The effect of new proteasome inhibitors, belactosin A and C, on protein metabolism in isolated rat skeletal muscle.

The proteasome inhibitors are used as research tools to study of the ATP-dependent ubiquitin-proteasome system. Some of them are at present undergoing clinical trials to be used as therapeutic agents for cancer or inflammation. These diseases are often accompanied by muscle wasting. We herein demonstrate findings about new proteasome inhibitors, belactosin A and C, and their direct effect on protein metabolism in rat skeletal muscle. M. soleus (SOL) and m. extensor digitorum longus (EDL) were dissected from both legs of male rats (40-60 g) and incubated in a buffer containing belactosin A or C (30 microM) or no inhibitor. The release of amino acids into the medium was estimated using high performance liquid chromatography to calculate total and myofibrillar proteolysis. Chymotrypsin-like activity (CTLA) of proteasome and cathepsin B, L activity were determined by fluorometric assay. Protein synthesis and leucine oxidation were detected using specific activity of L-[1-14C] leucine added to medium. Inhibited and control muscles from the same rat were compared using paired t-test. The results indicate that after incubation with both belactosin A and C total proteolysis and CTLA of proteasome decreased while cathepsin B, L activity did not change in both SOL and EDL. Leucine oxidation was significantly enhanced in SOL, protein synthesis decreased in EDL. Myofibrillar proteolysis was reduced in both muscles in the presence of belactosin A only. In summary, belactosin A and C affected basic parameters of protein metabolism in rat skeletal muscle. The response was both muscle- and belactosin-type-dependent.

The effect of new proteasome inhibitors, belactosin A and C, on protein metabolism in isolated rat skeletal muscle

The proteasome inhibitors are used as research tools to study of the ATP-dependentubiquitin-proteasome system. Some of them are at present undergoing clinicaltrials to be used as therapeutic agents for cancer or inflammation. These diseases areoften accompanied by muscle wasting. We herein demonstrate findings about newproteasome inhibitors, belactosin A and C, and their direct effect on protein metabolismin rat skeletal muscle. M. soleus (SOL) and m. extensor digitorum longus(EDL) were dissected from both legs of male rats (40-60g) and incubated in a buffercontaining belactosin A or C (30 ìM) or no inhibitor. The release of amino acids intothe medium was estimated using high performance liquid chromatography to calculatetotal and myofibrillar proteolysis. Chymotrypsin-like activity (CTLA) of proteasomeand cathepsin B, L activity were determined by fluorometric assay. Proteinsynthesis and leucine oxidation were detected using specific activity of L-[1-14C]leucine added to medium. Inhibited and control muscles from the same rat were comparedusing paired t-test. The results indicate that after incubation with both belactosinA and C total proteolysis and CTLA of proteasome decreased while cathepsinB, L activity did not change in both SOL and EDL. Leucine oxidation was significantlyenhanced in SOL, protein synthesis decreased in EDL. Myofibrillar proteolysiswas reduced in both muscles in the presence of belactosin A only. In summary,belactosin A and C affected basic parameters of protein metabolism in rat skeletalmuscle. The response was both muscle- and belactosin-type-dependent(AU)

Effect of beta-hydroxy-beta-methylbutyrate (HMB) on protein metabolism in whole body and in selected tissues.

Beta-hydroxy-beta-methylbutyrate (HMB) is a leucine metabolite with protein anabolic effect. The aim of the study was to examine the role of exogenous HMB on leucine and protein metabolism in whole body and selected tissues. Rats were administered by HMB (0.1 g/kg b.w.) or by saline. The parameters of whole-body protein metabolism were evaluated 24 h later using L-[1-14C]leucine and L-[3,4,5-3H]phenylalanine. Changes in proteasome dependent proteolysis and protein synthesis were determined according the "chymotrypsin-like" enzyme activity and labeled leucine and phenylalanine incorporation into the protein. A decrease in leucine clearance and whole-body protein turnover (i.e., a decrease in whole-body proteolysis and protein synthesis) was observed in HMB treated rats. Proteasome-dependent proteolysis decreased significantly in skeletal muscle, changes in heart, liver, jejunum, colon, kidney, and spleen were insignificant. Decrease in protein synthesis was observed in the heart, colon, kidney, and spleen, while an increase was observed in the liver. There were no significant changes in leucine oxidation. We conclude that protein anabolic effect of HMB in skeletal muscle is related to inhibition of proteolysis in proteasome. Alterations in protein synthesis in visceral tissues may affect several important functions and the metabolic status of the whole body.