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. 2009 Oct;297(4):E935-40.
doi: 10.1152/ajpendo.00179.2009. Epub 2009 Aug 4.

Relation Between Extent of Myostatin Depletion and Muscle Growth in Mature Mice

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Free PMC article

Relation Between Extent of Myostatin Depletion and Muscle Growth in Mature Mice

Stephen Welle et al. Am J Physiol Endocrinol Metab. .
Free PMC article

Abstract

Myostatin is a negative regulator of muscle growth and fiber size. Changes in myostatin expression might contribute to changes in muscle mass associated with various conditions, and reducing the amount of active myostatin is a potential strategy for preventing or reversing muscle atrophy. The present study was done to determine the extent to which myostatin levels must decline to induce growth of mature muscles. Myostatin expression was reduced by activating Cre recombinase in adult mice with floxed myostatin genes. The duration of Cre activation varied from 1 to 6 wk, and the residual myostatin mRNA expression after Cre activation varied from 3 to 63% of the normal level. Promyostatin levels declined in parallel with myostatin mRNA. There was no increase in muscle mass over the 3 mo following Cre activation if residual myostatin expression was >or=40% of normal. In mice with <40% of normal myostatin expression, muscle mass increased in proportion to the extent of myostatin depletion. In mice with <or=10% of normal myostatin expression, muscle mass increased approximately 25%. Myostatin depletion increased myonuclear domain volumes and the ratio of RNA to myonuclei probably by enhancing DNA transcription rather than by inhibiting RNA decay. There was no evidence that maintenance of the hypertrophy during chronic myostatin deficiency requires altered activity of Akt/mTOR or p38 MAPK signaling pathways. These data suggest that anabolic therapies based on reducing the concentration of active myostatin will be effective only if a very large proportion of the myostatin is removed or inactivated.

Figures

Fig. 1.
Fig. 1.
Myostatin mRNA expression in gastrocnemius muscles before exposure to tamoxifen (tmx) and 3 mo after 1, 2, or 6 wk of tmx ingestion. There were no significant differences across conditions in the Mstn/GAPDH gene expression ratios in Mstnw/w mice, so data from these mice were pooled to define the normal ratio (open bar). Gray bars show values from Mstnf/f mice. P refers to comparison with mean value of Mstnw/w mice. Bars represent mean values. Error bars represent 1 SE.
Fig. 2.
Fig. 2.
Loss of promyostatin is proportional to loss of myostatin mRNA expression. Immunoblots of samples taken from muscles of tmx-treated Mstnf/f (f) and Mstnw/w (w) mice after extraction from equal amounts of total protein with concanavalin A. The %mRNA and %protein values were calculated by defining 100% as the average value for Mstnw/w samples on a particular blot. Scatter plot with line of identity summarizes data shown under blots.
Fig. 3.
Fig. 3.
Muscle fiber cross-sectional areas and myonuclear domain size in quadriceps muscles of myostatin-deficient (Mstnf/f; gray bars) and control mice (Mstnw/w; open bars) at 9 mo of age. The mice were fed tmx to activate Cre recombinase activity for three 2-wk periods starting at 4 mo of age. Cryosections of quadriceps muscles were obtained from 3 mice of each genotype, and then 5–7 microscope fields/section (each with ∼30–50 fibers, >200 fibers/mouse) were selected by a technician who did not know the genotypes of the mice. The box plots show the distribution [median, 25th and 75th percentiles (box borders), 10th and 90th percentiles (whiskers), and highest/lowest values (•)] of mean fiber cross-sectional areas and mean myonuclear domain sizes in the individual microscope fields.
Fig. 4.
Fig. 4.
Relation between myostatin expression and muscle mass 3 mo after induction of Cre recombinase activity with tmx. Open symbols illustrate the variability in muscle mass (normalized to body mass) and myostatin gene expression (normalized to GAPDH gene expression) in Mstnw/w mice, in which Cre recombinase should have no specific effect on myostatin gene expression. Filled symbols illustrate relation between decline in myostatin mRNA induced by Cre recombinase-mediated knockout of floxed myostatin exon 3 and subsequent muscle growth in Mstnf/f mice. Circles (Rosa26-CreERT2 transgene) represent data from the present study, and triangles (CAGG-CreER transgene) represent data published previously (39).

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