HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text (PDF) All Versions of this Article: 16/2/207 01-0544fjev1    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by CARSON, J. A. Articles by REECY, J. M. Search for Related Content PubMed PubMed Citation Articles by CARSON, J. A. Articles by REECY, J. M.

Differential gene expression in the rat soleus muscle during early work overload-induced hypertrophy ^{1 ,2}

JAMES A. CARSON, DAN NETTLETON^* and JAMES M. REECY³

Department of Exercise Science, University of South Carolina, Columbia, South Carolina 29208, USA; and
^* Departments of Statistics and of
Animal Science, Iowa State University, Ames, Iowa 50011, USA

³Correspondence: Department of Animal Science, 2255 Kildee Hall, Iowa State University, Ames, IA 50011-3150, USA. E-mail: jreecy{at}iastate.edu

SPECIFIC AIMS

The aim of this study was to identify work overload-responsive genes in skeletal muscle. Understanding the molecular basis underlying the adaptive mechanisms of skeletal muscle in response to work load will provide insight into the processes of hypertrophy and atrophy, which control skeletal muscle mass.

PRINCIPAL FINDINGS

We isolated total RNA from the soleus muscle of rats that had the synergistic gastrocnemius muscle ablated and from sham-operated rats. We analyzed global gene expression with the Affymetrix U34A GeneChip. Pairwise comparisons were made between all ablated and sham animals to identify differentially expressed genes. Of the 8799 gene and EST sequences analyzed with the rat genomic U34A GeneChip, on average 3634 (41.3%) ± 129 (SE) genes or EST were expressed in the soleus muscle. Of these (based on the Affymetrix absolute call data), the expression of 78 (0.89% of expressed genes) genes and 10 (0.11%) EST increased >1.9-fold in response to 3 days of work overload (Table 1 ). The expression of 20 (0.23%) genes and 5 (0.06%) EST decreased >1.8-fold in response to 3 days of work overload (data not shown).

1. Work overload alters the expression of metabolism genes
We identified 11 genes involved in cellular metabolism whose mRNA abundance decreased in response to work overload. For example, lactate dehydrogenase (-16.7-fold), acetyl-CoA dehydrogenase (-4.2-fold), and muscle-specific ß enolase (-64.5-fold) expression decreased in work overload muscle vs. sham-operated muscle. In contrast, work overload increased the expression of genes like apolipoprotein E (7-fold) and stearyl-CoA desaturase (29.2-fold).

2. Work overload increases the expression of genes encoding for autocrine/paracrine proteins
The expression levels of five growth factors and chemokines were up-regulated during skeletal muscle hypertrophy. For instance, the mRNA abundance of insulin-like growth factor 1 and osteopontin increased 16-fold and 9.7-fold, respectively. Similarly, the expression level of interleukin 6 increased 15.3-fold.

3. Work overload increases the expression of genes encoding for extracellular matrix and matrix modifying proteins
The mRNA abundance level of genes encoding for extracellular matrix proteins or matrix modifying proteins was increased during skeletal muscle hypertrophy; two extracellular genes were versican V3 (3.6-fold) and collagen alpha 1 type V (2.3-fold). In contrast, examples of matrix-modifying enzymes were lysyl hydrolase (2.5-fold) and lysyl oxidase (4.4-fold).

4. Work overload altered the expression of intracellular signaling genes
The mRNA abundance of many intracellular signaling genes changed in response to work overload. The expression level of Janus kinase 2 increased 8.7-fold and angiotensin receptor 1 increased 7.2-fold. Similarly, the mRNA abundance of phosphodiesterase 1 and 4 increased 4- and 3.1-fold, respectively. In contrast, the mRNA abundance of protein phosphatase 1ß decreased 7.9-fold.

5. Work overload increases the expression of transcription factors
Work overload increased the expression of 10 transcription factors, one being the muscle-specific transcription factor myogenin (32-fold). Furthermore, work overload increased the expression of smooth muscle cell LIM (5.0-fold) and cardiac Adriamycin-responsive protein (6.9-fold).

6. Work overload increases the expression of immune response genes
Five immune response genes were up-regulated in response to work overload. Three were major histocompatibility complex proteins such as MHC-associated invariant chain gamma (5.4-fold); one was a complement protein C4 (18-fold).

7. Work overload increases the expression of genes involved in cell cycle regulation
The expression of four genes involved in cell cycle regulation increased in response to work overload. These proteins are involved in cell cycle progression, like the cell cycle inhibitor p21 (52.3-fold) and GADD45 (35.8-fold) also increased.

8. Work overload increases the expression of genes involved in protein metabolism
The mRNA abundance of numerous genes involved in protein turnover (protein synthesis and degradation) changed during work overload—for example, the lysosomal proteases cathepsin B (threefold) and S (fivefold). The expression level of the serine proteases kallikrein-like RSKG-50 (22.5-fold) and renal kallikrein (31.4-fold) increased.

CONCLUSIONS

These findings provide a global assessment of the genes whose expression level change in response to work overload. A majority of the genes down-regulated in the hypertrophying skeletal muscle was involved in cellular metabolism. In contrast, up-regulated genes were involved in cell proliferation, cell signaling, metabolism, neural transmission, protein turnover, transcriptions, and structural (Fig. 1 ).

View larger version (41K): [in this window] [in a new window]   Figure 1. Scheme depicting the proposed molecular events and related functional alterations occurring in skeletal muscle in response to work overload.

We observed an up-regulation of genes involved in cell proliferation. This finding is consistent with the fact that satellite cells, fibroblasts, and endothelial cells proliferated in response to work overload. We also observed increased mRNA level for genes encoding for extracellular/secreted growth factor, interleukin, and chemokine proteins, which are likely involved in cell proliferation, cell activation, and cell migration, respectively. Furthermore, work overload increased the expression of growth factor and interleukin receptors. This would suggest that not only does work overload increase the level of signaling molecules produced, but also the cells’ responsiveness to those molecules.

Work overload increased autocrine/paracrine signaling molecules and dramatically increased the expression of intracellular signaling molecules. Some changes are similar to those observed during cardiac muscle hypertrophy and identical to previously published results. However, we also observed increased Wnt and angiotensin signaling. The potential roles of these two intracellular signaling pathways have not received much attention as to their possible role in skeletal muscle hypertrophy. We also observed increased JAK2 signaling. These changes in intracellular signaling probably result in the increased expression levels of transcription factors in response to work overload. Thus, consistent with the theory of mechano-transduction, work overload resulted in altered expression autocrine/paracrine factors, transmembrane protein, intracellular signaling pathways, transcription factors, and late response genes.

We believe that many of the changes observed are probably early signs of fiber type switching, even though myosin isoform switching is not observed until the late stages of skeletal muscle hypertrophy.

There were some lysosomal proteases whose expression level increased in response to increased workload. This increased expression is probably due to macrophage infiltration and subsequent wound repair of the injured myofibers.

In summary, work overload resulted in major changes in skeletal muscle including cell proliferation, extracellular matrix, autocrine/paracrine signaling, intracellular signaling, transcription, protein turnover, metabolism, and neural. These findings help to define the molecular mechanisms underlying the biological adaptation of skeletal muscle in response to an increased workload. They should help in determining the mechanisms regulating skeletal muscle mass, which may lead to strategies to combat muscle atrophy in at-risk individuals: those in spaceflight as well as the bedridden and aged.

FOOTNOTES

¹ To read the full text of this article, go to http://www.fasebj.org/cgi/doi/10.1096/fj.01-0544fje; to cite this article, use FASEB J. (December 14, 2001) 10.1096/fj.01-0544fje

² Journal Paper No. J-19429 of the Iowa Agriculture and Home Economics Experiment Station, Ames, IA; Project 3600.

This article has been cited by other articles:

				C. Hayashi, Y. Ono, N. Doi, F. Kitamura, M. Tagami, R. Mineki, T. Arai, H. Taguchi, M. Yanagida, S. Hirner, et al. Multiple Molecular Interactions Implicate the Connectin/Titin N2A Region as a Modulating Scaffold for p94/Calpain 3 Activity in Skeletal Muscle J. Biol. Chem., May 23, 2008; 283(21): 14801 - 14814. [Abstract] [Full Text] [PDF]

				I. A. Barash, M.-L. Bang, L. Mathew, M. L. Greaser, J. Chen, and R. L. Lieber Structural and regulatory roles of muscle ankyrin repeat protein family in skeletal muscle Am J Physiol Cell Physiol, July 1, 2007; 293(1): C218 - C227. [Abstract] [Full Text] [PDF]

				K. A. Huey, G. E. McCall, H. Zhong, and R. R. Roy Modulation of HSP25 and TNF-{alpha} during the early stages of functional overload of a rat slow and fast muscle J Appl Physiol, June 1, 2007; 102(6): 2307 - 2314. [Abstract] [Full Text] [PDF]

				N. Juretic, P. Garcia-Huidobro, J. A. Iturrieta, E. Jaimovich, and N. Riveros Depolarization-induced slow Ca2+ transients stimulate transcription of IL-6 gene in skeletal muscle cells Am J Physiol Cell Physiol, May 1, 2006; 290(5): C1428 - C1436. [Abstract] [Full Text] [PDF]

				S. Choi, X. Liu, P. Li, T. Akimoto, S. Y. Lee, M. Zhang, and Z. Yan Transcriptional profiling in mouse skeletal muscle following a single bout of voluntary running: evidence of increased cell proliferation J Appl Physiol, December 1, 2005; 99(6): 2406 - 2415. [Abstract] [Full Text] [PDF]

				D. D. Armstrong and K. A. Esser Wnt/{beta}-catenin signaling activates growth-control genes during overload-induced skeletal muscle hypertrophy Am J Physiol Cell Physiol, October 1, 2005; 289(4): C853 - C859. [Abstract] [Full Text] [PDF]

				C. M. Westerkamp and S. E. Gordon Angiotensin-converting enzyme inhibition attenuates myonuclear addition in overloaded slow-twitch skeletal muscle Am J Physiol Regulatory Integrative Comp Physiol, October 1, 2005; 289(4): R1223 - R1231. [Abstract] [Full Text] [PDF]

				C. S. Lin and C. W. Hsu Differentially transcribed genes in skeletal muscle of Duroc and Taoyuan pigs J Anim Sci, September 1, 2005; 83(9): 2075 - 2086. [Abstract] [Full Text] [PDF]

				M. Fluck, S. Schmutz, M. Wittwer, H. Hoppeler, and D. Desplanches Transcriptional reprogramming during reloading of atrophied rat soleus muscle Am J Physiol Regulatory Integrative Comp Physiol, July 1, 2005; 289(1): R4 - R14. [Abstract] [Full Text] [PDF]

				J. M. McClung, K. A. Mehl, R. W. Thompson, L. L. Lowe, and J. A. Carson Nandrolone decanoate modulates cell cycle regulation in functionally overloaded rat soleus muscle Am J Physiol Regulatory Integrative Comp Physiol, June 1, 2005; 288(6): R1543 - R1552. [Abstract] [Full Text] [PDF]

				T. A. Washington, J. M. Reecy, R. W. Thompson, L. L. Lowe, J. M. McClung, and J. A. Carson Lactate dehydrogenase expression at the onset of altered loading in rat soleus muscle J Appl Physiol, October 1, 2004; 97(4): 1424 - 1430. [Abstract] [Full Text] [PDF]

				I. A. Barash, L. Mathew, A. F. Ryan, J. Chen, and R. L. Lieber Rapid muscle-specific gene expression changes after a single bout of eccentric contractions in the mouse Am J Physiol Cell Physiol, February 1, 2004; 286(2): C355 - C364. [Abstract] [Full Text]

				Y.-W. Chen, M. J. Hubal, E. P. Hoffman, P. D. Thompson, and P. M. Clarkson Molecular responses of human muscle to eccentric exercise J Appl Physiol, December 1, 2003; 95(6): 2485 - 2494. [Abstract] [Full Text]

				L. Bey, N. Akunuri, P. Zhao, E. P. Hoffman, D. G. Hamilton, and M. T. Hamilton Patterns of global gene expression in rat skeletal muscle during unloading and low-intensity ambulatory activity Physiol Genomics, April 16, 2003; 13(2): 157 - 167. [Abstract] [Full Text] [PDF]

				W. J. Lee, J. McClung, G. A. Hand, and J. A. Carson Overload-induced androgen receptor expression in the aged rat hindlimb receiving nandrolone decanoate J Appl Physiol, March 1, 2003; 94(3): 1153 - 1161. [Abstract] [Full Text] [PDF]

				Z. Yan, S. Choi, X. Liu, M. Zhang, J. J. Schageman, S. Y. Lee, R. Hart, L. Lin, F. A. Thurmond, and R. S. Williams Highly Coordinated Gene Regulation in Mouse Skeletal Muscle Regeneration J. Biol. Chem., February 28, 2003; 278(10): 8826 - 8836. [Abstract] [Full Text] [PDF]

				J. M. Reecy, S. A. Miller, and M. Webster Recent Advances That Impact Skeletal Muscle Growth and Development Research J Anim Sci, January 1, 2003; 81(13_suppl_1): E1 - 8. [Abstract] [Full Text] [PDF]

				B. S. Tseng, P. Zhao, J. S. Pattison, S. E. Gordon, J. A. Granchelli, R. W. Madsen, L. C. Folk, E. P. Hoffman, and F. W. Booth Regenerated mdx mouse skeletal muscle shows differential mRNA expression J Appl Physiol, August 1, 2002; 93(2): 537 - 545. [Abstract] [Full Text] [PDF]

This Article Full Text (PDF) All Versions of this Article: 16/2/207 01-0544fjev1    most recent Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by CARSON, J. A. Articles by REECY, J. M. Search for Related Content PubMed PubMed Citation Articles by CARSON, J. A. Articles by REECY, J. M.