated rats. Results from these experiments demonstrate that stretching muscle liberates HGF, in a NO-dependent manner, which can activate satellite cells.
Impacts
This research provides new information on how muscle growth is regulated. The rate and efficency of growth are key determinants of meat animal productivity.
Publications
- Wagner, K.R., Liu, X., Chang, X., and Allen, R.E. (2005). Muscle regeneration in the prolonged absence of myostatin. Proc. Nat. Acad. Sci. USA 102:2519-2524.
- Tatsumi, R., Liu, X., Pulido, A., Morales, M., Sakata, T., Hattori, A., Ikeuchi, Y., Dial, S. and Allen, R.E. (2006). Satellite cell activation in stretched skeletal muscle and the role of nitric oxide and hepatocyte growth factor. Am. J. Physiol. (in press).
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