Optimization of Culture Conditions for Determining Protein Degradation in Myoblasts Using Extracts of Adult Bovine Muscle Treated with Muscle Enhancing Compounds1

The objective of this study was to optimize a methodology in which muscle extracts from animals treated with growth agents are added to growth media and then applied to muscle cell cultures to determine effects of growth agents on indirect cellular protein degradation. Experiments were designed as a 2 × 2 × 2 × 2 factorial arrangement of cell type (C2C12 myoblasts or primary bovine muscle cell cultures), growth medium [(Dulbecco's Modified Eagle's Medium (DMEM)or skeletal muscle cell basal medium (SKBM)], muscle extract medium [potassium phosphate buffer (KPB) or prerigor extraction buffer], and β-adrenergic agonist treatments of the bovine (control or treated) to determine differences in cellular protein degradation. There was a treatment media × β-adrenergic agonist treatment × cell type interaction (P < 0.0001). C2C12 myoblasts with the β-adrenergic agonist in DMEM media had less protein degradation than controls in DMEM (P < 0.05). Primary bovine muscle cell cultures treated with the β-adrenergic agonist treatment in DMEM media had greater protein degradation than did controls in DMEM (P < 0.05). However, primary bovine muscle cell cultures treated with the β-adrenergic agonist treatment in SKBM media had reduced protein degradation than did controls in SKBM (P < 0.05). There was a treatment media × β-adrenergic agonist treatment × muscle cell extraction buffer interaction (P = 0.04). The β-agonist treatment decreased protein degradation when muscle samples were extracted with KPB or prerigor extraction buffer and made with SKBM culture media (P < 0.05). The results indicated SKBM and C2C12 myoblasts had the most consistent differences for showing relative β-adrenergic agonist treatment effects on indirect protein degradation.