Denervation stimulates apoptosis but not Id2 expression in hindlimb muscles of aged rats.

Inhibitors of differentiation (Id) proteins are repressors of myogenic regulatory factors and have been implicated in apoptosis and muscle atrophy during aging. Indeed, we have previously found that Id levels are elevated in muscles from old rodents, possibly as a consequence of loss of alpha-motoneurons during senescence. To determine if Id2 proteins increase after denervation and if this is accompanied by increased apoptosis in aged as compared with adult animals, the gastrocnemius and soleus muscles were denervated in 1 limb of Fischer 344 x Brown Norway rats aged 9 months (adult, n = 12) and 33 months (aged, n = 9), while the contralateral limb served as the intra-animal control. After 14 days, the muscles in each limb were removed. The levels of Id1, Id2, and Id3 mRNA and protein were significantly greater in muscles of old as compared with young adult rats. Denervation, however, did not significantly increase Id1, Id2, and Id3 mRNA in soleus or gastrocnemius muscles from either young or old rats. Also Id2 protein levels were similar in denervated and control muscles from young adult and old rats. In young adult rats only, denervation induced an increase in Id1 and Id3 protein levels in both the soleus (Id1 113%; Id3 900%) and gastrocnemius (Id1 86%; Id3 80%). Denervation induced a significant increase in caspase 8 in both soleus and gastrocnemius muscles from young (101% and 147%, respectively) and old rats (167% and 190%, respectively). Bax protein levels, as estimated by western blots, increased by 726% and 1087% after denervation in the soleus and by 368% and 49% in the gastrocnemius muscles of young and old rats, respectively. The data suggest that the denervation-induced muscle loss was at least partly due to apoptosis as indicated by elevated caspase 8 and Bax levels in denervated muscles. While Id2 may have a role in aging-induced sarcopenia, Id2 does not appear to directly regulate apoptosis during denervation. The elevated Id expression in muscles from aged animals is therefore not a direct consequence of loss of alpha-motoneurons during senescence.

Id2 expression during apoptosis and satellite cell activation in unloaded and loaded quail skeletal muscles.

Inhibitor of differentiation-2 (Id2) is a basic helix-loop-helix protein that acts as a negative regulator of the myogenic regulatory transcription factor family, but Id2 has also been implicated in apoptosis in several cell lines. In this study, we tested the hypothesis that Id2 has a role in both apoptosis-associated muscle atrophy and muscle hypertrophy. A weight corresponding to 12% of the body weight was attached to one wing of Japanese quail to induce hypertrophy in the patagialis (PAT) muscle. Birds in group 1 were killed after 5 (n = 8), 7 (n = 10), or 14 days (n = 10) of loading. The left wing was loaded for 14 days in group 2 birds, and then the weight was removed and the PAT was examined after 7 (n = 10), 14 (n = 10), or 21 (n = 5) days of unloading. A time-released bromodeoxyuridine (BrdU) pellet was implanted subcutaneously with wing weighting to identify activated satellite cells during loading. The left wing was loaded for 14 days, unloaded for 14 days, and then the weight was reattached for a subsequent 7 (n = 10) or 14 days (n = 10) in group 3 birds. BrdU was implanted on the second loading phase in this group. Id2 mRNA as measured by kinetic PCR increased by 3.9-, 2.7-, and 1.6-fold, relative to control levels after 7, 14, and 21 days of unloading (group 2). Id2 protein as estimated by Western blots increased by 1.5-, 1.4-, and 0.75-fold after 7, 14, and 21 days of unloading (group 2). Muscle unloading induced apoptosis, because poly(ADP-ribose) polymerase-(PARP)-positive nuclei increased and caspase 8 levels increased by 2.6- and 1.7-fold after 7 or 14 days of unloading, respectively (group 2). Although BrdU-positive nuclei increased during loading (groups 1 and 3), 50% failed to survive during unloading (group 2). Id2 mRNA increased by 2.2- and 1.8-fold after 5 and 7 days of loading, respectively, but decreased to control levels by 14 days of loading in group 1. Id2 protein levels increased 2.1-fold after 5 days of loading (group 1). In contrast, Id2 did not increase in reloaded muscles of group 3 birds. These data suggest that Id2 may have a role in apoptosis-associated atrophy of skeletal muscles, but its role in muscle hypertrophy is less clear.