Exercise Dose Equalization in High-Intensity Interval Training: A Scoping Review.

Based on comparisons to moderate continuous exercise (MICT), high-intensity interval training (HIIT) is becoming a worldwide trend in physical exercise. This raises methodological questions related to equalization of exercise dose when comparing protocols. The present scoping review aims to identify in the literature the evidence for protocol equalization and the soundness of methods used for it. PubMed and Scopus databases were searched for original investigations comparing the effects of HIIT to MICT. A total of 2041 articles were identified, and 169 were included. Of these, 98 articles equalized protocols by utilizing energy-based methods or exercise volume (58 and 31 articles, respectively). No clear consensus for protocol equalization appears to have evolved over recent years. Prominent equalization methods consider the exercise dose (i.e., energy expenditure/production or total volume) in absolute values without considering the nonlinear nature of its relationship with duration. Exercises resulting from these methods induced maximal exertion in HIIT but low exertion in MICT. A key question is, therefore, whether exercise doses are best considered in absolute terms or relative to individual exercise maximums. If protocol equalization is accepted as an essential methodological prerequisite, it is hypothesized that comparison of program effects would be more accurate if exercise was quantified relative to intensity-related maximums.

Acceleration-based training: A new mode of training in senescent rats improving performance and left ventricular and muscle functions.

High intensity training (HIT) has been shown to improve maximal aerobic capacity and muscle protein synthesis but has not yet been investigated in senescent rats. We hypothesized that the change of speed (acceleration) during each bout of HIT acts as a stimulus responsible for the adaptations of the organism to exercise. Twenty two month-old (mo) rats (n=13) were subjected to a short acceleration protocol (20-30min) of exercise, comprising 3 independent bouts of acceleration and compared to an age-matched sedentary group (n=14). The protocol was repeated twice a week for two months. Following the protocol, performance, cardiac function, muscle mechanics, and the cellular and molecular pathways that are implicated in exercise adaptations were investigated. This new training, comprising only 16 sessions, improved maximal oxygen uptake (⩒O2peak; +6.6%, p<0.05), running distance (+95.2%; p<0.001), speed (+29.7%; p<0.01) and muscle function of 24mo rats in only 8weeks. This new training protocol induced cardiac hypertrophy and improved fractional shortening (47.3% vs. 41.1% in the control group, p<0.01) and ejection fraction. Moreover, it also improved the mechanics of skeletal muscle by increasing developed force (+31% vs. the control group, p<0.05) and maximal mechanical efficiency, activated the IGF1/mTOR/Akt pathway, and reduced the Smad2/3 pathway. Our results clearly show that the change in speed is a stimulus to control cardiac and skeletal muscle mass. This acceleration-based training is not time-consuming and may be adaptable for athletes, the elderly or chronic disease patients in order to improve strength, oxidative capacity, and quality of life.

Case Studies in Physiology: Maximal oxygen consumption and performance in a centenarian cyclist.

The purpose of this study was to examine the physiological characteristics of an elite centenarian cyclist who, at 101 yr old, established the 1-h cycling record for individuals ≥100 yr old (24.25 km) and to determine the physiological factors associated with his performance improvement 2 yr later at 103 yr old (26.92 km; +11%). Before each record, he performed an incremental test on a cycling ergometer. For 2 yr, he trained 5,000 km/yr with a polarized training that involved cycling 80% of mileage at "light" rate of perceived exertion (RPE) ≤12 and 20% at "hard" RPE ≥15 at a cadence between 50 and 70 rpm. His body weight and lean body mass did not change, while his maximal oxygen consumption (V̇o2max) increased (31-35 ml·kg-1·min-1; +13%). Peak power output increased from 90 to 125 W (+39%), mainly because of increasing the maximal pedaling frequency (69-90 rpm; +30%). Maximal heart rate did not change (134-137 beats/min) in contrast to the maximal ventilation (57-70 l/min, +23%), increasing with both the respiratory frequency (38-41 cycles/min; +8%) and the tidal volume (1.5-1.7 liters; +13%). Respiratory exchange ratio increased (1.03-1.14) to the same extent as tolerance to V̇co2 In conclusion, it is possible to increase performance and V̇o2max with polarized training focusing on a high pedaling cadence even after turning 100 yr old.NEW & NOTEWORTHY This study shows, for the first time, that maximal oxygen consumption (+13%) and performance (+11%) can still be increased between 101 and 103 yr old with 2 yr of training and that a centenarian is able, at 103 yr old, to cover 26.9 km/h in 1 h.

Erythropoietin and the use of a transgenic model of erythropoietin-deficient mice.

Despite its well-known role in red blood cell production, it is now accepted that erythropoietin (Epo) has other physiological functions. Epo and its receptors are expressed in many tissues, such as the brain and heart. The presence of Epo/Epo receptors in these organs suggests other roles than those usually assigned to this protein. Thus, the aim of this review is to describe the effects of Epo deficiency on adaptation to normoxic and hypoxic environments and to suggest a key role of Epo on main physiological adaptive functions. Our original model of Epo-deficient (Epo-TAgh) mice allowed us to improve our knowledge of the possible role of Epo in O2 homeostasis. The use of anemic transgenic mice revealed Epo as a crucial component of adaptation to hypoxia. Epo-TAgh mice survive well in hypoxic conditions despite low hematocrit. Furthermore, Epo plays a key role in neural control of ventilatory acclimatization and response to hypoxia, in deformability of red blood cells, in cerebral and cardiac angiogenesis, and in neuro- and cardioprotection.

Regulation of myogenesis by environmental hypoxia.

In aerobic organisms, oxygen is a critical factor for tissue and organ morphogenesis from embryonic development throughout the adult life. It regulates various intracellular pathways involved in cellular metabolism, proliferation, cell survival and fate. Organisms or tissues rapidly respond to changes in oxygen availability by activating complex signalling networks, which culminate in the control of mRNA translation and/or gene expression. This Commentary presents the effects of hypoxia during embryonic development, myoblasts and satellite cell proliferation and differentiation in vertebrates. We also outline the relationship between Notch, Wnt and growth factor signalling pathways, as well as the post-transcriptional regulation of myogenesis under conditions of hypoxia.

Oxygen modulates the glutathione peroxidase activity during the L6 myoblast early differentiation process.

AIM: This work aims to study the regulation of the glutathione peroxidase and catalase activities in myoblasts from the L6 line exposed to 21%, 5% and 1% O2 during the cell differentiation. MATERIAL AND METHODS: Rat L6 myoblasts were grown in 1%, 5% or 21% O2 in the presence or absence of N-acetyl cysteine. The cell proliferation was evaluated by determining the doubling time and kinetics of cultures by counting cells. The cell differentiation was analyzed by determining the myogenic fusion index using antibodies against the myosin heavy chain. The glutathione peroxidase and catalase activities were assayed. The p110-PI3K/Thr308-Akt pathway was studied using western blotting. The oxidative status of the cells was carried out by determining TBARS. RESULTS: 5% O2 improves the glutathione peroxidase activity, p110-PI3K/Thr308-Akt pathway and differentiation while 1% O2 alters all these parameters compared to 21% O2. NAC (0.5 mM) can prevent the deleterious effects of hypoxia (1% O2) on the L6 myoblast proliferation and enhances the myoblast differentiation when exposed to 21% O2. TBARS are reduced in 5% O2 compared to both 21% and 1% O2. CONCLUSION: The glutathione peroxidase activity and p110-PI3K/Thr308-Akt are both modulated in the same way by oxygen.

Epo is relevant neither for microvascular formation nor for the new formation and maintenance of mice skeletal muscle fibres in both normoxia and hypoxia.

Erythropoietin (Epo) and vascular growth factor (VEGF) are known to be involved in the regulation of cellular activity when oxygen transport is reduced as in anaemia or hypoxic conditions. Because it has been suggested that Epo could play a role in skeletal muscle development, regeneration, and angiogenesis, we aimed to assess Epo deficiency in both normoxia and hypoxia by using an Epo-deficient transgenic mouse model (Epo-TAg(h)). Histoimmunology, ELISA and real time RT-PCR did not show any muscle fiber atrophy or accumulation of active HIF-1alpha but an improvement of microvessel network and an upregulation of VEGFR2 mRNA in Epo-deficient gastrocnemius compared with Wild-Type one. In hypoxia, both models exhibit an upregulation of VEGF120 and VEGFR2 mRNA but no accumulation of Epo protein. EpoR mRNA is not up-regulated in both Epo-deficient and hypoxic gastrocnemius. These results suggest that muscle deconditioning observed in patients suffering from renal failure is not due to Epo deficiency.

Skeletal muscle intrinsic functional properties are preserved in a model of erythropoietin deficient mice exposed to hypoxia.

Erythropoietin (Epo)-induced polycythemia is the main factor of adaptation to hypoxia. In this study, we analysed the effects of Epo deficiency on intrinsic functional properties of slow and fast twitch muscles in a model of erythropoietin deficient mice (Epo-TAg(h)) exposed to hypoxia. We hypothesised that Epo deficiency would be deleterious for skeletal muscle structure and phenotype, which could change its functional properties and alters the adaptive response to ambient hypoxia. Wild-type (WT) and Epo-TAg(h) mice were left in hypobaric chamber at 420 mm Hg pressure for 14 days. Soleus (SOL) and extensor digitorum longus (EDL) were analysed in vitro by mechanical measurements, immunohistological and biochemical analyses. The results were compared to those obtained in corresponding muscles of age-matched normoxic groups. Our data did not show any difference between the groups whatever the Epo deficiency and/or hypoxic conditions for twitch force, tetanic force, fatigue, typology and myosin heavy chain composition. Normoxic Epo-TAg(h) mice exhibit improved capillary-to-fibre ratio compared to WT mice in both SOL and EDL whereas no angiogenic effects of hypoxia or combined Epo-deficiency/hypoxia were observed. These results suggest that skeletal muscles possess a great capacity of adaptation to Epo deficiency. Then Epo deficiency is not a sufficient factor to modify intrinsic functional properties of skeletal muscles.

Cerebral adaptations to chronic anemia in a model of erythropoietin-deficient mice exposed to hypoxia.

Anemia and hypoxia in rats result in an increase in factors potentially involved in cerebral angiogenesis. Therefore, the aim of this study was to assess the effect of chronic anemia and/or chronic hypoxia on cerebral cellular responses and angiogenesis in wild-type and anemic transgenic mice. These studies were done in erythropoietin-deficient mice (Epo-TAg(h)) in normoxia and following acute (one day) and chronic (14 days, barometric pressure = 420 mmHg) hypoxia. In normoxia, Epo-TAg(h) mice showed an increase in transcript and protein levels of hypoxia-inducible factor 1alpha (HIF-1alpha), vascular endothelial growth factor (VEGF), erythropoietin receptors (EpoR), phospho-STAT-5/STAT-5 ratio, and neuronal neuronal nitric oxide synthase (nNOS) along with a higher cerebral capillary density. In wild-type (WT) mice, acute hypoxia increased all of the studied factors, while in chronic hypoxia, HIF-1alpha, EpoR, phospho-STAT-5/STAT-5 ratio, nNOS, and inducible NOS remained elevated, with an increase in capillary density. Surprisingly, in Epo-TAg(h) mice, chronic hypoxia did not further increase any factor except the nitric oxide metabolites, while HIF-1alpha, EpoR, and phospho-STAT-5/STAT-5 ratio were reduced. Normoxic Epo-TAg(h) mice developed cerebral angiogenesis through the HIF-1alpha/VEGF pathway. In acute hypoxia, WT mice up-regulated all of the studied factors, including cerebral NO. Polycythemia and angiogenesis occurred with acclimatization to chronic hypoxia only in WT mice. In Epo-TAg(h), the decrease in HIF-1alpha, VEGF proteins, and phospho-STAT-5 ratio in chronic hypoxia suggest that neuroprotective and angiogenesis pathways are altered.

Prevotella timonensis sp. nov., isolated from a human breast abscess.

Gram-negative anaerobic rods were isolated from a human breast abscess. Based on genotypic and phenotypic characteristics, the novel strain belonged to the genus Prevotella. Phylogenetic analysis based on 16S rRNA gene sequence comparisons showed that it was closely related to Prevotella buccalis (94 % 16S rRNA gene sequence similarity), Prevotella salivae (90 %) and Prevotella oris (89.1 %). The major cellular fatty acid was C(14 : 0) (19.5 %). The new isolate represents a novel species in the genus Prevotella, for which the name Prevotella timonensis sp. nov. is proposed. The type strain is strain 4401737(T) (=CIP 108522(T)=CCUG 50105(T)).

Expression of slow myosin heavy chain during muscle regeneration is not always dependent on muscle innervation and calcineurin phosphatase activity.

In the literature, there is an ambiguity as to the respective roles played by calcineurin phosphatase activity (CPA) and muscle innervation in the reestablishment of the slow-twitch muscle phenotype after muscle regeneration in different species. In this study, we wanted to determine the role of calcineurin and muscle innervation on the appearance and maintenance of the slow phenotype during mouse muscle regeneration. The pattern of myosin expression and CPA was analyzed in adult (n=15), regenerating (n=45) and denervated-regenerating (n=32) slow-twitch soleus and fast-twitch extensor digitorum longus (EDL) muscles. Moreover, in a second group of denervated-regenerating mice (n=9), the animals were treated with a calcineurin inhibitor. Regeneration was induced by injection of cardiotoxin and in the denervated-regenerating group, denervation was carried out by cutting the sciatic nerve before the administration of cardiotoxin. In innervated-regenerating soleus muscle, CPA increased continuously after 10 days postinjury and by 21 days, there was a 3.5-fold increase in CPA compared with adult basal level, whereas in innervated-regenerating EDL muscle, CPA remained unchanged. Moreover, our results show that in denervated-regenerating muscles, the MyHC profile was identical in spite of the functional differences inherent in these muscles. In long-term denervated-regenerating muscles, a slow muscle phenotype was reexpressed both in the presence or absence of calcineurin inhibitor. Our results show that although in innervated-regenerating mouse muscle, the appearance of a slow phenotype is correlated with a peak of CPA, in denervated-regenerating muscles, a slow phenotype is triggered and maintained in a calcineurin- and nerve-independent manner.

FGF6 regulates muscle differentiation through a calcineurin-dependent pathway in regenerating soleus of adult mice.

Important functions in myogenesis have been proposed for FGF6, a member of the fibroblast growth factor family accumulating almost exclusively in the myogenic lineage, but its precise role in vivo remains mostly unclear. Here, using FGF6 (-/-) mice and rescue experiments by injection of recombinant FGF6, we dissected the functional role of FGF6 during in vivo myogenesis. We found that the appearance of myotubes was accelerated during regeneration of the soleus of FGF6 (-/-) mice versus wild type mice. This accelerated differentiation was correlated with increased expression of differentiation markers such as CdkIs and calcineurin, as well as structural markers such as MHCI and slow TnI. We showed that an elevated transcript level for calcineurin Aalpha subunit correlated with a positive regulation of calcineurin A activity in regenerating soleus of the FGF6 (-/-) mice. Cyclin D1 and calcineurin were up- and down-regulated, respectively in a dose-dependent manner upon injection of rhFGF6 in regenerating soleus of the mutant mice. We showed an increase of the number of slow oxidative (type I) myofibers, whereas fast oxidative (type IIa) myofibers were decreased in number in regenerating soleus of FGF6 (-/-) mice versus that of wild type mice. In adult soleus, the number of type I myofibers was also higher in FGF6 (-/-) mice than in wild type mice. Taken together these results evidenced a specific phenotype for soleus of the FGF6 (-/-) mice and led us to propose a model accounting for a specific dose-dependent effect of FGF6 in muscle regeneration. At high doses, FGF6 stimulates the proliferation of the myogenic stem cells, whereas at lower doses it regulates both muscle differentiation and muscle phenotype via a calcineurin-signaling pathway.

IGF-II is up-regulated and myofibres are hypertrophied in regenerating soleus of mice lacking FGF6.

Important functions in myogenesis have been proposed for FGF6, a member of the fibroblast growth factor family accumulating almost exclusively in the myogenic lineage. However, the use of FGF6(-/-) mutant mice gave contradictory results and the role of FGF6 during myogenesis remains largely unclear. Using FGF6(-/-) mice, we first analysed the morphology of the regenerated soleus following cardiotoxin injection and showed hypertrophied myofibres in soleus of the mutant mice as compared to wild-type mice. Secondly, to examine the function of the IGF family in the hypertrophy process, we used semiquantitative and real-time RT-PCR assays and Western blots to monitor the expression of the insulin-like growth factors (IGF-I and IGF-II), their receptors [type I IGF receptor (IGF1R) and IGF-II receptor (IGF2R)], and of a binding protein IGFBP-5 in regenerating soleus muscles of FGF6(-/-) knockout mice vs. wild-type mice. In the mutant, both IGF-II and IGF2R, but not IGF-I and IGF1R, were strongly up-regulated, whereas IGFBP5 was down-regulated, strongly suggesting that, in the absence of FGF6, the mechanisms leading to myofibre hypertrophy were mediated specifically by an IGF-II/IGF2R signalling pathway distinct from the classic mechanism involving IGF-I and IGF1R previously described for skeletal muscle hypertrophy. The potential regulating role of IGFBP5 on IGF-II expression is also discussed. This report shows for the first time a specific role for FGF6 in the regulation of myofibre size during a process of in vivo myogenesis.

Injection of FGF6 accelerates regeneration of the soleus muscle in adult mice.

FGF6, a member of the fibroblast growth factor (FGF) family, accumulated almost exclusively in the myogenic lineage, supporting the finding that FGF6 could specifically regulate myogenesis. Using FGF6 (-/-) mutant mice, important functions in muscle regeneration have been proposed for FGF6 but remain largely controversial. Here, we examined the effect of a single injection of recombinant FGF6 (rhFGF6) on the regeneration of mouse soleus subjected to cardiotoxin injection, specifically looking for molecular and morphological phenotypes. The injection of rhFGF6 has two effects. First, there is an up-regulation of cyclin D1 mRNA, accounting for the regulating role of a high FGF6 concentration on proliferation, and second, differentiation markers such as CdkIs and MHC I and Tn I increase and cellular differentiation is accelerated. We also show a down-regulation of endogenous FGF6, acceleration of FGFR1 receptor expression and deceleration of the FGFR4 receptor expression, possibly accounting for biphasic effects of exogenous FGF6 on muscle regeneration.

Specific activation of the acetylcholine receptor subunit genes by MyoD family proteins.

Whether the myogenic regulatory factors (MRFs) of the MyoD family can discriminate among the muscle gene targets for the proper and reproducible formation of skeletal muscle is a recurrent question. We have previously shown that, in Xenopus laevis, myogenin specifically transactivated muscle structural genes in vivo. In the present study, we used the Xenopus model to examine the role of XMyoD, XMyf5, and XMRF4 for the transactivation of the (nicotinic acetylcholine receptor) nAChR genes in vivo. During early Xenopus development, the expression patterns of nAChR subunit genes proved to be correlated with the expression patterns of the MRFs. We show that XMyf5 specifically induced the expression of the delta-subunit gene in cap animal assays and in endoderm cells of Xenopus embryos but was unable to activate the expression of the gamma-subunit gene. In embryos, overexpression of a dominant-negative XMyf5 variant led to the repression of delta-but not gamma-subunit gene expression. Conversely, XMyoD and XMRF4 activated gamma-subunit gene expression but were unable to activate delta-subunit gene expression. Finally, all MRFs induced expression of the alpha-subunit gene. These findings strengthen the concept that one MRF can specifically control a subset of muscle genes that cannot be activated by the other MRFs.

Expression of MRF4 protein in adult and in regenerating muscles in Xenopus.

In Xenopus, previous studies showed that the transcripts of the myogenic regulatory factor (MRF) MRF4 accumulate during skeletal muscle differentiation, but nothing is known about the accumulation of XMRF4 protein during myogenesis. In this report, an affinity-purified polyclonal antibody against Xenopus MRF4 was developed and used to describe the pattern of expression of this myogenic factor in the adult and in regenerating muscles. From young forming myotubes, XMRF4 protein persistently accumulated in nuclei during the regeneration process and was strongly expressed in nuclei of adult muscles. No selective accumulation of XMRF4 protein was detectable at neuromuscular junctions, but XMRF4 immunoreactivity was observed in sole plate nuclei as well as in extrasynaptic myofiber nuclei. We also report that XMRF4 protein accumulated before the establishment of neuromuscular connections, showing that innervation is not necessary for the appearance of XMRF4 protein during muscle regeneration.

Expression and neural control of follistatin versus myostatin genes during regeneration of mouse soleus.

Follistatin and myostatin are two secreted proteins involved in the control of muscle mass during development. These two proteins have opposite effects on muscle growth, as documented by genetic models. The aims of this work were to analyze in mouse, by using in situ hybridization, the spatial and temporal expression patterns of follistatin and myostatin mRNAs during soleus regeneration after cardiotoxin injury, and to investigate the influence of innervation on the accumulation of these two transcripts. Follistatin transcripts could be detected in activated satellite cells as early as the first stages of regeneration and were transiently expressed in forming myotubes. In contrast, myostatin mRNAs accumulated persistently throughout the regeneration process as well as in adult control soleus. Denervation significantly affected both follistatin and myostatin transcript accumulation, but in opposite ways. Muscle denervation persistently reduced the levels of myostatin transcripts as early as the young myotube stage, whereas the levels of follistatin mRNA were strongly increased in the small myotubes in the late stages of regeneration. These results are discussed with regard to the potential functions of both follistatin, as a positive regulator of muscle differentiation, and myostatin, as a negative regulator of skeletal muscle growth. We suggest that the belated up-regulation of the follistatin mRNA level in the small myotubes of the regenerating soleus as well as the down-regulation of the myostatin transcript level after denervation contribute to the differentiation process in denervated regenerating muscle.

Two myogenin-related genes are differentially expressed in Xenopus laevis myogenesis and differ in their ability to transactivate muscle structural genes.

Among the myogenic regulatory factors, myogenin is a transcriptional activator situated at a crucial position for terminal differentiation in muscle development. It is unclear at present whether myogenin exhibits unique specificities to transactivate late muscular markers. During Xenopus development, the accumulation of myogenin mRNA is restricted to secondary myogenesis, at the onset of the appearance of adult isoforms of beta-tropomyosin and myosin heavy chain. To determine the role of myogenin in the isoform switch of these contractile proteins, we characterized and directly compared the functional properties of myogenin with other myogenic regulatory factors in Xenopus embryos. Two distinct cDNAs related to myogenin, XmyogU1 and XmyogU2, were differentially expressed during myogenesis and in adult tissues, in which they preferentially accumulated in oxidative myofibers. Animal cap assays in Xenopus embryos revealed that myogenin, but not the other myogenic regulatory factors, induced expression of embryonic/larval isoforms of the beta-tropomyosin and myosin heavy chain genes. Only XmyogU1 induced expression of the adult fast isoform of the myosin heavy chain gene. This is the first demonstration of a specific transactivation of one set of muscle structural genes by myogenin.