Muscle Metabolites Research Articles

Intracellular PO2 Does Not Limit Skeletal Muscle Bioenergetics During Fatiguing Contractions In Vivo

Perturbations to the intracellular milieu during muscular work can include the accumulation of fatigue‐inducing metabolites in skeletal muscle. It has been suggested that oxygen availability contributes to fatigue by limiting the bioenergetic pathways that support contractile activity. Our purpose was to examine the role of intracellular oxygen (PO2) in the disruption of muscle bioenergetics and consequent fatigue in vivo. We hypothesized that fatigue (decrease in peak power) would be associated with acidosis and inorganic phosphate (Pi), but not with PO2, which would remain above the critical threshold for limiting mitochondrial respiration. Seven untrained adults (35 ± 3 yr, mean ± SD; 5 F) performed 120 maximal, isotonic contractions (1 every 2 s) at 20% of peak torque on an MR‐compatible knee extension ergometer in a 3‐tesla magnetic resonance (MR) system. Phosphorus (2000‐ms repetition time) and proton (30‐ms repetition time) MR spectra were collected concurrently with a dual‐tuned (1H/31P) surface coil on the vastus lateralis muscle during 60 s of rest and 4 min of contractions; spectra were averaged to obtain 20‐s temporal resolution. Peaks corresponding to phosphocreatine (PCr), Pi, ATP and deoxymyoglobin (dMb) were fit using jMRUI. [PCr] and [Pi] were calculated assuming ATP = 8.2 mM at rest. Oxidative ATP synthesis (ATPOX, mM·s‐1) was calculated as Vmax·(1+K·([ADP][Pi][ATP]‐1))‐1 where Vmax was determined from PCr resynthesis kinetics, [ADP] was calculated using the creatine kinase equilibrium, and K = 1.1 mM. PO2 was calculated using the oxygen binding curve for myoglobin (Mb; P50 · [(1‐□)/□]), where □ is fractional Mb desaturation relative to maximal dMb during cuff occlusion, assuming a P50 for Mb of 2.39 Torr. Statistics: paired t‐tests and Pearson correlations on individual data. PO2 was fit using bilinear regression to obtain the time to PO2 plateau. Peak power declined to 63.1 ± 16.2% of initial (p < 0.01), indicating significant fatigue. PCr declined from 31.6 ± 1.7 mM at rest to 5.6 ± 4.4 mM, Pi increased from 2.9 ± 0.4 mM to 22.0 ± 3.1 mM, and pH declined to 6.61 ± 0.17 (p < 0.01, all). PO2was 36.5 ± 6.4 Torr at rest and decreased rapidly at the onset of contractions. By 27.4 ± 2.4 s, PO2 plateaued at 1.54 ± 0.6 Torr, well above the critical threshold of 0.35 Torr. ATPOX increased steadily from the start of contractions to 0.49 ± 0.16 mM·s‐1 at 100 s, which is well beyond the time when PO2 plateaued, further indicating that PO2 did not limit oxidative phosphorylation. Changes in pH and Pi were linearly associated with changes in peak power (r = 0.74 ± 0.2 and r = ‐0.66 ± 0.1, respectively), consistent with their role in myosin cross‐bridge impairment in fatigue. In contrast, there was no linear relationship between PO2 and fatigue, pH, or [Pi] during or at the end of the contraction protocol (p > 0.22, all). Overall, the maintenance of PO2 above its critical threshold, as well as its dissociation from ATPOX, fatigue and the metabolites that cause it, indicate that an oxygen limitation does not impair bioenergetic pathways or power production during maximal knee extension contractions in vivo.

Effect of short-term dietary protein restriction before slaughter on meat quality and skeletal muscle metabolomic profile in culled ewes

Tenderness is one of the most important meat quality traits. It has been suggested that a short-term dietary protein restriction prior to slaughter could change muscle protein turnover and improve meat tenderness without affecting animal performance. Therefore, the aim of this study was to evaluate the effect of a short-term dietary protein restriction in the end of the finishing phase on skeletal muscle metabolome and meat quality traits of culled ewes. Fourteen crossbred ewes (3 years of age) averaging 39.3 ± 9.4 kg of body weight (BW) were enrolled in this study. Culled ewes were grouped (n = 7) and randomly assigned to one of the following treatments, consisting of: continuously fed the control diet with 13.3% of crude protein (CTRL); fed the control diet to reach the target body weight (45 kg) followed by 14 days feeding a low protein diet (4.33% crude protein; STR). The experiment lasted 46 days. Short-term dietary protein restriction increased meat shear-force at 1 day postmortem (3.67 vs. 3.02 kgf; P = 0.04), which could be partially explained by the higher total collagen content (P = 0.02) in STR compared to CTRL group (2.35 vs. 2.05 mg/g). The protein restriction did not affect final body weight nor carcass weight (P > 0.05), but decreased backfat thickness (3.19 vs. 5.10 mm; P = 0.04). Excreted and retained nitrogen were respectively 80.4% and 70% lower in STR (P < 0.01) than in the CTRL group. A lower abundance of valine (P = 0.02) and asparagine (P = 0.01) was observed in skeletal muscle of STR ewes compared to the CTRL group. Of the muscle metabolites identified, progesterone, succinic acid, and tartronic acid were positively correlated with meat shear-force (toughness), while lysine was negatively correlated (P < 0.05) with the same trait. Collectively, our data shows that despite the changes observed in the skeletal muscle for the abundance of metabolites involved in energy and protein metabolism, the short-term dietary protein restriction near maintenance level prior to slaughter did not improve quality of ewes’ meat quality traits.

Independent and Interactive Effects of Genetic Background and Sex on Tissue Metabolomes of Adipose, Skeletal Muscle, and Liver in Mice.

Genetics play an important role in the development of metabolic diseases. However, the relative influence of genetic variation on metabolism is not well defined, particularly in tissues, where metabolic dysfunction that leads to disease occurs. We used inbred strains of laboratory mice to evaluate the impact of genetic variation on the metabolomes of tissues that play central roles in metabolic diseases. We chose a set of four common inbred strains that have different levels of susceptibility to obesity, insulin resistance, and other common metabolic disorders. At the ages used, and under standard husbandry conditions, these lines are not overtly diseased. Using global metabolomics profiling, we evaluated water-soluble metabolites in liver, skeletal muscle, and adipose from A/J, C57BL/6J, FVB/NJ, and NOD/ShiLtJ mice fed a standard mouse chow diet. We included both males and females to assess the relative influence of strain, sex, and strain-by-sex interactions on metabolomes. The mice were also phenotyped for systems level traits related to metabolism and energy expenditure. Strain explained more variation in the metabolite profile than did sex or its interaction with strain across each of the tissues, especially in liver. Purine and pyrimidine metabolism and pathways related to amino acid metabolism were identified as pathways that discriminated strains across all three tissues. Based on the results from ANOVA, sex and sex-by-strain interaction had modest influence on metabolomes relative to strain, suggesting that the tissue metabolome remains largely stable across sexes consuming the same diet. Our data indicate that genetic variation exerts a fundamental influence on tissue metabolism.

Acetaminophen pharmacokinetics in geese.

To evaluate the pharmacokinetics of acetaminophen (APAP) after single-dose IV and PO in the goose; to quantify APAP and its main metabolites in goose muscle, heart, lung, liver, and kidney; and to perform a histopathologic evaluation of goose stomach, duodenum, liver, and kidney tissues for potential signs of toxicity. 24 geese. Geese were randomly divided into 3 groups (n = 8). Group I received APAP (10 mg/kg) IV, and groups II and III received the same dose PO. Groups I and II were used for the pharmacokinetic assessment, and group III was used for the residue analysis and histopathologic evaluation. APAP and its metabolites were quantified in plasma and tissues by ultra-high-performance liquid chromatography-tandem mass spectrometry, and the pharmacokinetic analysis was performed using a noncompartmental approach. APAP plasma concentrations were lower than those of the metabolites in similar selected time points after both treatments. After IV treatment, the APAP area under the curve value was statistically higher than that after PO administration, resulting in an oral bioavailability of 46%. In contrast, the area under the curve of the metabolites following PO administration was statistically higher than those found after IV administration. Tissue residues of APAP were highest in the liver, with an accumulation index > 1. Fatty degeneration of hepatocytes was observed 24 hours after administration of APAP. In geese, treatment by PO administration of APAP shows incomplete absorption and a slight accumulation in lung and liver. Tissue alterations occurred in the liver at 24 hours, while no signs of toxicity were found in the other tested organs.

Gut Microbiota Modulate Rabbit Meat Quality in Response to Dietary Fiber.

Antibiotics are widely used in gastrointestinal diseases in meat rabbit breeding, which causes safety problems for meat products. Dietary fiber can regulate the gut microbiota of meat rabbits, but the mechanism of improving meat quality is largely unknown. The objective of this study was to evaluate the effects of adding different fiber sources to rabbit diets on the growth performance, gut microbiota composition, and muscle metabolite composition of meat rabbits. A total of 18 New Zealand white rabbits of similar weight (40 ± 1 day old) were randomly assigned to beet pulp treatment (BP), alfalfa meal treatment (AM), and peanut vine treatment (PV). There were 6 repeats in each treatment and all were raised in a single cage. The predictive period was 7 days and the experimental period was 40 days. The results revealed that AM and PV supplementation increased growth performance, slaughter performance, and intestinal development of meat rabbits compared with the BP treatment, and especially the effect of AM treatment was better. The content of butyric acid was increased in PV and AM treatments compared with the BP treatment. The expression of mitochondrial biosynthesis genes of liver, cecum, and muscle showed that AM treatment increased gene expression of CPT1b compared to the BP treatment. In addition, AM and PV treatments significantly increased the microbial diversity and richness compared with BP treatment, and their bacterial community composition was similar, and there were some differences between AM and PV treatments and BP treatment. Metabonomics analysis of muscle showed that AM treatment significantly increased amino acid and fatty acid metabolites compared with BP treatment, which were mainly concentrated in energy metabolism, amino acid metabolism, and fatty acid regulation pathways. Furthermore, through correlation analysis, it was found that there was a significant correlation between rumenococci in the cecum and amino acid metabolites in the muscle. Overall, these findings indicate that AM may affect the body's health by changing its gut microbiota, and then improving meat quality, and the intestinal–muscle axis provides a theoretical basis.

The Effect of Breed and Age on the Growth Performance, Carcass Traits and Metabolic Profile in Breast Muscle of Chinese Indigenous Chickens.

Indigenous chickens possess desirable characteristics and account for considerable proportions of the total chicken production in China. The current study examined the growth performance, carcass characteristics and muscle metabolites among a crossbred broiler and two indigenous, yellow-feathered chickens (Mahuang and Tuer) with different ages (60 and 75 days). Results indicated that the crossbred broiler had better feed efficiency, higher breast and thigh muscle yield, as well as a lower abdominal fat percentage than Mahuang and Tuer chickens (p < 0.05). Gas chromatography–mass spectrometry-based metabolomics and multivariate analysis revealed sugars, amino acids and organic acids were the predominant metabolites that differed among the three chicken breeds. Growth performance and carcass traits of yellow-feathered chickens exhibited significant differences with the extension of the feeding period (p < 0.05). Moreover, differential metabolites reflected altered aminoacyl-tRNA biosynthesis, ATP-binding cassette transporters, pantothenate and CoA biosynthesis, as well as glutathione metabolism in yellow-feathered chickens affected by age. Collectively, this study contributes to a deeper understanding of the production efficiency and chemical composition of precursor flavor in Chinese indigenous, yellow-feathered chicken.

Impact of protocol-based physiotherapy on insulin sensitivity and peripheral glucose metabolism in critically ill patients.

BackgroundThe impact of physiotherapy on insulin sensitivity and peripheral glucose metabolism in critically ill patients is not well understood.MethodsThis pooled analysis investigates the impact of different physiotherapeutic strategies on insulin sensitivity in critically ill patients. We pooled data from two previous trials in adult patients with sequential organ failure assessment score (SOFA)≥ 9 within 72 h of intensive care unit (ICU) admission, who received hyperinsulinaemic euglycaemic (HE) clamps. Patients were divided into three groups: standard physiotherapy (sPT, n = 22), protocol‐based physiotherapy (pPT, n = 8), and pPT with added muscle activating measures (pPT+, n = 20). Insulin sensitivity index (ISI) was determined by HE clamp. Muscle metabolites lactate, pyruvate, and glycerol were measured in the M. vastus lateralis via microdialysis during the HE clamp. Histochemical visualization of glucose transporter‐4 (GLUT4) translocation was performed in surgically extracted muscle biopsies. All data are reported as median (25th/75th percentile) (trial registry: ISRCTN77569430 and ISRCTN19392591/ethics approval: Charité‐EA2/061/06 and Charité‐EA2/041/10).ResultsFifty critically ill patients (admission SOFA 13) showed markedly decreased ISIs on Day 17 (interquartile range) 0.029 (0.022/0.048) (mg/min/kg)/(mU/L) compared with healthy controls 0.103 (0.087/0.111), P < 0.001. ISI correlated with muscle strength measured by medical research council (MRC) score at first awakening (r = 0.383, P = 0.026) and at ICU discharge (r = 0.503, P = 0.002). Different physiotherapeutic strategies showed no effect on the ISI [sPT 0.029 (0.019/0.053) (mg/min/kg)/(mU/L) vs. pPT 0.026 (0.023/0.041) (mg/min/kg)/(mU/L) vs. pPT+ 0.029 (0.023/0.042) (mg/min/kg)/(mU/L); P = 0.919]. Regardless of the physiotherapeutic strategy metabolic flexibility was reduced. Relative change of lactate/pyruvate ratio during HE clamp is as follows: sPT 0.09 (−0.13/0.27) vs. pPT 0.07 (−0.16/0.31) vs. pPT+ −0.06 (−0.19/0.16), P = 0.729, and relative change of glycerol concentration: sPT −0.39 (−0.8/−0.12) vs. pPT −0.21 (−0.33/0.07) vs. pPT+ −0.21 (−0.44/−0.03), P = 0.257. The majority of ICU patients showed abnormal localization of GLUT4 with membranous GLUT4 distribution in 37.5% (3 of 8) of ICU patients receiving sPT, in 42.9% (3 of 7) of ICU patients receiving pPT, and in 53.8% (7 of 13) of ICU patients receiving pPT+ (no statistical testing possible).ConclusionsOur data suggest that a higher duration of muscle activating measures had no impact on insulin sensitivity or metabolic flexibility in critically ill patients with sepsis‐related multiple organ failure.

Glutamine Regulates Skeletal Muscle Immunometabolism in Type 2 Diabetes.

Dysregulation of skeletal muscle metabolism influences whole-body insulin sensitivity and glucose homeostasis. We hypothesized that type 2 diabetes–associated alterations in the plasma metabolome directly contribute to skeletal muscle immunometabolism and the subsequent development of insulin resistance. To this end, we analyzed the plasma and skeletal muscle metabolite profile and identified glutamine as a key amino acid that correlates inversely with BMI and insulin resistance index (HOMA-IR) in men with normal glucose tolerance or type 2 diabetes. Using an in vitro model of human myotubes and an in vivo model of diet-induced obesity and insulin resistance in male mice, we provide evidence that glutamine levels directly influence the inflammatory response of skeletal muscle and regulate the expression of the adaptor protein GRB10, an inhibitor of insulin signaling. Moreover, we demonstrate that a systemic increase in glutamine levels in a mouse model of obesity improves insulin sensitivity and restores glucose homeostasis. We conclude that glutamine supplementation may represent a potential therapeutic strategy to prevent or delay the onset of insulin resistance in obesity by reducing inflammatory markers and promoting skeletal muscle insulin sensitivity.

Reduced Levels of NAD in Skeletal Muscle and Increased Physiologic Frailty Are Associated With Viral Coinfection in Asymptomatic Middle-Aged Adults.

Background:People living with HIV (PLWH) are disproportionately burdened with multimorbidity and decline in physiologic function compared with their uninfected counterparts, but biological mechanisms that differentially contribute to the decline in muscle function in PLWH compared with uninfected people remain understudied.Setting:The study site was Brigham and Women's Hospital, Harvard Medical School, Boston, MA.Methods:We evaluated skeletal muscle tissue for levels of total nicotinamide adenine dinucleotide (NAD), NAD+, and nicotinamide adenine dinucleotide (NADH) in middle-aged asymptomatic PLWH, coinfected with hepatitis C virus and/or cytomegalovirus and compared them with uninfected control participants.Results:Of the 54 persons with muscle biopsy data, the mean age was 57 years with 33% women. Total NAD levels declined in skeletal muscle in association with HIV infection and was exacerbated by hepatitis C virus and cytomegalovirus coinfection, with lowest levels of total NAD, NAD+, and NADH among persons who were coinfected with all 3 viruses (P = 0.015, P = 0.014, and P = 0.076, respectively). Levels of total NAD, NAD+, and NADH in skeletal muscle were inversely associated with inflammation (P = 0.014, P = 0.013, and P = 0.055, respectively). Coinfections were also associated with measures of inflammation (CD4/CD8 ratio: P < 0.001 and sCD163: P < 0.001) and immune activation (CD38 and human leukocyte antigen-DR expression on CD8 T cells: P < 0.001). In addition, coinfection was associated with increased physiologic frailty based on the Veteran Aging Cohort Study 1.0 index assessment (P = 0.001).Conclusions:Further research is warranted to determine the clinical relevance of preclinical deficits in NAD metabolites in skeletal muscle in association with viral coinfection and inflammation, as well as the observed association between viral coinfection and physiologic frailty.

The Effect of Long-Lasting Swimming on Rats Skeletal Muscles Energy Metabolism after Nine Days of Dexamethasone Treatment.

This study investigates the effect of Dexamethasone (Dex) treatment on blood and skeletal muscle metabolites level and skeletal muscle activity of enzymes related to energy metabolism after long-duration swimming. To evaluate whether Dex treatment, swimming, and combining these factors act on analyzed data, rats were randomly divided into four groups: saline treatment non-exercise and exercise and Dex treatment non-exercised and exercised. Animals in both exercised groups underwent long-lasting swimming. The concentration of lipids metabolites, glucose, and lactate were measured in skeletal muscles and blood according to standard colorimetric and fluorimetric methods. Also, activities of enzymes related to aerobic and anaerobic metabolism were measured in skeletal muscles. The results indicated that Dex treatment induced body mass loss and increased lipid metabolites in the rats’ blood but did not alter these changes in skeletal muscles. Interestingly, prolonged swimming applied after 9 days of Dex treatment significantly intensified changes induced by Dex; however, there was no difference in skeletal muscle enzymatic activities. This study shows for the first time the cumulative effect of exercise and Dex on selected elements of lipid metabolism, which seems to be essential for the patient’s health due to the common use of glucocorticoids like Dex.

Up-regulation of thioesterase superfamily member 2 in skeletal muscle promotes hepatic steatosis and insulin resistance in mice.

Thioesterase superfamily member 2 (Them2) is highly expressed in liver and oxidative tissues, where it hydrolyzes long-chain fatty acyl-CoA esters to free fatty acids and CoA. Although mice globally lacking Them2 (Them2-/- ) are protected against diet-induced obesity, hepatic steatosis (HS), and insulin resistance (IR), liver-specific Them2-/- mice remain susceptible. The aim of this study was to test whether Them2 activity in extrahepatic oxidative tissues is a primary determinant of HS and IR. Upon observing IR and up-regulation of Them2 in skeletal, but not cardiac, muscle of high-fat-diet (HFD)-fed wild-type compared to Them2-/- mice, we created mice with Them2 specifically deleted in skeletal (S-Them2-/- ) and cardiac muscle (C-Them2-/- ), as well as in adipose tissue (A-Them2-/- ). When fed an HFD, S-Them2-/- , but not C-Them2-/- or A-Them2-/- , mice exhibited reduced weight gain and improved glucose homeostasis and insulin sensitivity. Reconstitution of Them2 expression in skeletal muscle of global Them2-/- mice, using adeno-associated virus, was sufficient to restore excess weight gain. Increased rates of fatty acid oxidation in skeletal muscle of S-Them2-/- mice contributed to protection from HFD-induced HS by increasing VLDL triglyceride secretion rates in response to greater demand. Increases in insulin sensitivity were further attributable to alterations in production of skeletal muscle metabolites, including short-chain fatty acids, branched-chain amino acids, and pentose phosphate pathway intermediates, as well as in expression of myokines that modulate insulin responsiveness. These results reveal a key role for skeletal muscle Them2 in the pathogenesis of HS and IR and implicate it as a target in the management of NAFLD.

Improved Peripheral and Hepatic Insulin Sensitivity after Lifestyle Interventions in Type 2 Diabetes Is Associated with Specific Metabolomic and Lipidomic Signatures in Skeletal Muscle and Plasma.

Lifestyle interventions with weight loss can improve insulin sensitivity in type 2 diabetes (T2D), but mechanisms are unclear. We explored circulating and skeletal muscle metabolite signatures of altered peripheral (pIS) and hepatic insulin sensitivity (hIS) in overweight and obese T2D individuals that were randomly assigned a 12-week Paleolithic-type diet with (diet-ex, n = 13) or without (diet, n = 13) supervised exercise. Baseline and post-intervention measures included: mass spectrometry-based metabolomics and lipidomics of skeletal muscle and plasma; pIS and hIS; ectopic lipid deposits in the liver and skeletal muscle; and skeletal muscle fat oxidation rate. Both groups lowered BMI and total % fat mass and increased their pIS. Only the diet-group improved hIS and reduced ectopic lipids in the liver and muscle. The combined improvement in pIS and hIS in the diet-group were associated with decreases in muscle and circulating branched-chain amino acid (BCAA) metabolites, specifically valine. Improved pIS with diet-ex was instead linked to increased diacylglycerol (34:2) and triacylglycerol (56:0) and decreased phosphatidylcholine (34:3) in muscle coupled with improved muscle fat oxidation rate. This suggests a tissue crosstalk involving BCAA-metabolites after diet intervention with improved pIS and hIS, reflecting reduced lipid influx. Increased skeletal muscle lipid utilization with exercise may prevent specific lipid accumulation at sites that perturb insulin signaling.

Effect of black soldier fly (Hermetia illucens) larvae meal on lipid and glucose metabolism of Pacific white shrimp Litopenaeus vannamei.

The present study investigated the effect of black soldier fly (Hermetia illucens) larvae meal (BSF) on haemolymph biochemical indicators, muscle metabolites as well as the lipid and glucose metabolism of Pacific white shrimp Litopenaeus vannamei. Four diets were formulated in which the control diet contained 25 % of fishmeal (FM) and 10 % (BSF10), 20 % (BSF20), and 30 % (BSF30) of FM protein were replaced with BSF. Four hundred and eighty shrimp (0·88 ± 0·00 g) were distributed to four groups of three replicates and fed for 7 weeks. Results showed that growth performance of shrimp fed BSF30 significantly decreased compared with those fed FM, but there was no significant difference in survival among groups. The whole shrimp crude lipid content, haemolymph TAG and total cholesterol were decreased with the increasing BSF inclusion. The results of metabolomics showed that the metabolite patterns of shrimp fed different diets were altered, with significant changes in metabolites related to lipid metabolism, glucose metabolism as well as TCA cycle. The mRNA expressions of hk, pfk, pk, pepck, ampk, mcd, cpt-1 and scd1 in hepatopancreas were downregulated in shrimp fed BSF30, but mRNA expression of acc1 was upregulated. Unlike BSF30, the mRNA expressions of fas, cpt-1, fbp and 6pgd in hepatopancreas were upregulated in shrimp fed BSF20. This study indicates that BSF20 diet promoted lipid synthesis and lipolysis, while BSF30 diet weakened β-oxidation and glycolysis as well as affected the unsaturated fatty acids synthesis, which may affect the growth performance and body composition of shrimp.

Biochemical Response to Freezing in the Siberian Salamander Salamandrella keyserlingii.

Simple SummaryThe Siberian salamander is a unique amphibian that is capable to survive long-term freezing at −55 °C. We used 1H-NMR analysis to study quantitative changes of multiple metabolites in liver and hindlimb muscle of the Siberian salamander in response to freezing. For the majority of molecules we observed significant changes in concentrations. Glycerol content in frozen organs was as high as 2% w/w, which confirms its role as a cryoprotectant. No other putative cryoprotectants were detected. Freezing resulted in increased concentrations of glycolysis products: lactate and alanine. Unexpectedly, we detected no increase in concentrations of succinate, which accumulates under ischemia in various tetrapods. Freezing proved to be a dramatic stress with high levels of nucleotide degradation products. There was also significant increase in the concentrations of choline and glycerophosphocholine, which may be interpreted as the degradation of biomembranes. Thus, we found that freezing results not only in macroscopical damage due to ice formation, but also to degradation of DNA and biomembranes.The Siberian salamander Salamandrella keyserlingii Dybowski, 1870 is a unique amphibian that is capable to survive long-term freezing at −55 °C. Nothing is known on the biochemical basis of this remarkable freezing tolerance, except for the fact that it uses glycerol as a low molecular weight cryoprotectant. We used 1H-NMR analysis to study quantitative changes of multiple metabolites in liver and hindlimb muscle of S. keyserlingii in response to freezing. For the majority of molecules we observed significant changes in concentrations. Glycerol content in frozen organs was as high as 2% w/w, which confirms its role as a cryoprotectant. No other putative cryoprotectants were detected. Freezing resulted in ischemia manifested as increased concentrations of glycolysis products: lactate and alanine. Unexpectedly, we detected no increase in concentrations of succinate, which accumulates under ischemia in various tetrapods. Freezing proved to be a dramatic stress with reduced adenosine phosphate pool and high levels of nucleotide degradation products (hypoxanthine, β-alanine, and β-aminoisobutyrate). There was also significant increase in the concentrations of choline and glycerophosphocholine, which may be interpreted as the degradation of biomembranes. Thus, we found that freezing results not only in macroscopical damage due to ice formation, but also to degradation of DNA and biomembranes.

Impacts of high-intensity exercise on the metabolomics profile of human skeletal muscle tissue.

The study aimed to identify and quantify the metabolites profile and metabolic pathways in human muscle tissue engaged during exhaustive high-intensity cycling exercise. Seven healthy physically active men performed a graded exercise test and an exhaustive supramaximal effort at 115% of maximal aerobic power with muscles biopsies performed in rest and immediately after exhaustion for quantifying of muscle metabolites changes by 1 H-NMR spectroscopy. The time until exhaustion (tlim) recorded was 224.7±35.5s whereas the muscle pH at exhaustion was 6.48±0.05. A total of 54metabolites were identified and quantified. The most enriched and impacted pathways included: beta oxidation of very long chain fatty acids, mitochondrial electron transport chain, alanine aspartate, and glutamate metabolism, citric acid cycle, arginine biosynthesis, propanoate metabolism, threonine and 2-oxobutanoate degradation and pyruvate metabolism. In addition, the muscle concentrations in Post exercise, compared to Pre increased significantly (p<0.0398) for fumarate (42.0%), succinate (101.2%), glucose (249.7%), lactate (122.8%), O-acetylcarnitine (164.7%), glycerol (79.3%), AMP (288.2%), 2-oxobutyrate (121.0%), and methanol (58.5%), whereas decreased significantly (p<0.010) for creatine phosphate (-70.2%), ADP (-56.5%), carnitine (-33.5%), and glutamate (-42.3%). Only the succinate was significantly correlated with tlim (r=-0.76; p=0.0497). Besides the classical expected contribution of glycolytic and phosphagen energetic pathways, it was demonstrated that the high-intensity exercise is also associated with pathways indicatives of amino acid and fatty acid oxidation metabolisms, highlighting the inverse relation between changes in the intramuscular succinate levels and tlim.

3D localized lactate detection in muscle tissue using double-quantum filtered 1 H MRS with adiabatic refocusing pulses at 7T.

Lactate is a key metabolite in skeletal muscle and whole-body physiology. Its MR visibility in muscle is affected by overlapping lipid signals and fiber orientation. Double-quantum filtered (DQF) 1 H MRS selectively detects lactate at 1.3ppm, but at ultra-high field the efficiency of slice-selective 3D-localization with conventional RF pulses is limited by bandwidth. This novel 3D-localized 1 H DQF MRS sequence uses adiabatic refocusing pulses to unambiguously detect lactate in skeletal muscle at 7T. Lactate double-quantum coherences were 3D-localized using slice-selective Shinnar-Le Roux optimized excitation and adiabatic refocusing pulses (similar to semi-LASER). DQF MR spectra were acquired at 7T from lactate phantoms, meat specimens with injected lactate (exploring multiple TEs and fiber orientations), and human gastrocnemius in vivo during and after exercise (without cuff ischemia). Lactate was readily detected, achieving the full potential of 50% signal with a DQF, in solution. The effects of fiber orientation and TE on the lactate doublet (peak splitting, amplitude, and phase) were in good agreement with theory and literature. Exercise-induced lactate accumulation was detected with 30s time resolution. This novel 3D-localized 1 H DQF MRS sequence can dynamically detect glycolytically generated lactate in muscle during exercise and recovery at 7T.

Corticosterone-Mediated Physiological Stress Alters Liver, Kidney, and Breast Muscle Metabolomic Profiles in Chickens.

Simple SummaryCorticosterone is the major stress hormone in birds and research has shown that an increase in corticosterone can have adverse effects on bird health (e.g., predisposition to disease) production performance metrics. However, it is not currently possible to monitor commercial flocks for stress before performance is affected. A popular model of chicken stress involves administering corticosterone to chickens though their drinking water. However, corticosterone is non-polar so it must first be dissolved in ethanol, which means that the chickens are also drinking ethanol. In this study, an untargeted nuclear magnetic resonance-based metabolomics approach was used to investigate the effects of this model of stress in chickens, as well as the effects corticosterone on the chicken kidney, liver, and breast muscle metabolomes. We hypothesized that physiological stress modulates the metabolome of liver, kidney, and breast muscle due to increases in catabolism and gluconeogenesis. The administration of corticosterone altered the chicken liver, kidney, and breast muscle metabolomes. However, the ethanol carrier affected the metabolome of all three tissues, which indicated that corticosterone should be administered in an alternate fashion in future metabolomics studies to remove the confounding effects of ethanol. Furthermore, future research should focus on relating metabolite changes in tissues to non-destructive markers like blood, feces, or feathers to develop new diagnostic tools to better monitor on-farm stress during production.The impact of physiological stress on the metabolomes of liver, kidney, and breast muscle was investigated in chickens. To incite a stress response, birds were continuously administered corticosterone (CORT) in their drinking water at three doses (0, 10, and 30 mg L−1), and they were sampled 1, 5, and 12 days after the start of the CORT administration. To solubilize CORT, it was first dissolved in ethanol and then added to water. The administration of ethanol alone significantly altered branched chain amino acid metabolism in both the liver and the kidney, and amino acid and nitrogen metabolism in breast muscle. CORT significantly altered sugar and amino acid metabolism in all three tissues, but to a much greater degree than ethanol alone. In this regard, CORT administration significantly altered 11, 46, and 14 unique metabolites in liver, kidney, and breast muscle, respectively. Many of the metabolites that were affected by CORT administration, such as mannose and glucose, were previously linked to increases in glycosylation and gluconeogenesis in chickens under conditions of production stress. Moreover, several of these metabolites, such as dimethylglycine, galactose, and carnosine were also previously linked to reduced quality meat. In summary, the administration of CORT in chickens significantly modulated host metabolism. Moreover, results indicated that energy potentials are diverted from muscle anabolism to muscle catabolism and gluconeogenesis during periods of stress.

Association Between Changes in Serum and Skeletal Muscle Metabolomics Profile With Maximum Power Output Gains in Response to Different Aerobic Training Programs: The Times Study.

Purpose: High heterogeneity of the response of cardiorespiratory fitness (CRF) to standardized exercise doses has been reported in different training programs, but the associated mechanisms are not widely known. This study investigated whether changes in the metabolic profile and pathways in blood serum and the skeletal muscle are associated with the inter-individual variability of CRF responses to 8-wk of continuous endurance training (ET) or high-intensity interval training (HIIT).Methods: Eighty men, young and sedentary, were randomized into three groups, of which 70 completed 8 wk of intervention (> 90% of sessions): ET, HIIT, or control. Blood and vastus lateralis muscle tissue samples, as well as the measurement of CRF [maximal power output (MPO)] were obtained before and after the intervention. Blood serum and skeletal muscle samples were analyzed by 600 MHz 1H-NMR spectroscopy (metabolomics). Associations between the pretraining to post-training changes in the metabolic profile and MPO gains were explored via three analytical approaches: (1) correlation between pretraining to post-training changes in metabolites' concentration levels and MPO gains; (2) significant differences between low and high MPO responders; and (3) metabolite contribution to significantly altered pathways related to MPO gains. After, metabolites within these three levels of evidence were analyzed by multiple stepwise linear regression. The significance level was set at 1%.Results: The metabolomics profile panel yielded 43 serum and 70 muscle metabolites. From the metabolites within the three levels of evidence (15 serum and 4 muscle metabolites for ET; 5 serum and 1 muscle metabolites for HIIT), the variance in MPO gains was explained: 77.4% by the intervention effects, 6.9, 2.3, 3.2, and 2.2% by changes in skeletal muscle pyruvate and valine, serum glutamine and creatine phosphate, respectively, in ET; and 80.9% by the intervention effects; 7.2, 2.2, and 1.2% by changes in skeletal muscle glycolate, serum creatine and creatine phosphate, respectively, in HIIT. The most changed and impacted pathways by these metabolites were: arginine and proline metabolism, glycine, serine and threonine metabolism, and glyoxylate and dicarboxylate metabolism for both ET and HIIT programs; and additional alanine, aspartate and glutamate metabolism, arginine biosynthesis, glycolysis/gluconeogenesis, and pyruvate metabolism for ET.Conclusion: These results suggest that regulating the metabolism of amino acids and carbohydrates may be a potential mechanism for understanding the inter-individual variability of CRF in responses to ET and HIIT programs.

Simultaneous Determination of Albendazole and Its Three Metabolites in Pig and Poultry Muscle by Ultrahigh-Performance Liquid Chromatography-Fluorescence Detection.

A fast, simple and efficient ultrahigh-performance liquid chromatography-fluorescence detection (UPLC-FLD) method for the determination of residues of albendazole (ABZ) and its three metabolites, albendazole sulfone (ABZ-SO2), albendazole sulfoxide (ABZ-SO), and albendazole-2-aminosulfone (ABZ-2NH2-SO2), in pig and poultry muscle (chicken, duck and goose) was established. The samples were extracted with ethyl acetate, and the extracts were further subjected to cleanup by utilizing a series of liquid–liquid extraction (LLE) steps. Then, extracts were purified by OASIS® PRiME hydrophilic-lipophilic balance (HLB) solid-phase extraction (SPE) cartridges (60 mg/3 mL). The target compounds were separated on an ACQUITY UPLC® BEH C18 (2.1 mm × 100 mm, 1.7 μm) chromatographic column, using a mobile phase composed of 31% acetonitrile and 69% aqueous solution (containing 0.2% formic acid and 0.05% triethylamine). The limits of detection (LODs) and limits of quantification (LOQs) of the four target compounds in pig and poultry muscle were 0.2–3.8 µg/kg and 1.0–10.9 µg/kg, respectively. The recoveries were all above 80.37% when the muscle samples were spiked with the four target compounds at the LOQ, 0.5 maximum residue limit (MRL), 1.0 MRL, and 2.0 MRL levels. The intraday relative standard deviations (RSDs) were less than 5.11%, and the interday RSDs were less than 6.29%.

Influence of an L-type SALMFamide neuropeptide on locomotory performance and muscle physiology in the sea cucumber Apostichopus japonicus.

Neuropeptides in the SALMFamide family serve as muscle relaxants in echinoderms and may affect locomotion, as the motor behavior in sea cucumbers involves alternating contraction and extension of the body wall, which is under the control of longitudinal muscle. We evaluated the effect of an L-type SALMFamide neuropeptide (LSA) on locomotory performance of Apostichopus japonicus. We also investigated the metabolites of longitudinal muscle tissue using ultra performance liquid chromatography and quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) to assess the potential physiological mechanisms underlying the effect of LSA. The hourly distance, cumulative duration and number of steps moved significantly increased in sea cucumbers in the fourth hour after injection with LSA. Also, the treatment enhanced the mean and maximum velocity by 9.8% and 17.8%, respectively, and increased the average stride by 12.4%. Levels of 27 metabolites in longitudinal muscle changed after LSA administration, and the increased concentration of pantothenic acid, arachidonic acid and lysophosphatidylethanolamine, and the altered phosphatidylethanolamine/phosphatidylcholine ratio are potential physiological mechanisms that could explain the observed effect of LSA on locomotor behavior in A. japonicus.