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We examined changes in skeletal muscle protein lactylation and acetylation in response to acute resistance exercise, chronic resistance training (RT), and a single endurance cycling bout. Additionally, we performed in vitro experiments to determine if different sodium lactate treatments affect myotube protein lactylation and acetylation. The acute and chronic RT study (12 college-aged participants) consisted of 10 weeks of unilateral leg extensor RT with vastus lateralis (VL) biopsies taken at baseline, 24 h following the first RT bout, and the morning of the last day of the RT bout. For the acute cycling study (9 college-aged participants), VL biopsies were obtained before, 2 h after, and 8 h after 60 min of cycling. For in vitro experiments, C2C12 myotubes were treated with varying levels of sodium lactate, including LOW (1 mM for 24 h), HIGH (10 mM for 24 h), and PULSE (10 mM for 30 min followed by 1 mM for 23.5-h). Neither acute nor chronic RT significantly affected nuclear or cytoplasmic protein lactylation. However, cytoplasmic protein acetylation was significantly reduced following one RT bout (−15%, p = 0.002) and chronic RT (−16%, p = 0.006). Cycling did not acutely alter post-exercise global protein lactylation or acetylation patterns. Lastly, varying 24 h lactate treatments did not alter nuclear or cytoplasmic protein lactylation or acetylation, cytoplasmic protein synthesis levels, or myotube diameters. These findings continue to support the idea that exercise induces more dynamic changes in skeletal muscle protein acetylation, but not lactylation. However, further human research with more sampling timepoints and a lactylomics approach are needed to determine if, at all, different exercise modalities affect skeletal muscle protein lactylation. [ABSTRACT FROM AUTHOR]

We examined how resistance exercise (RE), cycling exercise and disuse atrophy affect myosin heavy chain (MyHC) protein fragmentation. The 1boutRE study involved younger men (n = 8; 5 ± 2 years of RE experience) performing a lower body RE bout with vastus lateralis (VL) biopsies being obtained prior to and acutely following exercise. With the 10weekRT study, VL biopsies were obtained in 36 younger adults before and 24 h after their first/naïve RE bout. Participants also engaged in 10 weeks of resistance training and donated VL biopsies before and 24 h after their last RE bout. VL biopsies were also examined in an acute cycling study (n = 7) and a study involving 2 weeks of leg immobilization (n = 20). In the 1boutRE study, fragmentation of all MyHC isoforms (MyHCTotal) increased 3 h post‐RE (∼200%, P = 0.018) and returned to pre‐exercise levels by 6 h post‐RE. Interestingly, a greater magnitude increase in MyHC type IIa versus I isoform fragmentation occurred 3 h post‐RE (8.6 ± 6.3‐fold vs. 2.1 ± 0.7‐fold, P = 0.018). In 10weekRT participants, the first/naïve and last RE bouts increased MyHCTotal fragmentation 24 h post‐RE (+65% and +36%, P < 0.001); however, the last RE bout response was attenuated compared to the first bout (P = 0.045). Although cycling exercise did not alter MyHCTotal fragmentation, ∼8% VL atrophy with 2 weeks of leg immobilization increased MyHCTotal fragmentation (∼108%, P < 0.001). Mechanistic C2C12 myotube experiments indicated that MyHCTotal fragmentation is likely due to calpain proteases. In summary, RE and disuse atrophy increase MyHC protein fragmentation. Research into how ageing and disease‐associated muscle atrophy affect these outcomes is needed. Highlights: What is the central question of this study?How different exercise stressors and disuse affect skeletal muscle myosin heavy chain fragmentation.What is the main finding and its importance?This investigation is the first to demonstrate that resistance exercise and disuse atrophy lead to skeletal muscle myosin heavy chain protein fragmentation in humans. Mechanistic in vitro experiments provide additional evidence that MyHC fragmentation occurs through calpain proteases. [ABSTRACT FROM AUTHOR]

Ribosomal DNA (rDNA) copies exist across multiple chromosomes, and interindividual variation in copy number is speculated to influence the hypertrophic response to resistance training. Thus, we examined if rDNA copy number was associated with resistance training-induced skeletal muscle hypertrophy. Participants (n = 53 male, 21 ± 1 yr old; n = 29 female, 21 ± 2 yr old) performed 10–12 wk of full-body resistance training. Hypertrophy outcomes were determined, as was relative rDNA copy number from preintervention vastus lateralis (VL) biopsies. Pre- and postintervention VL biopsy total RNA was assayed in all participants, and mRNA/rRNA markers of ribosome content and biogenesis were also assayed in the 29 female participants before training, 24 h following training bout 1, and in the basal state after 10 wk of training. Across all participants, no significant associations were evident between relative rDNA copy number and training-induced changes in whole body lean mass (r = −0.034, P = 0.764), vastus lateralis thickness (r = 0.093, P = 0.408), mean myofiber cross-sectional area (r = −0.128, P = 0.259), or changes in muscle RNA concentrations (r = 0.026, P = 0.818), and these trends were similar when examining each gender. However, all Pol-I regulon mRNAs as well as 45S pre-rRNA, 28S rRNA, and 18S rRNA increased 24 h following the first training bout in female participants. Follow-up studies using LHCN-M2 myotubes demonstrated that a reduction in relative rDNA copy number induced by bisphenol A did not significantly affect insulin-like-growth factor-induced myotube hypertrophy. These findings suggest that relative rDNA copy number is not associated with myofiber hypertrophy. NEW & NOTEWORTHY: We examined ribosomal DNA (rDNA) copy numbers in men and women who resistance trained for 10–12 wk and found no significant associations with skeletal muscle hypertrophy outcomes. These data, along with in vitro data in immortalized human myotubes whereby rDNA copy number was reduced, provide strong evidence that relative rDNA copy number is not associated with anabolism. [ABSTRACT FROM AUTHOR]

Purpose: Resistance training (RT) induces muscle growth at varying rates across RT phases, and evidence suggests that the muscle-molecular responses to training bouts become refined or attenuated in the trained state. This study examined how proteolysis-related biomarkers and extracellular matrix (ECM) remodeling factors respond to a bout of RT in the untrained (UT) and trained (T) state. Methods: Participants (19 women and 19 men) underwent 10 weeks of RT. Biopsies of vastus lateralis were collected before and after (24 h) the first (UT) and last (T) sessions. Vastus lateralis cross-sectional area (CSA) was assessed before and after the experimental period. Results: There were increases in muscle and type II fiber CSAs. In both the UT and T states, calpain activity was upregulated and calpain-1/-2 protein expression was downregulated from Pre to 24 h. Calpain-2 was higher in the T state. Proteasome activity and 20S proteasome protein expression were upregulated from Pre to 24 h in both the UT and T. However, proteasome activity levels were lower in the T state. The expression of poly-ubiquitinated proteins was unchanged. MMP activity was downregulated, and MMP-9 protein expression was elevated from Pre to 24 h in UT and T. Although MMP-14 protein expression was acutely unchanged, this marker was lower in T state. TIMP-1 protein levels were reduced Pre to 24 h in UT and T, while TIMP-2 protein levels were unchanged. Conclusion: Our results are the first to show that RT does not attenuate the acute-induced response of proteolysis and ECM remodeling-related biomarkers. [ABSTRACT FROM AUTHOR]

Although several reports have hypothesized that exercise may increase skeletalmuscle protein lactylation, empirical evidence in humans is lacking. Thus, we adopted a multifaceted approach to examine if acute and subchronic resistance training (RT) altered skeletal muscle protein lactylation levels. Inmice, we also sought to examine if surgical ablation-induced plantaris hypertrophy coincided with increases in muscle protein lactylation. To examine acute responses, participants' blood lactate concentrations were assessed before, during, and after eight sets of an exhaustive lower body RT bout (n = 10 trained college-agedmen). Vastus lateralis biopsies were also taken before, 3-h post, and 6-h post-exercise to assess muscle protein lactylation. To identify training responses, another cohort of trained college-aged men (n = 14) partook in 6 weeks of lower-body RT (3x/week) and biopsies were obtained before and following the intervention. Five-month-old C57BL/6 mice were subjected to 10 days of plantaris overload (OV, n = 8) or served as age-matched sham surgery controls (Sham, n = 8). Although acute resistance training significantly increased blood lactate responses ~7.2-fold (p < 0.001), cytoplasmic and nuclear protein lactylation levels were not significantly altered at the post-exercise time points, and no putative lactylation-dependent mRNA was altered following exercise. Six weeks of RT did not alter cytoplasmic protein lactylation (p = 0.800) despite significantly increasing VLmuscle size (+3.5%, p=0.037), and again, no putative lactylation-dependent mRNA was significantly affected by training. Plantaris muscles were larger in OV versus Sham mice (+43.7%, p < 0.001). However, cytoplasmic protein lactylation was similar between groups (p = 0.369), and nuclear protein lactylation was significantly lower in OV versus Sham mice (p < 0.001). The current null findings, along with other recent null findings in the literature, challenge the thesis that lactate has an appreciable role in promoting skeletalmuscle hypertrophy. [ABSTRACT FROM AUTHOR]

We investigated the effects of performing a period of resistance training (RT) on the performance and molecular adaptations to a subsequent period of endurance training (ET). Twenty‐five young adults were divided into an RT+ET group (n = 13), which underwent 7 weeks of RT followed by 7 weeks of ET, and an ET‐only group (n = 12), which performed 7 weeks of ET. Body composition, endurance performance and muscle biopsies were collected before RT (T1, baseline for RT+ET), before ET (T2, after RT for RT+ET and baseline for ET) and after ET (T3). Immunohistochemistry was performed to determine fibre cross‐sectional area (fCSA), myonuclear content, myonuclear domain size, satellite cell number and mitochondrial content. Western blots were used to quantify markers of mitochondrial remodelling. Citrate synthase activity and markers of ribosome content were also investigated. RT improved body composition and strength, increased vastus lateralis thickness, mixed and type II fCSA, myonuclear number, markers of ribosome content, and satellite cell content (P < 0.050). In response to ET, both groups similarly decreased body fat percentage (P < 0.0001) and improved endurance performance (e.g. V̇O2max${\dot V_{{{\mathrm{O}}_2}\max }}$, and speed at which the onset of blood lactate accumulation occurred, P < 0.0001). Levels of mitochondrial complexes I–IV in the ET‐only group increased 32–66%, while those in the RT+ET group increased 1–11% (time, P < 0.050). Additionally, mixed fibre relative mitochondrial content increased 15% in the ET‐only group but decreased 13% in the RT+ET group (interaction, P = 0.043). In conclusion, RT performed prior to ET had no additional benefits to ET adaptations. Moreover, prior RT seemed to impair mitochondrial adaptations to ET. Key points: Resistance training is largely underappreciated as a method to improve endurance performance, despite reports showing it may improve mitochondrial function.Although several concurrent training studies are available, in this study we investigated the effects of performing a period of resistance training on the performance and molecular adaptations to subsequent endurance training.Prior resistance training did not improve endurance performance and impaired most mitochondrial adaptations to subsequent endurance training, but this effect may have been a result of detraining from resistance training. [ABSTRACT FROM AUTHOR]

Limited research exists examining how resistance training to failure affects applied outcomes and single motor unit characteristics in previously trained individuals. Herein, resistance‐trained adults (24 ± 3 years old, self‐reported resistance training experience was 6 ± 4 years, 11 men and 8 women) were randomly assigned to either a low‐repetitions‐in‐reserve (RIR; i.e., training near failure, n = 10) or high‐RIR (i.e., not training near failure, n = 9) group. All participants implemented progressive overload during 5 weeks where low‐RIR performed squat, bench press, and deadlift twice weekly and were instructed to end each training set with 0–1 RIR. high‐RIR performed identical training except for being instructed to maintain 4–6 RIR after each set. During week 6, participants performed a reduced volume‐load. The following were assessed prior to and following the intervention: (i) vastus lateralis (VL) muscle cross‐sectional area (mCSA) at multiple sites; (ii) squat, bench press, and deadlift one‐repetition maximums (1RMs); and (iii) maximal isometric knee extensor torque and VL motor unit firing rates during an 80% maximal voluntary contraction. Although RIR was lower in the low‐ versus high‐RIR group during the intervention (p < 0.001), total training volume did not significantly differ between groups (p = 0.222). There were main effects of time for squat, bench press, and deadlift 1RMs (all p‐values < 0.05), but no significant condition × time interactions existed for these or proximal/middle/distal VL mCSA data. There were significant interactions for the slope and y‐intercept of the motor unit mean firing rate versus recruitment threshold relationship. Post hoc analyses indicated low‐RIR group slope values decreased and y‐intercept values increased after training suggesting low‐RIR training increased lower‐threshold motor unit firing rates. This study provides insight into how resistance training in proximity to failure affects strength, hypertrophy, and single motor unit characteristics, and may inform those who aim to program for resistance‐trained individuals. [ABSTRACT FROM AUTHOR]

Although transcriptome profiling has been used in several resistance training studies, the associated analytical approaches seldom provide in-depth information on individual genes linked to skeletal muscle hypertrophy. Therefore, a secondary analysis was performed herein on a muscle transcriptomic dataset we previously published involving trained college-aged men (n = 11) performing two resistance exercise bouts in a randomized and crossover fashion. The lower-load bout (30 Fail) consisted of 8 sets of lower body exercises to volitional fatigue using 30% one-repetition maximum (1 RM) loads, whereas the higher-load bout (80 Fail) consisted of the same exercises using 80% 1 RM loads. Vastus lateralis muscle biopsies were collected prior to (PRE), 3 h, and 6 h after each exercise bout, and 58 genes associated with skeletal muscle hypertrophy were manually interrogated from our prior microarray data. Select targets were further interrogated for associated protein expression and phosphorylation induced-signaling events. Although none of the 58 gene targets demonstrated significant bout x time interactions, ~57% (32 genes) showed a significant main effect of time from PRE to 3 h (15↑ and 17↓, p < 0.01), and ~26% (17 genes) showed a significant main effect of time from PRE to 6 h (8↑ and 9↓, p < 0.01). Notably, genes associated with the myostatin (9 genes) and mammalian target of rapamycin complex 1 (mTORC1) (9 genes) signaling pathways were most represented. Compared to mTORC1 signaling mRNAs, more MSTN signaling-related mRNAs (7 of 9) were altered post-exercise, regardless of the bout, and RHEB was the only mTORC1-associated mRNA that was upregulated following exercise. Phosphorylated (phospho-) p70S6K (Thr389) (p = 0.001; PRE to 3 h) and follistatin protein levels (p = 0.021; PRE to 6 h) increased post-exercise, regardless of the bout, whereas phospho-AKT (Thr389), phospho-mTOR (Ser2448), and myostatin protein levels remained unaltered. These data continue to suggest that performing resistance exercise to volitional fatigue, regardless of load selection, elicits similar transient mRNA and signaling responses in skeletal muscle. Moreover, these data provide further evidence that the transcriptional regulation of myostatin signaling is an involved mechanism in response to resistance exercise. [ABSTRACT FROM AUTHOR]

We sought to determine the skeletal muscle genome-wide DNA methylation and mRNA responses to one bout of lower load (LL) versus higher load (HL) resistance exercise. Trained college-aged males (n = 11, 23 ± 4 years old, 4 ± 3 years self-reported training) performed LL or HL bouts to failure separated by one week. The HL bout (i.e., 80 Fail) consisted of four sets of back squats and four sets of leg extensions to failure using 80% of participants estimated one-repetition maximum (i.e., est. 1-RM). The LL bout (i.e., 30 Fail) implemented the same paradigm with 30% of est. 1-RM. Vastus lateralis muscle biopsies were collected before, 3 h, and 6 h after each bout. Muscle DNA and RNA were batch-isolated and analyzed using the 850k Illumina MethylationEPIC array and Clariom S mRNA microarray, respectively. Performed repetitions were significantly greater during the 30 Fail versus 80 Fail (p < 0.001), although total training volume (sets × reps × load) was not significantly different between bouts (p = 0.571). Regardless of bout, more CpG site methylation changes were observed at 3 h versus 6 h post exercise (239,951 versus 12,419, respectively; p < 0.01), and nuclear global ten-eleven translocation (TET) activity, but not global DNA methyltransferase activity, increased 3 h and 6 h following exercise regardless of bout. The percentage of genes significantly altered at the mRNA level that demonstrated opposite DNA methylation patterns was greater 3 h versus 6 h following exercise (~75% versus ~15%, respectively). Moreover, high percentages of genes that were up- or downregulated 6 h following exercise also demonstrated significantly inversed DNA methylation patterns across one or more CpG sites 3 h following exercise (65% and 82%, respectively). While 30 Fail decreased DNA methylation across various promoter regions versus 80 Fail, transcriptome-wide mRNA and bioinformatics indicated that gene expression signatures were largely similar between bouts. Bioinformatics overlay of DNA methylation and mRNA expression data indicated that genes related to "Focal adhesion," "MAPK signaling," and "PI3K-Akt signaling" were significantly affected at the 3 h and 6 h time points, and again this was regardless of bout. In conclusion, extensive molecular profiling suggests that post-exercise alterations in the skeletal muscle DNA methylome and mRNA transcriptome elicited by LL and HL training bouts to failure are largely similar, and this could be related to equal volumes performed between bouts. [ABSTRACT FROM AUTHOR]

New Findings: What is the central question of this study?Do changes in myofibre cross‐sectional area, pennation angle and fascicle length predict vastus lateralis whole‐muscle cross‐sectional area changes following resistance training?What is the main finding and its importance?Changes in vastus lateralis mean myofibre cross‐sectional area, fascicle length and pennation angle following a period of resistance training did not collectively predict changes in whole‐muscle cross‐sectional area. Despite the limited sample size in this study, these data reiterate that it remains difficult to generalize the morphological adaptations that predominantly drive tissue‐level vastus lateralis muscle hypertrophy. Myofibre hypertrophy during resistance training (RT) poorly associates with tissue‐level surrogates of hypertrophy. However, it is underappreciated that, in pennate muscle, changes in myofibre cross‐sectional area (fCSA), fascicle length (Lf) and pennation angle (PA) likely coordinate changes in whole‐muscle cross‐sectional area (mCSA). Therefore, we determined if changes in fCSA, PA and Lf predicted vastus lateralis (VL) mCSA changes following RT. Thirteen untrained college‐aged males (23 ± 4 years old, 25.4 ± 5.2 kg/m2) completed 7 weeks of full‐body RT (twice weekly). Right leg VL ultrasound images and biopsies were obtained prior to (PRE) and 72 h following (POST) the last training bout. Regression was used to assess if training‐induced changes in mean fCSA, PA and Lf predicted VL mCSA changes. Correlations were also performed between PRE‐to‐POST changes in obtained variables. Mean fCSA (+18%), PA (+8%) and mCSA (+22%) increased following RT (P < 0.05), but not Lf (0.1%, P = 0.772). Changes in fCSA, Lf and PA did not collectively predict changes in mCSA (R2= 0.282, adjusted R2= 0.013, F3,8 = 1.050, P = 0.422). Moderate negative correlations existed for percentage changes in PA and Lf (r = −0.548, P = 0.052) and changes in fCSA and Lf (r = −0.649, P = 0.022), and all other associations were weak (|r| < 0.500). Although increases in mean fCSA, PA and VL mCSA were observed, inter‐individual responses for each variable and limitations for each technique make it difficult to generalize the morphological adaptations that predominantly drive tissue‐level VL muscle hypertrophy. However, the small subject pool is a significant limitation, and more research in this area is needed. [ABSTRACT FROM AUTHOR]

We sought to determine if manipulating resistance training (RT) variables differentially altered the expression of select sarcoplasmic and myofibril proteins as well as myofibrillar spacing in myofibers. Resistance-trained men (n = 20; 26 ± 3 years old) trained for 8 weeks where a randomized leg performed either a standard (CON) or variable RT protocol (VAR: manipulation of load, volume, muscle action, and rest intervals at each RT session). A pre-training (PRE) vastus lateralis biopsy was obtained from a randomized single leg, and biopsies were obtained from both legs 96 h following the last training bout. The sarcoplasmic protein pool was assayed for proteins involved in energy metabolism, and the myofibril protein pool was assayed for relative myosin heavy chain (MHC) and actin protein abundances. Sections were also histologically analyzed to obtain myofibril spacing characteristics. VAR resulted in ~12% greater volume load (VL) compared to CON (p < 0.001). The mean fiber cross-sectional area increased following both RT protocols [CON: 14.6% (775.5 μm2), p = 0.006; VAR: 13.9% (743.2 μm2), p = 0.01 vs. PRE for both], but without significant differences between protocols (p = 0.79). Neither RT protocol affected a majority of assayed proteins related to energy metabolism, but both training protocols increased hexokinase 2 protein levels and decreased a mitochondrial beta-oxidation marker (VLCAD protein; p < 0.05). Citrate synthase activity levels increased with CON RT (p < 0.05), but not VAR RT. The relative abundance of MHC (summed isoforms) decreased with both training protocols (p < 0.05). However, the relative abundance of actin protein (summed isoforms) decreased with VAR only (13.5 and 9.0%, respectively; p < 0.05). A decrease in percent area occupied by myofibrils was observed from PRE to VAR (−4.87%; p = 0.048), but not for the CON (4.53%; p = 0.979). In contrast, there was an increase in percent area occupied by non-contractile space from PRE to VAR (10.14%; p = 0.048), but not PRE to CON (0.72%; p = 0.979). In conclusion, while both RT protocols increased muscle fiber hypertrophy, a higher volume-load where RT variables were frequently manipulated increased non-contractile spacing in resistance-trained individuals. [ABSTRACT FROM AUTHOR]

Resistance training increases muscle fiber hypertrophy, but the morphological adaptations that occur within muscle fibers remain largely unresolved. Fifteen males with minimal training experience (24±4years, 23.9±3.1kg/m2 body mass index) performed 10weeks of conventional, full-body resistance training (2× weekly). Body composition, the radiological density of the vastus lateralis muscle using peripheral quantitative computed tomography (pQCT), and vastus lateralis muscle biopsies were obtained 1week prior to and 72h following the last training bout. Quantification of myofibril and mitochondrial areas in type I (positive for MyHC I) and II (positive for MyHC IIa/IIx) fibers was performed using immunohistochemistry (IHC) techniques. Relative myosin heavy chain and actin protein abundances per wet muscle weight as well as citrate synthase (CS) activity assays were also obtained on tissue lysates. Training increased whole-body lean mass, mid-thigh muscle cross-sectional area, mean and type II fiber cross-sectional areas (fCSA), and maximal strength values for leg press, bench press, and deadlift (p <0.05). The intracellular area occupied by myofibrils in type I or II fibers was not altered with training, suggesting a proportional expansion of myofibrils with fCSA increases. However, our histological analysis was unable to differentiate whether increases in myofibril number or girth occurred. Relative myosin heavy chain and actin protein abundances also did not change with training. IHC indicated training increased mitochondrial areas in both fiber types (p =0.018), albeit CS activity levels remained unaltered with training suggesting a discordance between these assays. Interestingly, although pQCT-derived muscle density increased with training (p =0.036), suggestive of myofibril packing, a positive association existed between training-induced changes in this metric and changes in mean fiber myofibril area (r =0.600, p =0.018). To summarize, our data imply that shorter-term resistance training promotes a proportional expansion of the area occupied by myofibrils and a disproportional expansion of the area occupied by mitochondria in type I and II fibers. Additionally, IHC and biochemical techniques should be viewed independently from one another given the lack of agreement between the variables assessed herein. Finally, the pQCT may be a viable tool to non-invasively track morphological changes (specifically myofibril density) in muscle tissue. [ABSTRACT FROM AUTHOR]