Cancer cell-derived exosomes promote cell proliferation and inhibit cell apoptosis of both normal lung fibroblasts and non-small cell lung cancer cell through delivering alpha-smooth muscle actin.

This study aimed to investigate the effect of non-small cell lung cancer (NSCLC) cell-derived exosome on cell proliferation and apoptosis in normal lung fibroblast cells and NSCLC cells, and whether it regulates cell functions through delivering alpha-smooth muscle actin (ASMA). NSCLC exosomes were extracted from A549 cells, then cocultured with normal lung fibroblasts (HLF1 cells) and NSCLC cells (A549 cells). Blank ShRNA and ASMA ShRNA plasmids were transferred into HLF1 cells/A549 cells with or without NSCLC exosomes, which were divided into 4 groups accordingly: Negative control (NC) group, SH group, Exosome group and Exosome+SH group. Western blot, immunofluorescence, qPCR, CCK-8 and AV/PI were used to detect protein level, gene expression, cell proliferation and cell apoptosis, respectively. In HLF1 cells, cell proliferation was enhanced while cell apoptosis rate was inhibited in Exosome group compared with NC group; and cell proliferation was attenuated while cell apoptosis rate was raised in Exosome+SH group than Exosome group in rescue experiment; the expressions of apoptotic markers C-caspase3 and Bcl-2 also revealed the same trends. Additionally, in A549 cells, cell proliferation was also increased while cell apoptosis was inhibited in Exosome group compared with NC group; and cell proliferation was reduced while cell apoptosis rate was elevated in Exosome+SH group than Exosome group in rescue experiment. In conclusion, NSCLC derived exosomes promote cell proliferation and inhibit cell apoptosis in both normal lung fibroblasts and NSCLC cells by delivering ASMA.

SIRT6 drives epithelial-to-mesenchymal transition and metastasis in non-small cell lung cancer via snail-dependent transrepression of KLF4.

BACKGROUND: Epithelial-to-mesenchymal transition (EMT) contributes to the invasion and metastasis of epithelial tumors. Sirtuin 6 (SIRT6), an NAD-dependent deacetylase, is known to promote metastasis of non-small cell lung cancer (NSCLC). METHODS: In this work, we determined the role of SIRT6 in the EMT of NSCLC cells and identified the key EMT-related genes involved in the oncogenic activity of SIRT6. RESULTS: We report that depletion of SIRT6 inhibits transforming growth factor-ß1 (TGF-ß1)-induced EMT in A549 and H1299 NSCLC cells, which is rescued by ectopic expression of SIRT6. Knockdown of SIRT6 leads to a reduction in Snail protein without affecting the mRNA level. Immunoprecipitation experiments demonstrate a physical association between SIRT6 and Snail. SIRT6 deacetylates Snail and prevents its proteasomal degradation. Silencing of Snail blunts SIRT6-induced NSCLC cell migration and invasion, while overexpression of Snail restores the invasion and EMT in SIRT6-depleted NSCLC cells. SIRT6 depletion leads to an upregulation of kruppel-like factor 4 (KLF4) and reduced Snail binding to the promoter of Klf4 in NSCLC cells. Knockdown of KLF4 rescues the invasive capacity in SIRT6-depleted NSCLC cells. Conversely, co-expression of KLF4 impairs SIRT6-induced aggressive behavior. In vivo data further demonstrate that SIRT6-induced NSCLC metastasis is antagonized by overexpression of KLF4. CONCLUSIONS: These findings provide mechanistic insights into the pro-metastatic activity of SIRT6 and highlight the role of the SIRT6/Snail/KLF4 axis in regulating EMT and invasion of NSCLC cells.

Coherent and Noncoherent Joint Processing of Sonar for Detection of Small Targets in Shallow Water.

A coherent-noncoherent joint processing framework is proposed for active sonar to combine diversity gain and beamforming gain for detection of a small target in shallow water environments. Sonar utilizes widely-spaced arrays to sense environments and illuminate a target of interest from multiple angles. Meanwhile, it exploits spatial diversity for time-reversal focusing to suppress reverberation, mainly strong bottom reverberation. For enhancement of robustness of time-reversal focusing, an adaptive iterative strategy is utilized in the processing framework. A probing signal is firstly transmitted and echoes of a likely target are utilized as steering vectors for the second transmission. With spatial diversity, target bearing and range are estimated using a broadband signal model. Numerical simulations show that the novel sonar outperforms the traditional phased-array sonar due to benefits of spatial diversity. The effectiveness of the proposed framework has been validated by localization of a small target in at-lake experiments.

Co-ingestion of carbohydrate and whey protein increases fasted rates of muscle protein synthesis immediately after resistance exercise in rats.

The objective of the study was to investigate whether co-ingestion of carbohydrate and protein as compared with protein alone augments muscle protein synthesis (MPS) during early exercise recovery. Two months old rats performed 10 repetitions of ladder climbing with 75% of body weight attached to their tails. Placebo (PLA), whey protein (WP), or whey protein plus carbohydrate (CP) was then given to rats by gavage. An additional group of sedentary rats (SED) was used as controls. Blood samples were collected immediately and at either 1 or 2 h after exercise. The flexor hallucis longus muscle was excised at 1 or 2 h post exercise for analysis of MPS and related signaling proteins. MPS was significantly increased by CP compared with PLA (p<0.05), and approached significance compared with WP at 1 h post exercise (p = 0.08). CP yielded a greater phosphorylation of mTOR compared with SED and PLA at 1 h post exercise and SED and WP at 2 h post exercise. CP also increased phosphorylation of p70S6K compared with SED at 1 and 2 h post exercise. 4E-BP1 phosphorylation was inhibited by PLA at 1 h but elevated by WP and CP at 2 h post exercise relative to SED. The phosphorylation of AMPK was elevated by exercise at 1 h post exercise, and this elevated level was sustained only in the WP group at 2 h. The phosphorylation of Akt, GSK3, and eIF2Bε were unchanged by treatments. Plasma insulin was transiently increased by CP at 1 h post exercise. In conclusion, post-exercise CP supplementation increases MPS post exercise relative to PLA and possibly WP, which may have been mediated by greater activation of the mTOR signaling pathway.

L-Alanylglutamine inhibits signaling proteins that activate protein degradation, but does not affect proteins that activate protein synthesis after an acute resistance exercise.

Sustamine™ (SUS) is a dipeptide composed of alanine and glutamine (AlaGln). Glutamine has been suggested to increase muscle protein accretion; however, the underlying molecular mechanisms of glutamine on muscle protein metabolism following resistance exercise have not been fully addressed. In the present study, 2-month-old rats climbed a ladder 10 times with a weight equal to 75 % of their body mass attached at the tail. Rats were then orally administered one of four solutions: placebo (PLA-glycine = 0.52 g/kg), whey protein (WP = 0.4 g/kg), low dose of SUS (LSUS = 0.1 g/kg), or high dose of SUS (HSUS = 0.5 g/kg). An additional group of sedentary (SED) rats was intubated with glycine (0.52 g/kg) at the same time as the ladder-climbing rats. Blood samples were collected immediately after exercise and at either 20 or 40 min after recovery. The flexor hallucis longus (FHL), a muscle used for climbing, was excised at 20 or 40 min post exercise and analyzed for proteins regulating protein synthesis and degradation. All supplements elevated the phosphorylation of FOXO3A above SED at 20 min post exercise, but only the SUS supplements significantly reduced the phosphorylation of AMPK and NF-kB p65. SUS supplements had no effect on mTOR signaling, but WP supplementation yielded a greater phosphorylation of mTOR, p70S6k, and rpS6 compared with PLA at 20 min post exercise. However, by 40 min post exercise, phosphorylation of mTOR and rpS6 in PLA had risen to levels not different than WP. These results suggest that SUS blocks the activation of intracellular signals for MPB, whereas WP accelerates mRNA translation.

The effect of an amino acid beverage on glucose response and glycogen replenishment after strenuous exercise.

PURPOSE: We previously reported that an amino acid mixture (AA) was able to lower the glucose response to an oral glucose challenge in both rats and humans. Increased glucose uptake and glycogen storage in muscle might be associated with the faster blood glucose clearance. We therefore tested the effect of two different doses of AA provided with a carbohydrate supplement on blood glucose homeostasis and muscle glycogen replenishment in human subjects after strenuous aerobic exercise. METHODS: Ten subjects received a carbohydrate (1.2 g/kg body weight, CHO), CHO/HAA (CHO + 13 g AA), or CHO/LAA (CHO + 6.5 g AA) supplement immediately and 2 h after an intense cycling bout. Muscle biopsies were performed immediately and 4 h after exercise. RESULTS: The glucose responses for CHO/HAA and CHO/LAA during recovery were significantly lower than CHO, as was the glucose area under the curve (CHO/HAA 1259.9 ± 27.7, CHO/LAA 1251.5 ± 47.7, CHO 1376.8 ± 52.9 mmol/L 4 h, p < 0.05). Glycogen storage rate was significantly lower in CHO/HAA compared with CHO, while it did not differ significantly between CHO/LAA or CHO (CHO/HAA 15.4 ± 2.0, CHO/LAA 18.1 ± 2.0, CHO 21.5 ± 1.4 µmol/g wet muscle 4 h). CHO/HAA caused a significantly higher insulin response and a greater effect on mTOR and Akt/PKB phosphorylation compared with CHO. Phosphorylation of AS160 and glycogen synthase did not differ across treatments. Likewise, there were no differences in blood lactate across treatments. CONCLUSIONS: The AA lowered the glucose response to a carbohydrate supplement after strenuous exercise. However, it was not effective in facilitating subsequent muscle glycogen storage.

An amino acid mixture improves glucose tolerance and lowers insulin resistance in the obese Zucker rat.

The purpose of this investigation was to test an amino acid mixture on glucose tolerance in obese Zucker rats [experiment (Exp)-1] and determine whether differences in blood glucose were associated with alterations in muscle glucose uptake [experiment (Exp)-2]. Exp-1 rats were gavaged with either carbohydrate (OB-CHO), carbohydrate plus amino acid mixture (OB-AA-1), carbohydrate plus amino acid mixture with increased leucine concentration (OB-AA-2) or water (OB-PLA). The glucose response in OB-AA-1 and OB-AA-2 were similar, and both were lower compared to OB-CHO. This effect of the amino acid mixtures did not appear to be solely attributable to an increase in plasma insulin. Rats in Exp-2 were gavaged with carbohydrate (OB-CHO), carbohydrate plus amino acid mixture (OB-AA-1) or water (OB-PLA). Lean Zuckers were gavaged with carbohydrate (LN-CHO). Fifteen minutes after gavage, a radiolabeled glucose analog was infused through a catheter previously implanted in the right jugular vein. Blood glucose was significantly lower in OB-AA-1 compared to OB-CHO while the insulin responses were similar. Glucose uptake was greater in OB-AA-1 compared with OB-CHO, and similar to that in LN-CHO in red gastrocnemius muscle (5.15 ± 0.29, 3.8 ± 0.27, 5.18 ± 0.34 µmol/100 g/min, respectively). Western blot analysis showed that Akt substrate of 160 kDa (AS160) phosphorylation was enhanced for OB-AA-1 and LN-CHO compared to OB-CHO. These findings suggest that an amino acid mixture improves glucose tolerance in an insulin resistant model and that these improvements are associated with an increase in skeletal muscle glucose uptake possibly due to improved intracellular signaling.

An amino acid mixture is essential to optimize insulin-stimulated glucose uptake and GLUT4 translocation in perfused rodent hindlimb muscle.

The purpose of this study was to investigate whether an amino acid mixture increases glucose uptake across perfused rodent hindlimb muscle in the presence and absence of a submaximal insulin concentration, and if the increase in glucose uptake is related to an increase in GLUT4 plasma membrane density. Sprague-Dawley rats were separated into one of four treatment groups: basal, amino acid mixture, submaximal insulin, or amino acid mixture with submaximal insulin. Glucose uptake was greater for both insulin-stimulated treatments compared with the non-insulin-stimulated treatment groups but amino acids only increased glucose uptake in the presence of insulin. Phosphatidylinositol 3-kinase (PI 3-kinase) activity was greater for both insulin-stimulated treatments with amino acids having no additional impact. Akt substrate of 160 kDa (AS160) phosphorylation, however, was increased by the amino acids in the presence of insulin, but not in the absence of insulin. AMPK was unaffected by insulin or amino acids. Plasma membrane GLUT4 protein concentration was greater in the rats treated with insulin compared with no insulin in the perfusate. In the presence of insulin, amino acids increased GLUT4 density in the plasma membrane but had no effect in the absence of insulin. AS160 phosphorylation and plasma membrane GLUT4 density accounted for 76% of the variability in muscle glucose uptake. Collectively, these findings suggest that the beneficial effects of an amino acid mixture on skeletal muscle glucose uptake, in the presence of a submaximal insulin concentration, are due to an increase in AS160 phosphorylation and plasma membrane-associated GLUT4, but independent of PI 3-kinase and AMPK activation.

Amino acid mixture acutely improves the glucose tolerance of healthy overweight adults.

Certain amino acids have been reported to influence carbohydrate metabolism and blood glucose clearance, as well as improve the glucose tolerance in animal models. We hypothesized that an amino acid mixture consisting of isoleucine and 4 additional amino acids would improve the glucose response of healthy overweight men and women to an oral glucose tolerance test (OGTT). Twenty-two overweight healthy subjects completed 2 OGTTs after consuming 2 different test beverages. The amino acid mixture beverage (CHO/AA) consisted of 0.088 g cystine 2HCl, 0.043 g methionine, 0.086 g valine, 12.094 g isoleucine, 0.084 g leucine, and 100 g dextrose. The control beverage (CHO) consisted of 100 g dextrose only. Venous blood samples were drawn 10 minutes before the start of ingesting the drinks and 15, 30, 60, 120, and 180 minutes after the completion of the drinks. During the OGTT, the plasma glucose response for the CHO/AA treatment was significantly lower than that of the CHO treatment (P < .01), as was the plasma glucose area under the curve (CHO/AA 806 ± 31 mmol/L·3 hours vs CHO 942 ± 40 mmol/L·3 hours). Differences in plasma glucose between treatments occurred at 30, 60, 120, and 180 minutes after supplement ingestion. Plasma glucagon during the CHO/AA treatment was significantly higher than during the CHO treatment. However, there were no significant differences in plasma insulin or C-peptide responses between treatments. These results suggest that the amino acid mixture lowers the glucose response to an OGTT in healthy overweight subjects in an insulin-independent manner.

Aerobic exercise training adaptations are increased by postexercise carbohydrate-protein supplementation.

Carbohydrate-protein supplementation has been found to increase the rate of training adaptation when provided postresistance exercise. The present study compared the effects of a carbohydrate and protein supplement in the form of chocolate milk (CM), isocaloric carbohydrate (CHO), and placebo on training adaptations occurring over 4.5 weeks of aerobic exercise training. Thirty-two untrained subjects cycled 60 min/d, 5 d/wk for 4.5 wks at 75-80% of maximal oxygen consumption (VO(2) max). Supplements were ingested immediately and 1 h after each exercise session. VO(2) max and body composition were assessed before the start and end of training. VO(2) max improvements were significantly greater in CM than CHO and placebo. Greater improvements in body composition, represented by a calculated lean and fat mass differential for whole body and trunk, were found in the CM group compared to CHO. We conclude supplementing with CM postexercise improves aerobic power and body composition more effectively than CHO alone.

Postexercise carbohydrate-protein supplementation improves subsequent exercise performance and intracellular signaling for protein synthesis.

Postexercise carbohydrate-protein (CHO + PRO) supplementation has been proposed to improve recovery and subsequent endurance performance compared to CHO supplementation. This study compared the effects of a CHO + PRO supplement in the form of chocolate milk (CM), isocaloric CHO, and placebo (PLA) on recovery and subsequent exercise performance. Ten cyclists performed 3 trials, cycling 1.5 hours at 70% VO2max plus 10 minutes of intervals. They ingested supplements immediately postexercise and 2 hours into a 4-hour recovery. Biopsies were performed at recovery minutes 0, 45, and 240 (R0, R45, REnd). Postrecovery, subjects performed a 40-km time trial (TT). The TT time was faster in CM than in CHO and in PLA (79.43 ± 2.11 vs. 85.74 ± 3.44 and 86.92 ± 3.28 minutes, p ≤ 0.05). Muscle glycogen resynthesis was higher in CM and in CHO than in PLA (23.58 and 30.58 vs. 7.05 µmol·g⁻¹ wet weight, p ≤ 0.05). The mammalian target of rapamycin phosphorylation was greater at R45 in CM than in CHO or in PLA (174.4 ± 36.3 vs. 131.3 ± 28.1 and 73.7 ± 7.8% standard, p ≤ 0.05) and at REnd in CM than in PLA (94.5 ± 9.9 vs. 69.1 ± 3.8%, p ≤ 0.05). rpS6 phosphorylation was greater in CM than in PLA at R45 (41.0 ± 8.3 vs. 15.3 ± 2.9%, p ≤ 0.05) and REnd (16.8 ± 2.8 vs. 8.4 ± 1.9%, p ≤ 0.05). FOXO3A phosphorylation was greater at R45 in CM and in CHO than in PLA (84.7 ± 6.7 and 85.4 ± 4.7 vs. 69.2 ± 5.5%, p ≤ 0.05). These results indicate that postexercise CM supplementation can improve subsequent exercise performance and provide a greater intracellular signaling stimulus for PRO synthesis compared to CHO and placebo.

Effect of carbohydrate-protein supplementation postexercise on rat muscle glycogen synthesis and phosphorylation of proteins controlling glucose storage.

To examine whether addition of protein to a carbohydrate supplement enhances muscle glycogen synthesis, we compared the muscle glycogen concentrations of rats that had been depleted of their muscle glycogen stores with a 3-hour swim and immediately supplemented with a placebo (Con), carbohydrate (CHO), or carbohydrate plus protein supplement (C+P). Rats were given either 0.9 g carbohydrate per kilogram body mass for the CHO group or 0.9 g carbohydrate + 0.3 g protein per kilogram body mass for the C+P groups. Muscle samples of the red and white quadriceps were excised immediately, 30 minutes, or 90 minutes postexercise. Glycogen concentration of the C+P group was greater than that of the CHO group at 90 minutes postexercise in both red (C+P, 28.3 ± 2.6 µmol/g vs CHO, 22.4 ± 2.0 µmol/g; P < .05) and white (C+P, 24.9 ± 2.4 µmol/g vs CHO, 17.64 ± 1.5 µmol/g; P < .01) quadriceps. Protein kinase B phosphorylation was greater in the C+P-30 group (the number following treatment group abbreviation refers to time [in minutes] of euthanasia following exercise) than the sedentary control and exercised control groups in red quadriceps at 30 minutes and in white quadriceps at 90 minutes postexercise. This difference was not observed in the CHO group. Phosphorylation of glycogen synthase was significantly reduced 30 minutes postexercise and returned to baseline levels by 90 minutes postexercise in both CHO- and C+P-supplemented groups, with no difference between supplements. These results demonstrated that the addition of protein to a carbohydrate supplement will enhance the rate of muscle glycogen restoration postexercise and may involve facilitation of the glucose transport process.

A low carbohydrate-protein supplement improves endurance performance in female athletes.

The purpose of this study was to investigate if a low mixed carbohydrate (CHO) plus moderate protein (PRO) supplement, provided during endurance exercise, would improve time to exhaustion (TTE) in comparison to a traditional 6% CHO supplement. Fourteen (n = 14) trained female cyclists and triathletes cycled on 2 separate occasions for 3 hours at intensities varying between 45 and 70% VO2max, followed by a ride to exhaustion at an intensity approximating the individual's ventilatory threshold average 75.06% VO2max. Supplements (275 mL) were provided every 20 minutes during exercise and were composed of a CHO mixture (1% each of dextrose, fructose, and maltodextrin) + 1.2% PRO (CHO + PRO) or 6% dextrose only (CHO). The TTE was significantly greater with CHO + PRO in comparison to with CHO (49.94 ± 7.01 vs. 42.36 ± 6.21 minutes, respectively, p < 0.05). Blood glucose was significantly lower during the CHO + PRO trial (4.07 ± 0.12 mmol · L(-1)) compared to during the CHO trial (4.47 ± 0.12 mmol · L(-1)), with treatment × time interactions occurring from 118 minutes of exercise until exhaustion (p < 0.05). Results from the present study suggest that the addition of a moderate amount of PRO to a low mixed CHO supplement improves endurance performance in women above that of a traditional 6% CHO supplement. Improvement in performance occurred despite CHO + PRO containing a lower CHO and caloric content. It is likely that the greater performance seen with CHO + PRO was a result of the CHO-PRO combination and the use of a mixture of CHO sources.

An amino acid mixture improves glucose tolerance and insulin signaling in Sprague-Dawley rats.

The aims of this investigation were to evaluate the effect of an amino acid supplement on the glucose response to an oral glucose challenge (experiment 1) and to evaluate whether differences in blood glucose response were associated with increased skeletal muscle glucose uptake (experimental 2). Experiment 1 rats were gavaged with either glucose (CHO), glucose plus an amino acid mixture (CHO-AA-1), glucose plus an amino acid mixture with increased leucine concentration (CHO-AA-2), or water (PLA). CHO-AA-1 and CHO-AA-2 had reduced blood glucose responses compared with CHO, with no difference in insulin among these treatments. Experiment 2 rats were gavaged with either CHO or CHO-AA-1. Fifteen minutes after gavage, a bolus containing 2-[(3)H]deoxyglucose and [U-(14)C]mannitol was infused via a tail vein. Blood glucose was significantly lower in CHO-AA-1 than in CHO, whereas insulin responses were similar. Muscle glucose uptake was higher in CHO-AA-1 compared with CHO in both fast-twitch red (8.36 ± 1.3 vs. 5.27 ± 0.7 µmol·g(-1)·h(-1)) and white muscle (1.85 ± 0.3 vs. 1.11 ± 0.2 µmol·g(-1)·h(-1)). There was no difference in Akt/PKB phosphorylation between treatment groups; however, the amino acid treatment resulted in increased AS160 phosphorylation in both muscle fiber types. Glycogen synthase phosphorylation was reduced in fast-twitch red muscle of CHO-AA-1 compared with CHO, whereas mTOR phosphorylation was increased. These differences were not noted in fast-twitch white muscle. These findings suggest that amino acid supplementation can improve glucose tolerance by increasing skeletal muscle glucose uptake and intracellular disposal through enhanced intracellular signaling.

The effect of a low carbohydrate beverage with added protein on cycling endurance performance in trained athletes.

Ingesting carbohydrate plus protein during prolonged variable intensity exercise has demonstrated improved aerobic endurance performance beyond that of a carbohydrate supplement alone. The purpose of the present study was to determine if a supplement containing a mixture of different carbohydrates (glucose, maltodextrin, and fructose) and a moderate amount of protein given during endurance exercise would increase time to exhaustion (TTE), despite containing 50% less total carbohydrate than a carbohydrate-only supplement. We also sought post priori to determine if there was a difference in effect based on percentage of ventilatory threshold (VT) at which the subjects cycled to exhaustion. Fifteen trained male and female cyclists exercised on 2 separate occasions at intensities alternating between 45 and 70% VO2max for 3 hours, after which the workload increased to ∼74-85% VO2max until exhaustion. Supplements (275 mL) were provided every 20 minutes during exercise, and these consisted of a 3% carbohydrate/1.2% protein supplement (MCP) and a 6% carbohydrate supplement (CHO). For the combined group (n = 15), TTE in MCP did not differ from CHO (31.06 ± 5.76 vs. 26.03 ± 4.27 minutes, respectively, p = 0.064). However, for subjects cycling at or below VT (n = 8), TTE in MCP was significantly greater than for CHO (45.64 ± 7.38 vs. 35.47 ± 5.94 minutes, respectively, p = 0.006). There were no significant differences in TTE for the above VT group (n = 7). Our results suggest that, compared to a traditional 6% CHO supplement, a mixture of carbohydrates plus a moderate amount of protein can improve aerobic endurance at exercise intensities near the VT, despite containing lower total carbohydrate and caloric content.

Added protein maintains efficacy of a low-carbohydrate sports drink.

The purpose of the present study was to investigate the aerobic capacity characteristics of an isocaloric carbohydrate (CHO) plus protein (PRO) drink and a low-calorie CHO plus PRO drink against a traditional 6% CHO sports beverage. Twelve male and female trained cyclists exercised on 4 separate occasions at intensities that varied between 55 and 75% V(O2)max for 2.5 hours and then at 80% V(O2)max until fatigued. Supplements (255.4 +/- 9.1 mL) were provided every 20 minutes and consisted of a 4.5% carbohydrate plus 1.15% protein complex (CHO/PRO H), a 3% carbohydrate plus 0.75% protein complex (CHO/PRO L), a 6% carbohydrate supplement (CHO), or a placebo (PLA). Time to fatigue at 80% V(O2)max was significantly longer (p < 0.05) during the CHO (26.9 +/- 6.1 minutes, mean +/- SE), the CHO/PRO H (30.5 +/- 5.9 minutes), and the CHO/PRO L (28.9 +/- 6.5 minutes) trials compared with the PLA trial (14.7 +/- 3.4 minutes), with no significant differences among the CHO, CHO/PRO H, and CHO/PRO L treatments. In general, blood glucose, plasma insulin, and carbohydrate oxidation were elevated above PLA during the CHO, CHO/PRO H, and CHO/PRO L trials, whereas plasma free fatty acids, rating of perceived exertion, and fat oxidation values were lower during the CHO, CHO/PRO H, and CHO/PRO L trials compared with the PLA trial. Only minor differences in blood parameters occurred among the CHO, CHO/PRO H, and CHO/PRO L treatments. In summary, partially substituting PRO for CHO in a sports drink did not enhance aerobic capacity, but substitution was able to occur without loss of efficacy. Thus, adding PRO to a low-calorie CHO sports drink may be an effective strategy to enhance aerobic capacity while limiting carbohydrate and caloric consumption.

Improved cycling time-trial performance after ingestion of a caffeine energy drink.

CONTEXT: Not all athletic competitions lend themselves to supplementation during the actual event, underscoring the importance of preexercise supplementation to extend endurance and improve exercise performance. Energy drinks are composed of ingredients that have been found to increase endurance and improve physical performance. PURPOSE: The purpose of the study was to investigate the effects of a commercially available energy drink, ingested before exercise, on endurance performance. METHODS: The study was a double-blind, randomized, crossover design. After a 12-hr fast, 6 male and 6 female trained cyclists (mean age 27.3 +/- 1.7 yr, mass 68.9 +/- 3.2 kg, and VO2 54.9 +/- 2.3 ml x kg-1 x min-1) consumed 500 ml of either flavored placebo or Red Bull Energy Drink (ED; 2.0 g taurine, 1.2 g glucuronolactone, 160 mg caffeine, 54 g carbohydrate, 40 mg niacin, 10 mg pantothenic acid, 10 mg vitamin B6, and 10 microg vitamin B12) 40 min before a simulated cycling time trial. Performance was measured as time to complete a standardized amount of work equal to 1 hr of cycling at 70% Wmax. RESULTS: Performance improved with ED compared with placebo (3,690 +/- 64 s vs. 3,874 +/- 93 s, p < .01), but there was no difference in rating of perceived exertion between treatments. b-Endorphin levels increased during exercise, with the increase for ED approaching significance over placebo (p = .10). Substrate utilization, as measured by open-circuit spirometry, did not differ between treatments. CONCLUSION: These results demonstrate that consuming a commercially available ED before exercise can improve endurance performance and that this improvement might be in part the result of increased effort without a concomitant increase in perceived exertion.

Cereal and nonfat milk support muscle recovery following exercise.

BACKGROUND: This study compared the effects of ingesting cereal and nonfat milk (Cereal) and a carbohydrate-electrolyte sports drink (Drink) immediately following endurance exercise on muscle glycogen synthesis and the phosphorylation state of proteins controlling protein synthesis: Akt, mTOR, rpS6 and eIF4E. METHODS: Trained cyclists or triathletes (8 male: 28.0 +/- 1.6 yrs, 1.8 +/- 0.0 m, 75.4 +/- 3.2 kg, 61.0 +/- 1.6 ml O2*kg-1*min-1; 4 female: 25.3 +/- 1.7 yrs, 1.7 +/- 0.0 m, 66.9 +/- 4.6 kg, 46.4 +/- 1.2 mlO2*kg-1*min-1) completed two randomly-ordered trials serving as their own controls. After 2 hours of cycling at 60-65% VO2MAX, a biopsy from the vastus lateralis was obtained (Post0), then subjects consumed either Drink (78.5 g carbohydrate) or Cereal (77 g carbohydrate, 19.5 g protein and 2.7 g fat). Blood was drawn before and at the end of exercise, and at 15, 30 and 60 minutes after treatment. A second biopsy was taken 60 minutes after supplementation (Post60). Differences within and between treatments were tested using repeated measures ANOVA. RESULTS: At Post60, blood glucose was similar between treatments (Drink 6.1 +/- 0.3, Cereal 5.6 +/- 0.2 mmol/L, p < .05), but after Cereal, plasma insulin was significantly higher (Drink 123.1 +/- 11.8, Cereal 191.0 +/- 12.3 pmol/L, p < .05), and plasma lactate significantly lower (Drink 1.4 +/- 0.1, Cereal 1.00 +/- 0.1 mmol/L, p < .05). Except for higher phosphorylation of mTOR after Cereal, glycogen and muscle proteins were not statistically different between treatments. Significant Post0 to Post60 changes occurred in glycogen (Drink 52.4 +/- 7.0 to 58.6 +/- 6.9, Cereal 58.7 +/- 9.6 to 66.0 +/- 10.0 mumol/g, p < .05) and rpS6 (Drink 17.9 +/- 2.5 to 35.2 +/- 4.9, Cereal 18.6 +/- 2.2 to 35.4 +/- 4.4 %Std, p < .05) for each treatment, but only Cereal significantly affected glycogen synthase (Drink 66.6 +/- 6.9 to 64.9 +/- 6.9, Cereal 61.1 +/- 8.0 to 54.2 +/- 7.2%Std, p < .05), Akt (Drink 57.9 +/- 3.2 to 55.7 +/- 3.1, Cereal 53.2 +/- 4.1 to 60.5 +/- 3.7 %Std, p < .05) and mTOR (Drink 28.7 +/- 4.4 to 35.4 +/- 4.5, Cereal 23.0 +/- 3.1 to 42.2 +/- 2.5 %Std, p < .05). eIF4E was unchanged after both treatments. CONCLUSION: These results suggest that Cereal is as good as a commercially-available sports drink in initiating post-exercise muscle recovery.

Adding protein to a carbohydrate supplement provided after endurance exercise enhances 4E-BP1 and RPS6 signaling in skeletal muscle.

To examine the role of both endurance exercise and nutrient supplementation on the activation of mRNA translation signaling pathways postexercise, rats were subjected to a 3-h swimming protocol. Immediately following exercise, the rats were provided with a solution containing either 23.7% wt/vol carbohydrates (CHO), 7.9% wt/vol protein (Pro), 31.6% wt/vol (23.7% wt/vol CHO + 7.9% wt/vol Pro) carbohydrates and Pro (CP), or a placebo (EX). The rats were then killed at 0, 30, and 90 min postexercise, and phosphorylation states of mammalian target of rapamycin (mTOR), ribosomal S6 kinase (p70(S6K)), ribosomal protein S6 (rpS6), and 4E-binding protein 1 (4E-BP1), were analyzed by immunoblot analysis in the red and white quadriceps muscle. Results demonstrated that rat groups provided with any of the three nutritional supplements (CHO, Pro, CP) transiently increased the phosphorylation states of mTOR, 4E-BP1, rpS6, and p70(S6K) compared with EX rats. Although CHO, Pro, and CP supplements phosphorylated mTOR and p70(S6K) after exercise, only CP elevated the phosphorylation of rpS6 above all other supplements 30 min postexercise and 4E-BP1 30 and 90 min postexercise. Furthermore, the phosphorylation states of 4E-BP1 (r(2) = 0.7942) and rpS6 (r(2) = 0.760) were highly correlated to insulin concentrations in each group. These results suggest that CP supplementation may be most effective in activating the mTOR-dependent signaling pathway in the postprandial state postexercise, and that there is a strong relationship between the insulin concentration and the activation of enzymes critical for mRNA translation.