Blunted hypertrophic response in aged skeletal muscle is associated with decreased ribosome biogenesis

Abstract

The ability of skeletal muscle to hypertrophy in response to a growth stimulus is known to be compromised in older individuals. We hypothesized that a change in the expression of protein-encoding genes in response to a hypertrophic stimulus contributes to the blunted hypertrophy observed with aging. To test this hypothesis, we determined gene expression by microarray analysis of plantaris muscle from 5- and 25-mo-old mice subjected to 1, 3, 5, 7, 10, and 14 days of synergist ablation to induce hypertrophy. Overall, 1,607 genes were identified as being differentially expressed across the time course between young and old groups; however, the difference in gene expression was modest, with cluster analysis showing a similar pattern of expression between the two groups. Despite ribosome protein gene expression being higher in the aged group, ribosome biogenesis was significantly blunted in the skeletal muscle of aged mice compared with mice young in response to the hypertrophic stimulus (50% vs. 2.5-fold, respectively). The failure to upregulate pre-47S ribosomal RNA (rRNA) expression in muscle undergoing hypertrophy of old mice indicated that rDNA transcription by RNA polymerase I was impaired. Contrary to our hypothesis, the findings of the study suggest that impaired ribosome biogenesis was a primary factor underlying the blunted hypertrophic response observed in skeletal muscle of old mice rather than dramatic differences in the expression of protein-encoding genes. The diminished increase in total RNA, pre-47S rRNA, and 28S rRNA expression in aged muscle suggest that the primary dysfunction in ribosome biogenesis occurs at the level of rRNA transcription and processing.

Keywords: age; hypertrophy; microarray; ribosome biogenesis; skeletal muscle.

Fig. 1.

Aged skeletal muscle demonstrates blunted hypertrophic growth following mechanical overload. Normalized (to body weight) plantaris muscle weight from 25-mo-old mice showed impaired hypertrophy compared with young mice following 14 days of synergist ablation (SA-14). *Significant age-effect; †significant increase relative to sham (P < 0.05).

Fig. 2.

Hierarchical clustering of differentially expressed genes in response to synergist ablation. The expression pattern of genes within cluster 12 demonstrated the most significant effect of age during hypertrophic growth. The majority of genes within this cluster belonged to either the major urinary protein family or the serine (or cysteine) peptidase inhibitor family. Solid lines indicate young animals; dashed lines indicate old animals.

Fig. 3.

Increased expression of ribosomal proteins during hypertrophy with old age. A: geometric mean expression of all differentially expressed ribosomal protein genes following synergist ablation. B: Rpl11 increases earlier during mechanical overload in both young and old animals; however, the aged animals maintain higher expression levels at the later time points following synergist ablation. C: RT-PCR confirmed a significant increase in Rpl11 expression in old animals following 10 days of synergist ablation (SA-10), which did not occur in young animals. *Significant age-effect; †significant increase relative to sham (P < 0.05).

Fig. 4.

Increased ribosome content during hypertrophy is blunted in aged animals. Total RNA content, of which >85% is ribosomal RNA, increased in response to synergist ablation (SA), indicating increased ribosome biogenesis. Conversely, skeletal muscle of old animals demonstrated a compromised ability to increase total RNA content in response to synergist ablation. Old animals demonstrated a delayed increase in RNA content, not occurring until at SA10 and SA14. Furthermore, at SA5 through SA14, RNA content was significantly lower in the old animals relative to the young animals. *Significant age-effect at that time-point; †significant increase from sham (P < 0.05).

Fig. 5.

Increased pre-47S and 28S rRNA expression during muscle hypertrophy are attenuated in old animals. A: pre-47S rRNA expression, a readout of polymerase I activity, increased in response to synergist ablation in young animals after 3 and 7 days of synergist ablation with old animals only showing a modest increase at day 3. Furthermore, pre-47S expression was significantly higher in young animals compared with old animals at both days 3 and 7 of synergist ablation. B: 28S rRNA expression increased in response to synergist ablation at from day 3 through day 10 in young animals, whereas it increased in old animals only at day 10. 28S expression was significantly attenuated at days 3 and 7 in old animals. *Significant age-effect at that time-point; †significant increase from sham (P < 0.05).