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.

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.

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."

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 13C6‐phenylalanine injected at time 0, and [15N]‐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.

Many individuals with end-stage osteoarthritis undergo elective total hip/knee arthroplasty (THA/TKA) to relieve pain and improve mobility and quality of life. However, ~35% suffer long-term mobility impairment following surgery. This may be in part due to muscle inflammation susceptibility (MuIS+), an overt pro-inflammatory pathology localized to skeletal muscle surrounding the diseased joint, present in some TKA/THA patients. PURPOSE: We are interrogating the hypotheses that a) MuIS+/- status will result in a differential perioperative expression profile that may partially explain low functional outcomes, and b) resistance training rehabilitation will more effectively overcome MuIS+ status than usual care for successful recovery post-surgery. METHODS: Muscle, serum, and functional data were leveraged from our ongoing two-site, randomized, controlled trial (N = 75 randomized). Participants were dichotomized based on surgical (SX) muscle gene expression of TNFRSF1A (TNFαR). MuIS+/- samples were probed for transcript expression and indices of skeletal muscle integrity. Preliminary perioperative comparisons were made using two-tailed T-tests; alpha P < 0.05. Preliminary miRNA-seq data were analyzed using Partek Flow; differentially expressed (DE) miRNAs were defined as Fold change (FC) ≥ 1.5 and P ≤ 0.05. RESULTS: 70 participants (27 M/43F; mean age 62 ± 9 yrs; mean BMI 31.3 ± 4.9 kg/m2) undergoing THA/TKA were clustered as MuIS+ (n = 24) or MuIS- (n = 46). MuIS+ muscle exhibited significantly higher pro-inflammatory (IL-6R) and catabolic (TRIM63) gene expression (P = 0.0001 and 0.03, respectively) compared to MuIS-. Further, significantly lower skeletal muscle integrity exists on the SX vs non-SX limb, illustrated by 20% higher fibrosis (P = 0.0003), and 3.5% lower thigh muscle mass (P = 0.005). Preliminary serum miRNA-seq (for N = 9) indicate 193 DE miRNAs (P ≤ 0.05, FC ≥ 1.5) following the intervention. CONCLUSIONS: Preliminary results suggest MuIS+ patients exhibit more inflammation and catabolic signaling in skeletal muscle of the SX limb, accompanied by decreased skeletal muscle integrity. This highlights the profound impact of MuIS+ status and emphasizes the potential value of perioperative MuIS assessment to inform optimal post-surgical care. Funding: T32HD071866, R01HD084124

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.

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.

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

Wild game consumption has been associated with health benefits, but the immediate influence on protein metabolism remains unknown. In order to address the direct effects on protein kinetics, we compared the feeding‐induced response to free‐range reindeer versus commercial beef using stable isotope methodology in humans. Eight male and female participants (age: 38±12 years; body mass index: 24±3 kg/m2) completed two studies using a randomized, crossover design in which they ingested reindeer or beef. L‐[2H5]phenylalanine & L‐[2H2]tyrosine were delivered via primed, continuous intravenous infusion. Blood samples were collected during a 3‐hour basal period and for 4 hrs after consumption of reindeer and beef. A priming dose of L‐[2H4]tyrosine was also delivered. Feeding‐induced changes in whole body protein synthesis (PS), protein breakdown (PB), and net protein balance (NB) were determined via analysis of plasma samples by liquid chromatography‐electrospray ionization‐tandem mass spectrometry (LC‐ESI‐MS). Post‐prandial essential amino acid (EAA) concentrations were higher with reindeer ingestion (P=0.02). PS was not different but PB was reduced with reindeer compared to beef ingestion (P=0.02), contributing to superior level of NB (P=0.001). Reindeer promoted higher plasma EAA levels and NB compared to beef. The amino acid and fatty acid profiles of reindeer may have an immediate and greater benefit on protein metabolism as compared to beef. These data support the potential health benefits of wild game in the preservation of whole‐body protein.Support or Funding InformationResearch reported in this publication was supported by an Institutional Development Award (IDeA) under grant number P20GM103395 and the Biomedical Learning and Student Training Program (UL1GM118991, TL4GM118992, or RL5GM118990) from the National Institute of General Medical Sciences from the National Institutes of Health. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

The original version of this article unfortunately contained a mistake. Unfortunately, the Figs. 3 and 4 overlapped. Figures 3 and 4 should be.

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.

Wild game consumption has been associated with health benefits, but the influence on protein metabolism remains unknown. We compared the feeding-induced response to 2 oz of free-range reindeer (FR) versus commercial beef (CB) using stable isotope methodology. Seven male and female participants (age: 38±12 years; body mass index: 24±3 kg/m2) completed two studies using a randomized, crossover design in which they ingested 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 analysis of plasma samples for phenyalanine and tyrosine enrichment by gas chromatography mass spectrometry; plasma essential amino acid concentrations were determined by liquid chromatography-electrospray ionization-mass spectrometry. Plasma post-prandial essential amino acid (EAA) concentrations were higher with the ingestion of FR compared to CB (P=0.02). The acute feeding-induced response in PS was not different in either trial, but PB was reduced with the ingestion of FR compared to CB (P<0.0001). The difference in PB contributed to a superior level of NB (P<0.0001). When protein kinetics were normalized relative to the amino acids ingested, PB/EAAs and total amino acids ingested were reduced (P<0.01 and 0.001, respectively) in FR compared to CB; contributing to greater NB/total amino acid ingested (P<0.0001) between FR and CB. We conclude that the nutrient profiles of FR may have a more favorable benefit on protein metabolism compared to CB. These data support the potential health benefits of wild game in the preservation of whole-body protein.

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 2 H 5 -phenylalnine and 2 H 2 -tyrosine. Muscle protein fractional synthetic rate (FSR) was also determined with the 2 H 5 -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.

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.

The Beef Checkoff, National Pork Board, and Egg Nutrition Center.

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/m(2)) female and male participants completed two studies using a randomized, crossover design in which they ingested each meal replacement. The isotopic tracers L-[H-2(5)]phenylalanine & L-[H-2(2)]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. (C) 2018 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

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.

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 4h 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.

Our recent acute study of dietary protein distribution indicated no difference in the anabolic response to differing patterns of dietary protein intake. However, it has been argued that acute metabolic studies may not predict functional changes over a longer period time. To examine the chronic effects of protein distribution pattern, older subjects (51– 69 yrs) consumed 1.1 g/kg/d in 2 distinct patterns. One group (n=7) consumed protein in a traditional, or uneven, manner (UNEVEN, 15/20/65%; breakfast, lunch, dinner), while the other group (n=7) consumed meals with an even distribution of protein (EVEN: 33/33/33%). Lean body mass (LBM), functional outcomes, and protein kinetics at whole‐body and muscle levels using stable isotope tracers were determined before and after 8 weeks of each respective protein intake pattern. There were no significant differences between the intake patterns in LBM, muscle strength, and other functional outcomes. Consistent with these findings, the 20‐h integrated whole body protein kinetics [net protein balance (NB), protein synthesis (PS), and breakdown (PB)] above basal states and muscle protein synthesis were not different between the intake patterns. We conclude that the distribution pattern of protein intake in mixed meals over an 8‐week period does not affect protein metabolism, lean body mass, muscle strength, or functional outcomes.Support or Funding InformationThe project was financially supported by a grant from National Dairy Council, Egg Nutrition Center, and National Cattlemen's Beef Association. This project was financially supported by Pepper Center Grant PG30‐AG‐028718 and Award Number UL1‐TR‐000039 and KL2‐TR‐000063 from the National Center for Advancing Translations Sciences (NCATS).

BackgroundSeveral studies have reported that skeletal muscle mitochondrial respiratory capacity is lower in older adults than in young adults, suggesting that mitochondrial dysfunction may occur as a consequence of the aging process. In addition, blood‐cell bioenergetics has been suggested as an indicator of systemic mitochondrial health. We have shown that supplementation with essential amino acids (EAAs) plus arginine decreases plasma and liver triacylglycerols in older adults (1). However, no studies to date have reported the impact of such supplementation on mitochondrial respiration within circulating cells. The purpose of this study was to assess the effect of eight weeks of EAA + arginine supplementation on circulating cell bioenergetics in older adults with hypertriacylglycerolaemia.MethodsSix (two men) older adults (age: 68.8±2.0 years) underwent testing before and after 8 weeks of supplementation with EAA + arginine (11 g of AAs ingested twice daily). Specifically, a skeletal muscle biopsy, peripheral blood mononuclear cells (PBMCs; n=4), and platelets (n=4) were collected pre‐ and post‐supplementation, in order to measure mitochondrial function by high‐resolution respirometry using the Oroboros and the Seahorse, respectively. Mitochondrial respiration was determined in permeabilized myofibers from the m. vastus lateralis after the addition of substrates, oligomycin, and FCCP. Thereafter, mitochondrial coupling control was calculated.ResultsMaximal coupled respiration (respiration linked to ATP production) in muscle was not significantly changed by 8 weeks of AA supplementation. On the other hand, ATP‐linked respiration (31.55±4.43 vs. 18.93±2.89 pmol/min, p=0.05), maximal respiration (127±25 vs. 60±9 pmol/min, p=0.05) and the reserve capacity (94.2±21.7 vs. 40.1±6.2, pmol/min, p=0.05) were decreased in isolated PBMCs following 8 weeks of supplementation. While no significant differences in mitochondrial respiration were observed for platelets, the glycolytic capacity of platelets was decreased (70.3±0.8 vs. 55.3±2.6 mpH/min, p=0.03) after AA supplementation. No significant alterations in glycolysis were observed in PBMCs.ConclusionsThese results indicate that EAA supplementation may lower respiration rates in PBMCs. The mechanisms underlying this may be that lowering lipids as previously observed decreases stress on cells and therefore lowers ATP demand, or that the substrate pool is altered and thus metabolism shifted. Future studies are needed in order to elucidate mechanisms and understand the implications of these novel findings.Support or Funding InformationFunding: NIH R01‐AG033761, NIH P30‐AG028718, Arkansas Biosciences Institute the major research component of the Arkansas Tobacco Settlement Proceeds Act of 2000, FEDER funds through the Operational Programme Competitiveness Factors ‐ COMPETE and UID/NEU/04539/2013