Acute dose toxicity evaluation of the food supplement calcium 3-hydroxy-3-methylbutyrate (HMB) in female Sprague Dawley rats.

The leucine metabolite, 3-hydroxy-3-methylbutyrate (HMB), primarily utilized as the calcium salt (CaHMB) has become one of the most widely used supplements and food ingredients to promote muscle health. While both CaHMB and HMB free acid have published sub-chronic toxicity studies, and CaHMB has published studies on genotoxicity, data are lacking on the acute dosing of HMB which is important for regulatory and transportation classification as well as in cases of accidental overconsumption. Therefore, an acute oral toxicity study was conducted with CaHMB following OECD 420 guidelines. One rat was used in the dosage sighting study and four rats were used in the main study. In both studies, rats were given a single oral dose of 2000 mg/kg body weight by gavage and monitored for 14 days following the dosage for changes in body weight, clinical signs as noted in OECD 420, and at the end of the study a necropsy was conducted to determine any gross tissue abnormalities. The dosage of CaHMB administered resulted in no deaths, no significant adverse clinical signs, and no findings of lesions or abnormal tissues. Under the Global Harmonized System of classification, CaHMB was found to be in the least toxic Category 5 or non-toxic.

ß-hydroxy-ß-methylbutyrate supplementation benefits the effects of resistance training on body fat reduction via increased irisin expression in white adipose tissue.

The effects of resistance training (RT) associated with calcium ß-hydroxyß-methylbutyrate (CaHMB) supplementation on the body composition and gene expression of cytokines related to skeletal muscle hypertrophy and adipose tissue metabolism were studied in rats. Male Wistar rats were divided into four groups of 12 animals: sedentary control (SC); sedentary supplemented (SS); resistance training control (RTC) and resistance training supplemented (RTS). Rats from RTC and RTS groups were submitted to an RT programme and those from SS and RTS groups received 1 mL of CaHMB (320 mg kg-1 day-1) by gavage, for 8 weeks. We evaluated: body composition; plasma lipid profile; the gene expression of interleukin (IL)-6, IL-10, IL-15 and fibronectin type III domain-containing protein 5 (FNDC-5) in skeletal muscle, and IL-6, mitochondrial uncoupling protein 1 (UCP-1) in white adipose tissue (WAT); and the concentration of irisin in WAT. Compared to RTC alone, the combination of CaHMB with RT (RTS) further reduced abdominal circumference (5.3%), Lee index (2.4%), fat percentage (24.4%), plasma VLDL cholesterol (16.8%) and triglycerides (17%) and increased the gene expression of FNDC-5 (78.9%) and IL-6 (47.4%) in skeletal muscle and irisin concentration (26.9%) in WAT. Neither RT nor CaHMB affected the protein percentage or the gene expression of IL-6 and UCP-1 in WAT and IL-10, IL-15 in skeletal muscle. In conclusion, CaHMB supplementation increased the beneficial effects of RT on body fat reduction and was associated with muscular genic expression of IL-6 and FNDC-5 and irisin concentration in WAT, despite the lack of change in protein mass and maximal strength.

Genotoxicity assessment of calcium ß-hydroxy-ß-methylbutyrate.

ß-Hydroxy-ß-methylbutyrate (HMB) is a leucine metabolite available in calcium salt (CaHMB) and free acid forms as a sports nutrition ergogenic aid. HMB has also been used to support muscle health in the elderly and other populations needing to maintain muscle mass. Several human studies have reported safety data for CaHMB, and rodent sub-chronic toxicity studies have been conducted; however, there are no published genotoxicity studies for HMB. Therefore, three studies (a bacterial reverse mutation test, an in vitro mammalian chromosomal aberration test, and an in vivo mammalian cell micronucleus test) were performed. In the Ames test, no changes in revertant colonies or background were noted with CaHMB concentrations up to 5000 µg per plate, either with or without metabolic activation in five bacterial strains. In the chromosomal aberration test, the number of aberrations associated with up to 2.5 mM CaHMB (long-term) or 10.0 mM (short-term) were similar to those observed for negative controls (<5%), and no polyploidy was observed. Lastly, in the mammalian micronucleus test, no changes in immature erythrocyte or micronuclei frequencies were observed in animals treated with up to 2000 mg·kg-1 body weight CaHMB. In conclusion, CaHMB was determined to have no genotoxic effects.

Subchronic (90-day) repeated dose toxicity study of 2-hydroxybenzylamine acetate in rats.

2-Hydroxybenzylamine (2-HOBA), a naturally occurring compound found in buckwheat, can protect cells and tissues from oxidative stress. In this study, 2-HOBA acetate was orally administered to male and female rats for 90 consecutive days at doses of 100, 500, and 1000 mg·kg BW-1·d-1 (n = 20 per sex/group). Subchronic administration of 2-HOBA was well tolerated at all dose levels. 2-HOBA-treated male rats were slightly heavier in the last weeks of the study, but this difference was very small (<5%), did not show a dose-response relationship, and was not observed in female rats. Similarly, some statistically significant changes in serum biochemistry and hematology parameters were noted, but these were not considered to be of biological or toxicological significance. Sporadic differences in organ weights were observed between groups, but all were small (<10%) and unlikely to indicate toxicity. The incidence of histopathological lesions was similar between treated and control groups across all organs. Based upon these findings, the no-observed-adverse-effect level was determined to be ≥ 1000 mg·kg BW-1·d-1, which was the highest dose tested. These results further support no toxicity associated with oral consumption of 2-HOBA acetate in rats and the continued development of 2-HOBA as a dietary supplement or functional food.

Subchronic (90-day) repeated dose oral toxicity study of 2-hydroxybenzylamine acetate in rabbit.

2-hydroxybenzylamine (2-HOBA), a naturally occurring compound found in buckwheat, has potential for use as a nutrition supplement due to its ability to protect against the damaging effects of oxidative stress. In a series of rodent toxicity studies, 2-HOBA acetate was well-tolerated and did not produce any toxic effects over 28 or 90 days of repeated oral administration. However, it remained necessary to test the potential toxicity of 2-HOBA acetate in a non-rodent species. In this investigation, 2-HOBA acetate was orally administered to male and female New Zealand White Rabbits for 90 days at doses of 100, 500, and 1000 mg·kg BW-1·day-1 (n = 5 per sex/group). As previously observed in rodents, 2-HOBA acetate administration was well tolerated. No toxic effects of 2-HOBA acetate were detected in body weight, feed consumption, hematology, blood chemistry, urine chemistry, organ weights, gross pathology or histopathology. Based on these findings, the no-observed-adverse-effect-level of 2-HOBA acetate in rabbits was determined to be 1000 mg·kg BW-1·day-1, which was the highest dose tested. These results provide further support for the safety of 2-HOBA acetate administration.

Acute and 28-day repeated dose toxicity evaluations of 2-hydroxybenzylamine acetate in mice and rats.

2-hydroxybenzylamine (2-HOBA), a compound naturally found in buckwheat, has been shown to protect cells and tissues from the damaging effects of oxidative stress. The purpose of this report was to evaluate 2-HOBA in preclinical oral rodent toxicity studies. This report includes the results from three oral toxicity studies in rodents: a preliminary 28-day feeding study in mice, a 14-day acute oral toxicity study in rats, and a 28-day repeated dose oral toxicity study in rats. The preliminary mouse feeding study showed no adverse effects of 2-HOBA at concentrations up to 0.456% by weight in feed, but decreased food intake and weight loss were observed at 1.56% 2-HOBA in the diet, likely due to poor palatability. In the acute dosing study, 2000 mg/kg BW 2-HOBA resulted in mortality in one of the six tested female rats, indicating a median lethal dose of 2500 mg/kg BW. In the 28-day repeated oral dose study, small differences were observed between 2-HOBA treated and control group rats, but none of these differences were determined to be of toxicological significance. Together, these studies support the lack of toxicity of oral administration of 2-HOBA acetate at doses up to 1000 mg/kg BW d-1 in rodents.

Oral Adenosine-5'-triphosphate (ATP) Administration Increases Postexercise ATP Levels, Muscle Excitability, and Athletic Performance Following a Repeated Sprint Bout.

OBJECTIVE: Oral adenosine-5'-triphosphate (ATP) administration has failed to increase plasma ATP levels; however, chronic supplementation with ATP has shown to increase power, strength, lean body mass, and blood flow in trained athletes. The purpose of this study was to investigate the effects of ATP supplementation on postexercise ATP levels and on muscle activation and excitability and power following a repeated sprint bout. METHODS: In a double-blind, placebo-controlled, randomized design, 42 healthy male individuals were given either 400 mg of ATP as disodium salt or placebo for 2 weeks prior to an exercise bout. During the exercise bout, muscle activation and excitability (ME, ratio of power output to muscle activation) and Wingate test peak power were measured during all sprints. ATP and metabolites were measured at baseline, after supplementation, and immediately following exercise. RESULTS: Oral ATP supplementation prevented a drop in ATP, adenosine-5'-diphosphate (ADP), and adenosine-5'-monophosphate (AMP) levels postexercise (p < 0.05). No group by time interaction was observed for muscle activation. Following the supplementation period, muscle excitability significantly decreased in later bouts 8, 9, and 10 in the placebo group (-30.5, -28.3, and -27.9%, respectively; p < 0.02), whereas ATP supplementation prevented the decline in later bouts. ATP significantly increased Wingate peak power in later bouts compared to baseline (bout 8: +18.3%, bout 10: +16.3%). CONCLUSIONS: Oral ATP administration prevents exercise-induced declines in ATP and its metabolite and enhances peak power and muscular excitability, which may be beneficial for sports requiring repeated high-intensity sprinting bouts.

Interaction of Beta-Hydroxy-Beta-Methylbutyrate Free Acid and Adenosine Triphosphate on Muscle Mass, Strength, and Power in Resistance Trained Individuals.

Lowery, RP, Joy, JM, Rathmacher, JA, Baier, SM, Fuller, JC Jr, Shelley, MC II, Jäger, R, Purpura, M, Wilson, SMC, and Wilson, JM. Interaction of beta-hydroxy-beta-methylbutyrate free acid and adenosine triphosphate on muscle mass, strength, and power in resistance trained individuals. J Strength Cond Res 30(7): 1843-1854, 2016-Adenosine-5'-triphosphate (ATP) supplementation helps maintain performance under high fatiguing contractions and with greater fatigue recovery demands also increase. Current evidence suggests that the free acid form of ß-hydroxy-ß-methylbutyrate (HMB-FA) acts by speeding regenerative capacity of skeletal muscle after high-intensity or prolonged exercise. Therefore, we investigated the effects of 12 weeks of HMB-FA (3 g) and ATP (400 mg) administration on lean body mass (LBM), strength, and power in trained individuals. A 3-phase double-blind, placebo-, and diet-controlled study was conducted. Phases consisted of an 8-week periodized resistance training program (phase 1), followed by a 2-week overreaching cycle (phase 2), and a 2-week taper (phase 3). Lean body mass was increased by a combination of HMB-FA/ATP by 12.7% (p < 0.001). In a similar fashion, strength gains after training were increased in HMB-FA/ATP-supplemented subjects by 23.5% (p < 0.001). Vertical jump and Wingate power were increased in the HMB-FA/ATP-supplemented group compared with the placebo-supplemented group, and the 12-week increases were 21.5 and 23.7%, respectively. During the overreaching cycle, strength and power declined in the placebo group (4.3-5.7%), whereas supplementation with HMB-FA/ATP resulted in continued strength gains (1.3%). In conclusion, HMB-FA and ATP in combination with resistance exercise training enhanced LBM, power, and strength. In addition, HMB-FA plus ATP blunted the typical response to overreaching, resulting in a further increase in strength during that period. It seems that the combination of HMB-FA/ATP could benefit those who continuously train at high levels such as elite athletes or military personnel.

Comparison of availability and plasma clearance rates of ß-hydroxy-ß-methylbutyrate delivery in the free acid and calcium salt forms.

ß-Hydroxy-ß-methylbutyrate (HMB), a leucine metabolite, has long been supplemented as a Ca salt (Ca-HMB) to increase strength and performance gains with exercise and to reduce recovery time. Recently, the free acid form of HMB (HMB-FA) has become commercially available in capsule form (gelcap). The current study was conducted to compare the bioavailability of HMB using the two commercially available capsule forms of HMB-FA and Ca-HMB. We also compared the pharmacokinetics of each form when administered mixed in water. Ten human subjects (five male and five female) were studied in a randomised crossover design. There was no significant sex by treatment interaction for any of the pharmacokinetic parameters measured. HMB-FA administered in capsules was more efficient than Ca-HMB capsule at HMB delivery with a 37 % increase in plasma clearance rate (74·8 (sem 4·0) v. 54·5 (sem 3·2) ml/min, P<0·0001) and a 76 % increase in peak plasma HMB concentration (270·2 (sem 17·8) v. 153·9 (sem 17·9) µmol/l, P<0·006), which was reached in one-third the time (P<0·009). When HMB-FA and Ca-HMB were administered in water, the differences still favoured HMB-FA, albeit to a lesser degree. Plasma HMB with HMB-FA administered in water was greater during the early phase of absorption (up to 45 min postadministration, P<0·05); this resulted in increased AUC during the first 60 min after administration, when compared with Ca-HMB mixed in water (P<0·03). In conclusion, HMB-FA in capsule form improves clearance rate and availability of HMB compared with Ca-HMB in capsule form.

Inflammation Biomarker Response to Oral 2-Hydroxybenzylamine (2-HOBA) Acetate in Healthy Humans.

Inflammation is associated with the formation of reactive oxygen species (ROS) and the formation of lipid-derived compounds, such as isolevuglandins (IsoLGs), malondialdehyde, 4-hydroxy-nonenal, and 4-oxo-nonenal. The most reactive of these are the IsoLGs, which form covalent adducts with lysine residues and other cellular primary amines leading to changes in protein function, immunogenicity, and epigenetic alterations and have been shown to contribute to a number of inflammatory diseases. 2-Hydroxybenzylamine (2-HOBA) is a natural compound found in buckwheat seeds and reacts with all IsoLG adducts preventing adduct formation with proteins and DNA. Therefore, 2-HOBA is well positioned as an agent for the prevention of inflammatory-prone diseases. In this study, we examined the potential beneficial effects of 2-HOBA on oxidative stress and inflammatory biomarkers in two cohorts of healthy younger and older adults. We utilized the Olink® targeted inflammation panel before and after an oral 15-day treatment regimen with 2-HOBA. We found significant relative changes in the plasma concentration of 15 immune proteins that may reflect the in vivo immune targets of 2-HOBA. Treatment of 2-HOBA resulted in significant increased levels of CCL19, IL-12ß, IL-20Rα, and TNFß, whereas levels of TWEAK significantly decreased. Ingenuity Pathway Analysis identified canonical pathways regulated by the differentially secreted cytokines, chemokines, and growth factors upon 2-HOBA treatment and further points to biofunctions related to the recruitment, attraction, and movement of different immune cell types. In conclusion, 2-HOBA significantly altered the protein biomarkers CCL19, IL-12ß, IL-20Rα, TNFß, and TWEAK, and these may be responsible for the protective effects of 2-HOBA against reactive electrophiles, such as IsoLGs, commonly expressed in conditions of excessive oxidative stress. 2-HOBA has a role as a IsoLG scavenger to proactively improve immune health in a variety of conditions.

The nutraceutical electrophile scavenger 2-hydroxybenzylamine (2-HOBA) attenuates gastric cancer development caused by Helicobacter pylori.

Stomach cancer is a leading cause of cancer death. Helicobacter pylori is a bacterial gastric pathogen that is the primary risk factor for carcinogenesis, associated with its induction of inflammation and DNA damage. Dicarbonyl electrophiles are generated from lipid peroxidation during the inflammatory response and form covalent adducts with amine-containing macromolecules. 2-hydroxybenzylamine (2-HOBA) is a natural compound derived from buckwheat seeds and acts as a potent scavenger of reactive aldehydes. Our goal was to investigate the effect of 2-HOBA on the pathogenesis of H. pylori infection. We used transgenic FVB/N insulin-gastrin (INS-GAS) mice as a model of gastric cancer. First, we found that 2-HOBA is bioavailable in the gastric tissues of these mice after supplementation in the drinking water. Moreover, 2-HOBA reduced the development of gastritis in H. pylori-infected INS-GAS mice without affecting the bacterial colonization level in the stomach. Further, we show that the development of gastric dysplasia and carcinoma was significantly reduced by 2-HOBA. Concomitantly, DNA damage were also inhibited by 2-HOBA treatment in H. pylori-infected mice. In parallel, DNA damage was inhibited by 2-HOBA in H. pylori-infected gastric epithelial cells in vitro. In conclusion, 2-HOBA, which has been shown to be safe in human clinical trials, represents a promising nutritional compound for the chemoprevention of the more severe effects of H. pylori infection.

Electrophilic reactive aldehydes as a therapeutic target in colorectal cancer prevention and treatment.

Colorectal cancer (CRC) is a major health problem worldwide. Dicarbonyl electrophiles, such as isolevuglandins (isoLGs), are generated from lipid peroxidation and form covalent adducts with amine-containing macromolecules. We have shown high levels of adducts of isoLGs in colonic epithelial cells of patients with CRC. We thus investigated the role of these reactive aldehydes in colorectal cancer development. We found that 2-hydroxybenzylamine (2-HOBA), a natural compound derived from buckwheat seeds that acts as a potent scavenger of electrophiles, is bioavailable in the colon of mice after supplementation in the drinking water and does not affect the colonic microbiome. 2-HOBA reduced the level of isoLG adducts to lysine as well as tumorigenesis in models of colitis-associated carcinogenesis and of sporadic CRC driven by specific deletion of the adenomatous polyposis coli gene in colonic epithelial cells. In parallel, we found that oncogenic NRF2 activation and signaling were decreased in the colon of 2-HOBA-treated mice. Additionally, the growth of xenografted human HCT116 CRC cells in nude mice was significantly attenuated by 2-HOBA supplementation. In conclusion, 2-HOBA represents a promising natural compound for the prevention and treatment of CRC.

Small-Scale Randomized Controlled Trial to Explore the Impact of ß-Hydroxy-ß-Methylbutyrate Plus Vitamin D3 on Skeletal Muscle Health in Middle Aged Women.

ß-Hydroxy-ß-methylbutyrate (HMB), a leucine metabolite, can increase skeletal muscle size and function. However, HMB may be less effective at improving muscle function in people with insufficient Vitamin D3 (25-OH-D < 30 ng/mL) which is common in middle-aged and older adults. Therefore, we tested the hypothesis that combining HMB plus Vitamin D3 (HMB + D) supplementation would improve skeletal muscle size, composition, and function in middle-aged women. In a double-blinded fashion, women (53 ± 1 yrs, 26 ± 1 kg/m2, n = 43) were randomized to take placebo or HMB + D (3 g Calcium HMB + 2000 IU D per day) during 12 weeks of sedentary behavior (SED) or resistance exercise training (RET). On average, participants entered the study Vitamin D3 insufficient while HMB + D increased 25-OH-D to sufficient levels after 8 and 12 weeks. In SED, HMB + D prevented the loss of arm lean mass observed with placebo. HMB + D increased muscle volume and decreased intermuscular adipose tissue (IMAT) volume in the thigh compared to placebo but did not change muscle function. In RET, 12-weeks of HMB + D decreased IMAT compared to placebo but did not influence the increase in skeletal muscle volume or function. In summary, HMB + D decreased IMAT independent of exercise status and may prevent the loss or increase muscle size in a small cohort of sedentary middle-aged women. These results lend support to conduct a longer duration study with greater sample size to determine the validity of the observed positive effects of HMB + D on IMAT and skeletal muscle in a small cohort of middle-aged women.

Free acid gel form of ß-hydroxy-ß-methylbutyrate (HMB) improves HMB clearance from plasma in human subjects compared with the calcium HMB salt.

The leucine metabolite, ß-hydroxy-ß-methylbutyrate (HMB), is a nutritional supplement that increases lean muscle and strength with exercise and in disease states. HMB is presently available as the Ca salt (CaHMB). The present study was designed to examine whether HMB in free acid gel form will improve HMB availability to tissues. Two studies were conducted and in each study four males and four females were given three treatments in a randomised, cross-over design. Treatments were CaHMB (gelatin capsule, 1 g), equivalent HMB free acid gel swallowed (FASW) and free acid gel held sublingual for 15 s then swallowed (FASL). Plasma HMB was measured for 3 h following treatment in study 1 and 24 h with urine collection in study 2. In both the studies, the times to peak plasma HMB were 128 (sem 11), 38 (sem 4) and 38 (sem 1) min (P < 0·0001) for CaHMB, FASW and FASL, respectively. The peak concentrations were 131 (sem 6), 249 (sem 14) and 239 (sem 14) µmol/l (P < 0·0001) for CaHMB, FASW and FASL, respectively. The areas under the curve were almost double for FASW and FASL (P < 0·0001). Daily urinary HMB excretion was not significantly increased resulting in more HMB retained (P < 0·003) with FASW and FASL. Half-lives were 3·17 (sem 0·22), 2·50 (sem 0·13) and 2·51 (sem 0·14) h for CaHMB, FASW and FASL, respectively (P < 0·004). Free acid gel resulted in quicker and greater plasma concentrations (+185%) and improved clearance (+25%) of HMB from plasma. In conclusion, HMB free acid gel could improve HMB availability and efficacy to tissues in health and disease.

Dose Response of Acute ATP Supplementation on Strength Training Performance.

Background: Chronic oral ATP supplementation benefits cardiovascular health, muscular performance, body composition, and recovery while attenuating muscle breakdown and fatigue. A single 400 mg dose of oral ATP supplementation improved lower body resistance training performance and energy expenditure in recreational resistance trained males, however, the minimal effective dose is currently unknown. Materials and Methods: Twenty recreationally trained men (age 28.6 ± 1.0 years, body mass 81.2 ± 2.0 kg, height 175.2 ± 1.4 cm, 1RM 141.5 ± 5.0 kg) consumed a single dose of either 400 mg, 200 mg, or 100 mg ATP (PEAK ATP®, TSI USA LLC, Missoula, MT, USA) or a placebo in a randomized, placebo-controlled crossover design, separated by a one week wash out between treatments. After warm-up, participants performed 4 sets of half-squats using free-weights until movement failure separated by 2 mins of rest between sets. Results: In comparison to placebo, 400 mg ATP significantly increased the number of set 1 repetitions (+13%, p = 0.04), and numerically increased total repetitions (+7%, p = 0.19) and total weight lifted (+6%, p = 0.22). 200 mg ATP numerically increased set 1 repetitions (+4% p = 0.47), while 100 mg ATP showed no improvements over placebo. 100 mg ATP (-4%, p < 0.05) and 400 mg ATP (-4%, p = 0.11) decreased the perceived rate of exertion compared to placebo. Conclusions: In this study, the effective minimal dose of acute oral ATP supplementation during resistance exercise to increase performance was determined to be 400 mg, while as little as 100 mg showed improvements in perceived exertion.

2-Hydroxybenzylamine (2-HOBA) to prevent early recurrence of atrial fibrillation after catheter ablation: protocol for a randomized controlled trial including detection of AF using a wearable device.

BACKGROUND: Although catheter ablation is an effective therapy for atrial fibrillation (AF), the most common cardiac arrhythmia encountered in clinical practice, AF ablation generates inflammation and oxidative stress in the early postoperative period predisposing to recurrence of AF. Isolevuglandins (IsoLGs) are reactive lipid mediators of oxidative stress injury that rapidly react with endogenous biomolecules to compromise their function. 2-Hydroxybenzylamine (2-HOBA), a potent small molecule scavenger of IsoLGs, sequesters the reactive species as inert adducts. This mechanism, coupled with reported safety in humans, supports the investigation of 2-HOBA as a novel therapeutic to reduce AF caused by oxidative stress, such as that which occurs after catheter ablation. Accordingly, we seek to test the hypothesis that treatment with 2-HOBA will decrease early recurrence of AF and other atrial arrhythmias following AF ablation by decreasing IsoLG adducts with native biomolecules. METHODS: The proposed trial will randomly assign 162 participants undergoing cryo- or radiofrequency catheter ablation for AF to 2-HOBA (N = 81) or placebo (N = 81). Individuals will begin the study drug 3 days prior to ablation and continue for 28 days. Participants will be given a wearable smartwatch capable of detecting and recording atrial arrhythmias. They will be instructed to record ECGs daily with additional ECGs if they experience symptoms of AF or when alerted by the smartwatch AF detection alarm. The primary clinical endpoint will be an episode of AF, atrial tachycardia, or atrial flutter lasting 30 s or more within 28 days post-AF ablation. Secondary measures will be the change in IsoLG adduct levels from blood samples collected immediately pre-ablation and post-ablation and reduction in AF burden as calculated from the smartwatch. DISCUSSION: The proposed trial will test the hypothesis that 2-HOBA reduces post-ablation atrial arrhythmias through sequestration of reactive IsoLG species. The results of this study may improve the understanding of the role of IsoLGs and oxidative stress in AF pathogenesis and provide evidence to advance 2-HOBA and related compounds as a new therapeutic strategy to treat AF. TRIAL REGISTRATION: ClinicalTrials.gov NCT04433091 . Registered on June 3, 2020.

Dileucine ingestion is more effective than leucine in stimulating muscle protein turnover in young males: a double blind randomized controlled trial.

Leucine is regarded as an anabolic trigger for the mTORC1 pathway and the stimulation muscle protein synthesis rates. More recently, there has been an interest in underpinning the relevance of branched-chain amino acid (BCAA)-containing dipeptides and their intact absorption into circulation to regulate muscle anabolic responses. We investigated the effects of dileucine and leucine ingestion on postprandial muscle protein turnover. Ten healthy young men (age: 23 ± 3 yr) consumed either 2 g of leucine (LEU) or 2 g of dileucine (DILEU) in a randomized crossover design. The participants underwent repeated blood and muscle biopsy sampling during primed continuous infusions of l-[ring-13C6]phenylalanine and l-[15N]phenylalanine to determine myofibrillar protein synthesis (MPS) and mixed muscle protein breakdown rates (MPB), respectively. LEU and DILEU similarly increased plasma leucine net area under the curve (AUC; P = 0.396). DILEU increased plasma dileucine AUC to a greater extent than LEU (P = 0.013). Phosphorylation of Akt (P = 0.002), rpS6 (P < 0.001), and p70S6K (P < 0.001) increased over time under both LEU and DILEU conditions. Phosphorylation of 4E-BP1 (P = 0.229) and eEF2 (P = 0.999) did not change over time irrespective of condition. Cumulative (0-180 min) MPS increased in DILEU (0.075 ± 0.032%·h-1), but not in LEU (0.047 ± 0.029%·h-1; P = 0.023). MPB did not differ between LEU (0.043 ± 0.030%·h-1) and DILEU conditions (0.051 ± 0.027%·h-1; P = 0.659). Our results showed that dileucine ingestion elevated plasma dileucine concentrations and muscle protein turnover by stimulating MPS in young men.NEW & NOTEWORTHY The role of dipeptides as anabolic agents remains unresolved in humans. We show that the ingestion of 2 g dileucine increased plasma dileucine concentrations and resulted in an enhancement of muscle protein turnover by stimulating an increase in muscle protein synthesis rates in healthy young males. The ingestion of 2 g leucine, however, did not stimulate an increase in muscle protein turnover. Our work provides the first insights into the effects of dipeptides on human protein metabolism.

Long-term Effects of Calcium ß-Hydroxy-ß-Methylbutyrate and Vitamin D3 Supplementation on Muscular Function in Older Adults With and Without Resistance Training: A Randomized, Double-blind, Controlled Study.

The primary aim of this study was to determine whether supplementation with calcium ß-hydroxy-ß-methylbutyrate (HMB) and vitamin D3 (D) would enhance muscle function and strength in older adults. Older adults over 60 years of age with insufficient circulating 25-hydroxy-vitamin D (25OH-D) levels were enrolled in a double-blinded controlled 12-month study. Study participants were randomly assigned to treatments consisting of: (a) Control + no exercise, (b) HMB+D + no exercise, (c) Control + exercise, and (d) HMB+D + exercise. The study evaluated 117 participants via multiple measurements over the 12 months that included body composition, strength, functionality, and questionnaires. HMB+D had a significant benefit on lean body mass within the nonexercise group at 6 months (0.44 ± 0.27 kg, HMB+D vs -0.33 ± 0.28 kg, control, p < .05). In nonexercisers, improvement in knee extension peak torque (60°/s) was significantly greater in HMB+D-supplemented participants than in the nonsupplemented group (p = .04) at 3 months, 10.9 ± 5.7 Nm and -5.2 ± 5.9 Nm, respectively. A composite functional index, integrating changes in handgrip, Get Up, and Get Up and Go measurements, was developed. HMB+D + no exercise resulted in significant increases in the functional index compared with those observed in the control + no exercise group at 3 (p = .03), 6 (p = .04), and 12 months (p = .04). Supplementation with HMB+D did not further improve the functional index within the exercising group. This study demonstrated the potential of HMB and vitamin D3 supplementation to enhance muscle strength and physical functionality in older adults, even in individuals not engaged in an exercise training program.