Whey protein supplementation, containing large amount of leucine, has been a traditional intervention to maintain net protein balance in the past decades. It has been recognized that leucine alone is able to stimulate protein synthesis by activating mTOR and its related downstream pathway without affecting protein degradation, whereas its metabolite β-hydroxy-β-methylbutyrate (HMB) is known to attenuate protein degradation when provided chronically. However, the mechanism of HMB’s benefit remains unclear. To address how HMB regulates protein synthesis and degradation signaling pathways, we compared one dose of whey protein (187.5mg/kg), HMB (400mg/kg) or leucine (1.4g/kg) by oral gavage. Blood was collected at 0, 45 and 90 min for blood glucose and plasma insulin analysis. Red and white gastrocnemius muscle was taken separately 90 min after gavage. Blood glucose was reduced by leucine at 45 and 90 min post gavage. Plasma insulin was enhanced by leucine at 45 min and then decreased at 90 min post gavage, whereas HMB decreased plasma insulin through 90 min post gavege. Western blot analysis showed that HMB phosphorylated Akt in red gastronemius, and enhanced phosphorylation of mTOR in both types of muscles. Leucine phosphorylated mTOR, p70s6k and 4E-BP1 in both red and white gastronemius. Regarding protein degradation signals, phosphorylation of FOXO3A was enhanced by HMB, but not in the other treatment groups. Whey protein had no effect on those cellular signaling. Our results indicate that both HMB and leucine may stimulate protein synthesis through the mTOR pathway in red and white gastrocnemius muscles by different degrees with leucine more effective than HMB. HMB may have a greater effect than leucine on limiting protein degradation by phosphorylating Akt and FOXO3A in red and white gastrocnemius muscles. A combination of HMB and leucine, as a new interventional strategy, is predicted to maximize protein accretion by increasing protein synthesis as well as inhibiting protein degradation. / text

Consumption of β-hydroxy β-methylbutyrate (HBM), a leucine metabolite, alters muscle composition and metabolism leading to strength and agility improvements in human athletes. To determine if HMB affects athletic performance and muscle function in horses, Thoroughbred geldings were fed a control (CON; n=5) or HMB (n=6) supplement (30 mg/kg/day) for 6 weeks prior to completing a standardized exercise test (SET). Gluteus medius (GM) muscle samples were obtained before the SET for fiber-typing and venous blood was collected before and immediately upon completion of the SET for lactate measurements. Heart rate (HR), biceps femoris (BF) and semitendinosus (ST) surface electromyograms, and fore- and hindlimb metacarpophalangeal joint angles were captured for the duration of the SET. Results demonstrate that HMB supplementation increased (P < 0.05) the percentage of type IIA muscle fibers in the GM with a corresponding decrease (P < 0.05) in type IIX fibers. The percentage of type I fibers was unaffected by diet. Supplementation with HMB did not result in any significant effects on performance, muscle function or biomechanical properties by comparison to CON. Increasing treadmill speed resulted in an increase (P < 0.05) in stride length and maximal extension angle of the fore fetlock, and a shortening (P < 0.05) of the stance phase of the gait cycle. Integrated EMG (iEMG) increased (P < 0.05) with increasing treadmill speeds for both the BF and ST, with the BF exhibiting greater iEMG values than the ST. In summary, HMB increased the percentage of type IIA fibers which did not translate into immediate, improved athletic performance / Master of Science / Muscles depend on their fibers, innervation, energy supply, and blood flow to contract. Failure to meet one or more of these requirements precludes muscle tissue from performing work, situation termed fatigue. Identification of fatigue indicators is of interest to the horse industry for a number of reasons, including horse and human safety, prevention of unnecessary expenses, and general public opinion of the sport disciplines. Diet supplementation with legal, performanceenhancing compounds is of interest to riders and horse owners alike. Molecules such as betahydroxy beta-methylbutyrate (HMB) improve muscle function, protein synthesis, and muscle tissue repair. Assessment of the athletic capacity and performance of horses by evaluating fatigue indicators favors responsible training regimes. Techniques to achieve this goal include muscle sampling, biochemical, electromyographic, and biomechanical analysis. We hypothesized that dietary supplementation of HMB would have positive effects on the athletic performance of horses. This study evaluated the effects of 45-day HMB supplementation on muscle fiber composition, muscle performance, and rates of fatigue in adult Thoroughbred horses by use of a high-speed treadmill. Muscle biopsies, blood lactate, high-speed video captures, and electromyography were analyzed. These analyses revealed that HMB supplementation increased the number of fatigue-resistant fibers in muscles but caused no substantial, immediate improvements on the athletic performance of horses.

Purpose: To examine the effects of 28 days of β-hydroxy-β-methylbutyrate free acid (HMB) and high-intensity interval training (HIIT) on maximal oxygen consumption (VO2peak), ventilatory threshold (VT), respiratory compensation point (RCP) and time to exhaustion (Tmax) in collegeaged men and women. Methods: Healthy men and women (n=34, age and VO2peak= 22.7+3.1yr and 39.3+5.0 mL. kg-1.min-1, respectively) participated in this study. All participants completed a series of tests prior and subsequent to treatment. A maximal oxygen consumption test was performed on a cycle ergometer to assess VO2peak, Tmax, VT, and RCP. The peak power output (Ppeak), power at VT (PVT) and power at RCP (PRCP) were also recorded from this test. Twenty-six subjects completed 12 HIIT (80-120% maximal workload) exercise sessions consisting of 5-6 bouts of a 2:1 minute cycling work to rest ratio protocol over a four-week period, while eight served as controls (CTL). In double-blind fashion, the HIIT groups were assigned into either a placebo (HIIT) or 3g per day of HMB (HMB-HIIT). Body composition was measured with dual energy x-ray absorptiometry (DEXA). Outcomes were assessed by ANCOVA with posttest means adjusted for pretest differences. Results: The HMB-HIIT intervention showed significant (p & lt 0.05) gains in VO2peak, VT, and PVT versus the CTL and HIIT group. Both HIIT and HMB-HIIT treatment groups demonstrated significant (p < 0.05) improvement over CTL for Ppeak, Tmax, RCP, and PRCP with no significant difference between the treatment groups. There were no significant differences observed for any measures of body composition. An independent-samples t-test confirmed that there were no significant differences between the training volumes for the HIIT and HMB-HIIT groups. Conclusion: These findings suggest that the addition of HMB supplementation may result in greater changes in VO2peak and VT than HIIT alone. Therefore, in college-aged men and women, the use of HMB supplementation may enhance the benefits of HIIT on aerobic performance measures.

This study investigated the effects of high intensity interval training (HIIT) and β-hydroxy-β-methylbutyric free acid (HMBFA) on the onset of neuromuscular fatigue in healthy young men and women. Thirty-seven subjects (22 men and 15 women; mean ± SD age = 22.8 ± 3.4yrs) completed an incremental cycle ergometer test (GXT) to exhaustion. During the GXT, electromyography (EMG) signals from the right vastus lateralis were recorded to determine the power output at the onset of neuromuscular fatigue (PWCFT), and peak wattage was used to assign individual training loads. After baseline testing (PRE), subjects were randomly assigned to one of three groups: control (C, n = 9), training with placebo (P, n = 14), or training with supplementation (S, n = 14). Subjects assigned to P and S completed 12 HIIT sessions over 4 weeks while subjects assigned to C were asked to maintain their normal diet and activity patterns. After 4 weeks, subjects returned for post-testing (POST). The PWCFT values (W) were determined using a DMAX method. The EMG amplitude root mean square (µVrms) values were used to generate a third-order polynomial regression (3PR) representing the increase in µVrms versus time of the GXT. The onset of fatigue (TF) was defined as the x-value (time, s) of the point on the 3PR that measured the maximal perpendicular distance from the line between the first and last data points. TF was used to estimate PWCFT according to the equation: PWCFT = PO + a · (n/N), where PO is the power output of the stage in which TF occurred, a is the increment in power output between GXT stages (25W), n is the difference (s) between TF and the beginning of the stage during which TF occurred, and N is the duration of a stage (120s). A two-way repeated measures ANOVA was used to identify group × time interaction for PWCFT. If a significant interaction occurred, one-way factorial ANOVAs were used. Fisher's least significant difference post hoc comparisons were performed between groups. If a significant main effect occurred, dependent samples t-tests with Bonferroni corrections (p = [0.05/3] = 0.017) were performed across time for each group. The two-way ANOVA resulted in a significant interaction (F = 6.69, p = 0.004). Follow-up analysis with one-way ANOVA resulted in no difference among groups at PRE (F = 0.87, p = 0.43), however a significant difference was shown for POST values (F = 5.46, p = 0.009). Post-hoc analysis among POST values showed significant differences between S and both P (p = 0.034) and C (p = 0.003). No differences (p = 0.226) were noted between P and C. Paired samples t-tests detected significant changes following HIIT for S (p < 0.001) and P (p = 0.016), but no change in C (p = 0.473). Results of this study indicate that HIIT was effective in delaying the onset of fatigue, but supplementation with HMBFA in conjunction with HIIT was more effective than HIIT alone. An increase in PWCFT represents an increase in the maximal power output an individual can sustain without eliciting fatigue. Therefore, HIIT can be used to improve performance in both endurance activities as well as intermittent sports. In addition, HMBFA supplementation is a simple method that can be used to maximize the benefits of HIIT. Acknowledgments: Metabolic Technologies, Inc. provided the supplement and funding for this study.