Philippe Patureau Mirand

Effect of glucocorticoid excess on skeletal muscle and heart protein synthesis in adult and old rats

This study was carried out to analyse glucocorticoid-induced muscle wasting and subsequent recovery in adult (6-8 months) and old (18-24 months) rats because the increased incidence of various disease states results in hypersecretion of glucocorticoids in ageing. Adult and old rats received dexamethasone in their drinking water for 5 or 6 d and were then allowed to recover for 3 or 7 d. As dexamethasone decreased food intake, all groups were pair-fed to dexamethasone-treated old rats (i.e. the group that had the lowest food intake). At the end of the treatment, adult and old rats showed significant increases in blood glucose and plasma insulin concentrations. This increase disappeared during the recovery period. Protein synthesis of different muscles was assessed in vivo by a flooding dose of [13C]valine injected subcutaneously 50 min before slaughter. Dexamethasone induced a significant decrease in protein synthesis in fast-twitch glycolytic and oxidative glycolytic muscles (gastrocnemius, tibialis anterior, extensor digitorum longus). The treatment affected mostly ribosomal efficiency. Adult dexamethasone-treated rats showed an increase in protein synthesis compared with their pair-fed controls during the recovery period whereas old rats did not. Dexamethasone also significantly decreased protein synthesis in the predominantly oxidative soleus muscle but only in old rats, and increased protein synthesis in the heart of adult but not of old rats. Thus, in skeletal muscle, the catabolic effect of dexamethasone is maintained or amplified during ageing whereas the anabolic effect in heart is depressed. These results are consistent with muscle atrophy occurring with ageing.

Differential variation of mitochondrial H2O2 release during aging in oxidative and glycolytic muscles in rats.

Mitochondrial free radical (ROS) production could be involved in sarcopenia. Our aim was to measure this production in various muscles during aging. Male Wistar rats aged 4.5 and 24 months were used. H(2)O(2) release and protein carbonyls were evaluated in isolated mitochondria from an oxidative (soleus) and a glycolytic (tibialis anterior) muscle. Total and Mn-superoxide dismutase (SOD), catalase, glutathione peroxidase (GPX) and glutathione reductase (GR) activities were measured in tibialis anterior. In soleus, glutamate/malate supported mitochondrial H(2)O(2) release was lower than in tibialis anterior in young rats, but increased significantly with age. In tibialis anterior, glutamate/malate or succinate supported H(2)O(2) release was unchanged with age. ROS generators were complexes I and III. Mitochondrial carbonyl content remained stable during aging in both muscles but tended to be higher in tibialis anterior than in soleus. Tibialis anterior total SOD (+17%), catalase (+84%), and GPX (-17%) activities varied significantly with age but Mn-SOD was unchanged, suggesting an increase in cytosolic ROS production. In conclusion, the higher life-long H(2)O(2) release observed in tibialis anterior is consistent with the known sensitivity of glycolytic muscles to sarcopenia. The fact that the rate of H(2)O(2) release increases with age in soleus seems to have little impact.

Age-related changes in protein synthesis measured in vivo in rat liver and gastrocnemius muscle

This study analyses in detail the effects of ageing on gastrocnemius muscle and liver protein synthesis measured in vivo at three ages, 1.5 months (young), 12 months (adult) and 24 months (old) in Sprague-Dawley rats. Comparing adult and old rats, muscle protein synthesis was decreased in old rats when expressed per unit of RNA and per day (translational efficiency), was unchanged when expressed in absolute terms and increased when expressed in fractional terms as a result of protein loss due to muscle atrophy. In the liver, only translational efficiency tended to decrease in old rats compared to adult rats. It is concluded that the decline in protein turnover described in vitro is consistent with a decrease in translational efficiency, but that absolute synthesis rates are maintained during ageing. Muscle atrophy is unlikely to result from alterations in protein synthesis pathways.

Age-related changes in glutathione availability and skeletal muscle carbonyl content in healthy rats.

The free radical theory of aging proposes that oxidative stress plays a key role in the aging process. By altering muscle protein degradation rates, it could accelerate the age-related loss of muscle proteins. Glutathione (GSH), one of the main body antioxidants, could prevent this phenomenon, but its concentration decreases during aging. Our aims were to have a better understanding of the mechanisms of the age-related decrease in glutathione availability and of the links with sarcopenia. Male Wistar rats aged 6, 9, 12, 15, 19, 22, 25 and 28 months (n = 6 per age) were used to measure plasma and skeletal muscle protein carbonyl content, plasma total and free cyst(e)ine content, liver and muscle glutathione content as well as liver GSSG reductase, GSH peroxidase, GSH transferase and gamma glutamyl cysteine synthetase (GCS) activities. Although tissue glutathione content decreased with age, the other markers of oxidative stress were little changed during aging. In particular, muscle protein carbonyl content was unchanged. Variations in glutathione availability were not explained by cyst(e)ine availability but depended on gamma GCS activity. The stability of skeletal muscle carbonyl content during aging suggests a very efficient degradation of oxidized proteins in muscle.

Daily protein intakes and eating patterns in young and elderly French

An adequate level of protein intake is required to limit the gradual body protein loss observed during ageing. Different factors (cohort age, sex, life conditions) may modify protein intake and distribution. However the precise amounts, as well as their daily distribution which affects protein utilisation and N retention, are unknown in both young and elderly individuals. The hypothesis was tested that protein intake and its distribution over daily meals could be different between the young and elderly. The consumption of six different protein-rich food groups by 292 healthy individuals aged 20-30 and 65-75 years was determined throughout each day for 1 week. The data of the total protein intake and protein intakes at each meal were analysed by ANOVA for each sex separately, using age group as the independent factor. The average protein intake of men was lower in the older age group whilst the opposite trend was seen in women. The distribution of protein intake was different between the two age groups: 56.5 % of the daily protein was eaten at lunch by the elderly but only 47 % (P<0.0001) by the younger subjects. In the elderly subjects, those eating larger amounts consumed a greater proportion of protein-rich foods at dinner than those eating small amounts (30.4 v. 26.2 %, P=0.05). A high level of protein intake was related to a higher meat-product consumption in both the elderly and young individuals. In conclusion, the pattern of protein intake differs significantly between age groups and sexes.

Peptides reproducibly released by in vivo digestion of beef meat and trout flesh in pigs

Characterisation and identification of peptides (800 to 5000 Da) generated by intestinal digestion of fish or meat were performed using MS analyses (matrix-assisted laser desorption ionisation time of flight and nano-liquid chromatography electrospray-ionisation ion trap MS/MS). Four pigs fitted with cannulas at the duodenum and jejunum received a meal exclusively made of cooked Pectoralis profundus beef meat or cooked trout fillets. A protein-free meal, made of free amino acids, starch and fat, was used to identify peptides of endogenous origin. Peptides reproducibly detected in digesta (i.e. from at least three pigs) were evidenced predominantly in the first 3 h after the meal. In the duodenum, most of the fish- and meat-derived peptides were characteristic of a peptic digestion. In the jejunum, the majority of peptides appeared to result from digestion by chymotrypsin and trypsin. Despite slight differences in gastric emptying kinetics and overall peptide production, possibly in relation to food structure and texture, six and four similar peptides were released after ingestion of fish or meat in the duodenum and jejunum. A total of twenty-six different peptides were identified in digesta. All were fragments of major structural (actin, myosin) or sarcoplasmic (creatine kinase, glyceraldehyde-3-phosphate dehydrogenase and myoglobin) muscle proteins. Peptides were short ( < 2000 Da) and particularly rich in proline residues. Nineteen of them contained bioactive sequences corresponding mainly to an antihypertensive activity. The present work showed that after fish or meat ingestion, among the wide variety of peptides produced by enzymic digestion, some of them can be reproducibly observed in intestinal digesta.

The inflammatory response to vaccination is altered in the elderly

To further explore whether immune function and acute phase response are altered during ageing, the response to a mild inflammatory stress (DT-Polio-Typhim vaccination) was studied in elderly and young subjects. Cytokine production (IFN-gamma, TNF-alpha, IL-6, IL-10) by whole blood cultures, circulating cytokines and acute phase proteins were analysed before and 2 days after vaccination. Prior to vaccination, only IFN-gamma production was lower in the elderly than in the young subjects due to a lower mononuclear cell number. In the same time, although in the normal range, several acute phase proteins were greater in elderly than in young subjects, suggesting a low-grade inflammatory state in the elderly. After vaccination, IFN-gamma production remained lower in the elderly than in the young, supporting an altered cell-mediated immunity with advancing age. TNF-alpha production was unaffected by either ageing or vaccination. IL-6 production was stimulated by vaccination in young subjects but not significantly in the elderly. IL-10 production was inhibited by vaccination in the elderly but not in the young. Acute phase proteins were less increased in elderly than in young subjects. Taken together, these results support a general lack of inflammatory response in the elderly exposed to an immune challenge and suggest that immune deficiency may concern both Th1 and Th2 responses. However, the interpretation must respect the limitation of small subjects number.

Type and timing of protein feeding to optimize anabolism.

&NA; The synchronization of variations in anabolic capability with amino acid supply partly explains the effects of the type and timing of protein feeding. This effect is modulated by the amount of amino acids required to increase whole‐body proteins and by the signaling properties of some amino acids to stimulate protein synthesis. Indeed, the anabolic effect of amino acids is determined by their interaction with other anabolic factors (other nutrients or physiological factors, whose efficiency is mainly related to their effect on protein degradation). It is clear that benefits can be obtained from adapted protein feeding patterns. Purpose of review The delivery rate of amino acids to an organism significantly affects protein anabolism. The rate can be controlled by the type and the timing of feeding. Our aim was to bring new insights to the way they may act. Recent findings During young and adult ages, when food supply is liberal, subjects can adapt to various modes of protein feeding. However, during food restriction, protein anabolism is favored when the delivery of amino acids is evenly distributed over the day, either with frequent meals, or through the use of slowly absorbed proteins like casein. In contrast, during aging, quickly absorbed protein sources become more efficient. During recovery after exercise, the timing of protein feeding after the end of exercise may or may not influence its anabolic effect, depending on the subjects age and the type of exercise.

Glutamate and CO2 production from glutamine in incubated enterocytes of adult and very old rats

Glutamine is the major fuel for enterocytes and promotes the growth of intestinal mucosa. Although oral glutamine exerts a positive effect on intestinal villus height in very old rats, how glutamine is used by enterocytes is unclear. Adult (8 months) and very old (27 months) female rats were exposed to intermittent glutamine supplementation for 50% of their age lifetime. Treated rats received glutamine added to their drinking water, and control rats received water alone. Jejunal epithelial cells (~300×10(6) cells) were incubated in oxygenated Krebs-Henseleit buffer for 30 min containing [1-(13)C] glutamine (~17 M) for analysis of glutamine metabolites by (13)C nuclear magnetic resonance ((13)C NMR). An aliquot fraction was incubated in the presence of [U-(14)C] glutamine to measure produced CO2. Glutamine pretreatment increased glutamate production and decreased CO2 production in very old rats. The ratio CO2/glutamate, which was very high in control very old rats, was similar at both ages after glutamine pretreatment, as if enterocytes from very old rats recovered the metabolic abilities of enterocytes from adult rats. Our results suggest that long-term treatment with glutamine started before advanced age (a) prevented the loss of rat body weight without limiting sarcopenia and (b) had a beneficial effect on enterocytes from very old rats probably by favoring the role of glutamate as a precursor for glutathione, arginine and proline biosynthesis, which was not detected in (13)C NMR spectra in our experimental conditions.

Differential Effects of Cooked Common Bean (Phaseolus vulgaris) and Lentil (Lens esculenta puyensis) Feeding on Protein and Nucleic Acid Contents in Intestines, Liver and Muscles in Rats

Aim: Our aim was to investigate the influence of legume feeding on the protein and nucleic acid content of intestinal tissues and muscles. Methods: Growing male Wistar rats were fed ad libitum on balanced diets containing cooked common bean or lentil as the unique protein source (180 g·kg–1 dry matter) for 20 days. The control group was pair-fed with an iso-energetic, iso-nitrogenous balanced casein diet. Results: Intestinal tissues were heavier in the legume-fed groups and higher relative mass (g per 100 g body mass) of protein, RNA and DNA were found in these tissues as compared to those of the control groups. In liver protein and RNA masses were significantly lower in the bean group than in the control group but the DNA content was not different in the legume and in the control groups. In gastrocnemius and soleus muscles, there was no significant effect of legume feeding on the fresh mass and on the protein and RNA contents, but the DNA content of the m. gastrocnemius was significantly lower in both legume groups than in their control group. The ribosomal capacity of intestines, liver and muscles was not significantly affected by legume feeding. Conclusion: Legume feeding had a trophic effect on both proximal and distal intestinal tissues; dietary fiber appears to be the main cause of this effect.