Transcriptional Regulation Of Apoptosis In Atrophied Fast Twitch Plantaris Muscles Of Young Adult And Aged Rats

Abstract

Hindlimb suspension in rats is a model of simulated microgravity that has been used to investigate skeletal muscle adaptations during non-weight-bearing conditions. The loss of muscle mass occurs quickly in hindlimb muscles that contain a high percentage of slow myosin; however, muscles with a high percentage of fast myosin containing fibers usually have less severe atrophy during unloading. Although apoptosis appears to explain part of the atrophy in slow muscles (e.g., soleus muscles), it is not known if pro-apoptotic markers are elevated in fast muscles (e.g., plantaris muscles) where atrophy is less severe. PURPOSE Because inhibitor of differentiation protein-2 (Id2) has been implicated in apoptosis and muscle atrophy during aging, we examined if Id2 and pro-apoptotic gene levels were increased during hypokinesia in the rat plantaris after hindlimb suspension. METHODS Young adult (n=8, 9 mo.) and aged (n=8, 33 mo.) Fischer Brown Norway rats were hindlimb suspended for 2 weeks to allow atrophy of the hindlimb musculature. Following suspension, rats were euthanized, the plantaris muscles were excised, and total RNA was isolated. RT-PCR for AIF, Apaf-1, Bax, Caspase-9, and Id2 genes was performed with the amplification products normalized to the 18S gene. The young adult and aged animals were compared to aged-matched controls using ANOVA with significance set at p<0.05 RESULTS Muscle wet weight significantly decreased by 17.1% in the aged animals, compared to only 7.8% in young animals. Compared to young adult rats, aged rats had significantly elevated mRNA levels of Apaf-1 (53%) in response to HS, with similar increases in AIF mRNA (52% vs. 53%) in both young and aged animals following HS. The increases in the mRNA for Caspase-9 or Bax were similar in muscles from young and aged rats following HS. When normalized to control muscles, the change in Id2 mRNA was less in unloaded muscles from young adult rats (30%) as compared with aged rats (52%), suggesting that an elevation of Id2 transcripts might contribute to the exaggerated muscle atrophy observed in response to HS in aged plantaris muscle. Significant positive correlations were observed between Id2 and Apaf-1 (p<0.01), Id2 and Caspase-9 (p<0.05), Bax and AIF (p<0.01) Bax and Caspase-9 (p<0.05). CONCLUSIONS These data suggest that unloading induced muscle atrophy involves an altered gene expression for pro-apoptotic transcripts even in muscles where atrophy is less severe, but aging exacerbates the increase in expression of these transcripts. Supported by NIH R01AG021530.