Proteomic Analysis Of Protein Hyper-metabolism In Skeletal Muscles Of Rats Induced By Sepsis

One of the most pronounced metabolic changes during sepsis is the catabolic response in skeletal muscle,characterized by increased protein breakdown.To define the skeletal muscle changes triggering muscle atrophy in sepsis induced by cecal ligation puncture(CLP),a proteomic analysis was performed by comparing extensor digitorum longus(EDL)and soleus muscles of sham-CLP groups to the corresponding muscles of CLP groups at 1,3,5 and 7 days post operation.Muscle mass measurements showed that the mass of EDL muscles of CLP rats significantly reduced from day 1 compared to that of sham-CLP rats(P＜0.05), while the significantly reduced mass of soleus muscles was observed from day 3 (P＜0.05).Sixteen spots were detected significantly altered in EDL muscles in the CLP compared to the sham-CLP rats by two-dimensional electrophoresis(2-DE), twelve proteins were identified by mass spectrometry,meanwhile,seventeen spots were significantly changed in soleus muscles and 13 proteins were identified. These identified proteins can be catalogued according to their functions as contractile apparatus proteins,cytoskeleton proteins,metabolic enzymes, oxidative phosphorylation proteins,oxidate-redox proteins,stress response proteins and apoptosis proteins.The significant changes in voltage-dependent anion channel 2(VDAC 2),αB-crystallin(Cry AB)and cofilin 2 were verified by Western blot analysis.VDAC 2 was downregulated in both EDL and soleus muscles of CLP rats,while cofilin 2 was upregulated in the two muscles after CLP. Cry AB was differently regulated in the two muscles.Following CLP,the protein levels of Cry AB in EDL muscles were upregulated from day 1 but downregulated at 1 day in soleus muscles.Using cultured L6 myotubes,we showed that dexamethasone(Dexa)induced protein degradation in vitro could be inhibited by prior heat shock treatment or in the presence of insulin-like growth factorⅠ(IGF-Ⅰ). We also found that Dexa suppressed VDAC 2 protein expression,but upregulated the expression of Cry AB and cofilin 2.Both heat shock and IGF-Ⅰenhanced the protein levels of VDAC 2,Cry AB and cofilin 2 and prevented the downregulation of VDAC 2 by Dexa.These results from the experiments on rats and cultured L6 myotubes indicate that:(1)the imbalance between oxidation and reduction induced by sepsis is one of the important factors for apoptosis;(2) increased proteolysis of skeletal muscle in sepsis may be related to the activation of mitochondria-initiated apoptotic pathways involving VDAC 2;(3)the upregulation of Cry AB and cofilin 2 in skeletal muscles appears to be an autoprotective mechanism for stabilizing myofibrillar components during sepsis or Dexa treatment and compared to soleus muscles,EDL muscles contain lower amounts of Cry AB,which might contribute to the phenomenon that EDL muscles are more susceptive to proteolysis in sepsis.