Fast-twitch Muscle Research Articles

Lack of cyclin D3 induces skeletal muscle fiber-type shifting, increased endurance performance and hypermetabolism

The mitogen-induced D-type cyclins (D1, D2 and D3) are regulatory subunits of the cyclin-dependent kinases CDK4 and CDK6 that drive progression through the G1 phase of the cell cycle. In skeletal muscle, cyclin D3 plays a unique function in controlling the proliferation/differentiation balance of myogenic progenitor cells. Here, we show that cyclin D3 also performs a novel function, regulating muscle fiber type-specific gene expression. Mice lacking cyclin D3 display an increased number of myofibers with higher oxidative capacity in fast-twitch muscle groups, primarily composed of myofibers that utilize glycolytic metabolism. The remodeling of myofibers toward a slower, more oxidative phenotype is accompanied by enhanced running endurance and increased energy expenditure and fatty acid oxidation. In addition, gene expression profiling of cyclin D3−/− muscle reveals the upregulation of genes encoding proteins involved in the regulation of contractile function and metabolic markers specifically expressed in slow-twitch and fast-oxidative myofibers, many of which are targets of MEF2 and/or NFAT transcription factors. Furthermore, cyclin D3 can repress the calcineurin- or MEF2-dependent activation of a slow fiber-specific promoter in cultured muscle cells. These data suggest that cyclin D3 regulates muscle fiber type phenotype, and consequently whole body metabolism, by antagonizing the activity of MEF2 and/or NFAT.

Muscle type-specific RNA polymerase II recruitment during PGC-1α gene transcription after acute exercise in adult rats.

Epigenetic regulation of gene expression differs between fast- and slow-twitch skeletal muscles in adult rats, although the precise mechanisms are still unknown. The present study investigates the differences in responses of RNA polymerase II (Pol II) and histone acetylation during transcriptional activation in the plantaris and soleus muscles of adult rats after acute treadmill running. We targeted the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) gene to analyze epigenomic changes by chromatin immunoprecipitation. The mRNA expression of the PGC-1α-b isoform was significantly upregulated in both plantaris and soleus muscles 2 h after acute running, although the magnitude of the upregulation was more pronounced in the plantaris muscle. The sequences of proximal exons of the PGC-1α locus were expressed more in the plantaris muscle after acute running. Accumulation of Pol II was noted near the alternative exon 1 in both plantaris and soleus muscles in association with the enhanced distribution of acetylated histone 3. Accumulation of Pol II was also observed at the transcription start site, exon 2, and exon 3 in the plantaris muscle, but not the soleus muscle. It was noted that in the soleus muscle, acetylation of histone 3 at lysine 27 was enhanced throughout the PGC-1α locus in response to transcriptional activation, suggesting that elongating Pol II was capable of traveling through to the end of the locus. These results indicate that the mobility of Pol II during PGC-1α transcription differed between fast- and slow-twitch skeletal muscles, affecting the strength of the transcriptional activity.NEW & NOTEWORTHY Fast- and slow-twitch skeletal muscles have distinct characteristics in both force production and metabolism. Epigenetic regulations also largely differ in these muscles. Here we show that RNA polymerase II is distributed extensively at the proximal regions downstream of transcription start site during the transcriptional activation of PGC-1α in fast-twitch muscles, but it accumulates at the first exon in slow-twitch muscles. These findings will provide a basis to understand type-specific mechanisms in skeletal muscle.

Integrated analysis of mRNA and miRNA expression profiles reveals muscle growth differences between adult female and male Chinese concave-eared frogs (Odorrana tormota)

The Chinese concave-eared torrent frog (Odorrana tormota) is the first known non-mammalian vertebrate that can communicate using ultrasound. In this species, females are approximately four times as large as males, in which the female growth rate is obviously higher than that of male. Until now, the molecular mechanisms underlying muscle growth development differences between male and female frogs have not been reported. Here, we integrated mRNA and miRNA expression profiles to reveal growth differences in the hindlimb muscles of 2-year-old frogs. Among 569 differentially expressed genes (DEGs), 69 were associated with muscle growth and regeneration. Fifty-one up-regulated genes in females were potentially involved in promoting muscle growth and regeneration, whereas 18 up-regulated genes in males may lead to muscle growth inhibition and fast-twitch muscle fiber contraction. 244 DEGs were enriched in mTOR and other protein synthesis signaling pathways, and protein degradation pathways, including lysosomal protease, calpain, caspase, and ubiquitin–proteasome system pathways. It may interpret why female muscles grow faster than males. Based on expression differences of genes involved in glycolysis and oxidative metabolism, we speculated that the proportion of slow muscle fiber was higher and that of fast muscle fiber was lower in female compared with male muscle. Additionally, 767 miRNAs were identified, including 217 new miRNAs, and 6248 miRNA-negatively regulated mRNAs were predicted. The miRNA target genes were enriched in pathways related to muscle growth, protein synthesis, and degradation. Thus, in addition to the identified mRNA differential expressions, miRNAs may play other important roles in the differential regulation of hindlimb muscle growth between female and male O. tormota.

Ingestion of soy protein isolate attenuates eccentric contraction-induced force depression and muscle proteolysis via inhibition of calpain-1 activation in rat fast-twitch skeletal muscle.

Eccentric contraction (ECC) is a contraction in which skeletal muscles are stretched while contracting. The aim of this study was to determine how ingestion of soy protein isolate (SPI) or animal-based proteins affect force deficit, calpain activation, and proteolysis of calcium ion (Ca2+)-regulatory proteins in rat fast-twitch muscles subjected to ECC. In the first experiment, male Wistar rats were randomly assigned to a control and an SPI group, which were fed a 20% casein and a 20% SPI diet, respectively, for 28 d before the ECC protocol. Anterior crural muscles underwent 200 repeated ECCs and were excised 3 d later. In the second experiment, half of the SPI rats were given water containing NG-nitro-l-arginine-methyl ester (L-NAME), an inhibitor of nitric oxide synthase, for 3 d of recovery after ECC. SPI ingestion attenuated ECC-induced force deficit, proteolysis of Ca2+-regulatory proteins, and autolysis of calpain-1. Co-ingestion of L-NAME inhibited SPI-associated increases in nitrite and nitrate levels and negated the force recovery effects of SPI. These results suggest that SPI ingestion inhibits ECC-elicited force deficit and proteolysis of Ca2+ regulatory proteins, which is caused by inhibited activation of calpain-1 via increased nitric oxide production.

Effects of streptozotocin-induced diabetes on leg muscle contractile properties and motor neuron morphology in rats.

Skeletal muscle fiber subtypes are differentially sensitive to diabetes-related pathology; For example, fast-twitch muscles exhibit severe decreases in contraction force while slow-twitch muscles demonstrate prolonged half-relaxation time. However, such alterations have only been examined after a relatively short period following diabetes onset, with no information available regarding muscle damage caused by longer disease periods (>20weeks). This study examined alterations in the contractile properties of the medial gastrocnemius (fast-twitch) and soleus (slow-twitch) muscles, as well as morphological changes in their motor neurons 12 and 22weeks after diabetes onset. Adult male Wistar rats were divided into diabetic (12- or 22-week post-streptozotocin injection) and age-matched control groups. Electrically evoked maximum twitch and tetanic tension were recorded from leg muscles. Additionally, motor neuron number and cell body size were examined. At 12weeks after diabetes onset, decreases in twitch force were observed predominantly in medial gastrocnemius muscles, while soleus muscles exhibited prolonged half-relaxation time. However, these differences became ambiguous at 22weeks, with decreased twitch force and prolonged half-relaxation time observed in both muscles. On the other hand, reduction in soleus motor neurons was observed 12weeks after diabetes onset, while medial gastrocnemius motor neurons were diminished at 22weeks. These data indicate that experimental diabetes induces differential damage to medial gastrocnemius and soleus muscles as well as motor neurons. These diabetes-induced differences may partly underlie the differential deficits observed in gastrocnemius and soleus.

ANKK1 is found in myogenic precursors and muscle fibers subtypes with glycolytic metabolism.

Ankyrin repeat and kinase domain containing 1 (ANKK1) gene has been widely related to neuropsychiatry disorders. The localization of ANKK1 in neural progenitors and its correlation with the cell cycle has suggested its participation in development. However, ANKK1 functions still need to be identified. Here, we have further characterized the ANKK1 localization in vivo and in vitro, by using immunolabeling, quantitative real-time PCR and Western blot in the myogenic lineage. Histologic investigations in mice and humans revealed that ANKK1 is expressed in precursors of embryonic and adult muscles. In mice embryos, ANKK1 was found in migrating myotubes where it shows a polarized cytoplasmic distribution, while proliferative myoblasts and satellite cells show different isoforms in their nuclei and cytoplasm. In vitro studies of ANKK1 protein isoforms along the myogenic progression showed the decline of nuclear ANKK1-kinase until its total exclusion in myotubes. In adult mice, ANKK1 was expressed exclusively in the Fast-Twitch muscles fibers subtype. The induction of glycolytic metabolism in C2C12 cells with high glucose concentration or treatment with berberine caused a significant increase in the ANKK1 mRNA. Similarly, C2C12 cells under hypoxic conditions caused the increase of nuclear ANKK1. These results altogether show a relationship between ANKK1 gene regulation and the metabolism of muscles during development and in adulthood. Finally, we found ANKK1 expression in regenerative fibers of muscles from dystrophic patients. Future studies in ANKK1 biology and the pathological response of muscles will reveal whether this protein is a novel muscle disease biomarker.

Recovery Interstitial PO2 Dynamics Following Contractions in Healthy Skeletal Muscle of Different Oxidative Capacity

The recovery of skeletal muscle from repetitive tasks is dependent, in part, on adequate O2 delivery and blood-mitochondrial O2 diffusion. Key steps in this pathway include the microvascular and interstitial space (is) and novel use of specific phosphorescence probes allow measurements of O2 partial pressures (PO2) at those sites near the myocytes. Given the disparate contribution of nitric oxide (NO) between fiber types we sought to resolve how NO impacts PO2is in recovery following rhythmic muscle contractions. PURPOSE: To determine the contribution of NO bioavailability to the temporal profiles of PO2is off-kinetics in two fast-twitch muscles with different oxidative capacities (citrate synthase: peroneal (PER) ~20 vs white gastrocnemius (WG) ~8-11 μmol/min/g). We tested the hypothesis that the more oxidative PER would display faster kinetics while recovering to baseline PO2is levels compared to the WG. In addition, NO synthase inhibition via L-nitroarginine methyl ester (L-NAME) would slow recovery off- kinetics whereas this process would be speeded by increased NO. METHODS: PO2is was determined via phosphorescence quenching (Oxyphor G4) in the exposed rat PER and WG (n=5) during and following electrically stimulated muscle contractions (1 Hz, 8 V, both 3 min) under control (CON), sodium nitroprusside (SNP, NO donor) and L-NAME (n= 3 PER, 5 WG) conditions. RESULTS: PER PO2is was higher than WG for CON at baseline (18.1 ± 1.8 vs 11.3 ± 1.2 mmHg), end contractions (11.5 ± 1.2 vs 5.2 ± 0.9 mmHg), and following recovery (19.6 ± 2.1 vs 10.1 ± 0.9 mmHg; p<0.05 for all). SNP elevated PO2is at all time points in both muscles compared to CON and L-NAME (p<0.05). PO2is recovered to baseline levels in both muscles in CON and L-NAME (p>0.05) but not SNP (p<0.05). Off-kinetics were faster in the PER compared to WG in CON (35 ± 6 vs 76 ± 6 s; p<0.05). L-NAME did not alter PER off-kinetics but prolonged recovery in WG (101 ± 8 s; p<0.05). CONCLUSIONS: Consistent with our hypothesis, PO2is returned to baseline levels faster in the CON PER than WG. This likely reflects greater O2 delivery in PER. However, NO synthase inhibition via L-NAME did not diminish the magnitude nor rate of recovery in PER indicating that the interstitial-mitochondrial pressure head for O2 delivery may be preserved via other pathways in more oxidative muscles in health.

Glycogen Enhancement Augments Overload-Induced Protein Synthesis, Growth, and Myogenesis in Aged Skeletal Muscle.

Age-related skeletal muscle (SkM) wasting is associated with elevated 5’-AMP-Activated Protein Kinase (AMPK) activity, which inhibits overload-induced (OI) SkM protein synthesis (MPS) and growth. Glycogen, an inhibitor of AMPK, is reduced in aged SkM. PURPOSE: To examine the effects of manipulating glycogen on AMPK, MPS and related signaling, and OI-growth in aged SkM. METHODS: Mutant glycogen synthase (GS; designed to enhance SkM glycogen content [GC]) or empty-vector plasmids were electrotransferred into fast-twitch plantaris muscles prior to 21-day synergist ablation-induced unilateral overload in young adult (8 mo.; empty vector; YE, n=9) and old (33 mo.; empty vector, OE, n=11; or mutant GS, OM, n=13) male FBN rats. Contralateral limbs underwent SHAM ablations with no plasmid. RESULTS: As expected, mutant GS expression and GC were significantly higher in OM overloaded muscles (the only muscles receiving the mutant GS plasmid) vs SHAM OM muscles or vs both SHAM or overloaded YE and OE muscles. There were significant increases in OI-(all vs SHAM) MPS and hypertrophy in all groups and OM was greater than OE. Markers of AMPK activity and other signaling intermediates affecting MPS were largely unaltered by glycogen enhancement. However, there was a strong and significant effect of enhancing GC (via mutant GS vs empty vector plasmid) on myogenic regulatory factors MyoD and myogenin, embryonic myosin heavy chain-positive fibers, and total fiber number in aged muscle under conditions of overload. CONCLUSIONS: Thus, enhancing GC may lead to enhanced MPS and OI growth in aged SkM. This effect may be due, in part, to an enhanced myogenesis.

MicroRNA-139-5p suppresses myosin heavy chain I and IIa expression via inhibition of the calcineurin/NFAT signaling pathway

MicroRNAs (miRNAs) are a class of small non-coding RNAs that are widely involved in a variety of biological processes. Different skeletal muscle fiber type composition exhibits characteristic differences in functional properties and energy metabolism of skeletal muscle. However, the molecular mechanism by which miRNAs control the different type of muscle fiber formation is still not fully understood. In the present study, we characterized the role of microRNA-139-5p (miR-139-5p) in the regulation of myosin heavy chain (MyHC) isoform expression and its underlying mechanisms. Here we found that the expression of miR-139-5p was significantly higher in mouse slow-twitch muscle than in fast-twitch muscle. Overexpression of miR-139-5p downregulated the expression of MyHC I and MyHC IIa, whereas inhibition of miR-139-5p upregulated them. We also found that the levels of calcineurin (CaN), NFATc1, MEF2C and MCIP1.4, which are the components of CaN/NFAT signaling pathway that has shown to positively regulate slow fiber-selective gene expression, were notably inhibited by miR-139-5p overexpression. Furthermore, treatment of phenylephrine (PE), a α1-adrenoceptor agonist, abolished the inhibitory effect of miR-139-5p on MyHC I and MyHC IIa expression. Together, our findings indicated that the role of miR-139-5p in regulating the MyHC isoforms, especially MyHC I and MyHC IIa, may be achieved through inhibiting CaN/NFAT signaling pathway.

Kir4.1-Dependent Astrocyte-Fast Motor Neuron Interactions Are Required for Peak Strength

SummaryDiversified neurons are essential for sensorimotor function, but whether astrocytes become specialized to optimize circuit performance remains unclear. Large fast α-motor neurons (FαMNs) of spinal cord innervate fast-twitch muscles that generate peak strength. We report that ventral horn astrocytes express the inward-rectifying K+ channel Kir4.1 (a.k.a. Kcnj10) around MNs in a VGLUT1-dependent manner. Loss of astrocyte-encoded Kir4.1 selectively altered FαMN size and function and led to reduced peak strength. Overexpression of Kir4.1 in astrocytes was sufficient to increase MN size through activation of the PI3K/mTOR/pS6 pathway. Kir4.1 was downregulated cell autonomously in astrocytes derived from amyotrophic lateral sclerosis (ALS) patients with SOD1 mutation. However, astrocyte Kir4.1 was dispensable for FαMN survival even in the mutant SOD1 background. These findings show that astrocyte Kir4.1 is essential for maintenance of peak strength and suggest that Kir4.1 downregulation might uncouple symptoms of muscle weakness from MN cell death in diseases like ALS.

Identification of CCL5/RANTES as a novel contraction-reducible myokine in mouse skeletal muscle

Skeletal muscle is an endocrine organ that secretes several proteins, which are collectively termed myokines. Although many studies suggest that exercise regulates myokine secretion, the underlying mechanisms remain unclear and all the exercise-dependent myokines have not yet been identified. Therefore, in this study, we attempted to identify novel exercise-dependent myokines by using our recently developed in vitro contractile model. Differentiated C2C12 myotubes were cultured with or without electrical pulse stimulation (EPS) for 24 h to induce cell contraction, and the myokines secreted in conditioned medium were analyzed using a cytokine array. Although most myokine secretions were not affected by EPS, the secretion of Chemokine (C-C motif) ligand 5 (CCL5) (regulated on activation, normal T cell expressed and secreted (RANTES)) was significantly reduced by EPS. This was further confirmed by ELISA and quantitative PCR. Contraction-dependent calcium transients and activation of 5′-AMP activating protein kinase (AMPK) appears to be involved in this decrease, as the chelating Ca2+ by EGTA blocked contraction-dependent CCL5 reduction, whereas the pharmacological activation of AMPK significantly reduced it. However, Ccl5 gene expression was increased by AMPK activation, suggesting that AMPK-dependent CCL5 decrease occurred via post-transcriptional regulation. Finally, mouse experiments revealed that voluntary wheel-running exercise reduced serum CCL5 levels and Ccl5 gene expression in the fast-twitch muscles. Overall, our study provides the first evidence of an exercise-reducible myokine, CCL5, in the mouse skeletal muscle. Although further studies are required to understand the precise roles of the skeletal muscle cell contraction-induced decrease in CCL5, this decrease may explain some exercise-dependent physiological changes such as those in immune responses.

Branched fibers from old fast-twitch dystrophic muscles are the sites of terminal damage in muscular dystrophy.

A striking pathological feature of dystrophinopathies is the presence of morphologically abnormal branched skeletal muscle fibers. The deterioration of muscle contractile function in Duchenne muscular dystrophy is accompanied by both an increase in number and complexity of these branched fibers. We propose that when number and complexity of branched fibers reaches a critical threshold, or "tipping point," the branches in and of themselves are the site of contraction-induced rupture. In the present study, we use the dystrophic mdx mouse and littermate controls to study the prediseased dystrophic fast-twitch extensor digitorum longus (EDL) muscle at 2-3 wk, the peak myonecrotic phase at 6-9 wk, and finally, "old," at 58-112 wk. Using a combination of isolated muscle function contractile measurements coupled with single-fiber imaging and confocal microscope imaging of cleared whole muscles, we identified a distinct pathophysiology, acute fiber rupture at branch nodes, which occurs in "old" fast-twitch EDL muscle approaching the end stage of the dystrophinopathy muscle disease, where the EDL muscles are entirely composed of complexed branched fibers. This evidence supports our concept of "tipping point" where the number and extent of fiber branching reach a level where the branching itself terminally compromises muscle function, irrespective of the absence of dystrophin.

Sarcopenia.

Sarcopenia is avery common, but frequently overlooked and undertreated geriatric syndrome comprising pronounced muscle mass and strength/performance loss. Estimated prevalence is between 5 and 40% in the general population, accompanied by an exponential incline with increasing age. Sarcopenia is connected to atrophy and loss of muscle fibers and motor units, affecting primarily the fast-twitch muscle fibers und their motor units. Fast-twitch muscle fibers seem to be more prone to failure of function and loss over time. Main causes for the development of sarcopenia are hormonal changes (reduced release of testosterone, estrogen, and growth hormone), nutritional deficiencies, chronic inflammation, and particularly adecrease in physical activity due to sedentary lifestyle with advancing age. Treatment options for sarcopenia comprise an active lifestyle with physical activity and exercise training, modifications of nutritional intake, and pharmacological therapies. Strength training and an adequate nutritional intake form the basis of successful sarcopenia treatment.

Effect of speed endurance training and reduced training volume on running economy and single muscle fiber adaptations in trained runners.

The aim of the present study was to examine whether improved running economy with a period of speed endurance training and reduced training volume could be related to adaptations in specific muscle fibers. Twenty trained male (n = 14) and female (n = 6) runners (maximum oxygen consumption (VO2‐max): 56.4 ± 4.6 mL/min/kg) completed a 40‐day intervention with 10 sessions of speed endurance training (5–10 × 30‐sec maximal running) and a reduced (36%) volume of training. Before and after the intervention, a muscle biopsy was obtained at rest, and an incremental running test to exhaustion was performed. In addition, running at 60% vVO 2‐max, and a 10‐km run was performed in a normal and a muscle slow twitch (ST) glycogen‐depleted condition. After compared to before the intervention, expression of mitochondrial uncoupling protein 3 (UCP3) was lower (P < 0.05) and dystrophin was higher (P < 0.05) in ST muscle fibers, and sarcoplasmic reticulum calcium ATPase 1 (SERCA1) was lower (P < 0.05) in fast twitch muscle fibers. Running economy at 60% vVO 2‐max (11.6 ± 0.2 km/h) and at v10‐km (13.7 ± 0.3 km/h) was ~2% better (P < 0.05) after the intervention in the normal condition, but unchanged in the ST glycogen‐depleted condition. Ten kilometer performance was improved (P < 0.01) by 3.2% (43.7 ± 1.0 vs. 45.2 ± 1.2 min) and 3.9% (45.8 ± 1.2 vs. 47.7 ± 1.3 min) in the normal and the ST glycogen‐depleted condition, respectively. VO 2‐max was the same, but vVO 2‐max was 2.0% higher (P < 0.05; 19.3 ± 0.3 vs. 18.9 ± 0.3 km/h) after than before the intervention. Thus, improved running economy with intense training may be related to changes in expression of proteins linked to energy consuming processes in primarily ST muscle fibers.

In praise of slow.

I huff and puff my way up the moderate slope. Even by my own abysmal standards, this is a poor run. In the past hour, I have been overtaken by both an octogenarian and a mum pushing her toddlers in a buggy. Yet I am smiling. I am a happy runner, despite my utter mediocrity at this sport. But at work, happiness had become elusive. After a relatively relaxed Ph.D. and postdoc, I had been thrilled when I landed a tenured job. But as I worked to establish myself as a group leader, I began to feel intense pressure to be more competitive and publish more. Recently, as I wondered why I felt so discontented at my job, I realized that I could apply some lessons from running to my research. ![Figure][1] ILLUSTRATION: ROBERT NEUBECKER > “Not being able to sprint makes me a better endurance runner.” Lesson one: In the right race, your weakness can become your strength. For more than 30 years I equated running to speed and so, lacking the fast-twitch muscles of a sprinter, I chose not to run at all. But once I finally started running, I learned that not being able to sprint makes me a better endurance runner. Similarly, I often perceived my inability to focus on a single research topic as a barrier to success as an academic scientist. But I've found that my desire to branch out to different fields helps me make connections across disciplines and see my work in new ways, which has led to unexpected and exciting insights. Lesson two: Choose the right pace for your race. Last Christmas, a silver-haired gentleman helped me beat my (lamentable) 5K personal best by whispering, “Go at your pace, not theirs!” when I got stuck behind slower runners. I now realize that this would have been excellent advice early in my career. Some Ph.D. students push themselves too hard and burn out, but I had the opposite problem. I was happy to trundle along at the slowest pace I could get away with—but it ultimately held me back. Even though I like my slow-but-steady pace, I still needed to learn to push myself rather than drag my feet. Lesson three: An honest race is the only race worth running. I invariably finish in the bottom quartile of the local 5K run. All I would have to do to move up a couple of hundred places is take a shortcut through the fields. Yet I don't. Nobody does. Scientists are usually like that, too—but not always. Principal investigators are pressured to keep their spot in the fast lane, postdocs are chasing the elusive permanent contract, and students are keen to make their mark. Several of my publications would have had a much easier ride through the reviewing system had I been slightly less honest about our findings. The temptation to cheat to get an advantage can be great. However, and this is something that is often overlooked, an advantage is only useful if you are, in fact, engaged in a competition. This brings me to the last lesson, which is the most important of all. Lesson four: There really is no race. For me, running isn't about being faster than other runners. Likewise, my goal in research is not to “beat” my colleagues. Mark Rowlands, a philosopher, academic, and runner, argues that running makes us happy because it is a form of play and as such has intrinsic value. I don't run just to eat more peanut butter or to save money on psychotherapy (although these are strong motivating factors in my case). I run because doing so offers a glimpse of life's real value. I now think this is the secret to being happy in research, too. I don't do research only to get invited to conferences, see my name in print, or be promoted. Like running, research is a game with its own intrinsic value. Playing this game of discovery gives me enough joy to keep me going. Do I recommend an academic career in the slow lane? It doesn't work for everybody. Letting go of ambition in academia is a bit like leaving your GPS watch at home when heading out for a run. Scientists are ambitious; they want to be the hares leading the race ahead. But the tortoise's secret is that there is a lot of fun to be had at the back of the pack. [1]: pending:yes

Aging impairs regulation of ryanodine receptors from extensor digitorum longus but not soleus muscles.

Because impaired excitation-contraction coupling and reduced sarcoplasmic reticulum (SR) Ca2+ release may contribute to the age-associated decline in skeletal muscle strength, we investigated the effect of aging on regulation of the skeletal muscle isoform of the ryanodine receptor (RyR1) by physiological channel ligands. [3 H]Ryanodine binding to membranes from 8- and 26-month-old Fischer 344 extensor digitorum longus (EDL) and soleus muscles was used to investigate the effects of age on RyR1 modulation by Ca2+ and calmodulin (CaM). Aging reduced maximal Ca2+ -stimulated binding to EDL membranes. In 0.3 μM Ca2+ , age reduced binding and CaM increased binding to EDL membranes. In 300 μM Ca2+ , CaM reduced binding, but the age effect was not significant. Aging did not affect Ca2+ or CaM regulation of soleus RyR1. In aged fast-twitch muscle, impaired RyR1 Ca2+ regulation may contribute to lower SR Ca2+ release and reduced muscle function. Muscle Nerve 57: 1022-1025, 2018.

Copy Number Variation in SOX6 Contributes to Chicken Muscle Development.

Copy number variations (CNVs), which cover many functional genes, are associated with complex diseases, phenotypic diversity and traits that are economically important to raising chickens. The sex-determining region Y-box 6 (Sox6) plays a key role in fast-twitch muscle fiber differentiation of zebrafish and mice, but it is still unknown whether SOX6 plays a role in chicken skeletal muscle development. We identified two copy number polymorphisms (CNPs) which were significantly related to different traits on the genome level in chickens by AccuCopy® and CNVplex® analyses. Notably, five white recessive rock (CN = 1, CN = 3) variant individuals and two Xinghua (CN = 3) variant individuals contain a CNP13 (chromosome5: 10,500,294–10,675,531) which overlaps with SOX6. There is a disordered region in SOX6 proteins 265–579 aa coded by a partial CNV overlapping region. A quantitative real-time polymerase chain reaction showed that the expression level of SOX6 mRNA was positively associated with CNV and highly expressed during the skeletal muscle cell differentiation in chickens. After the knockdown of the SOX6, the expression levels of IGFIR1, MYF6, SOX9, SHOX and CCND1 were significantly down-regulated. All of them directly linked to muscle development. These results suggest that the number of CNVs in the CNP13 is positively associated with the expression level of SOX6, which promotes the proliferation and differentiation of skeletal muscle cells by up-regulating the expression levels of the muscle-growth-related genes in chickens as in other animal species.

Novel Sarcopenia-related Alterations in Sarcomeric Protein Post-translational Modifications (PTMs) in Skeletal Muscles Identified by Top-down Proteomics

Sarcopenia, the age-related loss of skeletal muscle mass and strength, is a significant cause of morbidity in the elderly and is a major burden on health care systems. Unfortunately, the underlying molecular mechanisms in sarcopenia remain poorly understood. Herein, we utilized top-down proteomics to elucidate sarcopenia-related changes in the fast- and slow-twitch skeletal muscles of aging rats with a focus on the sarcomeric proteome, which includes both myofilament and Z-disc proteins-the proteins that constitute the contractile apparatuses. Top-down quantitative proteomics identified significant changes in the post-translational modifications (PTMs) of critical myofilament proteins in the fast-twitch skeletal muscles of aging rats, in accordance with the vulnerability of fast-twitch muscles to sarcopenia. Surprisingly, age-related alterations in the phosphorylation of Cypher isoforms, proteins that localize to the Z-discs in striated muscles, were also noted in the fast-twitch skeletal muscle of aging rats. This represents the first report of changes in the phosphorylation of Z-disc proteins in skeletal muscle during aging. In addition, increased glutathionylation of slow skeletal troponin I, a novel modification that may help protect against oxidative damage, was observed in slow-twitch skeletal muscles. Furthermore, we have identified and characterized novel muscle type-specific proteoforms of myofilament proteins and Z-disc proteins, including a novel isoform of the Z-disc protein Enigma. The finding that the phosphorylation of Z-disc proteins is altered in response to aging in the fast-twitch skeletal muscles of aging rats opens new avenues for the investigation of the role of Z-discs in age-related muscle dysfunction.

Effect of Dietary Nitrate on Force Production and Sarcoplasmic Reticulum Ca&amp;lt;sup&amp;gt;2+&amp;lt;/sup&amp;gt; Handling in Rat Fast-Twitch Muscles Following Eccentric Contraction

Impaired excitation-contraction coupling occurs in eccentric contraction (ECC)-induced damaged muscles. It has been suggested that sarcoplasmic reticulum (SR) is susceptible to damage in the overstretched regions possibly marking the basis of excitation-contraction coupling damage. Recent studies have shown that dietary nitrate supplementation enhances SR function in fast-twitch muscles. In this study, we aimed to investigate whether dietary nitrate supplementation can alleviate a decline in muscle contractile properties and SR function following ECC. To this end, force production, Ca2+ uptake, Ca2+ release, and Ca2+-ATPase activity of the SR were examined in rat fast-twitch muscles immediately following ECC for 200 repetitions. In comparison with contralateral resting muscles, nitrate supplementation for up to 3 days resulted in an obvious decline in force production. However, there were no differences in terms of force production between 6-day nitrate-treated and contralateral muscles. Similar to the observations regarding force production, the SR Ca2+ release rate changed from an obvious decrease following the 0- and 3-day dietary nitrate supplementation to no difference following the 6-day nitrate supplementation. In contrast, ECC decreased the Ca2+-ATPase activity and Ca2+ uptake rate, irrespective of the period of dietary nitrate supplementation. Overall, these results indicate that dietary nitrate supplementation can alleviate ECC-related decreases in force production mediated through inhibited reductions in the SR Ca2+ release function.

Effects of a High-fat Diet on Mitochondrial Fission-Related Protein Drp1 in Slow- and Fast-Twitch Muscles

Effects of a High-fat Diet on Mitochondrial Fission-Related Protein Drp1 in Slow- and Fast-Twitch Muscles