Prolonged unloading of rat soleus muscle causes distinct adaptations of the gene profile (original) (raw)
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Analysis of altered gene expression in rat soleus muscle atrophied by disuse
Journal of Cellular Biochemistry, 2001
The present study involved a global analysis of genes whose expression was modi®ed in rat soleus muscle atrophied after hindlimb suspension (HS). HS muscle unloading is a common model for muscle disuse that especially affects antigravity slow-twitch muscles such as the soleus muscle. A cDNA cloning strategy, based on suppression subtractive hybridization technology, led to the construction of two normalized soleus muscle cDNA libraries that were subtracted in opposite directions, i.e., atrophied soleus muscle cDNAs subtracted by control cDNAs and vice versa. Differential screening of the two libraries revealed 34 genes with altered expression in HS soleus muscle, including 11 novel cDNAs, in addition to the 2X and 2B myosin heavy chain genes expressed only in soleus muscles after HS. Gene up-and down-regulations were quanti®ed by reverse Northern blot and classical Northern blot analysis. The 25 genes with known functions fell into seven important functional categories. The homogeneity of gene alterations within each category gave several clues for unraveling the interplay of cellular events implied in the muscle atrophy phenotype. In particular, our results indicate that modulations in slow-and fast-twitch-muscle component balance, the protein synthesis/secretion pathway, and the extracellular matrix/cytoskeleton axis are likely to be key molecular mechanisms of muscle atrophy. In addition, the cloning of novel cDNAs underlined the ef®ciency of the chosen technical approach and gave novel possibilities to further decipher the molecular mechanisms of muscle atrophy.
Transcriptional reprogramming and ultrastructure during atrophy and recovery of mouse soleus muscle
Physiological Genomics, 2004
This study investigated the use of the hindlimb suspension (HS) and reloading model of mice for the mapping of ultrastructural and gene expressional alterations underlying loaddependent muscular adaptations. Mice were hindlimb suspended for seven days or kept as controls (C, n=12). Soleus muscles were harvested after HS (HS7, n=23) or after resuming ambulatory cage activity (reloading) for either one (R1, n=13) or seven days (R7, n=9). Using electron microscopy, a reduction in mean fiber area (-37%) and in capillary-to-fiber ratio (from 1.83 to 1.42) was found for HS7. Subsequent reloading caused an increase in interstitial cells (+96%) and in total capillary length (+57%) while mean fiber area and capillary-to-fiber ratio did not significantly change compared to HS. Total RNA in the soleus muscle was altered with both HS (-63%) and reloading (+108% in R7 compared to C). This is seen as an important adaptive mechanism. Gene expression alterations were assessed by a muscle-specific low-density cDNA microarray. The transcriptional adjustments indicate an early increase of myogenic factors during reloading together with an overshoot of contractile (MyHC I and IIa) and metabolic (glycolytic and oxidative) mRNA amounts and suggest mechano-sensitivity of factors keeping the sarcomeres in register (desmin, titin, integrin β1).
Transcriptional reprogramming during reloading of atrophied rat soleus muscle
AJP: Regulatory, Integrative and Comparative Physiology, 2005
The hypothesis was tested that differential, coregulated transcriptional adaptations of various cellular pathways would occur early with increased mechanical loading of atrophied skeletal muscle and relate to concurrent damage of muscle fibers. Atrophy and slow-to-fast fiber transformation of rat soleus muscle was provoked by 14 days of hindlimb suspension (HS). Subsequent reloading of hindlimbs caused a fourfold increase in the percentage of muscle fibers, demonstrating endomysial tenascin-C staining. Five days after reloading, when 10% of the fibers were damaged, the normal muscle weight and slow-type fiber percentage were reestablished. Microarray analysis revealed major, biphasic patterns of gene expressional alterations with reloading that distinguish between treatments and gene ontologies. Transcript levels of factors involved in protein synthesis and certain proteasomal mRNAs were increased after 1 day of reloading and correlated to the percentage of fibers surrounded by tena...
Acute molecular response of mouse hindlimb muscles to chronic stimulation
AJP: Cell Physiology, 2009
LaFramboise WA, Jayaraman RC, Bombach KL, Ankrapp DP, Krill-Burger JM, Sciulli CM, Petrosko P, Wiseman RW. Acute molecular response of mouse hindlimb muscles to chronic stimulation. tion of the mouse hindlimb via the sciatic nerve was performed for a 4-h period to investigate acute muscle gene activation in a model of muscle phenotype conversion. Initial force production (1.6 Ϯ 0.1 g/g body wt) declined 45% within 10 min and was maintained for the remainder of the experiment. Force returned to initial levels upon study completion. An immediate-early growth response was present in the extensor digitorum longus (EDL) muscle (FOS, JUN, activating transcription factor 3, and musculoaponeurotic fibrosarcoma oncogene) with a similar but attenuated pattern in the soleus muscle. Transcript profiles showed decreased fast fiber-specific mRNA (myosin heavy chains 2A and 2B, fast troponins T3 and I, ␣-tropomyosin, muscle creatine kinase, and parvalbumin) and increased slow transcripts (myosin heavy chain-1/slow, troponin C slow, and tropomyosin 3y) in the EDL versus soleus muscles. Histological analysis of the EDL revealed glycogen depletion without inflammatory cell infiltration in stimulated versus control muscles, whereas ultrastructural analysis showed no evidence of myofiber damage after stimulation. Multiple fiber typespecific transcription factors (tea domain family member 1, nuclear factor of activated T cells 1, peroxisome proliferator-activated receptor-␥ coactivator-1␣ and -, circadian locomotor output cycles kaput, and hypoxia-inducible factor-1␣) increased in the EDL along with transcription factors characteristic of embryogenesis (Kruppel-like factor 4; SRY box containing 17; transcription factor 15; PBX/knotted 1 homeobox 1; and embryonic lethal, abnormal vision). No established in vivo satellite cell markers or genes activated in our parallel experiments of satellite cell proliferation in vitro (cyclins A 2, B2, C, and E1 and MyoD) were differentially increased in the stimulated muscles. These results indicated that the molecular onset of fast to slow phenotype conversion occurred in the EDL within 4 h of stimulation without injury or satellite cell recruitment. This conversion was associated with the expression of phenotype-specific transcription factors from resident fiber myonuclei, including the activation of nascent developmental transcriptional programs.
Acta Physiologica, 2009
Aim: To determine the adaptive changes in several molecules regulating muscle hypertrophy and atrophy after unloading, we examined whether unilateral lower limb suspension changes the mRNA and protein levels of SRF-linked (RhoA, RhoGDI, STARS and SRF), myostatin-linked (myostatin, Smad2, Smad3 and FLRG) and Foxo-linked (P-Akt, Foxo1, Foxo3a and Atrogin-1) mediators. Methods: A single lower limb of each of eight healthy men was suspended for 20 days. Biopsy specimens were obtained from the vastus lateralis muscle pre-and post-suspension. Results: The volume of the vastus lateralis muscle was significantly decreased after unloading. The amount of RhoA, RhoGDI or SRF protein in the muscle was not significantly changed post-suspension. An RT-PCR semiquantitative analysis showed increased levels of myostatin mRNA but not Smad2, Smad3 or FLRG mRNA. Unloading did not elicit significant changes in the amount of p-Smad3 or myostatin protein in the muscle. The amount of p-Akt protein was markedly reduced in the unloaded muscle. Lower limb suspension did not influence the expression pattern of Foxo1, Foxo3a or Atrogin-1. Conclusion: Unloading inducing a mild degree of muscle atrophy may decrease p-Akt and increase myostatin but not SRF-linked mediators.
Journal of Applied Physiology, 2003
After cessation of hindlimb immobilization, which resulted in a 27-37% loss in soleus mass, the atrophied soleus muscle of young but not old rats regrows to its mass before treatment. We hypothesized that during remobilization the mRNA levels of growth potentiating factor(s) would be present in the soleus muscle of young (3- to 4-mo-old) but absent in old (30- to 31-mo-old) Fischer 344 × Brown Norway rats or that mRNAs for growth inhibitory factor(s) would be absent in young but present in old. Gene expression levels of >24,000 transcripts were determined by using Affymetrix RGU34A-C high-density oligonucleotide microarrays in soleus muscles at 3, 6, 10, and 30 days of remobilization after cessation of a 10-day period of hindlimb immobilization. Each muscle sample was applied to an independent set of arrays. Recovery-related differences were determined by using a three-factor ANOVA with a false discovery rate-adjustment of P = 0.01, which yielded 64 significantly different probe ...
Temporal effects of inactivty on myosin heavy chain gene expression in rat slow muscle
Muscle & Nerve, 2001
Myosin heavy chain (MHC) mRNA and protein profiles in adult rat soleus and adductor longus were determined after 4, 8, 15, 30, 60, and 90 days of spinal cord isolation (SI). SI results in complete neuromuscular inactivity while leaving the motoneuron-muscle fiber connections intact. From 15 to 90 days, type I MHC mRNA was significantly decreased, whereas type I MHC protein did not significantly decrease until 30 and 60 days in the soleus and adductor longus, respectively. However, in both muscles, slow MHC downregulation was offset by significant upregulation of the faster MHC isoforms, primarily IIx. From 60 to 90 days, type I MHC was almost completely replaced with faster isoforms at the mRNA and protein levels. Thus, chronic inactivity and unloading of slow rat hindlimb muscles shifted the MHC profile from predominately type I to type IIx MHC mRNA and protein.
European Journal of Applied Physiology and Occupational Physiology, 1999
We have used a new approach to study the eects of overload on skeletal muscle phenotype in mice. The method used avoids any traumatising contact with muscles and the in¯ammatory reaction that this may provoke. Blocks of lead embedded in silicone were inserted under the skin of the lower part of the back. After 1 month, a 17% hypertrophy was found to have occurred in the tonic soleus muscle, but no change was observed in the fast-twitch extensor digitorum longus (EDL) muscle. The main eects on the contractile properties of the soleus muscle were a decrease in the tetanic relaxation rate and a reduction in the maximal velocity of shortening. Immunohistological analysis of the soleus muscles revealed an increase in the proportion of ®bres that express myosin heavy chain (MHC) 1, from 54.2% to 73.9%, with a reduction in the proportion of MHC2a-positive ®bres, from 45.8% to 30.2%. These changes were accompanied by an increase in the proportion of ®bres that express the slow type of sarcoplasmic reticulum calcium pump (SERCA2a), from 61.8% to 84.7%. In EDL muscles, overload induced only minor changes. Thus, this method of overload affected the soleus muscle in particular. The observed changes in the control of muscle contraction were sig-ni®cantly larger than the changes in typical myo®brillar properties that were observed. These results indicate that there is a temporal dissociation between the relative expression of MHCs and SERCAs.