Global and targeted gene expression and protein content in skeletal muscle of young men following short-term creatine monohydrate supplementation (original) (raw)
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To test the hypothesis that creatine supplementation would enhance the anabolic responses of muscle cell signalling and gene expression to exercise, we studied nine subjects who received either creatine or a placebo (maltodextrin) for 5d in a doubleblind fashion before undergoing muscle biopsies: at rest; immediately after exercise (10×10 repetitions of one leg-extension at 80% 1-RM); and 24h and 72h later (all in the morning after fasting overnight). Creatine supplementation decreased the phosphorylation state of protein kinase B (PKB) on Thr308 at rest by 60% (P<0.05) and that of eukaryotic initiation factor 4E-binding protein on Thr37/46 (4E-BP1) by 30% 24h post-exercise (P<0.05). Creatine increased mRNA for collagen 1( 1), glucose transporter-4 (GLUT-4) and myosin heavy chain I at rest by 250%, 45% and 80% respectively, and myosin heavy chain IIA (MHCIIA) mRNA immediately after exercise by 70% (all P<0.05). Immediately after exercise, and independent of creatine, mRNA for muscle atrophy F-box (MAFbx), MHCIIA, peroxisome proliferator-activated receptor coactivator-1 and interleukin-6 were up-regulated (60-350%, P<0.05); the phosphorylation state of p38 both in the sarcoplasm and nucleus were increased (12 and 25 fold respectively, both P<0.05). Concurrently, the phosphorylation states of PKB (Thr308) and 4E-BP1 (Thr37/46) were decreased by 50% and 75% respectively (P<0.05). Twenty-four hours post-exercise, MAFbx, myostatin and GLUT-4 mRNA expression decreased below pre-exercise values (-35 to -50%, P<0.05); calpain 1 mRNA increased 70% 72h post-exercise (P<0.05) and at no other time. In conclusion, 5d of creatine supplementation does not enhance anabolic signalling but increases the expression of certain targeted genes.
Physiological genomics, 2003
The present study examined the validity and reliability of measuring the expression of various genes in human skeletal muscle using quantitative real-time RT-PCR on a GeneAmp 5700 sequence detection system with SYBR Green 1 chemistry. In addition, the validity of using some of these genes as endogenous controls (i.e., housekeeping genes) when human skeletal muscle was exposed to elevated total creatine levels and exercise was also examined. For all except 28S, linear relationships between the logarithm of the starting RNA concentrations and the cycle threshold (C(T)) values were established for beta-actin, beta2-microglobulin (beta2M), cyclophilin (CYC), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found a linear response between C(T) values and the logarithm of a given amount of starting cDNA for all the genes tested. The overall intra-assay coefficient of variance for these genes was 1.3% and 21% for raw C(T) values and the linear value of 2(-C(T)), respectively. Inter...
Nutrients
Creatine (Cr) and phosphocreatine (PCr) are physiologically essential molecules for life, given they serve as rapid and localized support of energy- and mechanical-dependent processes. This evolutionary advantage is based on the action of creatine kinase (CK) isozymes that connect places of ATP synthesis with sites of ATP consumption (the CK/PCr system). Supplementation with creatine monohydrate (CrM) can enhance this system, resulting in well-known ergogenic effects and potential health or therapeutic benefits. In spite of our vast knowledge about these molecules, no integrative analysis of molecular mechanisms under a systems biology approach has been performed to date; thus, we aimed to perform for the first time a convergent functional genomics analysis to identify biological regulators mediating the effects of Cr supplementation in health and disease. A total of 35 differentially expressed genes were analyzed. We identified top-ranked pathways and biological processes mediating...
Molecular and metabolic insights of creatine supplementation on resistance training
Revista Colombiana de Química, 2016
In recent years the research problem in the field of sports supplementation has changed to explain the metabolic mechanisms by which creatine (Cr) administration enhances the performance of certain sports or simply benefits the muscular adaptation. In this review for first time the biochemical mechanisms of Cr ingestion in a cell signaling insight were analyzed, focusing on energetic bioavailability enhancement and optimization of the temporal and spatial buffering of Cr/ PCr/CK system. Moreover, intensification in proliferation and differentiation processes of muscle cells (IGF-I/PI3K/Akt-PKB, SPHK1/MAPK/p38/MRFs, mTOR, cellular swelling, mitotic activity of satellite cells, actin polymerization, and myoblast fusion) and inactivation and/or reduction in the expression of ergolitic metabolites (GSK3β, myostatin and AMPK regulation) were examined. In this way, we explained from a metabolic point of view the increase in muscle mass, strength, fatigue resistance, and performance of high intensity sports after Cr monohydrate supplementation.
RT-PCRin human skeletal muscle using real-time Effects of creatine supplementation on housekeeping
2015
; 10.1152/physiolgenomics. 00060.2002.-The present study examined the validity and reliability of measuring the expression of various genes in human skeletal muscle using quantitative real-time RT-PCR on a GeneAmp 5700 sequence detection system with SYBR Green 1 chemistry. In addition, the validity of using some of these genes as endogenous controls (i.e., housekeeping genes) when human skeletal muscle was exposed to elevated total creatine levels and exercise was also examined. For all except 28S, linear relationships between the logarithm of the starting RNA concentrations and the cycle threshold (C T) values were established for -actin, 2-microglobulin (2M), cyclophilin (CYC), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). We found a linear response between CT values and the logarithm of a given amount of starting cDNA for all the genes tested. The overall intra-assay coefficient of variance for these genes was 1.3% and 21% for raw CT values and the linear value of 2 ϪCT , respectively. Interassay variability was 2.3% for raw CT values and 34% for the linear value of 2 ϪCT. We also examined the expression of various housekeeping genes in human skeletal muscle at days 0, 1, and 5 following oral supplementation with either creatine or a placebo employing a double-blind crossover study design. Treatments were separated by a 5-wk washout period. Immediately following each muscle sampling, subjects performed two 30-s all-out bouts on a cycle ergometer. Creatine supplementation increased (P Ͻ 0.05) muscle total creatine content above placebo levels; however, there were no changes (P Ͼ 0.05) in CT values across the supplementation periods for any of the genes. Nevertheless, 95% confidence intervals showed that GAPDH was variable, whereas -actin, 2M, and CYC were the least varying genes. Normalization of the data to these housekeeping genes revealed variable behavior for 2M with more stable expressions for both -actin and CYC. We conclude that, using real-time RT-PCR, -actin or CYC may be used as housekeeping genes to study gene expression in human muscle in experiments employing short-term creatine supplementation combined with highintensity exercise. gene expression; quantitative RT-PCR; endogenous controls; biological variability; PCR efficiency
Increased IGF mRNA in Human Skeletal Muscle after Creatine Supplementation
Medicine & Science in Sports & Exercise, 2005
DELDICQUE, L.. M. LOUIS, D. THEISEN, H. NIELENS. M. DEHOUX. J.-P. THISSEN. M. J. RENNIE, and M. FRANCAUX. Increased IGF mRNA in Human Skeletal Muscle after Creatine Supplementation. Med. Sci. Sports Exerc., Vol. 37, No. 5, pp. 731-736, 2005. Purpose: We hypothesized that creatine supplementation would facilitate muscle anabolism by increasing the expression of growth factors and the phosphorylation of anabolic signaling molecules; we therefore tested the responses of mRNA for IGF-I and IGF-II and the phosphorylation state of components of anabolic signaling pathways p70,6k and 4E-BPI to a bout of high-intensity resistance exercise after 5 d of creatine supplementation. Methods: In a double-blind cross-over design, muscle biopsies were taken from the m. vastus lateralis at rest and 3 and 24 h postexercise in subjects who had taken creatine or placebo for 5 d (21 g-d-'). For the first 3 h postexercise, the subjects were fed with a drink containing maltodextrin (0.3 g.kg-' body weight-h-') and protein (0.08 g.kg-2 body weight-h-'). Results: After creatine supplementation, resting muscle expressed more mRNA for IGF-I (+30%, P < 0.05) and IGF-II (+40%, P = 0.054). Exercise caused an increase by 3 h postexercise in IGF-I (+24%, P < 0.05) and IGF-11 (+48%, P < 0.05) and by 24 h postexercise in IGF-I (+29%, P < 0.05). but this effect was not potentiated by creatine supplementation. The phosphorylation states of p70,6k and 4E-BPI were not affected by creatine at rest; phosphorylation of both increased (150-400%, P < 0.05) to similar levels under placebo and creatine conditions at 3 h postexercise plus feeding. However, the phosphorylation state of 4E-BPI was higher in the creatine versus placebo condition at 24 h postexercise. Conclusion: The increase in lean body mass often reported after creatine supplementation could be mediated by signaling pathway(s) involving IGF and 4E-BPI.
Clinical Science, 2004
It has been speculated that creatine supplementation affects muscle glucose metabolism in humans by increasing muscle glycogen storage and up-regulating GLUT-4 protein expression. In the present study, we assessed the effects of creatine loading and prolonged supplementation on muscle glycogen storage and GLUT-4 mRNA and protein content in humans. A total of 20 subjects participated in a 6-week supplementation period during which creatine or a placebo was ingested. Muscle biopsies were taken before and after 5 days of creatine loading (20 g · day −1 ) and after 6 weeks of continued supplementation (2 g · day −1 ). Fasting plasma insulin concentrations, muscle creatine, glycogen and GLUT-4 protein content as well as GLUT-4, glycogen synthase-1 (GS-1) and glycogenin-1 (Gln-1) mRNA expression were determined. Creatine loading significantly increased total creatine, free creatine and creatine phosphate content with a concomitant 18 + − 5 % increase in muscle glycogen content (P < 0.05). The subsequent use of a 2 g · day −1 maintenance dose for 37 days did not maintain total creatine, creatine phosphate and glycogen content at the elevated levels. The initial increase in muscle glycogen accumulation could not be explained by an increase in fasting plasma insulin concentration, muscle GLUT-4 mRNA and/or protein content. In addition, neither muscle GS-1 nor Gln-1 mRNA expression was affected. We conclude that creatine ingestion itself stimulates muscle glycogen storage, but does not affect muscle GLUT-4 expression.
2003
No effect of creatine supplementation on human myofibrillar and sarcoplasmic protein synthesis after resistance exercise. Muscle hypertrophy during resistance training is reportedly increased by creatine supplementation. Having previously failed to find an anabolic effect on muscle protein turnover at rest, either fed or fasted, we have now examined the possibility of a stimulatory effect of creatine in conjunction with acute resistance exercise. Seven healthy men (body mass index, 23 Ϯ 2 kg/m 2 , 21 Ϯ 1 yr, means Ϯ SE) performed 20 ϫ 10 repetitions of leg extension-flexion at 75% one-repetition maximum in one leg, on two occasions, 4 wk apart, before and after ingesting 21 g/day creatine for 5 days. The subjects ate ϳ21 g maltodextrin ϩ 6 g protein/h for 3 h postexercise. We measured incorporation of [1-13 C]leucine into quadriceps muscle proteins in the rested and exercised legs. Leg protein breakdown (as dilution of [ 2 H5]phenylalanine) was also assessed in the exercised and rested leg postexercise. Creatine supplementation increased muscle total creatine by ϳ21% (P Ͻ 0.01). Exercise increased the synthetic rates of myofibrillar and sarcoplasmic proteins by two-to threefold (P Ͻ 0.05), and leg phenylalanine balance became more positive, but creatine was without any anabolic effect. skeletal muscle; protein turnover Address for reprint requests and other correspondence: M. J. Rennie,
Short-term creatine supplementation changes protein metabolism signaling in hindlimb suspension
BRAZILIAN JOURNAL OF MEDICAL AND BIOLOGICAL RESEARCH, 2019
Abstract The effect of a short-term creatine supplementation on hindlimb suspension (HS)-induced muscle atrophy was investigated. Creatine monohydrate (5 g/kg b.w. per day) or placebo, divided in 2 daily doses, was given by oral gavage for 5 days. Rats were maintained in HS with dietary supplementation concomitantly for 5 days. Body weight, soleus and EDL muscle masses, and cross-sectional areas (CSA) of the muscle fibers were measured. Signaling pathways associated with skeletal muscle mass regulation (FST, MSTN, FAK, IGF-1, MGF, Akt, mTOR, atrogin-1, and MuRF1 expressions, and Akt, S6, GSK3B, and 4EBP1 proteins) were evaluated in the muscles. Soleus muscle exhibited more atrophy than the EDL muscle due to HS. Creatine supplementation attenuated the decrease of wet weight and increased p-4EBP1 protein in the EDL muscle of HS rats. Also, creatine increased mTOR and atrogin-1 expressions in the same muscle and condition. In the absence of HS, creatine sup-plementation increased FAK and decreased MGF expressions in the EDL muscle. Creatine attenuated the increase in FST expression due to HS in the soleus muscle. MuRF1 expression increased in the soleus muscle due to creatine supplementation in HS animals whereas atrogin-1 expression increased still further in this group compared with untreated HS rats. In conclusion, short-term creatine supplementation changed protein metabolism signaling in soleus and EDL muscles. However, creatine supplementation only slightly attenuated the mass loss of both muscles and did not prevent the CSA reduction and muscle strength decrease induced by HS for 5 days.
Creatine metabolism and the consequences of creatine depletion in muscle
Mol Cell Biochem, 1994
Currently, considerable research activities are focussing on biochemical, physiological and pathological aspects of the creatine kinase (CK) -phosphorylcreatine (PCr) -creatine (Cr) system (for reviews see ), but only little effort is directed towards a thorough investigation of Cr metabolism as a whole. However, a detailed knowledge of Cr metabolism is essential for a deeper understanding of bioenergetics in general and, for example, of the effects of muscular dystrophies, atrophies, CK deficiencies (e.g. in transgenic animals) or Cr analogues on the energy metabolism of the tissues involved. Therefore, the present article provides a short overview on the reactions and enzymes involved in Cr biosynthesis and degradation, on the organization and regulation of Cr metabolism within the body, as well as on the metabolic consequences of 3-guanidinopropionate (GPA) feeding which is known to induce a Cr deficiency in muscle. In addition, the phenotype of muscles depleted of Cr and PCr by GPA feeding is put into context with recent investigations on the muscle phenotype of 'gene knockout' mice deficient in the cytosolic muscle-type M-CK. (Mol Cell Biochem 133/134: 51-66, 1994).