Effect of beta agonists on protein turnover in isolated chick skeletal and atrial muscle

Various beta-adrenergic agonists were found to inhibit rates of protein degradation and net protein breakdown in isolated chick extensor digitorum communis (EDC) and atrial muscles. Rates of protein synthesis were not altered by these compounds. The beta-agonist cimaterol inhibited rates of protein degradation in EDC muscles incubated with or without amino acids and insulin. Cimaterol also inhibited the increased proteolysis induced by injury to muscle or by incubating muscles at body temperature (42 degrees C) versus 37 degrees C. Thus, beta-agonists may help promote skeletal muscle accretion in vivo even under conditions of severe negative nitrogen balance by slowing muscle proteolysis.     

The effect of glutamine on protein turnover in chick skeletal muscle in vitro.

The effect of glutamine on the rates of protein synthesis and degradation was studied in isolated chick extensor digitorum communis muscles incubated in the presence of plasma concentrations of amino acids. Addition of 0.5-15 mM-glutamine increases (P less than 0.01) intracellular glutamine concentrations by 31-670%. There is a positive relationship (r = 0.975, P less than 0.01) between intracellular glutamine concentration and the rate of muscle protein synthesis measured by the incorporation of [3H]phenylalanine. The stimulating effect of 15 mM-glutamine on protein synthesis was decreased from 58 to 19% in muscles incubated in the absence of tyrosine. The rates of protein degradation, estimated from [3H]phenylalanine release from muscle proteins prelabelled in vivo, decreased (P less than 0.05) by 15-30% in the presence of 4-15 mM-glutamine when compared with muscles incubated in the presence of physiological concentrations of glutamine (0.5-1 mM). Glutamine concentrations ranging from 2 to 15 mM appear to have an overall anabolic effect on chick skeletal muscles incubated in vitro.     

The effect of ketone bodies on protein turnover in isolated skeletal muscle from the fed and fasted chick

1. The addition of 4 mM acetoacetate or DL-beta-hydroxybutyrate to the incubation medium decreased the rate of protein synthesis without influencing the rate of protein degradation in extensor digitorum communis (EDC) muscles from fed chicks and decreased the rates of protein synthesis and degradation in muscles from fasted chicks. 2. Ketone bodies markedly decreased intracellular concentrations of glutamine in EDC muscles from fed chicks by increasing glutamine oxidation. 3. The addition of 0.5 mM glutamine to incubation media containing 1.0 mM glutamine reversed the ketone body-induced decrease in intracellular glutamine concentration to the control value and blocked the inhibiting effect of ketone bodies on protein synthesis in skeletal muscles from fed chicks. 4. The addition of 5 mM pyruvate blocked the ability of ketone bodies to increase glutamine oxidation and prevented the associated decrease in intracellular glutamine concentration and the rate of protein synthesis in EDC muscles from fed chicks. 5. These results suggest that ketone bodies can act directly on skeletal muscle to inhibit the rate of protein synthesis in muscles from fed chicks by decreasing intracellular glutamine concentration by increasing its oxidation.     

The effect of protein depletion and repletion on muscle-protein turnover in the chick.

Rates of growth and protein turnover in the breast muscle of young chicks were measured in order to assess the roles of protein synthesis and degradation in the regulation of muscle mass. Rates of protein synthesis were measured in vivo by injecting a massive dose of L-[1-14C]valine, and rates of protein degradation were estimated as the difference between the synthesis rate and the growth rate of muscle protein. In chicks fed on a control diet for up to 7 weeks of age, the fractional rate of synthesis decreased from 1 to 2 weeks of age and then changed insignificantly from 2 to 7 weeks of age, whereas DNA activity was constant for 1 to 7 weeks. When 4-week-old chicks were fed on a protein-free diet for 17 days, the total amount of breast-muscle protein synthesized and degraded per day and the amount of protein synthesized per unit of DNA decreased. Protein was lost owing to a greater decrease in the rate of protein synthesis, as a result of the loss of RNA and a lowered RNA activity. When depleted chicks were re-fed the control diet, rapid growth was achieved by a doubling of the fractional synthesis rate by 2 days. Initially, this was a result of increased RNA activity; by 5 days, the RNA/DNA ratio also increased. There was no evidence of a decrease in the fractional degradation rate during re-feeding. These results indicate that dietary-protein depletion and repletion cause changes in breast-muscle protein mass primarily through changes in the rate of protein synthesis.     

Accelerated protein turnover in the skeletal muscle of dystrophic mice

The excretion of 3-methylhistidine increased in the urine of dystrophic mice C57BL/6J. The content of 3-methylhistidine residue decreased in the muscle proteins of dystrophic mice, but not in other organs. Methylated proteins in the skeletal muscle, actin and myosin, were partially purified from the dystrophic and control muscles. The amount of 3-methylhistidine residue in unit weight of the actin and myosin preparations was normal in dystrophic muscle. These three facts indicate that the turnover rates of actin and myosin are increased in the muscle of the dystrophic mice.     

An in vitro preparation of the extensor digitorum communis muscle from the chick (Gallus domesticus) for studies of protein turnover

1. Extensor digitorum communis (EDC) muscles from the chick synthesized and degraded proteins at linear rates, and maintained high levels of ATP, creatine phosphate and glycogen, for 9 hr of incubation. 2. The apparent viability of the EDC preparation was improved by the provision of a continuous supply of oxygen, glucose, insulin and amino acids at the levels found in chick plasma, and incubation at 33.5 degrees C. 3. Incubated muscles were in net negative protein balance; however, under optimal conditions, this represented only 0.03% of total protein/hr. Rates of protein synthesis in the EDC preparation were 65% of those determined in vivo, comparing favourably with values obtained by other workers in mammalian muscle. 4. Rates of protein synthesis responded to insulin, amino acids, and to the nutritional status of the bird, as had been observed in prior studies with mammalian muscle. By contrast to mammalian muscle protein degradation was relatively insensitive to these modifiers.     

Antioxidant alpha-lipoic acid and protein turnover in insulin-resistant rat muscle

We have shown previously that the antioxidant alpha-lipoic acid (ALA) can stimulate glucose transport and can enhance the stimulation of this process by insulin in skeletal muscle from insulin-resistant obese Zucker rats. As insulin can also acutely activate general protein synthesis and inhibit net protein degradation in skeletal muscle, we hypothesized that ALA could directly affect protein turnover and also increase the effect of insulin on protein turnover in isolated skeletal muscle from developing obese Zucker rats. In epitrochlearis muscles isolated from obese Zucker rats, insulin (2 mU/ml) significantly (p < 0.05) increased in vitro protein synthesis (phenylalanine incorporation into protein) and decreased net protein degradation (tyrosine release), whereas a racemic mixture of ALA (2 mM) had no effect on either process. Interestingly, rates of protein synthesis in muscle from obese Zucker rats were substantially lower compared to those values observed in age-matched insulin-sensitive Wistar rats, whereas rates of protein degradation were comparable. Obese Zucker rats were also treated chronically with either vehicle or ALA (50 mg/kg/d for 10 d). Again, insulin significantly increased net protein synthesis and decreased net protein degradation in epitrochlearis muscles isolated from vehicle-treated obese Zucker rats; however, this stimulatory effect of insulin was not improved by prior in vivo ALA treatment. These results indicate that the previously described effect of the antioxidant ALA to increase insulin-stimulated glucose transport in skeletal muscle of obese, insulin-resistant rats does not apply to another important insulin-regulatable process, protein turnover. These findings imply that the cellular mode of action for ALA is restricted to signaling factors unique to the activation of glucose transport, and does not involve the pathway of stimulation of general protein synthesis and net protein degradation.     

Resistance-training-induced adaptations in skeletal muscle protein turnover in the fed state

Resistance training changes the balance of muscle protein turnover, leading to gains in muscle mass. A longitudinal design was employed to assess the effect that resistance training had on muscle protein turnover in the fed state. A secondary goal was investigation of the potential interactive effects of creatine (Cr) monohydrate supplementation on resistance-training-induced adaptations. Young (N = 19, 23.7 +/- 3.2 year), untrained (UT), healthy male subjects completed an 8-week resistance-training program (6 d/week). Supplementation with Cr had no impact on any of the variables studied; hence, all subsequent data were pooled. In the UT and trained (T) state, subjects performed an acute bout of resistance exercise with a single leg (exercised, EX), while their contralateral leg acted as a nonexercised (NE) control. Following exercise, subjects were fed while receiving a primed constant infusion of [d5]- and [15N]-phenylalanine to determine the fractional synthetic and breakdown rates (FSR and FBR), respectively, of skeletal muscle proteins. Acute exercise increased FSR (UT-NE, 0.065 +/- 0.025 %/h; UT-EX, 0.088 +/- 0.032 %/h; P < 0.01) and FBR (UT-NE, 0.047 +/- 0.023 %/h; UT-EX, 0.058 +/- 0.026 %/h; P < 0.05). Net balance (BAL = FSR - FBR) was positive in both legs (P < 0.05) but was significantly greater (+65%) in the EX versus the NE leg (P < 0.05). Muscle protein FSR and FBR were greater at rest following T (FSR for T-NE vs. UT-NE, +46%, P < 0.01; FBR for T-NE vs. UT-NE, +81%, P < 0.05). Resistance training attenuated the acute exercise-induced rise in FSR (T-NE vs. T-EX, +20%, P = 0.65). The present results demonstrate that resistance training resulted in an elevated resting muscle protein turnover but an attenuation of the acute response of muscle protein turnover to a single bout of resistance exercise.     

Rat muscle protein turnover and redox state in progressive diabetes

Protein synthesis and degradation, and redox state were measured in soleus and extensor digitorum longus muscles of rats up to 12 days after injection of streptozotocin. Muscle growth was slower in these animals apparently due to slower protein synthesis throughout the duration of diabetes. Up to day 4 after injection of streptozotocin or withdrawal of insulin from treated, diabetic animals, the muscle ratio of lactate/pyruvate, an indicator of the cytoplasmic NAD+ redox couple, was lower and protein degradation was faster than in control muscles. Thereafter, the ratio of lactate/pyruvate was greater and protein degradation was slower than in size- or age-matched control muscles. Insulin treatment in vitro or in vivo increased lactate/pyruvate and decreased proteolysis. Therefore, in muscles of streptozotocin-diabetic rats, the initial increase and later fall in proteolysis, and the inhibition of proteolysis by insulin, may correlate with opposite changes in NADH/NAD+.     

Phospholipid turnover in hamster and chick embryo fibroblasts

The stability of the glycerol backbone of phosphatidyl choline, phosphatidyl ethanolamine and phosphatidyl serine was measured in growing and non-growing hamster and chick embryo fibroblasts. Major differences were found for the rates of degradation of the individual glycerophospholipids in both hamster and chick embryo fibroblasts: considerable degradation of phosphatidyl choline was detected over a 24 h period while at the same time no degradation of the glycerol backbone of phosphatidyl ethanolamine and phosphatidyl serine was observed. The patterns of stability of these glycerophospholipids were similar in growing and non-growing cells.     

Growth rate correlates negatively with protein turnover in Arabidopsis accessions

Previous studies with Arabidopsis accessions revealed that biomass correlates negatively to dusk starch content and total protein, and positively to the maximum activities of enzymes in photosynthesis. We hypothesized that large accessions have lower ribosome abundance and lower rates of protein synthesis, and that this is compensated by lower rates of protein degradation. This would increase growth efficiency and allow more investment in photosynthetic machinery. We analysed ribosome abundance and polysome loading in 19 accessions, modelled the rates of protein synthesis and compared them with the observed rate of growth. Large accessions contained less ribosomes than small accessions, due mainly to cytosolic ribosome abundance falling at night in large accessions. The modelled rates of protein synthesis resembled those required for growth in large accessions, but were up to 30% in excess in small accessions. We then employed ¹³CO₂ pulse‐chase labelling to measure the rates of protein synthesis and degradation in 13 accessions. Small accessions had a slightly higher rate of protein synthesis and much higher rates of protein degradation than large accessions. Protein turnover was negligible in large accessions but equivalent to up to 30% of synthesised protein day^?1 in small accessions. We discuss to what extent the decrease in growth in small accessions can be quantitatively explained by known costs of protein turnover and what factors may lead to the altered diurnal dynamics and increase of ribosome abundance in small accessions, and propose that there is a trade‐off between protein turnover and maximisation of growth rate.     

Genetic studies on the muscle protein turnover rate of coturnix quail

The validation of the urinary excretion of N-methylhistidine (N-MH) by quail as an index of the muscle protein turnover rate was tested using the criterion of the rate of recovery of radioactivity in urine following an intraperitoneal dose of l-[3-¹⁴C]methylhistidine. A genetic study on muscle protein turnover in quail was conducted using three genetically diverse lines (LL, large body size; SS, small body size; RR, random-bred control line) selected for body size. When l-[3-¹⁴C]methylhistidine was administered to 20-week-old male and female coturnix quail by direct intraperitoneal injection, approximately 90% of the l-[3-¹⁴C]methylhistidine was recovered by 96 hr postinjection. Recoveries were low in the egg and muscle. These results show that N-MH released from myofibrillar protein is not reutilized and the excretion of N-MH is a satisfactory index of muscle protein breakdown. In all lines, the amount of urinary N-MH excretion and fractional synthesis (K_s) and degradation (K_d) rates at the high growing period were higher than those at the low growing period. The K_s and K_d are significantly different among selected lines at both 3 and 6 weeks of age. At 3 weeks of age, the fractional rate of synthesis of the LL line (13.2%/day) was higher than that of the RR line (11.5%/day), whereas the SS (8.1%/day) was lower than that of the RR line (11.5%/day). The fractional rates of degradation of both the LL line (4.1%/day) and the SS line (5.6%/day) were lower than that of the RR line (7.0%/day) at 3 weeks of age. From these results, it was recognized that selection for body size gave rise to the changes in the muscle protein turnover rate.     

Skeletal and cardiac muscle protein turnover during cold acclimation in young rats

The effect of long-term cold exposure on skeletal and cardiac muscle protein turnover was investigated in young growing animals. Two groups of 36 male 28-day-old rats were maintained at either 5 degrees C (cold) or 25 degrees C (control). Rates of protein synthesis and degradation were measured in vivo on days 5, 10, 15, and 20. Protein mass by day 20 was approximately 28% lower in skeletal muscle (gastrocnemius and soleus) and approximately 24% higher in heart in cold compared with control rats (P < 0.05). In skeletal muscle, the fractional rates of protein synthesis (k(syn)) and degradation (k(deg)) were not significantly different between cold and control rats, although k(syn) was lower (approximately -26%) in cold rats on day 5; consequent to the lower protein mass, the absolute rates of protein synthesis (approximately -21%; P < 0. 05) and degradation (approximately -13%; P < 0.1) were lower in cold compared with control rats. In heart, overall, k(syn) (approximately +12%; P < 0.1) and k(deg) (approximately +22%; P < 0.05) were higher in cold compared with control rats; consequently, the absolute rates of synthesis (approximately +44%) and degradation (approximately +54%) were higher in cold compared with control rats (P < 0.05). Plasma triiodothyronine concentration was higher (P < 0.05) in cold compared with control rats. These data indicate that long-term cold acclimation in skeletal muscle is associated with the establishment of a new homeostasis in protein turnover with decreased protein mass and normal fractional rates of protein turnover. In heart, unlike skeletal muscle, rates of protein turnover did not appear to immediately return to normal as increased rates of protein turnover were observed beyond day 5. These data also indicate that increased rates of protein turnover in skeletal muscle are unlikely to contribute to increased metabolic heat production during cold acclimation.