Greater Protein Intake Emphasizing Lean Beef Does Not Affect Resistance Training-Induced Adaptations in Skeletal Muscle and Tendon of Older Women: A Randomized Controlled Feeding Trial.

BACKGROUND: Although experimental research supports that resistance training (RT), especially with greater dietary protein intake, improves muscle mass and strength in older adults, comparable research on tendons is needed. OBJECTIVES: We assessed the effects of a protein-rich diet emphasizing lean beef, compared with 2 control diets, on RT-induced changes in skeletal muscle and tendon size and strength in older women. METHODS: We randomly assigned women [age: 66 ± 1 y, body mass index (BMI): 28 ± 1] to groups that consumed 1) 0.8 g total protein/kg body weight/day from mixed food sources (normal protein control, n = 16); 2) 1.4 g/kg/d protein from mixed food sources (high protein control, n = 17); or 3) 1.4 g/kg/d protein emphasizing unprocessed lean beef (high protein experimental group, n = 16). Participants were provided with all foods and performed RT 3 times/wk, 70% of 1-repetition maximum for 12 wk. We measured quadriceps muscle volume via magnetic resonance imaging (MRI). We estimated patellar tendon biomechanical properties and cross-sectional area (CSA) using ultrasound and MRI. RESULTS: Dietary intake did not influence RT-induced increases in quadriceps strength (P < 0.0001) or muscle volume (P < 0.05). We noted a trend for an RT effect on mean tendon CSA (P = 0.07), with no differences among diets (P > 0.05). Proximal tendon CSA increased with RT (P < 0.05) with no difference between dietary groups (P > 0.05). Among all participants, midtendon CSA increased with RT (P ≤ 0.05). We found a decrease in distal CSA in the 0.8 g group (P < 0.05) but no change in the 1.4 g group (P > 0.05). Patellar tendon MRI signal or biomechanical properties were unchanged. CONCLUSIONS: Our findings indicated that greater daily protein intake, emphasizing beef, did not influence RT-induced changes in quadriceps muscle strength or muscle volume of older women. Although we noted trends in tendon CSA, we did not find a statistically significant impact of greater daily protein intake from beef on tendon outcomes. This trial was registered at clinicaltrials.gov as NCT04347447.

Postabsorptive muscle protein synthesis is higher in outpatients as compared to inpatients.

Several factors affect muscle protein synthesis (MPS) in the postabsorptive state. Extreme physical inactivity (e.g., bedrest) may reduce basal MPS, whereas walking may augment basal MPS. We hypothesized that outpatients would have a higher postabsorptive MPS than inpatients. To test this hypothesis, we conducted a retrospective analysis. We compared 152 outpatient participants who arrived at the research site the morning of the MPS assessment with 350 Inpatient participants who had an overnight stay in the hospital unit before the MPS assessment the following morning. We used stable isotopic methods and collected vastus lateralis biopsies ∼2 to 3 h apart to assess mixed MPS. MPS was ∼12% higher (P < 0.05) for outpatients than inpatients. Within a subset of participants, we discovered that after instruction to limit activity, outpatients (n = 13) took 800 to 900 steps in the morning to arrive at the unit, seven times more steps than inpatients (n = 12). We concluded that an overnight stay in the hospital as an inpatient is characterized by reduced morning activity and causes a slight but significant reduction in MPS compared with participants studied as outpatients. Researchers should be aware of physical activity status when designing and interpreting MPS results.NEW & NOTEWORTHY The postabsorptive muscle protein synthesis rate is lower in the morning after an overnight inpatient hospital stay compared with an outpatient visit. Although only a minimal amount of steps was conducted by outpatients (∼900), this was enough to increase postabsorptive muscle protein synthesis rate.

Impact of essential amino acid intake, resistance exercise, and aging on the concentration of Achilles peritendinous amino acids and procollagen Iα1 in humans.

Recent studies have shown that consuming amino acid-rich compounds improves tendon collagen content and biomechanical properties. Yet, it is unclear if the consumption of amino acids alters local (peritendinous) amino acid concentrations. If aging or exercise influence local amino acid concentrations in conjunction with an amino acid bolus is also not known. We conducted two studies. In Study 1, young women (n = 7, 25 ± 2 years) completed two identical resistance training sessions with either essential amino acid (EAA) or placebo consumption. In Study 2, an EAA bolus identical to Study 1 was given to younger (n = 7; 27 ± 1 year) and older adults (n = 6; 68 ± 2 years). Microdialysis was used to determine Achilles peritendinous amino acid and pro-collagen Iα1 (a marker of collagen synthesis) concentrations. In Study 1, amino acid consumption increased peritendinous concentrations of all EAA except histidine (p < 0.05). In Study 2, the peritendinous concentration of EAAs except for methionine, histidine, and lysine (p > 0.05) increased with time (p < 0.05). Further, the concentrations of most measured amino acids were greater in older adults (p < 0.05). Pro-collagen Iα1 concentration (p > 0.05) was unaffected by exercise, EAA, or aging (p > 0.05). Our findings demonstrate the following: (1) when not combined with exercise, an oral EAA bolus leads to only modest increases in Achilles peritendinous amino acid concentrations; (2) when combined with resistance exercise, EAA consumption resulted in greater peritendinous amino acid concentrations compared to no exercise; (3) the basal concentrations of most amino acids were greater in older adults, and (4) neither the EAA bolus nor exercise altered peritendinous pro-collagen concentrations.

Impact of elevated serum advanced glycation end products and exercise on intact and injured murine tendons.

OVERVIEW: Delayed tendon healing is a significant clinical challenge for those with diabetes. We explored the role of advanced glycation end-products (AGEs), a protein modification present at elevated levels in serum of individuals with diabetes, on injured and intact tendons using a mouse model. Cell proliferation following tissue injury is a vital component of healing. Based on our previous work demonstrating that AGEs limit cell proliferation, we proposed that AGEs are responsible for the delayed healing process commonly observed in diabetic patients. Further, in pursuit of interventional strategies, we suggested that moderate treadmill exercise may support a healing environment in the presence of AGEs as exercise has been shown to stimulate cell proliferation in tendon tissue. MATERIALS AND METHODS: Mice began receiving daily intraperitoneal injections of bovine serum albumin (BSA)-Control or AGE-BSA injections (200µg/ml) at 16-weeks of age. A tendon injury was created in the central third of both patellar tendons. Animals assigned to an exercise group began a moderate treadmill protocol one week following injury. The intact Achilles tendon and soleus muscle were also evaluated to assess the effect of BSA and AGE-BSA on un-injured muscle and tendon. RESULTS: We demonstrate that our injection dosing and schedule lead to an increase in serum AGEs. Our findings imply that AGEs indeed modulate gene expression following a patellar tendon injury and have modest effects on gene expression in intact muscle and tendon. CONCLUSIONS: While additional biomechanical analysis is warranted, these data suggest that elevated serum AGEs in persons with diabetes may impact tendon health.

The effects of acute aerobic and resistance exercise on mTOR signaling and autophagy markers in untrained human skeletal muscle.

PURPOSE: Aerobic (AE) and resistance (RE) exercise elicit unique adaptations in skeletal muscle. The purpose here was to compare the post-exercise response of mTOR signaling and select autophagy markers in skeletal muscle to acute AE and RE. METHODS: In a randomized, cross-over design, six untrained men (27 ± 3 years) completed acute AE (40 min cycling, 70% HRmax) and RE (8 sets, 10 repetitions, 65% 1RM). Muscle biopsies were taken at baseline, and at 1 h and 4 h following each exercise. Western blot analyses were performed to examine total and phosphorylated protein levels. Upstream regulator analyses of skeletal muscle transcriptomics were performed to discern the predicted activation states of mTOR and FOXO3. RESULTS: Compared to AE, acute RE resulted in greater phosphorylation (P < 0.05) of mTORSer2448 at 4 h, S6K1Thr389 at 1 h, and 4E- BP1Thr37/46 during the post-exercise period. However, both AE and RE increased mTORSer2448 and S6K1Thr389 phosphorylation at 4 h (P < 0.05). Upstream regulator analyses revealed the activation state of mTOR was increased for both AE (z score, 2.617) and RE (z score, 2.789). No changes in LC3BI protein were observed following AE or RE (P > 0.05), however, LC3BII protein was decreased after both AE and RE at 1 h and 4 h (P < 0.05). p62 protein content was also decreased at 4 h following AE and RE (P < 0.05). CONCLUSION: Both acute AE and RE stimulate mTOR signaling and similarly impact select markers of autophagy. These findings indicate the early adaptive response of untrained human skeletal muscle to divergent exercise modes is not likely mediated through large differences in mTOR signaling or autophagy.

Nerve growth factor receptor TrkA signaling in streptozotocin-induced type 1 diabetes rat brain.

Previous work from our lab demonstrated a new role of TrkA in the insulin signaling pathway. The kinase activity of TrkA is essential for its interaction with the insulin receptor (IR) and insulin receptor substrate-1 (IRS-1) and activation of Akt and Erk5 in PC12 cells. Here we show in brain from streptozotocin (STZ)-induced type 1 diabetic rats that the expression of the inactive proNGF is elevated, whereas the expression of mature NGF is reduced. In addition, tyrosine phosphorylation of TrkA is decreased in STZ-induced diabetes compared to control. Results of the co-immunoprecipitation experiments indicate that the interaction of TrkA with the IR and IRS-1 is also reduced in the brain of diabetic rats. Moreover, tyrosine phosphorylation of the IR and IRS-1, and Akt activation is decreased in STZ diabetes compared to control. Our results suggest that the NGF-TrkA receptor is involved in insulin signaling and is impaired in the brain of STZ-induced diabetic rats.

Streptozotocin-induced diabetes alters transcription of multiple genes necessary for extracellular matrix remodeling in rat patellar tendon.

OVERVIEW: Tendon collagen fibril degradation is commonly seen in tendons of diabetics, but the mechanisms responsible for these changes remain to be elucidated. We have demonstrated that streptozotocin (STZ)-induced diabetes increases tendon cell proliferation and collagen content. In the present study, we evaluated that impact of STZ-induced diabetes on mRNA transcripts involved with collagen fibril organization, extracellular matrix (ECM) remodeling, apoptosis, and proliferation. MATERIALS AND METHODS: Rats were divided into four groups: nondiabetic (control, n = 9), 1 week (acute, n = 8) or 10 weeks of diabetes (chronic, n = 7), and 10 weeks of diabetes with insulin (insulin, n = 8). RNA was isolated from the patellar tendon for determination of mRNA transcripts using droplet digital PCR (ddPCR). RESULTS: Transcripts for Col1a1, Col3a1, Mmp2, Timp1, Scx, Tnmd, Casp3, Casp8, and Ager were lower in acute relative to control and insulin rats (p ≤ 0.05). With the exception of Scx, transcripts for Col1a1, Col3a1, Mmp2, Timp1, Tnmd, Casp3, Casp8, and Ager were also lower in chronic when compared to control (p < 0.05). Transcripts for Col1a1, Col3a1, Mmp2, Timp1, Tnmd, Casp3, Casp8, and Ager were not different between control and insulin (p > 0.05). Transcripts for Dcn, Mmp1a, Mmp9, Pcna, Tgfbr3, Ptgs2, Ptger2, Ptges, and iNos were not altered by diabetes or insulin (p > 0.05). CONCLUSION: Our findings indicated that STZ-induced diabetes results in rapid and large changes in the expression of several genes that are key to ECM remodeling, maintenance, and maturation.

Impact of acetaminophen consumption and resistance exercise on extracellular matrix gene expression in human skeletal muscle.

Acetaminophen (APAP) given during chronic exercise reduces skeletal muscle collagen and cross-linking in rats. We propose that the effect of APAP on muscle extracellular matrix (ECM) may, in part, be mediated by dysregulation of the balance between matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs). The purpose of this study was to evaluate the impact of APAP consumption during acute resistance exercise (RE) on several regulators of the ECM in human skeletal muscle. In a double-blinded, placebo-controlled, randomized crossover design, recreationally active men (n = 8, 25 ± 2 yr) performed two trials of knee extension. Placebo (PLA) or APAP (1,000 mg/6 h) was given for 24 h before and immediately following RE. Vastus lateralis biopsies were taken at baseline and 1 and 3 h post-RE. Quantitative RT-PCR was used to determine differences in mRNA expression. MMP-2, type I collagen, and type III collagen mRNA expression was not altered by exercise or APAP (P > 0.05). When compared with PLA, TIMP-1 expression was lower at 1 h post-RE during APAP conditions but greater than PLA at 3 h post-RE (P < 0.05). MMP-9 expression and protein levels were elevated at 3 h post-RE independent of treatment (P < 0.05). Lysyl oxidase expression was greater at 3 h post-RE during APAP consumption (P < 0.05) compared with PLA. MMP-2 and TIMP-1 protein was not altered by RE or APAP (P > 0.05). Phosphorylation of ERK1/2 and p38-MAPK increased (P < 0.05) with RE but was not influenced by APAP. Our findings do not support our hypothesis and suggest that short-term APAP consumption before RE has a small impact on the measured ECM molecules in human skeletal muscle following acute RE.

Impact of TGF-ß inhibition during acute exercise on Achilles tendon extracellular matrix.

The purpose of this study was to evaluate the role of TGF-ß1 in regulating tendon extracellular matrix after acute exercise. Wistar rats exercised (n = 15) on a treadmill for four consecutive days (60 min/day) or maintained normal cage activity. After each exercise bout, the peritendinous space of each Achilles tendon was injected with a TGF-ß1 receptor inhibitor or sham. Independent of group, tendons injected with inhibitor exhibited ~50% lower Smad 3 (Ser423/425) (P < 0.05) and 2.5-fold greater ERK1/2 phosphorylation (P < 0.05) when compared with sham (P < 0.05). Injection of the inhibitor did not alter collagen content in either group (P > 0.05). In exercised rats, hydroxylyslpyridinoline content and collagen III expression were lower (P < 0.05) in tendons injected with inhibitor when compared with sham. In nonexercised rats, collagen I and lysyl oxidase (LOX) expression was lower (P < 0.05) in tendons injected with inhibitor when compared with sham. Decorin expression was not altered by inhibitor in either group (P > 0.05). On the basis of evaluation of hematoxylin and eosin (H&E) stained cross sections, cell numbers were not altered by inhibitor treatment in either group (P > 0.05). Evaluation of H&E-stained sections revealed no effect of inhibitor on collagen fibril morphology. In contrast, scores for regional variation in cellularity decreased in exercised rats (P < 0.05). No differences in fiber arrangement, structure, and nuclei form were noted in either group (P > 0.05). Our findings suggest that TGF-ß1 signaling is necessary for the regulation of tendon cross-link formation, as well as collagen and LOX gene transcription in an exercise-dependent manner.

The influence of chronic IL-6 exposure, in vivo, on rat Achilles tendon extracellular matrix.

When compared to placebo, acetaminophen (APAP) reduces tendon stiffness and collagen cross-linking. APAP also enhances the exercise-induced increase in peritendinous levels of IL-6. Elevated levels of IL-6 are associated with tendinopathy, thus we hypothesized that chronic, elevated peritendinous IL-6 would alter tendon extracellular matrix (ECM). IL-6 (∼3000pgml-1) was injected (3dwk-1 for 8-wks) into the Achilles peritendinous region of male Wistar rats (n=16) with the opposite leg serving as a sham. Fractional synthesis rates (FSR) were determined using deuterium oxide. Collagen (hydroxyproline) and hydroxylysl pyridinoline (HP) cross-linking were analyzed by HPLC. ECM and IL-6 related genes were evaluated using qRT-PCR. Relative to sham, collagen (Col) 1a1 but not Col3a1 expression was suppressed (47%) in tendons exposed to IL-6 (p<0.05). Lysyl oxidase (LOX) and MMP-1 expression were also reduced (37%) in IL-6 treated tendons (p<0.05). Relative to sham the expression of MMP-2, -3, -9, and TIMP-1 were not altered by IL-6 treatment (p>0.05). Interleukin-6 receptor subunit beta precursor (IL6st) was lower (16%) in IL-6 treated tendons when compared to sham (p<0.05). Suppressor of cytokine signaling 3 (Socs3), signal transducer and activator of transcription 3 (STAT3), and protein inhibitor of activated STAT 1 (Pias1) were not altered by IL-6 exposure (p>0.05). Neither collagen nor cross-linking content were altered by IL-6 (p>0.05). Additionally, IL-6 treatment did not alter tendon FSR. Chronic treatment with physiologically relevant levels of IL-6 suppresses expression of Col1a1 and LOX while also altering expression of select MMPs but does not alter Achilles tendon collagen synthesis.

Insulin does not stimulate muscle protein synthesis during increased plasma branched-chain amino acids alone but still decreases whole body proteolysis in humans.

Insulin stimulates muscle protein synthesis when the levels of total amino acids, or at least the essential amino acids, are at or above their postabsorptive concentrations. Among the essential amino acids, branched-chain amino acids (BCAA) have the primary role in stimulating muscle protein synthesis and are commonly sought alone to stimulate muscle protein synthesis in humans. Fourteen healthy young subjects were studied before and after insulin infusion to examine whether insulin stimulates muscle protein synthesis in relation to the availability of BCAA alone. One half of the subjects were studied in the presence of postabsorptive BCAA concentrations (control) and the other half in the presence of increased plasma BCAA (BCAA). Compared with that prior to the initiation of the insulin infusion, fractional synthesis rate of muscle protein (%/h) did not change (P > 0.05) during insulin in either the control (0.04 ± 0.01 vs 0.05 ± 0.01) or the BCAA (0.05 ± 0.02 vs. 0.05 ± 0.01) experiments. Insulin decreased (P < 0.01) whole body phenylalanine rate of appearance (µmol·kg-1·min-1), indicating suppression of muscle proteolysis, in both the control (1.02 ± 0.04 vs 0.76 ± 0.04) and the BCAA (0.89 ± 0.07 vs 0.61 ± 0.03) experiments, but the change was not different between the two experiments (P > 0.05). In conclusion, insulin does not stimulate muscle protein synthesis in the presence of increased circulating levels of plasma BCAA alone. Insulin's suppressive effect on proteolysis is observed independently of the levels of circulating plasma BCAA.

Analgesic Drugs Alter Connective Tissue Remodeling and Mechanical Properties.

Exercising individuals commonly consume analgesics, but these medications alter tendon and skeletal muscle connective tissue properties, possibly limiting a person from realizing the full benefits of exercise training. I detail the novel hypothesis that analgesic medications alter connective tissue structure and mechanical properties by modifying fibroblast production of growth factors and matrix enzymes, which are responsible for extracellular matrix remodeling.

The effect of chronic treadmill exercise and acetaminophen on collagen and cross-linking in rat skeletal muscle and heart.

The purpose of this study was to determine whether exercise and/or acetaminophen (APAP) alter collagen and cross-linking in the rat gastrocnemius muscle, soleus muscle, and heart. Male Wistar rats (n = 50; 8 wk old) were divided into placebo (PLA) or APAP groups and sedentary (SED) or exercised (RUN) groups. APAP (200 mg/kg) was administered daily by oral gavage. Exercised groups ran on a treadmill 5 days/wk for 8 wk with progression to 60 min/day, 20 m/min, and 8° incline. Tissues were assayed for collagen (hydroxyproline) and hydroxylyslpyridinoline (HP) and lysylpyridinoline (LP) cross-links by HPLC. Collagen content (µg/mg dry weight) was greater in both the gastrocnemius (SED-PLA: 114 ± 16 vs. RUN-PLA: 244 ± 32; P < 0.001) and soleus (SED-PLA: 51 ± 7 vs. RUN-PLA: 99 ± 27; P = 0.005) of exercised animals. In contrast, collagen content was not significantly greater in exercised animals treated with APAP (SED-APAP: 113 ± 16 vs. RUN-APAP: 145 ± 21) and soleus (SED-APAP: 55 ± 8 vs. RUN-APAP: 57 ± 10). HP cross-linking (mmol/mol collagen) in the gastrocnemius (SED-PLA: 126 ± 28, RUN-PLA: 50 ± 7, SED-APAP: 41 ± 7, and RUN-APAP: 30 ± 4) and soleus muscles (SED-PLA: 547 ± 107, RUN-PLA: 318 ± 92, SED-APAP: 247 ± 64, and RUN-APAP: 120 ± 17) was lower in exercised rats compared with sedentary rats (P < 0.05). Cross-linking was further reduced in animals treated with APAP (P < 0.05). Neither heart collagen nor cross-linking was influenced by exercise or APAP (P > 0.05). Our findings suggest that exercise and APAP have tissue-specific effects on muscle collagen. Given the widespread use of APAP as an analgesic and antipyretic, further work in humans is warranted.

Influence of acute and chronic streptozotocin-induced diabetes on the rat tendon extracellular matrix and mechanical properties.

Diabetes is a major risk factor for tendinopathy, and tendon abnormalities are common in diabetic patients. The purpose of the present study was to evaluate the effect of streptozotocin (60 mg/kg)-induced diabetes and insulin therapy on tendon mechanical and cellular properties. Sprague-Dawley rats (n = 40) were divided into the following four groups: nondiabetic (control), 1 wk of diabetes (acute), 10 wk of diabetes (chronic), and 10 wk of diabetes with insulin treatment (insulin). After 10 wk, Achilles tendon and tail fascicle mechanical properties were similar between groups (P > 0.05). Cell density in the Achilles tendon was greater in the chronic group compared with the control and acute groups (control group: 7.8 ± 0.5 cells/100 µm(2), acute group: 8.3 ± 0.4 cells/100 µm(2), chronic group: 10.9 ± 0.9 cells/100 µm(2), and insulin group: 9.2 ± 0.8 cells/100 µm(2), P < 0.05). The density of proliferating cells in the Achilles tendon was greater in the chronic group compared with all other groups (control group: 0.025 ± 0.009 cells/100 µm(2), acute group: 0.019 ± 0.005 cells/100 µm(2), chronic group: 0.067 ± 0.015, and insulin group: 0.004 ± 0.004 cells/100 µm(2), P < 0.05). Patellar tendon collagen content was ∼32% greater in the chronic and acute groups compared with the control or insulin groups (control group: 681 ± 63 µg collagen/mg dry wt, acute group: 938 ± 21 µg collagen/mg dry wt, chronic: 951 ± 52 µg collagen/mg dry wt, and insulin group: 596 ± 84 µg collagen/mg dry wt, P < 0.05). In contrast, patellar tendon hydroxylysyl pyridinoline cross linking and collagen fibril organization were unchanged by diabetes or insulin (P > 0.05). Our findings suggest that 10 wk of streptozotocin-induced diabetes does not alter rat tendon mechanical properties even with an increase in collagen content. Future studies could attempt to further address the mechanisms contributing to the increase in tendon problems noted in diabetic patients, especially since our data suggest that hyperglycemia per se does not alter tendon mechanical properties.

Patellar tendon biomechanical and morphologic properties and their relationship to serum clinical variables in persons with prediabetes and type 2 diabetes.

Tendon biomechanical properties and fibril organization are altered in patients with diabetes compared to healthy individuals, yet few biomarkers have been associated with in vivo tendon properties. We investigated the relationships between in vivo imaging-based tendon properties, serum variables, and patient characteristics across healthy controls (n = 14, age: 45 ± 5 years, body mass index [BMI]: 24 ± 1, hemoglobin A1c [HbA1c]: 5.3 ± 0.1%), prediabetes (n = 14, age: 54 ± 5 years, BMI: 29 ± 2; HbA1c: 5.7 ± 0.1), and type 2 diabetes (n = 13, age: 55 ± 3 years, BMI: 33 ± 2, HbA1c: 6.7 ± 0.3). We used ultrasound speckle-tracking and measurements from magnetic resonance imaging (MRI) to estimate the patellar tendon in vivo tangent modulus. Analysis of plasma c-peptide, interleukin-1ß (IL-1ß), IL-6, IL-8, tumor necrosis factor-α (TNF-α), adiponectin, leptin, insulin-like growth factor 1 (IGF-1), and C-reactive protein (CRP) was completed. We built regression models incorporating statistically significant covariates and indicators for the clinically defined groups. We found that tendon cross-sectional area normalized to body weight (BWN CSA) and modulus were lower in patients with type 2 diabetes than in healthy controls (p < 0.05). Our regression analysis revealed that a model that included BMI, leptin, high-density lipoprotein (HDL), low-density lipoprotein (LDL), age, and group explained ~70% of the variability in BWN CSA (R2 = 0.70, p < 0.001). For modulus, including the main effects LDL, groups, HbA1c, age, BMI, cholesterol, IGF-1, c-peptide, leptin, and IL-6, accounted for ~54% of the variability in modulus (R2 = 0.54, p < 0.05). While BWN CSA and modulus were lower in those with diabetes, group was a poor predicter of tendon properties when considering the selected covariates. These data highlight the multifactorial nature of tendon changes with diabetes and suggest that blood variables could be reliable predictors of tendon properties.

Prostaglandin and myokine involvement in the cyclooxygenase-inhibiting drug enhancement of skeletal muscle adaptations to resistance exercise in older adults.

Twelve weeks of resistance training (3 days/wk) combined with daily consumption of the cyclooxygenase-inhibiting drugs acetaminophen (4.0 g/day; n = 11, 64 ± 1 yr) or ibuprofen (1.2 g/day; n = 13, 64 ± 1 yr) unexpectedly promoted muscle mass and strength gains 25-50% above placebo (n = 12, 67 ± 2 yr). To investigate the mechanism of this adaptation, muscle biopsies obtained before and ∼72 h after the last training bout were analyzed for mRNA levels of prostaglandin (PG)/cyclooxygenase pathway enzymes and receptors [arachidonic acid synthesis: cytosolic phospholipase A(2) (cPLA(2)) and secreted phospholipase A(2) (sPLA(2)); PGF(2α) synthesis: PGF(2α) synthase and PGE(2) to PGF(2α) reductase; PGE(2) synthesis: PGE(2) synthase-1, -2, and -3; PGF(2α) receptor and PGE(2) receptor-4], cytokines and myokines involved in skeletal muscle adaptation (TNF-α, IL-1ß, IL-6, IL-8, IL-10), and regulators of muscle growth [myogenin, myogenic regulatory factor-4 (MRF4), myostatin] and atrophy [Forkhead box O3A (FOXO3A), atrogin-1, muscle RING finger protein 1 (MuRF-1), inhibitory κB kinase ß (IKKß)]. Training increased (P < 0.05) cPLA(2), PGF(2α) synthase, PGE(2) to PGF(2α) reductase, PGE(2) receptor-4, TNF-α, IL-1ß, IL-8, and IKKß. However, the PGF(2α) receptor was upregulated (P < 0.05) only in the drug groups, and the placebo group upregulation (P < 0.05) of IL-6, IL-10, and MuRF-1 was eliminated in both drug groups. These results highlight prostaglandin and myokine involvement in the adaptive response to exercise in older individuals and suggest two mechanisms underlying the enhanced muscle mass gains in the drug groups: 1) The drug-induced PGF(2α) receptor upregulation helped offset the drug suppression of PGF(2α)-stimulated protein synthesis after each exercise bout and enhanced skeletal muscle sensitivity to this stimulation. 2) The drug-induced suppression of intramuscular PGE(2) production increased net muscle protein balance after each exercise bout through a reduction in PGE(2)-induced IL-6 and MuRF-1, both promoters of muscle loss.

The influence of acute resistance exercise on cyclooxygenase-1 and -2 activity and protein levels in human skeletal muscle.

This study evaluated the activity and content of cyclooxygenase (COX)-1 and -2 in response to acute resistance exercise (RE) in human skeletal muscle. Previous work suggests that COX-1, but not COX-2, is the primary COX isoform elevated with resistance exercise in human skeletal muscle. COX activity, however, has not been assessed after resistance exercise in humans. It was hypothesized that RE would increase COX-1 but not COX-2 activity. Muscle biopsies were taken from the vastus lateralis of nine young men (25 ± 1 yr) at baseline (preexercise), 4, and 24 h after a single bout of knee extensor RE (three sets of 10 repetitions at 70% of maximum). Tissue lysate was assayed for COX-1 and COX-2 activity. COX-1 and COX-2 protein levels were measured via Western blot analysis. COX-1 activity increased at 4 h (P < 0.05) compared with preexercise, but returned to baseline at 24 h (PRE: 60 ± 10, 4 h: 106 ± 22, 24 h: 72 ± 8 nmol PGH2·g total protein(-1)·min(-1)). COX-2 activity was elevated at 4 and 24 h after RE (P < 0.05, PRE: 51 ± 7, 4 h: 100 ± 19, 24 h: 98 ± 14 nmol PGH2·g total protein(-1)·min(-1)). The protein level of COX-1 was not altered (P > 0.05) with acute RE. In contrast, COX-2 protein levels were nearly 3-fold greater (P > 0.05) at 4 h and 5-fold greater (P = 0.06) at 24 h, compared with preexercise. In conclusion, COX-1 activity increases transiently with exercise independent of COX-1 protein levels. In contrast, both COX-2 activity and protein levels were elevated with exercise, and this elevation persisted to at least 24 h after RE.

Effects of Consuming Ounce-Equivalent Portions of Animal- vs. Plant-Based Protein Foods, as Defined by the Dietary Guidelines for Americans on Essential Amino Acids Bioavailability in Young and Older Adults: Two Cross-Over Randomized Controlled Trials.

BACKGROUND: The Dietary Guidelines for Americans (DGA) recommends consuming a variety of "Protein Foods" based on "ounce-equivalent" (oz-eq) portions. No study has assessed the same oz-eq portions of animal- vs. plant-based protein foods on essential amino acid (EAA) bioavailability for protein anabolism in young and older adults. OBJECTIVES: We assessed the effects of consuming two oz-eq portions of pork, eggs, black beans, and almonds on postprandial EAA bioavailability in young and older adults. METHODS: We conducted two investigator-blinded, randomized crossover trials in young (n = 30; mean age ± SD: 26.0 ± 4.9 y) and older adults (n = 25; mean age ± SD: 64.2 ± 6.6 y). Participants completed four testing sessions where they consumed a standardized meal with two oz-eq of either unprocessed lean pork, whole eggs, black beans, or sliced almonds. Blood samples were taken at baseline and 30, 60, 120, 180, 240, and 300 min postprandially. Plasma EAA bioavailability was based on postprandial integrated positive areas under the curve. RESULTS: Participant age did not affect EAA bioavailability among the four protein foods tested. Two oz-eq portions of pork (7.36 g EAA) and eggs (5.38 g EAA) resulted in greater EAA bioavailability than black beans (3.02 g EAA) and almonds (1.85 g EAA) in young and older adults, separately or combined (p < 0.0001 for all). Pork resulted in greater EAA bioavailability than eggs in young adults (p < 0.0001), older adults (p = 0.0007), and combined (p < 0.0001). There were no differences in EAA bioavailability between black beans and almonds. CONCLUSIONS: The same "oz-eq" portions of animal- and plant-based protein foods do not provide equivalent EAA content and postprandial bioavailability for protein anabolism in young and older adults.

Combined effects of heavy ion exposure and simulated Lunar gravity on skeletal muscle.

BACKGROUND: The limitations to prolonged spaceflight include unloading-induced atrophy of the musculoskeletal system which may be enhanced by exposure to the space radiation environment. Previous results have concluded that partial gravity, comparable to the Lunar surface, may have detrimental effects on skeletal muscle. However, little is known if these outcomes are exacerbated by exposure to low-dose rate, high-energy radiation common to the space environment. Therefore, the present study sought to determine the impact of highly charge, high-energy (HZE) radiation on skeletal muscle when combined with partial weightbearing to simulate Lunar gravity. We hypothesized that partial unloading would compromise skeletal muscle and these effects would be exacerbated by radiation exposure. METHODS: For month old female BALB/cByJ mice were -assigned to one of 2 groups; either full weight bearing (Cage Controls, CC) or partial weight bearing equal to 1/6th bodyweight (G/6). Both groups were then divided to receive either a single whole body absorbed dose of 0.5 Gy of 300 MeV 28Si ions (RAD) or a sham treatment (SHAM). Radiation exposure experiments were performed at the NASA Space Radiation Laboratory (NSRL) located at Brookhaven National Laboratory on Day 0, followed by 21 d of CC or G/6 loading. Muscles of the hind limb were used to measure protein synthesis and other histological measures. RESULTS: Twenty-one days of Lunar gravity (G/6) resulted in lower soleus, plantaris, and gastrocnemius muscle mass. Radiation exposure did not further impact muscle mass. 28Si exposure in normal ambulatory animals (RAD+CC) did not impact gastrocnemius muscle mass when compared to SHAM+CC (p>0.05), but did affect the soleus, where mass was higher following radiation compared to SHAM (p<0.05). Mixed gastrocnemius muscle protein synthesis was lower in both unloading groups. Fiber type composition transitioned towards a faster isoform with partial unloading and was not further impacted by radiation. The combined effects of partial loading and radiation partially mitigated fiber cross-sectional area when compared to partial loading alone. Radiation and G/6 reduced the total number of myonuclei per fiber while leading to elevated BrdU content of skeletal muscle. Similarly, unloading and radiation resulted in higher collagen content of muscle when compared to controls, but the effects of combined exposure were not additive. CONCLUSIONS: The results of this study confirm that partial weightbearing causes muscle atrophy, in part due to reductions of muscle protein synthesis in the soleus and gastrocnemius as well as reduced peripheral nuclei per fiber. Additionally, we present novel data illustrating 28Si exposure reduced nuclei in muscle fibers despite higher satellite cell fusion, but did not exacerbate muscle atrophy, CSA changes, or collagen content. In conclusion, both partial loading and HZE radiation can negatively impact muscle morphology.

Influence of acetaminophen consumption and exercise on Achilles tendon structural properties in male Wistar rats.

Chronic consumption of acetaminophen (APAP) during exercise training leads to a reduction in tendon stiffness and modulus compared with a placebo. We explored whether this effect could be due to a reduction in tendon collagen content or cross-linking. Ten-week-old male Wistar rats (n = 50) were divided into placebo or APAP groups and into sedentary or treadmill-exercised groups. APAP (200 mg/kg) or saline was administered once daily by oral gavage. Rats in the exercise groups ran on a treadmill 5 days per week for 8 wk with progression to 60 min per day, 20 m/min, and 8° incline. After 8 wk, lyophilized Achilles tendon samples were assayed for the collagen-specific amino acid hydroxyproline and cross-linking [hydroxylyslpyridinoline (HP)] content by high-performance liquid chromatrography. Collagen content was not influenced by exercise or APAP (P > 0.05). Compared with placebo, tendon water content was 7% (P = 0.006, main effect) lower in animals consuming APAP (placebo: 54.79 ± 0.8%, APAP: 50.89 ± 1.2%). HP in the Achilles tendon was 36% greater (sedentary: 141 ± 15, exercise: 204 ± 26 mmol/mol collagen) in the exercise-trained rats independent of drug treatment (P = 0.020, main effect). Independent of exercise, HP content was 33% lower (P = 0.032, main effect) in the animals consuming APAP (placebo: 195 ± 21, APAP: 140 ± 19 mmol/mol collagen). Our data suggests that chronic consumption of APAP results in a reduction in collagen cross-linking and a loss of tissue water independent of chronic exercise. This reduction in cross-linking and water content could contribute to the decrease in tendon stiffness noted in humans chronically consuming APAP.