INTRODUCTION:The present investigation aimed at identifying differences in muscle structural composition, substrate selection, and performance capacity in highly trained endurance athletes as a consequence of consuming a high-fat or a low-fat diet. METHODS:Eleven duathletes ingested high-fat (53% fat; HF) or high-carbohydrate diets (17% fat; LF) for 5 wk in a randomized crossover design. RESULTS:In m. vastus lateralis, oxidative capacity estimated as volume of mitochondria per volume of muscle fiber (HF: 9.86 +/- 0.36 vs LF: 9.79 +/- 0.52%, mean +/- SE) was not different after the two diet periods. Intramyocellular lipid (IMCL) was significantly increased after HF compared with LF (1.54 +/- 0.27% vs 0.69 +/- 0.09%, P = 0.0076). Glycogen content was lower after HF than after LF, but this difference was not statistically significant (487.8 +/- 38.2 vs 534.4 +/- 32.6 mmol x kg-1 dry weight, P = 0.2454). Maximal power and [OV0312]O(2max) (63.6 +/- 0.9 vs 63.9 +/- 1.2 mL O(2) x min-1 x kg-1 on HF and LF) during an incremental exercise test to exhaustion were not different between the two diet periods. Total work output during a 20-min all-out time trial (298 +/- 6 vs 297 +/- 7 W) on a bicycle ergometer as well as half-marathon running time (80 min 12 s +/- 86 s vs 80 min 24 s +/- 82 s) were not different between HF and LF. Blood lactate concentrations and respiratory exchange ratios (RER) were significantly lower after HF than after LF at rest and during all submaximal exercise loads. CONCLUSIONS:Muscle glycogen stores were maintained after a 5-wk high-fat diet period whereas IMCL content was more than doubled. Endurance performance capacity was maintained at moderate to high-exercise intensities with a significantly larger contribution of lipids to total energy turnover.

The effects of short-term exercise training on vascular endothelial growth factor (VEGF) and one of its regulatory transcription factors, the hypoxia inducible factor 1 (HIF-1) subunit, were studied in eight healthy males. Muscle and blood samples were obtained before the 1st, and 24 h after the 7th training session. VEGF and HIF-1 mRNA were analysed using RT-PCR, VEGF mRNA localization with in situ hybridization and VEGF protein with ELISA. Concurrent increases in VEGF mRNA and protein levels were observed in skeletal muscle, and the mRNA was expressed within the skeletal muscle fibres and in cells in the interstitium. These data support the idea of a pretranslational regulation of exercise-induced changes in VEGF mRNA, and indicate that increased VEGF protein expression is an early event in skeletal muscle adaptation to training. Furthermore, different cell types may act as sources for the production of angiogenic factors in response to exercise. The levels of HIF-1 mRNA subunits did not change, suggesting no change in HIF-1 mRNA transcript levels in the regulation of training-induced VEGF expression. In contrast to increased tissue VEGF expression, the arterial and femoral venous plasma levels of VEGF were decreased by training, which may indicate an exercise-induced enhancement of the peripheral uptake of VEGF.

This study was performed to explore changes in gene expression as a consequence of exercise training at two levels of intensity under normoxic and normobaric hypoxic conditions (corresponding to an altitude of 3,850 m). Four groups of human subjects trained five times a week for a total of 6 wk on a bicycle ergometer. Muscle biopsies were taken, and performance tests were carried out before and after the training period. Similar increases in maximal O(2) uptake (8.3-13.1%) and maximal power output (11.4-20.8%) were found in all groups. RT-PCR revealed elevated mRNA concentrations of the alpha-subunit of hypoxia-inducible factor 1 (HIF-1) after both high- (+82.4%) and low (+78.4%)-intensity training under hypoxic conditions. The mRNA of HIF-1alpha(736), a splice variant of HIF-1alpha newly detected in human skeletal muscle, was shown to be changed in a similar pattern as HIF-1alpha. Increased mRNA contents of myoglobin (+72.2%) and vascular endothelial growth factor (+52.4%) were evoked only after high-intensity training in hypoxia. Augmented mRNA levels of oxidative enzymes, phosphofructokinase, and heat shock protein 70 were found after high-intensity training under both hypoxic and normoxic conditions. Our findings suggest that HIF-1 is specifically involved in the regulation of muscle adaptations after hypoxia training. Fine-tuning of the training response is recognized at the molecular level, and with less sensitivity also at the structural level, but not at global functional responses like maximal O(2) uptake or maximal power output.

Acta Physiologica ScandinavicaVolume 165, Issue 3 p. 335-336 Related expression of vascular endothelial growth factor and hypoxia-inducible factor-1 mRNAs in human skeletal muscle GUSTAFSSON, GUSTAFSSON Section of Environmental Physiology, Department of Physiology & Pharmacology, Karolinska Institutet, Sweden Section of Clinical Physiology, Department of Laboratory Sciences and Technology, Huddinge sjukhus, Karolinska Institutet, SwedenSearch for more papers by this author PUNTSCHART, PUNTSCHART Department of Anatomy, University of Bern, SwitzerlandSearch for more papers by this author SUNDBERG, SUNDBERG Section of Environmental Physiology, Department of Physiology & Pharmacology, Karolinska Institutet, SwedenSearch for more papers by this author JANSSON, JANSSON Section of Clinical Physiology, Department of Laboratory Sciences and Technology, Huddinge sjukhus, Karolinska Institutet, SwedenSearch for more papers by this author GUSTAFSSON, GUSTAFSSON Section of Environmental Physiology, Department of Physiology & Pharmacology, Karolinska Institutet, Sweden Section of Clinical Physiology, Department of Laboratory Sciences and Technology, Huddinge sjukhus, Karolinska Institutet, SwedenSearch for more papers by this author PUNTSCHART, PUNTSCHART Department of Anatomy, University of Bern, SwitzerlandSearch for more papers by this author SUNDBERG, SUNDBERG Section of Environmental Physiology, Department of Physiology & Pharmacology, Karolinska Institutet, SwedenSearch for more papers by this author JANSSON, JANSSON Section of Clinical Physiology, Department of Laboratory Sciences and Technology, Huddinge sjukhus, Karolinska Institutet, SwedenSearch for more papers by this author First published: 24 December 2001 https://doi.org/10.1046/j.1365-201x.1999.00515.xCitations: 6 ThomasGustafsson Section of Environmental Physiology, Department of Physiology and Pharmacology, Karolinska Institutet, S-171 77 Stockholm, Sweden. Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article.Citing Literature Volume165, Issue3March 1999Pages 335-336 RelatedInformation

When studying the effect of various treatments on gene expression in humans, one occasionally is faced with the problem of detecting small changes in transcript levels in minute tissue samples. In addition, interindividual variations can be quite large and may even be the major source of variation (1). Therefore, numerous samples usually have to be analyzed to detect such small variations in gene expression.

It is believed that the induction of the fos and jun gene family of transcription factors might be at the origin of genetic events leading to the differential regulation of muscle-specific genes. We have investigated the effect of a 30-min running bout in untrained subjects on the expression of the mRNAs of all members of the fos and jun gene families, including c- fos, fosB, fosBdel, fra-1, and fra-2 as well as c- jun, junB, and junD. While the fos family members were transiently upregulated 10- to 20-fold (an exception being fra-2), the induction of the jun family members was up to 3-fold only. The induction of c- fos could also be demonstrated at the protein level. Both c- fos and c- jun mRNAs were coinduced in muscle fiber nuclei. The induction was not restricted to a particular fiber type, as expected from established muscle fiber recruitment schemes, but followed a “patchy” pattern confined to certain regions of the muscle. The signals leading to the expression of these immediate early genes are therefore unclear.

The fibre-type specific expression patterns of fast and slow isoforms of essential (alkali) myosin light chains (ELC) was analysed in trained, untrained and pathological human muscles. Biopsies from m. vastus lateralis of moderately trained and untrained persons, as well as highly trained endurance and strength athletes were analysed, by in situ hybridization, for the expression of the 'fast' ELC 1f/3f and the 'slow' ELC 1 sb. We wanted to investigate if changes in the fibre-type specific ELC mRNA pattern could be used as markers for training adaptation, especially, if the mRNA of the slow ELC 1sb isoform would appear in type IIA fibres as a result of endurance training (Baumann et al. 1987). We found the fast/slow ELC expression patterns in the fibre types to be remarkably stable. Physiological stress, even high training loads, did not affect it. No IIA fibres expressing ELC 1sb mRNA were found. They could be detected, however, in pathological muscle samples, where fast/slow ELC patterns not found in normal muscles were frequent. Our data suggest that in healthy muscles, only a subset of the theoretically possible combinations of myosin heavy and light chain isoforms are expressed at the level of their mRNAs.

We studied the expression patterns of the essential (alkali) myosin light-chain isoforms in adult human skeletal muscles, using in situ hybridization and single-fiber protein analysis. In analogy to other species, we found that the fiber type-specific expression of essential myosin light chains is regulated via the availability of the respective mRNAs in a given fiber. In contrast to other species, the slow isoform 1sa was only expressed in the most oxidative Type I fibers (Subtype IA) in addition to 1sb. These fibers also contained high levels of carbonic anhydrase III. Within the fibers, the essential myosin light-chain mRNAs were located preferentially in the perinuclear regions and to a lesser extent in the intermyofibrillar spaces, a distribution that excludes cotranslational assembly of these light chains into the myofibrils as the main mechanism. In comparing leg and shoulder muscles, we found less distinct fiber typing in the expression patterns of the essential myosin light chains in the leg muscles than in muscles from the shoulder region.

Prolonged exercise of a sufficiently high intensity is thought to create physiological stress and to disturb cellular homeostasis, ultimately inducing cellular adaptations which enable the organism to better deal with any future exercise challenge. Heat shock proteins (hsp) are expressed when cells are exposed to different types of stress. In this study, we have investigated whether the expression of the heat inducible form of hsp70 is increased in human skeletal muscle cells after a single bout of exercise. Five untrained subjects performed an exercise bout at their individual anaerobic threshold for 30 min on a treadmill. Hsp70 mRNA concentration was significantly increased by a factor of four at 4 min post-exercise. Similarly high levels were also observed 30 min and 3 h after the end of exercise. Hsp70 protein concentration, on the contrary, did not change within 3 h after cessation of exercise. Thus, a single exercise bout in humans is able to increase the steady state concentration of hsp70 mRNA, but is probably not sufficient to have an effect on the already high basal level of its protein. The analysis of hsp70 mRNA is potentially useful as a method to detect stress in tissues with a high basal level of heat shock proteins.

Improvements in endurance capacity by training are associated with structural and biochemical adaptations of working muscles that affect the mitochondrial compartment. We investigated whether the 1.8-fold higher mitochondrial volume density in a group of endurance-trained athletes compared with untrained subjects was reflected by higher steady-state levels of mRNAs coding for components of the oxidative phosphorylation pathway using a quantitative polymerase chain reaction approach. We found that mitochondrially encoded RNAs (cytochrome-c oxidase subunit I, NADH reductase subunit 6, 16S rRNA), as well as nuclear-encoded RNAs (cytochrome-c oxidase subunit IV, succinate dehydrogenase, fumarase) are all increased coordinately in the athletes (1.54- to 1.94-fold). In addition, mitochondrial (mt) DNA concentration was also 1.55-fold higher in the trained athletes, whereas genomic DNA was not changed. Our findings thus show similar RNA expression of mitochondrially encoded genes in sedentary and endurance-trained subjects, whereas pretranslational control mechanisms account for higher levels of nuclear-encoded RNAs in the athletes.

We describe a PCR quantitation approach that we have set up to study gene expression in human skeletal muscle biopsies. It is characterized by the independent standardization of the individual steps and ignores attempts to control for the efficiency of the PCR. Different RNA extraction/reverse transcription efficiencies are normalized by the addition of an unrelated cRNA. Quantitation is achieved by parallel amplifications of reference samples containing known amounts of PCR products. Precision was achieved by multiple measurements of several samples. Our approach allows for the detection of less than twofold differences (27-88%, depending on the RNA species studied) among samples. A comparison of biopsies from highly trained endurance runners with biopsies from untrained subjects showed that the increased mitochondrial density in the runners' samples is accompanied by a proportional increase in the concentration of the mRNA of cytochrome c oxidase subunit IV.

33P-labelled probes were used to localize the mRNAs coding for the myosin alkali light-chain isoforms MLC 1f/3f and MLC 1sb in adult human muscles, which are distributed in characteristic fibre type specific patterns. In situ hybridizations of 33P-labelled probes were compared with probes carrying 35S or digoxigenin labels. Signals of equal strength were obtained with each of the three labels. The preferentially peripheral localization of these mRNAs in the muscle fibres could be clearly seen with all three probes, with digoxigenin probes providing the best resolution. 33P can serve as a viable alternative in this type of experiment.