Muscle transcriptomic circuits linked to periarticular physiology in end-stage osteoarthritis.

The ability of individuals with end-stage osteoarthritis (OA) to functionally recover from total joint arthroplasty is highly inconsistent. The molecular mechanisms driving this heterogeneity have yet to be elucidated. Furthermore, OA disproportionately impacts females, suggesting a need for identifying female-specific therapeutic targets. We profiled the skeletal muscle transcriptome in females with end-stage OA (n = 20) undergoing total knee or hip arthroplasty using RNA-Seq. Single-gene differential expression (DE) analyses tested for DE genes between skeletal muscle overlaying the surgical (SX) joint and muscle from the contralateral (CTRL) leg. Network analyses were performed using Pathway-Level Information ExtractoR (PLIER) to summarize genes into latent variables (LVs), i.e., gene circuits, and link them to biological pathways. LV differences in SX versus CTRL muscle and across sources of muscle tissue (vastus medialis, vastus lateralis, or tensor fascia latae) were determined with ANOVA. Linear models tested for associations between LVs and muscle phenotype on the SX side (inflammation, function, and integrity). DE analysis revealed 360 DE genes (|Log2 fold-difference| ≥ 1, FDR ≤ 0.05) between the SX and CTRL limbs, many associated with inflammation and lipid metabolism. PLIER analyses revealed circuits associated with protein degradation and fibro-adipogenic cell gene expression. Muscle inflammation and function were linked to an LV associated with endothelial cell gene expression highlighting a potential regulatory role of endothelial cells within skeletal muscle. These findings may provide insight into potential therapeutic targets to improve OA rehabilitation before and/or following total joint replacement.

Perioperative assessment of muscle inflammation susceptibility in patients with end-stage osteoarthritis.

Many individuals with end-stage osteoarthritis (OA) undergo elective total hip/knee arthroplasty (THA/TKA) to relieve pain, improve mobility and quality of life. However, ∼30% suffer long-term mobility impairment following surgery. This may be in part due to muscle inflammation susceptibility (MuIS+), an overt proinflammatory pathology localized to skeletal muscle surrounding the diseased joint, present in some patients with TKA/THA. We interrogated the hypothesis that MuIS+ status results in a perturbed perioperative gene expression profile and decreases skeletal muscle integrity in patients with end-stage OA. Samples were leveraged from the two-site, randomized, controlled trial R01HD084124, NCT02628795. Participants were dichotomized based on surgical (SX) muscle gene expression of TNFRSF1A (TNF-αR). MuIS+/- samples were probed for gene expression and fibrosis. Paired and independent two-tailed t tests were used to determine differences between contralateral (CTRL) and surgical (SX) limbs and between-subject comparisons, respectively. Significance was declared at P < 0.05. Seventy participants (26M/44F; mean age 62.41 ± 8.86 yr; mean body mass index 31.10 ± 4.91 kg/m2) undergoing THA/TKA were clustered as MuIS+ (n = 24) or MuIS- (n = 46). Lower skeletal muscle integrity (greater fibrosis) exists on the SX versus CTRL limb (P < 0.001). Furthermore, MuIS+ versus MuIS- muscle exhibited higher proinflammatory (IL-6R and TNF-α) and catabolic (TRIM63) gene expression (P < 0.001, P = 0.004, and 0.024 respectively), with a trend for greater fibrosis (P = 0.087). Patients with MuIS+ exhibit more inflammation and catabolic gene expression in skeletal muscle of the SX limb, accompanied by decreased skeletal muscle integrity (Trend). This highlights the impact of MuIS+ status emphasizing the potential value of perioperative MuIS assessment to inform optimal postsurgical care.NEW & NOTEWORTHY This study assessed the skeletal muscle molecular characteristics associated with end-stage osteoarthritis and refined an important phenotype, in some patients, termed muscle inflammation susceptibility (MuIS+) that may be an important consideration following surgery. Furthermore, we provide evidence of differential inflammatory and catabolic gene expression between the contralateral and surgical limbs along with differences between the skeletal muscle surrounding the diseased hip versus knee joints.

Perioperative amino acid infusion reestablishes muscle net balance during total hip arthroplasty.

Surgery and anesthesia induce a catabolic response that leads to skeletal muscle protein loss. Previous investigations have observed positive effects of perioperative nutrition. Furthermore, the benefits of exogenous amino acids on muscle protein kinetics are well established. However, no investigation has focused on muscle protein kinetics with and without perioperative amino acid infusion. Thus, we aimed to assess the effect of perioperative amino acid (AA) infusion on muscle protein balance in individuals undergoing elective total hip arthroplasty (THA). Elective THA patients were randomized to undergo a metabolic study prior to surgery (n = 5; control [CON]), intraoperative AA infusion (n = 9), or no AA (n = 13; standard of care [SC]). The CON group was studied prior to surgery to provide nonoperative/non-anesthesia muscle protein kinetic reference values. The bolus infusion method with 13 C6 -phenylalanine injected at time 0, and [15 N]-phenylalanine 30 min later was used to calculate muscle protein synthesis (MPS), protein breakdown (MPB), and net balance (MPS-MPB). Perioperative AA significantly improved muscle net balance as compared to SC (-0.005 ± 0.018%/h vs. -0.052 ± 0.011%/h) but not CON (0.003 ± 0.013%/h). The AA infusion significantly increased muscle net balance via a significant increase in MPS (AA = 0.062 ± 0.007%/h; SC = 0.037 ± 0.004%/h; CON = 0.072% ± 0.005%/h), and a nonsignificant attenuation of MPB (AA = 0.067 ± 0.012%/h; SC = 0.089 ± 0.014%/h; CON = 0.075 ± 0.011%/h). Our data support the use of perioperative AA infusion during elective THA as pragmatic strategy to offset the loss of surgically induced skeletal muscle protein.

Metabolic Evaluation of the Dietary Guidelines' Ounce Equivalents of Protein Food Sources in Young Adults: A Randomized Controlled Trial.

BACKGROUND: The Dietary Guidelines for Americans (DGAs) published an "ounce equivalents" recommendation to help consumers meet protein requirements with a variety of protein food sources. However, the metabolic equivalency of these varied protein food sources has not been established. OBJECTIVE: We have investigated the hypothesis that the anabolic responses to consumption of ounce equivalents of protein food sources would be directly related to the essential amino acid (EAA) content of the protein food source. METHODS: Following 3 d of dietary control, a total of 56 healthy young adults underwent an 8.5-h metabolic study using stable isotope tracer methodology. The changes from baseline following consumption of 1 of 7 different protein food sources were compared with the baseline value for that individual (n = 8 per group). RESULTS: Consumption of ounce equivalents of animal-based protein food sources (beef sirloin, pork loin, eggs) resulted in a greater gain in whole-body net protein balance above baseline than the ounce equivalents of plant-based protein food sources (tofu, kidney beans, peanut butter, mixed nuts; P < 0.01). The improvement in whole-body net protein balance was due to an increase in protein synthesis (P < 0.05) with all the animal protein sources, whereas the egg and pork groups also suppressed protein breakdown compared with the plant protein sources (P < 0.01). The magnitude of the whole-body net balance (anabolic) response was correlated with the EAA content of the protein food source (P < 0.001). CONCLUSION: The "ounce equivalents" of protein food sources as expressed in the DGAs are not metabolically equivalent in young healthy individuals. The magnitude of anabolic response to dietary proteins should be considered as the DGAs develop approaches to establish healthy eating patterns.

Equivalent servings of free-range reindeer promote greater net protein balance compared to commercial beef.

Wild game consumption has been associated with health benefits but the acute influence on human protein metabolism remains unknown. We compared feeding-induced responses of equivalent amounts of free-range reindeer (FR) and commercial beef (CB) on protein kinetics using stable isotope methodology. Seven participants (age: 40 ± 14 years; body mass index: 24 ± 3 kg/m2) completed two randomised studies, ingesting 2 oz of FR or CB. L-[ring 2H5]phenylalanine & L-[ring 2H2]tyrosine were delivered via primed, continuous intravenous infusion. Blood samples were collected during the basal period and following consumption of FR or CB. Feeding-induced changes in whole-body protein synthesis (PS), protein breakdown (PB), and net protein balance (NB) were determined via plasma sample isotope enrichment analysis by gas chromatography-mass spectrometry; plasma essential amino acid (EAA) concentrations were determined by liquid chromatography-electrospray ionisation-mass spectrometry. Plasma post-prandial EAA concentrations were higher with FR compared to CB (P < 0.05). The acute feeding-induced PS response was not different, but PB was reduced and contributed to a superior level of NB (P < 0.00001) in FR compared to CB. Our results demonstrate that FR may influence more favourable protein metabolism than CB. These data support potential health benefits of wild game onf whole-body protein.Abbreviations: BMI: body mass index; DIAAS: digestible indispensable amino acid score; CB: commercial beef; EAA: essential amino acids; FR: free-range reindeer; Ra: rate of appearance; UAF: University of Alaska Fairbanks; USDA: USA Department of Agriculture.

Daily Consumption of a Specially Formulated Essential Amino Acid-Based Dietary Supplement Improves Physical Performance in Older Adults With Low Physical Functioning.

We have investigated the hypothesis that nutritional supplementation of the diet in low-physical-functioning older individuals with a specially formulated composition based on essential amino acids (EAAs) would improve physical function as compared to supplementation with the same amount of whey protein. A third group of comparable volunteers were given nutrition education but no supplementation of the diet. After 6 weeks of whey protein supplementation (n = 32), there was no effect on the distance walked in 6 minutes, but the distance walked improved significantly from the pre-value after 12 weeks of whey supplementation. EAA consumption (n = 28) significantly improved walking distance at both 6 and 12 weeks. The distance walked at 12 weeks (419.0 ± 25.0 m) was 35.4 m greater than the pre-value of 384.0 ± 23.0 m (p < .001). The increase in distance walked by the EAA group was also significantly greater than that in the whey group at both 6 and 12 weeks (p < .01). In contrast, a decrease in distance walked was observed in the control group (n = 32) (not statistically significant, NS). EAA supplementation also improved grip strength and leg strength, and decreased body weight and fat mass. Plasma low-density lipoprotein concentration was significantly reduced in the EAA group, as well as the concentration of macrophage migration inhibitory factor. There were no adverse responses in any groups, and compliance was greater than 95% in all individuals consuming supplements. We conclude that dietary supplementation with an EAA-based composition may be a beneficial therapy in older individuals with low physical functional capacity. Clinical Trials Registration Number: This study was registered with ClinicalTrials.gov: NCT03424265-"Nutritional interventions in heart failure."

The impact of Hayward green kiwifruit on dietary protein digestion and protein metabolism.

PURPOSE: The purpose of the study was to determine if an actinidin protease aids gastric digestion and the protein anabolic response to dietary protein. METHODS: Hayward green kiwifruit (containing an actinidin protease) and Hort 16A gold kiwifruit (devoid of actinidin protease) were given in conjunction with a beef meal to healthy older subjects. Twelve healthy older males (N = 6) and females (N = 6) were studied with a randomized, double-blinded, crossover design to assess muscle and whole-body protein metabolism before and after ingestion of kiwifruit and 100 g of ground beef. Subjects consumed 2 of each variety of kiwifruit daily for 14 d prior to each metabolic study, and again during each study with beef intake. RESULTS: Hayward green kiwifruit consumption with beef resulted in a more rapid increase in peripheral plasma essential amino acid concentrations. There were significant time by kiwifruit intake interactions for plasma concentrations of EAAs, branched chain amino acids (BCAAs), and leucine (P < 0.01). However, there was no difference in the total amount of EAAs absorbed. As a result, there were no differences between kiwifruit in any of the measured parameters of protein kinetics. CONCLUSION: Consumption of Hayward green kiwifruit, with a beef meal facilitates protein digestion and absorption of the constituent amino acids as compared to Hort 16A gold kiwifruit. CLINICAL TRIAL: NCT04356573, April 21, 2020 "retrospectively registered".

Effects of Fortetropin on the Rate of Muscle Protein Synthesis in Older Men and Women: A Randomized, Double-Blinded, Placebo-Controlled Study.

BACKGROUND: Fortetropin is a proteo-lipid complex made from fertilized egg yolk and, in young men, has been shown to increase lean body mass. METHODS: The purpose of this study was to examine the effects of 21 days of Fortetropin supplementation on the fractional synthetic rate (FSR) of muscle protein in 10 healthy, older men and 10 women (66.4 ± 4.5 y). We used 2H2O labeling to measure FSR of multiple muscle protein ontologies. D3-creatine dilution was used to determine muscle mass at baseline. Subjects ingested 70% 2H2O for 21 day and saliva samples were collected to determine body 2H2O enrichment. A microbiopsy was obtained from the m. vastus lateralis on Day 21. Subjects were randomly assigned to Fortetropin (19.8 g/d) or placebo (cheese powder, 19.8 g/d). RESULTS: Restricting kinetic data to proteins with ≥2 peptides measured in at least 4 subjects per group resulted in 117 proteins meeting these criteria. The mean FSR for a majority of proteins in several muscle gene ontologies was higher in the Fortetropin group compared to placebo (32/38 myofibril proteins, 33/44 sarcoplasmic proteins, and 12/17 mitochondrial proteins) and this proportion was significantly different between groups using a binomial test and were independent of sex or baseline muscle mass. CONCLUSIONS: The overall magnitude of the difference in muscle protein FSR of Fortetropin from placebo was 18%, with multiple gene ontologies affected. While these results should be confirmed in larger cohorts, they suggest that Fortetropin supplementation is effective for promoting muscle protein synthesis in older people.

The Anabolic Response to Dietary Protein Is Not Limited by the Maximal Stimulation of Protein Synthesis in Healthy Older Adults: A Randomized Crossover Trial.

We have recently demonstrated in young adults that an anabolic response with mixed meal protein intake above ~35 g/meal, previously recognized as an "optimal" protein dose, was further stimulated. However, it is unknown if this applies to older adults. We therefore examined anabolic response to a mixed meal containing either 35 g (MOD, moderate amount of protein) or 70 g (HIGH, high amount of protein) in a randomized cross-over metabolic study in older adults (n = 8). Primed continuous infusions of L-[2H5] phenylalanine and L-[2H2]tyrosine were performed to determine whole-body protein kinetics and muscle protein fractional synthesis rate (MPS) in basal fasted and fed states. Whole-body protein kinetics (NB, net protein balance; PS, protein synthesis; PB, protein breakdown) and MPS was expressed as changes from the baseline post-absorptive state. Consistent with our previous findings in young adults, both feedings resulted in a positive NB, with HIGH being more positive than MOD. Furthermore, NB (expressed as g protein∙240 min) increased linearly with an increasing amount of protein intake, expressed relative to lean body mass. The positive NB was achieved due mainly to the suppression of PB in both MOD and to a greater extent HIGH, while PS was only increased in HIGH. Consistent with the whole-body data, MPS was significantly higher in HIGH than MOD. Plasma concentrations of essential amino acids and insulin were greater in HIGH vs. MOD. We conclude that in the context of mixed meals, whole-body anabolic response linearly increases with increasing protein intake primarily through the suppression of PB, and MPS was further stimulated with protein intake above the previously considered "optimal" protein dose in older adults.

Anabolic response to essential amino acid plus whey protein composition is greater than whey protein alone in young healthy adults.

BACKGROUND: We have determined the acute response of protein kinetics to one or two servings (6.3 g and 12.6 g) of a proprietary composition containing free-form essential amino acids (EAA) (3.2 g EAA per serving) and whey protein (2.4 g per serving), as well as the response to consumption of a popular whey-based protein supplement (Gatorade Recover) (17 g; 12.6 g protein). METHODS: Whole-body rates of protein synthesis, breakdown and net balance (taken to be the anabolic response) were determined using primed-constant infusions of 2H5-phenylalnine and 2H2-tyrosine. Muscle protein fractional synthetic rate (FSR) was also determined with the 2H5-phenylalanine tracer. RESULTS: Plasma EAA levels increased following consumption of all beverages, with the greatest response in the high-dose EAA/protein composition. Similarly, the increase in net balance between whole-body protein synthesis and breakdown was greatest following consumption of the high-dose EAA/protein composition, while the low-dose EAA/protein composition and Gatorade Recover induced similar increases in net balance. When the net balance response was normalized for the total amount of product given, the high- and low-dose EAA/protein beverages were approximately 6- and 3-fold more anabolic than the Gatorade Recover, respectively. The greater anabolic response to the EAA/protein composition was due to greater increases in whole-body protein synthesis with both doses, and a markedly greater suppression of whole-body protein breakdown in the high-dose group. Muscle protein FSR after beverage consumption reflected changes in whole-body protein synthesis, with the larger EAA/protein dose significantly increasing FSR. CONCLUSION: We conclude that a composition of a balanced EAA formulation combined with whey protein is highly anabolic as compared to a whey protein-based recovery product, and that the response is dose-dependent. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03502941. This trial was registered on April 19, 2018.

Consumption of a Specially-Formulated Mixture of Essential Amino Acids Promotes Gain in Whole-Body Protein to a Greater Extent than a Complete Meal Replacement in Older Women with Heart Failure.

Heart failure in older individuals is normally associated with a high body mass index and relatively low lean body mass due to, in part, a resistance to the normal anabolic effect of dietary protein. In this study we have investigated the hypothesis that consumption of a specially-formulated composition of essential amino acids (HiEAAs) can overcome anabolic resistance in individuals with heart failure and stimulate the net gain of body protein to a greater extent than a commercially popular protein-based meal replacement beverage with greater caloric but lower essential amino acid (EAA) content (LoEAA). A randomized cross-over design was used. Protein kinetics were determined using primed continuous infusions of L-(2H5)phenylalanine and L-(2H2)tyrosine in the basal state and for four hours following consumption of either beverage. Both beverages induced positive net protein balance (i.e., anabolic response). However, the anabolic response was more than two times greater with the HiEAA than the LoEAA (p < 0.001), largely through a greater suppression of protein breakdown (p < 0.001). Net protein accretion (g) was also greater in the HiEAA when data were normalized for either amino acid or caloric content (p < 0.001). We conclude that a properly formulated EAA mixture can elicit a greater anabolic response in individuals with heart failure than a protein-based meal replacement. Since heart failure is often associated with obesity, the minimal caloric value of the HiEAA formulation is advantageous.

Essential amino acid-enriched meal replacement promotes superior net protein balance in older, overweight adults.

BACKGROUND & AIMS: Older individuals are susceptible to the loss of muscle and accumulation of fat. To address this problem, we have compared protein kinetics following consumption of an essential amino acid (EAA)-enriched meal replacement (EMR) to consumption of a high-protein meal replacement beverage (Bariatric Advantage, BA) using stable isotope methodology. METHODS: Eight older (67 ± 2), obese (35 ± 2 kg/m2) female and male participants completed two studies using a randomized, crossover design in which they ingested each meal replacement. The isotopic tracers L-[2H5]phenylalanine & L-[2H2]tyrosine were delivered via primed, continuous intravenous infusion throughout a basal period and following consumption of EMR or BA. We determined changes in whole body protein synthesis (PS), protein breakdown (PB), and net protein balance (NB) from fasted states via analysis of plasma samples by LC-ESI-MS. RESULTS: PS was higher (P = 0.03) and PB was less (P = 0.005) with EMR in comparison to BA. As a result, NB was much greater (P = 0.00003) following the ingestion of EMR as compared to BA. CONCLUSIONS: In comparison with BA, which has a higher amount of intact protein that any other meal replacement, EMR promoted a greater increment in NB. These data support the potential efficacy of EMR as a meal replacement for the preservation of lean tissue mass during weight loss in older, overweight individuals.

Quality of meal protein determines anabolic response in older adults.

BACKGROUND & AIMS: It has been demonstrated that the relative content and profile of essential amino acids (EAA) play a determining role for stimulation of muscle protein synthesis (MPS) following intake of pure EAA or protein alone. METHODS: To test if this also holds in the context of mixed meals at both whole body and muscle levels, twelve older subjects (57-74 yrs) received primed continuous infusion of L-[ring-2H5]phenylalanine and L-[ring-2H2]tyrosine over a 9-h experimental period to determine whole body protein kinetics and MPS in the fasted state and following consumption of egg-based (EGG) or cereal-based (CEREAL) isocaloric and isonitrogenous breakfast. A standardized lunch, primarily consisting of beef protein was also consumed by each group. Whole body protein kinetics [protein synthesis (PS), breakdown (PB), and net balance (NB)] were expressed as changes from basal fasted period. RESULTS: We found that EGG breakfast resulted in a greater NB through a greater suppression of PB compared with the CEREAL breakfast. The greater NB during the post-breakfast period with the EGG was normalized following the standard lunch despite the sustained elevations in plasma EAA concentrations. However, the EGG breakfast stimulated both PS and PB compared with the CEREAL breakfast during the post-lunch period. MPS was not different between meals despite larges differences in the plasma EAA responses. CONCLUSIONS: We conclude that in the context of mixed meals, quality of protein affects NB through changes in protein breakdown and affects protein turnover following subsequent meal intake.

Protein intake distribution pattern does not affect anabolic response, lean body mass, muscle strength or function over 8 weeks in older adults: A randomized-controlled trial.

BACKGROUND & AIMS: In our recent acute metabolic study, we found no differences in the anabolic response to differing patterns of dietary protein intake. To confirm this in a chronic study, we investigated the effects of protein distribution pattern on functional outcomes and protein kinetics in older adults over 8 weeks. METHODS: To determine chronic effects of protein intake pattern at 1.1 g protein/kg/day in mixed meals on lean body mass (LBM), functional outcomes, whole body protein kinetics and muscle protein fractional synthesis rate (MPS) over 8-week respective dietary intervention, fourteen older subjects were randomly divided into either EVEN or UNVEN group. The UNEVEN group (n = 7) consumed the majority of dietary protein with dinner (UNEVEN, 15/20/65%; breakfast, lunch, dinner), while the EVEN group (n = 7) consumed dietary protein evenly throughout the day (EVEN: 33/33/33%). RESULTS: We found no significant differences in LBM, muscle strength, and other functional outcomes between EVEN and UNEVEN before and after 8-week intervention. Consistent with these functional outcomes, we did not find significant differences in the 20-h integrated whole body protein kinetics [net protein balance (NB), protein synthesis (PS), and breakdown (PB)] above basal states and MPS between EVEN and UNEVEN intake patterns. CONCLUSIONS: We conclude that over an 8-week intervention period, the protein intake distribution pattern in mixed meals does not play an important role in determining anabolic response, muscle strength, or functional outcomes. This trial is registered at https://ClinicalTrials.gov as NCT02787889.

Do anabolic nutritional supplements stimulate human growth hormone secretion in elderly women with heart failure?

Growth hormone treatment has gained attention over the past decade as a treatment for heart failure. Human growth hormone (HGH) must be administered by injections (usually daily), so there is considerable advantage to stimulation of endogenous secretion by amino acid-based nutritional supplementation. However, studies investigating the effect of amino acid (AA) supplementation show conflicting results. Therefore, in this study we aimed to investigate the effect of nutritional supplementation on HGH production in elderly women with heart failure. Eight elderly women with heart failure participated in this randomized cross-over study. Plasma HGH concentration was measured before and for 4 h following ingestion of a mixture of protein, carbohydrate, and fat or an AA beverage. HGH concentration was determined with ELISA kits and AA concentrations were analyzed by Liquid Chromatography-Mass Spectrometry (LCMS). Linear mixed models was performed to analyze the effect of time, treatment, and interaction. Plasma arginine and lysine concentrations were significantly higher after consumption of the AA drink compared to the mixture of protein, carbohydrate, and fat. Nonetheless, only ingestion of the protein, carbohydrate, and fat mixture (meal replacement) increased HGH concentration. HGH concentration was increased in elderly women with heart failure following consumption of a meal replacement containing protein, carbohydrate, and fat. Consumption of a mixture of amino acids failed to increase HGH concentration despite significantly greater elevations in plasma amino acid concentrations, including arginine and lysine. The stimulatory effect of the protein/carbohydrate/fat mixture was presumably mediated by factors other than increases in free amino acid concentrations.

Short term elevation in dietary protein intake does not worsen insulin resistance or lipids in older adults with metabolic syndrome: a randomized-controlled trial.

BACKGROUND: There is a great deal of controversy as to whether higher protein intake improves or worsens insulin sensitivity in humans. The purpose of the study was to determine the influence of a short-term elevation in dietary protein on hepatic and peripheral insulin sensitivity in twelve older subjects (51-70 yrs) with metabolic syndrome. METHODS: Individuals were randomly assigned to one of the dietary groups: recommended protein intake (RPI, 10% of daily calorie intake) or elevated protein intake (EPI, 20% of daily calorie intake) for 4 weeks. Prior to and immediately following the dietary intervention, subjects were studied with primed continuous infusion of [6,6-2H2]glucose and [1-3C]glucose dissolved in drink during the dual tracer oral glucose tolerance test (DT OGTT) to determine hepatic and peripheral insulin sensitivity. Plasma lipids were measured pre- and post-dietary intervention. RESULTS: In both intervention groups: 1) hepatic insulin sensitivity as assessed by the endogenous glucose rate of appearance (glucose Ra), 2) peripheral insulin sensitivity as assessed by the metabolic clearance rate of glucose normalized to plasma glucose concentration (MCR) and/or the rate of glucose utilization (Rd) or 3) glucose/insulin AUC were unaffected by the diets. Moreover, fasting lipid was not affected by RPI or EPI. CONCLUSION: Our findings suggest that a short-term elevation in EPI with correspondingly higher branched chain amino acid (BCAA) contents has no detrimental impact on hepatic and peripheral insulin sensitivity or plasma lipid parameters in older adults with metabolic syndrome. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02885935; This trial was registered retrospectively (Study start date, April 01, 2013, date of registration, August 26, 2016).

Skeletal Muscle Acute and Chronic Metabolic Response to Essential Amino Acid Supplementation in Hypertriglyceridemic Older Adults.

Background: Supplementation with essential amino acids (EAAs) + arginine is a promising nutritional approach to decrease plasma triglyceride (TG) concentrations, which are an independent risk factor for ischemic heart disease. Objective: The objective of this study was to examine the effects of 8 wk of EAA supplementation on skeletal muscle basal metabolite concentrations and changes in metabolic response to acute EAA intake, with an emphasis on mitochondrial metabolism, in adults with elevated TGs to better understand the mechanisms of lowering plasma TGs. Methods: Older adults with elevated plasma TG concentrations were given 22 g EAAs to ingest acutely before and after an 8-wk EAA supplementation period. Skeletal muscle biopsy samples were collected before and after acute EAA intake, both pre- and postsupplementation (4 biopsy samples), and targeted metabolomic analyses of organic acids and acylcarnitines were conducted on the specimens. Results: Acute EAA intake resulted in increased skeletal muscle acylcarnitine concentrations associated with oxidative catabolism of the supplement components, with the largest increases found in acylcarnitines of branched-chain amino acid oxidative catabolism, including isovaleryl-carnitine (2200%) and 2-methylbutyryl-carnitine (2400%). The chronic EAA supplementation resulted in a 19% decrease in plasma TGs along with accumulation of long-chain acylcarnitines myristoyl- (90%) and stearoyl- (120%) carnitine in skeletal muscle and increases in succinyl-carnitine (250%) and the late-stage tricarboxylic acid cycle intermediates fumarate (44%) and malate (110%). Conclusions: Supplementation with EAAs shows promise as an approach for moderate reduction in plasma TGs. Changes in skeletal muscle metabolites suggest incomplete fatty acid oxidation and increased anaplerosis, which suggests a potential bottleneck in fatty acid metabolism.

The anabolic response to a meal containing different amounts of protein is not limited by the maximal stimulation of protein synthesis in healthy young adults.

We have determined whole body protein kinetics, i.e., protein synthesis (PS), breakdown (PB), and net balance (NB) in human subjects in the fasted state and following ingestion of ~40 g [moderate protein (MP)], which has been reported to maximize the protein synthetic response or ~70 g [higher protein (HP)] protein, more representative of the amount of protein in the dinner of an average American diet. Twenty-three healthy young adults who had performed prior resistance exercise (X-MP or X-HP) or time-matched resting (R-MP or R-HP) were studied during a primed continuous infusion of l-[(2)H5]phenylalanine and l-[(2)H2]tyrosine. Subjects were randomly assigned into an exercise (X, n = 12) or resting (R, n = 11) group, and each group was studied at the two levels of dietary protein intake in random order. PS, PB, and NB were expressed as increases above the basal, fasting values (mg·kg lean body mass(-1)·min(-1)). Exercise did not significantly affect protein kinetics and blood chemistry. Feeding resulted in positive NB at both levels of protein intake: NB was greater in response to the meal containing HP vs. MP (P < 0.00001). The greater NB with HP was achieved primarily through a greater reduction in PB and to a lesser extent stimulation of protein synthesis (for all, P < 0.0001). HP resulted in greater plasma essential amino acid responses (P < 0.01) vs. MP, with no differences in insulin and glucose responses. In conclusion, whole body net protein balance improves with greater protein intake above that previously suggested to maximally stimulating muscle protein synthesis because of a simultaneous reduction in protein breakdown.

Acute ingestion of citrulline stimulates nitric oxide synthesis but does not increase blood flow in healthy young and older adults with heart failure.

To determine if age-associated vascular dysfunction in older adults with heart failure (HF) is due to insufficient synthesis of nitric oxide (NO), we performed two separate studies: 1) a kinetic study with a stable isotope tracer method to determine in vivo kinetics of NO metabolism, and 2) a vascular function study using a plethysmography method to determine reactive hyperemic forearm blood flow (RH-FBF) in older and young adults in the fasted state and in response to citrulline ingestion. In the fasted state, NO synthesis (per kg body wt) was ∼ 50% lower in older vs. young adults and was related to a decreased rate of appearance of the NO precursor arginine. Citrulline ingestion (3 g) stimulated de novo arginine synthesis in both older [6.88 ± 0.83 to 35.40 ± 4.90 µmol · kg body wt(-1) · h(-1)] and to a greater extent in young adults (12.02 ± 1.01 to 66.26 ± 4.79 µmol · kg body wt(-1) · h(-1)). NO synthesis rate increased correspondingly in older (0.17 ± 0.01 to 2.12 ± 0.36 µmol · kg body wt(-1) · h(-1)) and to a greater extent in young adults (0.36 ± 0.04 to 3.57 ± 0.47 µmol · kg body wt(-1) · h(-1)). Consistent with the kinetic data, RH-FBF in the fasted state was ∼ 40% reduced in older vs. young adults. However, citrulline ingestion (10 g) failed to increase RH-FBF in either older or young adults. In conclusion, citrulline ingestion improved impaired NO synthesis in older HF adults but not RH-FBF, suggesting that factors other than NO synthesis play a role in the impaired RH-FBF in older HF adults, and/or it may require a longer duration of supplementation to be effective in improving RH-FBF.