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

Abstract

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.