Effects of proteasome inhibitors MG132, ZL3VS and AdaAhx3L3VS on protein metabolism in septic rats.

Proteasome inhibitors are novel therapeutic agents for the treatment of cancer and other severe disorders. One of the possible side effects is influencing the metabolism of proteins. The aim of our study was to evaluate the influence of three proteasome inhibitors MG132, ZL(3)VS and AdaAhx(3)L(3)VS on protein metabolism and leucine oxidation in incubated skeletal muscle of control and septic rats. Total proteolysis was determined according to the rates of tyrosine release into the medium during incubation. The rates of protein synthesis and leucine oxidation were measured in a medium containing L-[1-(14)C]leucine. Protein synthesis was determined as the amount of L-[1-(14)C]leucine incorporated into proteins, and leucine oxidation was evaluated according to the release of (14)CO(2) during incubation. Sepsis was induced in rats by means of caecal ligation and puncture. MG132 reduced proteolysis by more than 50% and protein synthesis by 10-20% in the muscles of healthy rats. In septic rats, proteasome inhibitors, except ZL(3)VS, decreased proteolysis in both soleus and extensor digitorum longus (EDL) muscles, although none of the inhibitors had any effect on protein synthesis. Leucine oxidation was increased by AdaAhx(3)L(3)VS in the septic EDL muscle and decreased by MG132 in intact EDL muscle. We conclude that MG132 and AdaAhx(3)L(3)VS reversed protein catabolism in septic rat muscles.

Acute effects of acidosis on protein and amino acid metabolism in perfused rat liver.

Acidosis is frequently associated with protein wasting and derangements in amino acid metabolism. As its effect on protein metabolism is significantly modulated by other abnormal metabolic conditions caused by specific illnesses, it is difficult to separate out the effects on protein metabolism solely due to acidosis. The aim of the present study was to evaluate, using a model of isolated perfused rat liver, the direct response of hepatic tissue to acidosis. We have compared hepatic response to perfusion with a solution of pH 7.2 and 7.4 (controls). Parameters of protein and amino acid metabolism were measured using both recirculation and single-pass technique with 4,5-[3H]leucine, [1-14C]leucine and [1-14C]ketoisocaproate (ketoleucine) as tracers and on the basis of difference of amino acid levels in perfusion solution at the beginning and end of perfusion. In liver perfused with a solution of pH 7.2, we observed higher rates of proteolysis, protein synthesis, amino acid utilization and urea production. Furthermore, the liver perfused with a solution of pH 7.2 released a higher amount of proteins to perfusate than the liver perfused with a solution of pH 7.4. Enhanced decarboxylation of ketoisocaproate in liver perfused by a solution of a lower pH indicates increased catabolism of branched-chain amino acids (leucine, valine and isoleucine), decreased reamination of branched-chain keto acids to corresponding essential amino acids and increased ketogenesis from leucine.

Acute effects of decreased glutamine supply on protein and amino acid metabolism in hepatic tissue: a study using isolated perfused rat liver.

Glutamine deficiency, a common finding in severe illness, has a negative influence on immune status, protein metabolism, and disease outcome. In several studies, a close relationship between glutamine, branched-chain amino acid (BCAA), and protein metabolism was demonstrated. The aim of the present study was to investigate the effect of glutamine deficiency on amino acid and protein metabolism in hepatic tissue using a model of isolated perfused rat liver (IPRL). Parameters of protein metabolism and amino acid metabolism were measured using both recirculation and single pass technique with L-[1-(14)C]leucine and [1-(14)C]ketoisocaproate (KIC) as a tracer. Glutamine concentration in perfusion solution was 0.5 mmol/L in control and 0 mmol/L in the glutamine-deficient group. The net release of glutamine (about 11 micromol/g/h) and higher net uptake of most of the amino acids was observed in the glutamine-deficient group. There was an insignificant effect of lack of glutamine on hepatic protein synthesis, proteolysis, and the release of urea. However, significantly lower release of proteins by the liver perfused with glutamine-deficient solution was observed. The lack of glutamine in perfusion solution caused a significant decrease in leucine oxidation (6.66 +/- 1.04 v 13.67 +/- 2.38, micromol/g dry liver/h, P <.05) and an increase in KIC oxidation (163.7 +/- 16.5 v 92.0 +/- 12.9 micromL/g dry liver/h, P <.05). We conclude that decreased delivery of glutamine to hepatic tissue activates glutamine synthesis, decreases resynthesis of essential BCAA from branched-chain keto acids (BCKA), increases catabolism of BCKA, and has an insignificant effect on protein turnover in hepatic tissue.

[Method of measurement of protein metabolism in isolated skeletal muscle of the rat]. / Metoda merení parametru metabolismu proteinu na izolovaných kosterních svalech laboratorního potkana.

The aim of our present research is to evaluate the direct effects of humoral factors on protein metabolism. For this purpose we have adopted a method that enables measuring of parameters of protein and amino acid metabolism in isolated m. soleus and m. extensor digitorum longus of the rat. The rate of tyrosine release was used to estimate protein breakdown. The oxidation of leucine was evaluated by the rate of 14CO2 production from [1-14C]leucine. Incorporation of [1-14C]leucine into muscle proteins was used to estimate protein synthesis. The metabolic state of the muscles was evaluated by measuring changes in ATP, ADP, AMP levels, and calculated energy charge.