Translocation and protein complex co-localization of mTOR is associated with postprandial myofibrillar protein synthesis at rest and after endurance exercise.

Translocation and colocalization of mechanistic target of rapamycin complex 1 (mTORC1) with regulatory proteins represents a critical step in translation initiation of protein synthesis in vitro. However, mechanistic insight into the control of postprandial skeletal muscle protein synthesis rates at rest and after an acute bout of endurance exercise in humans is lacking. In crossover trials, eight endurance-trained men received primed-continuous infusions of L-[ring-2 H5 ]phenylalanine and consumed a mixed-macronutrient meal (18 g protein, 60 g carbohydrates, 17 g fat) at rest (REST) and after 60 min of treadmill running at 70% VO2peak (EX). Skeletal muscle biopsies were collected to measure changes in phosphorylation and colocalization in the mTORC1-pathway, in addition to rates of myofibrillar (MyoPS) and mitochondrial (MitoPS) protein synthesis. MyoPS increased (P < 0.05) above fasted in REST (~2.1-fold) and EX (~twofold) during the 300 min postprandial period, with no corresponding changes in MitoPS (P > 0.05). TSC2/Rheb colocalization decreased below fasted at 60 and 300 min after feeding in REST and EX (P < 0.01). mTOR colocalization with Rheb increased above fasted at 60 and 300 min after feeding in REST and EX (P < 0.01), which was consistent with an increased phosphorylation 4E-BP1Thr37/46 and rpS6ser240/244 at 60 min. Our data suggest that MyoPS, but not MitoPS, is primarily nutrient responsive in trained young men at rest and after endurance exercise. The postprandial increase in MyoPS is associated with an increase in mTOR/Rheb colocalization and a reciprocal decrease in TSC2/Rheb colocalization and thus likely represent important regulatory events for in vivo skeletal muscle myofibrillar mRNA translation in humans.

Endurance Exercise Attenuates Postprandial Whole-Body Leucine Balance in Trained Men.

PURPOSE: Endurance exercise increases indices of small intestinal damage and leucine oxidation, which may attenuate dietary amino acid appearance and postprandial leucine balance during postexercise recovery. Therefore, the purpose of this study was to examine the effect of an acute bout of endurance exercise on postprandial leucine kinetics and net leucine balance. METHODS: In a crossover design, seven trained young men (age = 25.6 ± 2.3 yr; V˙O2peak = 61.4 ± 2.9 mL·kg·min; mean ± SEM) received a primed constant infusion of L-[1-C]leucine before and after ingesting a mixed macronutrient meal containing 18 g whole egg protein intrinsically labeled with L-[5,5,5-H3]leucine, 17 g fat, and 60 g carbohydrate at rest and after 60 min of treadmill running at 70% V˙O2peak. RESULTS: Plasma intestinal fatty acid binding protein concentrations and leucine oxidation both increased (P < 0.01) to peaks that were ~2.5-fold above baseline values during exercise with a concomitant decrease (P < 0.01) in nonoxidative leucine disposal. Meal ingestion attenuated (P < 0.01) endogenous leucine rates of appearance at rest and after exercise. There were no differences (both, P > 0.05) in dietary leucine appearance rates or in the amount of dietary protein-derived leucine that appeared into circulation over the 5-h postprandial period at rest and after exercise (62% ± 2% and 63% ± 2%, respectively). Leucine balance over the 5-h postprandial period was positive (P < 0.01) in both conditions but was negative (P < 0.01) during the exercise trial after accounting for exercise-induced leucine oxidation. CONCLUSIONS: We demonstrate that endurance exercise does not modulate dietary leucine availability from a mixed meal but attenuates postprandial whole-body leucine balance in trained young men.

Development of Intrinsically Labeled Eggs and Poultry Meat for Use in Human Metabolic Research.

BACKGROUND: Stable isotope amino acids are regularly used as tracers to examine whole-body and muscle protein metabolism in humans. To accurately assess in vivo dietary protein digestion and absorption kinetics, the amino acid tracer is required to be incorporated within the dietary protein food source (i.e., intrinsically labeled protein). OBJECTIVE: We assessed the practicality of producing eggs and poultry meat intrinsically labeled with l-[5,5,5-(2)H3]leucine through noninvasive oral tracer administration. METHODS: A specifically formulated diet containing 0.52% leucine was supplemented with 0.3% l-[5,5,5-(2)H3]leucine and subsequently fed to 3 laying hens (Lohmann LSL Whites) for 55 d. On day 55, the hens were slaughtered and their meat, bones, and organs were harvested to determine tissue labeling. In Expt. 1, 2 healthy young men [mean ± SEM age: 22 ± 1.5 y; mean ± SEM body mass index (BMI; in kg/m(2)): 23.7 ± 0.5] ingested 18 g l-[5,5,5-(2)H3]leucine-labeled egg protein. In Expt. 2, 2 healthy young men (mean ± SEM age: 20.0 ± 0.0 y; mean ± SEM BMI: 26.4 ± 3.1) ingested 28 g l-[5,5,5-(2)H3]leucine-labeled poultry meat protein. Plasma samples (Expts. 1 and 2) and muscle biopsies (Expt. 1) were collected before and after labeled-food ingestion. RESULTS: High tracer labeling [>20 mole percent excess (MPE)] in the eggs was obtained after 7 d and maintained throughout the feeding protocol (P < 0.05). Over a 55-d period, ∼850 g egg protein (145 eggs) was produced, with a mean ± SEM tracer enrichment of 22.0 ± 0.8 MPE. Mean ± SEM l-[5,5,5-(2)H3]leucine enrichment in the meat was 9.6 ± 0.1 MPE. In Expts. 1 and 2, the consumption of labeled eggs and poultry meat protein increased plasma l-[5,5,5-(2)H3]leucine enrichment, with mean ± SEM peak values of 6.7 ± 0.1 MPE and 4.0 ± 0.9 MPE, respectively. The mean ± SEM 5-h postprandial increase in myofibrillar l-[5,5,5-(2)H3]leucine enrichment after egg ingestion in healthy young men was 0.051 ± 0.008 MPE (Expt. 1). CONCLUSION: We demonstrated the feasibility of producing intrinsically labeled eggs and poultry meat for use in human metabolic research.