Physiological and muscle histochemical assessment of men and women 10 km runners matched for race performance or training volume.

Women have a disadvantage for performance in long-distance running compared with men. To elaborate on inherent characteristics, 12 subelite women were matched with 12 men for training volume (M-Tm) (56.6 ± 18 vs. 55.7 ± 17 km/wk). The women were also matched to other men for a 10 km staged outdoor time trial (M-Pm) (42:36 min:s) to determine which factors could explain equal running performance. Anthropometry and treadmill tests were done. Fiber type (% Type I and Type IIA) and citrate synthase activities were analyzed in muscle biopsy samples. Consistent sex differences for both comparisons included height, weight, % body fat (P < 0.01), and hematocrit (P < 0.05). Women had lower V̇o2max and peak treadmill speed (PTS) compared with both M-Tm and M-Pm (P < 0.01). Training matched pairs had no sex difference in % PTS at race pace but compared with M-Pm women ran at a higher % PTS (P < 0.05) and %HRmax (P < 0.01) at race pace. On average, the women trained 22.9 km/wk more than M-Pm (+67.5%, P < 0.01). This training was not associated with higher V̇o2max or better running economy. Muscle morphology and oxidative capacity did not differ between groups. Percentage body fat remained significantly higher in women. In conclusion, women matched to men for training volume had slower 10 km performance (-10.5% P < 0.05). Higher training volume, more high-intensity sessions/wk, and time spent training in the 95%-100% HRmax zone may explain the higher % PTS and %HRmax at race pace in women compared with performance-matched men.NEW & NOTEWORTHY When subelite women 10 km runners were matched with male counterparts for 10 km race performance, inherent differences in % body fat, V̇o2max, Hct, and peak treadmill speed were counteracted by significantly higher training volume, more time training at higher %HRmax and consequently, higher %HRmax and %PTS at race pace. Citrate synthase activity and muscle fiber types did not differ. When women and men matched for training, 10 km performance of men was 10.5% faster.

Skeletal muscle fibre type and enzymatic activity in adult offspring following placental and peripheral malaria exposure in foetal life.

Background: Maternal malaria may restrict foetal growth. Impaired utero-placental blood flow due to malaria infection may cause hypoxia-induced altered skeletal muscle fibre type distribution in the offspring, which may contribute to insulin resistance and impaired glucose metabolism. This study assessed muscle fibre distribution 20 years after placental and/or peripheral in-utero malaria exposure compared to no exposure, i.e., PPM+, PM+, and M-, respectively. Methods: We traced 101 men and women offspring of mothers who participated in a malaria chemosuppression study in Muheza, Tanzania. Of 76 eligible participants, 50 individuals (29 men and 21 women) had skeletal muscle biopsy taken from m. vastus lateralis in the right leg. As previously reported, fasting and 30 min post-oral glucose challenge plasma glucose values were higher, and insulin secretion disposition index was lower, in the PPM+ group. Aerobic capacity (fitness) was estimated by an indirect VO2max test on a stationary bicycle. Muscle fibre sub-type (myosin heavy chain, MHC) distribution was analysed, as were muscle enzyme activities (citrate synthase (CS), 3-hydroxyacyl-CoA dehydrogenase, myophosphorylase, phosphofructokinase, lactate dehydrogenase, and creatine kinase activities. Between-group analyses were adjusted for MHC-I %. Results: No differences in aerobic capacity were found between groups. Despite subtle elevations of plasma glucose levels in the PPM+ group, there was no difference in MHC sub-types or muscle enzymatic activities between the malaria-exposed and non-exposed groups. Conclusion: The current study did not show differences in MHC towards glycolytic sub-types or enzymatic activity across the sub-groups. The results support the notion of the mild elevations of plasma glucose levels in people exposed to placental malaria in pregnancy being due to compromised pancreatic insulin secretion rather than insulin resistance.

Therapeutic Benefit in Rheumatoid Cachexia Illustrated Using a Novel Primary Human Triple Cell Coculture Model.

Background: The loss of muscle mass in rheumatoid arthritis (RA), termed rheumatoid cachexia, is predicted to result from the complex interactions between different cell types involved in the maintenance of skeletal muscle mass, namely, myoblasts, fibroblasts, and macrophages. The complexity within the muscle is further highlighted by the incidence of nonresponsiveness to current RA treatment strategies. Method: This study aimed at determining differences in the cellular responses in a novel human primary cell triple coculture model exposed to serum collected from nonarthritic controls (NC), RA treatment naïve (RATN), and RA treatment-nonresponding (RATNR) patients. Bone morphogenetic protein-7 (BMP-7) was investigated as a treatment option. Results: Plasma analysis indicated that samples were indeed representative of healthy and RA patients-notably, the RATNR patients additionally exhibited dysregulated IL-6/IL-10 correlations. Coculture exposure to serum from RATNR patients demonstrated increased cellular growth (p < 0.001), while both hepatocyte growth factor (p < 0.01) and follistatin (p < 0.001) were reduced when compared to NC. Furthermore, decreased concentration of markers of extracellular matrix formation, transforming growth factor-ß (TGF-ß; p < 0.05) and fibronectin (p < 0.001), but increased collagen IV (p < 0.01) was observed following RATNR serum exposure. Under healthy conditions, BMP-7 exhibited potentially beneficial results in reducing fibrosis-generating TGF-ß (p < 0.05) and fibronectin (p < 0.05). BMP-7 further exhibited protective potential in the RA groups through reversing the aberrant tendencies observed especially in the RATNR serum-exposed group. Conclusion: Exposure of the triple coculture to RATN and RATNR serum resulted in dysregulated myoblast proliferation and growth, and ECM impairment, which was reversed by BMP-7 treatment.

Muscle Histopathological Abnormalities in a Patient With a CCT5 Mutation Predicted to Affect the Apical Domain of the Chaperonin Subunit.

Recognition of diseases associated with mutations of the chaperone system genes, e.g., chaperonopathies, is on the rise. Hereditary and clinical aspects are established, but the impact of the mutation on the chaperone molecule and the mechanisms underpinning the tissue abnormalities are not. Here, histological features of skeletal muscle from a patient with a severe, early onset, distal motor neuropathy, carrying a mutation on the CCT5 subunit (MUT) were examined in comparison with normal muscle (CTR). The MUT muscle was considerably modified; atrophy of fibers and disruption of the tissue architecture were prominent, with many fibers in apoptosis. CCT5 was diversely present in the sarcolemma, cytoplasm, and nuclei in MUT and in CTR and was also in the extracellular space; it colocalized with CCT1. In MUT, the signal of myosin appeared slightly increased, and actin slightly decreased as compared with CTR. Desmin was considerably delocalized in MUT, appearing with abnormal patterns and in precipitates. Alpha-B-crystallin and Hsp90 occurred at lower signals in MUT than in CTR muscle, appearing also in precipitates with desmin. The abnormal features in MUT may be the consequence of inactivity, malnutrition, denervation, and failure of protein homeostasis. The latter could be at least in part caused by malfunction of the CCT complex with the mutant CCT5 subunit. This is suggested by the results of the in silico analyses of the mutant CCT5 molecule, which revealed various abnormalities when compared with the wild-type counterpart, mostly affecting the apical domain and potentially impairing chaperoning functions. Thus, analysis of mutated CCT5 in vitro and in vivo is anticipated to provide additional insights on subunit involvement in neuromuscular disorders.

Total mRNA and primary human myoblasts' in vitro cell cycle progression distinguishes between clones.

Satellite cells are generally quiescent in vivo. Once activated, progression through the cell cycle begins. Immortalised myoblasts from a single cell line are fairly homogenous in culture, but primary human myoblasts (PHMs) demonstrate heterogeneity. This phenomenon is poorly understood however may impact on PHM expansion. This study aimed to evaluate cell cycle transition from growth to synthesis phases of the cell cycle (G1 to S phase) and total mRNA relevant to this transition in PHM clones derived from 2 donor biopsies. Proportions of cells transitioning from G1 to S phase were evaluated at 2-hourly intervals for 24 h (n = 3 for each) and total mRNA quantified. Both PHM clones revealed an exponential transition from G1 to S phase over time, with a significantly slower rate for PHMs from S9.1 compared to S6.3, which had a higher proportion of PHMs in S phase for most time-points (p < 0.05). After 24 h the proportion of PHMs in S phase was ∼13% (S6.3) compared to ∼22% (S9.1). Gene transcription increased as cells progressed from G1 to S phase. Although total RNA increased with similar linearity in both clones, S6.3 PHMs had consistently (10 out of 12 time points) significantly higher concentrations. Validating the 2-hourly assessment over 24 h, a 4-hourly assessment from 8 to 32 h revealed similar differences but included the beginning of a plateau. This study demonstrates that PHMs from different donors differ in both cell cycle progression and overall transcriptome revealing new aspects in the heterogeneity of isolated satellite cells in vitro.

Unresolved intramuscular inflammation, not diminished skeletal muscle regenerative capacity, is at the root of rheumatoid cachexia: insights from a rat CIA model.

Rheumatoid arthritis targets numerous organs in patients, including the skeletal muscle, resulting in rheumatoid cachexia. In the muscle niche, satellite cells, macrophages, and myofibroblasts may be affected and the factors they release altered. This study aimed to assess these cell types, cytokines, and growth factors and their relationships to muscle fiber size and number in a rodent collagen-induced arthritis (CIA) model, in order to identify new therapeutic targets. Fiber cross-sectional area (CSA) was 57% lower in CIA than controls (p < 0.0001), thus smaller but more fibers visible per field of view. Immunostaining indicated the increased presence of satellite cells, macrophages, myofibroblasts, and myonuclei per field of view in CIA (p < 0.01), but this finding was not maintained when taking fiber number into consideration. Western blots of gastrocnemius samples indicated that tumor necrosis factor-α was significantly elevated (p < 0.01) while interleukin-10 (IL-10) was decreased (p < 0.05) in CIA. This effect was maintained (and heightened for IL-10) when expressed per fiber number. Myogenic regulatory factors (MyoD and myogenin), transforming growth factor-ß and inhibitor of differentiation were significantly elevated in CIA muscle and levels correlated significantly with CSA. Several of these factors remained elevated, but bone morphogenetic protein-7 decreased when considering fiber number per area. In conclusion, CIA-muscle demonstrated a good regenerative response. Myoblast numbers per fiber were not elevated, suggesting their activity results from the persistent inflammatory signaling which also significantly hampered maintenance of muscle fiber size. A clearer picture of signaling events at cellular level in arthritis muscle may be derived from expressing data per fiber.

Sarcopenic Obesity in Africa: A Call for Diagnostic Methods and Appropriate Interventions.

This perspective aims to highlight the lack of current knowledge on sarcopenic obesity in Africa and to call for diagnostic methods and appropriate interventions. Sarcopenic obesity has been defined as obesity that occurs in combination with low muscle mass and function, which is typically evident in older adults. However, there has been no clear consensus on population-specific diagnostic criterion, which includes both gold-standard measures that can be used in a more advanced health care system, and surrogate measures that can be used in low-income settings with limited resources and funding. Importantly, low and middle-income countries (LMICs) across Africa are in an ongoing state of economic and social transition, which has contributed to an increase in the aging population, alongside the added burden of poverty, obesity, and associated co-morbidities. It is anticipated that alongside the increased prevalence of obesity, these countries will further experience an increase in age-related musculoskeletal diseases such as sarcopenia. The developmental origins of health and disease (DOHaD) approach will allow clinicians and researchers to consider developmental trajectories, and the influence of the environment, for targeting high-risk individuals and communities for treatment and/or prevention-based interventions that are implemented throughout all stages of the life course. Once a valid and reliable diagnostic criterion is developed, we can firstly assess the prevalence and burden of sarcopenic obesity in LMICs in Africa, and secondly, develop appropriate and sustainable interventions that target improved dietary and physical activity behaviors throughout the life course.

Food Security, Dietary Intake, and Foodways of Urban Low-Income Older South African Women: An Exploratory Study.

This cross-sectional study explored the differences in sociodemographics, dietary intake, and household foodways (cultural, socioeconomic practices that affect food purchase, consumption, and preferences) of food secure and food insecure older women living in a low-income urban setting in South Africa. Women (n = 122) aged 60-85 years old were recruited, a sociodemographic questionnaire was completed, and food security categories were determined. The categories were dichotomised into food secure (food secure and mild food insecurity) and food insecure (moderate and severe). A one-week quantified food frequency questionnaire was administered. Height and weight were measured to calculate body mass index (BMI, kg/m2). Most participants (>90%) were overweight/obese, unmarried/widowed, and breadwinners with a low monthly household income. Food insecure participants (36.9%) more frequently borrowed money for food (57.8% vs. 39.0%, p = 0.04), ate less so that their children could have more to eat (64.4%. vs. 27.3%, p = 0.001), and had higher housing density (1.2 vs. 1.0, p = 0.03), compared to their food-secure counterparts. Overall, <30% of participants met the WHO (Geneva, Switzerland) recommended daily servings of healthy foods (fruits, vegetables, and dairy products), but >60% perceived that they consumed an adequate amount of healthy foods. The overall low-quality diet of our cohort was associated with poor nutritional perceptions and choices, coupled with financial constraints.

Understanding factors associated with sarcopenic obesity in older African women from a low-income setting: a cross-sectional analysis.

BACKGROUND: High rates of food insecurity, obesity and obesity-related comorbidities in ageing South African (SA) women may amplify the risk of developing sarcopenic obesity. This study aimed to investigate the prevalence and correlates of sarcopenic obesity and its diagnostic components [grip strength, appendicular skeletal muscle mass (ASM) and body mass index (BMI)] in older SA women from a low-income setting. METHODS: This cross-sectional study recruited black SA women between the ages of 60-85 years (n = 122) from a low-income community. Testing included a fasting blood sample (markers of cardiometabolic risk, HIV), whole body and regional muscle and fat mass (dual-energy absorptiometry x-ray), anthropometry, blood pressure, functional movement tests, current medication use, demographic and health questionnaires, physical activity (PA; accelerometery), household food insecurity access scale, and a one-week quantified food frequency questionnaire. Foundation for the National Institutes of Health (FNIH) criteria (grip strength and ASM, adjusted for BMI) were used to classify sarcopenia. Participants with sarcopenia alongside a BMI of > 30.0 kg/m2 were classified as having sarcopenic obesity. Prevalence using other criteria (European Working Group on Sarcopenia in Older People, Asian Working Group for Sarcopenia and the International Working Group for Sarcopenia) were also explored. RESULTS: The prevalence of sarcopenia was 27.9%, which comprised of sarcopenia without obesity (3.3%) and sarcopenic obesity (24.6%). Other classification criteria showed that sarcopenia ranged from 0.8-14.7%, including 0.8-9.8% without obesity and 0-4.9% with sarcopenic obesity. Using multivariate-discriminant analysis (OPLS-DA) those with sarcopenic obesity presented with a descriptive profile of higher C-reactive protein, waist circumference, food security and sedentary time than women without sarcopenic obesity (p = 0.046). A similar profile described women with low BMI-adjusted grip strength (p < 0.001). CONCLUSIONS: The majority of women with sarcopenia were also obese (88%). We show a large discrepancy in the diagnostic criteria and the potential for significantly underestimating the prevalence of sarcopenia if BMI is not adjusted for. The main variables common to women with sarcopenic obesity were higher food security, lower PA and chronic inflammation. Our data highlights the importance of addressing obesity within these low-income communities to ensure the prevention of sarcopenic obesity and that quality of life is maintained with ageing.

Methods to Mimic In Vivo Muscle Cell Biology in Primary Human Myoblasts Using Quiescence as a Guidepost in Regenerative Medicine Research.

Regenerative medicine research and testing of new therapeutics for muscle-related human diseases call for a deeper understanding of how human myoblasts gain and maintain quiescence in vitro versus in vivo. The more closely we can experimentally simulate the in vivo environment, the more relevance in vitro research on myoblasts will have. In this context, isolation of satellite cells from muscle tissue causes activation while myoblasts remain activated in culture, thus not simulating quiescence as in their in vivo niche. Cells synchronized for cell cycle present a good starting point for experimental intervention. In the past, myoblast quiescence has been induced using suspension culture (SuCu) and, recently, by knockout serum replacement (KOSR)-supplemented culture media. We assessed the proportion of cells in G0 and molecular regulators after combining the two quiescence-inducing approaches. Quiescence was induced in primary human myoblasts (PHMs) in vitro using KOSR-treatment for 10 days or suspension in viscous media for 2 days (SuCu), or suspension combined with KOSR-treatment for 2 days (blended method, SuCu-KOSR). Quiescence and synchronization were achieved with all three protocols (G0/G1 cell cycle arrest >90% cells). Fold-change of cell cycle controller p21 mRNA for KOSR and SuCu was 3.23 ± 0.30 and 2.86 ± 0.15, respectively. Since this was already a significant change (p < 0.05), no further change was gained with the blended method. But SuCu-KOSR significantly decreased Ki67 (p = 0.0019). Myogenic regulatory factors, Myf5 and MyoD gene expression in PHMs were much more suppressed (p = 0.0004 and p = 0.0034, respectively) in SuCu-KOSR, compared to SuCu alone. In conclusion, a homogenous pool of quiescent primary myoblasts synchronized in the G0 cell cycle phase was achieved with cells from three different donors regardless of the experimental protocol. Myogenic dedifferentiation at the level of Myogenic Regulatory Factors was greater when exposed to the blend of suspension and serum-free culture. We suggest that this blended new protocol can be considered in future biomedical research if differentiation is detected too early during myoblast expansion. This shall also inform new ways to bridge the in vitro and in vivo divides in regenerative medicine research.

Co-culture of pro-inflammatory macrophages and myofibroblasts: Evaluating morphological phenotypes and screening the effects of signaling pathway inhibitors.

Skeletal muscle regeneration is a complex process influenced by non-myogenic macrophages and fibroblasts, which acquire different phenotypes in response to changes in the injury milieu or changes in experimental conditions. In vitro, serum stimulates the differentiation of fibroblasts into myofibroblasts, while lipopolysaccharide (LPS) stimulates the polarization of unstimulated (M0) macrophages to acquire an M1 pro-inflammatory phenotype. We characterized these phenotypes using morphology (with circularity as shape descriptor; perfect circularity = 1.0) and phenotype-specific markers. Myofibroblasts (high α-smooth muscle actin [SMA] expression) had high circularity (mean 0.60 ± 0.03). Their de-differentiation to fibroblasts (low α-SMA expression) significantly lessened circularity (0.47 ± 0.01 and 0.35 ± 0.02 in 2% or 0% serum culture media respectively (p < 0.05). Unstimulated (M0) macrophages (no CD86 expression) had high circularity (0.72 ± 0.02) which decreased when stimulated to M1 macrophages (CD86 expression) (LPS; 0.61 ± 0.02; p < 0.05). Utilizing these established conditions, we then co-cultured M1 macrophages with myofibroblasts or myoblasts. M1 macrophages significantly decreased relative myofibroblast numbers (from 223 ± 22% to 64 ± 7%), but not myoblast numbers. This pro-inflammatory co-culture model was used to rapidly screen the following four compounds for ability to prevent M1 macrophage-mediated decrease in myofibroblast numbers: L-NAME (inducible nitric oxide synthase inhibitor), SB203580 (p38 mitogen-activated protein kinase inhibitor), SP600125 (c-Jun N-terminal kinase inhibitor) and LY294002 (phosphoinositide 3-kinase [PI3K] inhibitor). We found that LY294002 rescued myofibroblasts and decreased macrophage numbers. Myofibroblast rescue did not occur with L-NAME, SB203580 or SP600125 incubation. In conclusion, these data suggest a PI3K-associated mechanism whereby myofibroblasts can be rescued, despite simulated pro-inflammatory conditions.

Dysregulated healing responses in diabetic wounds occur in the early stages postinjury.

Chronic wounds are a serious and debilitating complication of diabetes. A better understanding of the dysregulated healing responses following injury will provide insight into the optimal time frame for therapeutic intervention. In this study, a direct comparison was done between the healing dynamics and the proteome of acute and obese diabetic wounds on days 2 and 7 following injury. Full thickness excisional wounds were induced on obese diabetic (B6.Cg-lepob/J, ob/ob, n = 14) (blood glucose 423.25 ± 127.92 mg/dL) and WT control (C57BL/6J, n = 14) (blood glucose 186.67 ± 24.5 mg/dL) mice. Histological analysis showed no signs of healing in obese DM wounds whereas complete wound closure/re-epithelisation, the formation of granulation tissue and signs of re-vascularisation, was evident in acute wounds on day 7. In obese DM wounds, substance P deficiency and increased MMP-9 activity on day 2 coincided with increased cytokine/chemokine levels within wound fluid. LC-MS/MS identified 906 proteins, of which 23 (Actn3, Itga6, Epb41, Sncg, Nefm, Rsp18, Rsp19, Rpl22, Macroh2a1, Rpn1, Ppib, Snrnp70, Ddx5, Eif3g, Tpt1, FABP5, Cavin1, Stfa1, Stfa3, Cycs, Tkt, Mb, Chmp2a) were differentially expressed in wounded tissue on day 2 (P < 0.05; more than two-fold) and 6 (Cfd, Ptms, Hp, Hmga1, Cbx3, Syap1) (P < 0.05; more than two-fold) on day 7. A large number of dysregulated proteins on day 2 was associated with an inability to progress into the proliferative stage of healing and suggest that early intervention might be pivotal for effective healing outcomes. The proteomic approach highlighted the complexity of obese DM wounds in which the dysregulation involves multiple regulatory pathways and biological processes.

In vitro induction of quiescence in isolated primary human myoblasts.

Adult skeletal muscle stem cells, satellite cells, remain in an inactive or quiescent state in vivo under physiological conditions. Progression through the cell cycle, including activation of quiescent cells, is a tightly regulated process. Studies employing in vitro culture of satellite cells, primary human myoblasts (PHMs), necessitate isolation myoblasts from muscle biopsies. Further studies utilizing these cells should endeavour to represent their native in vivo characteristics as closely as possible, also considering variability between individual donors. This study demonstrates the approach of utilizing KnockOut™ Serum Replacement (KOSR)-supplemented culture media as a quiescence-induction media for 10 days in PHMs isolated and expanded from three different donors. Cell cycle analysis demonstrated that treatment resulted in an increase in G1 phase and decreased S phase proportions in all donors (p < 0.005). The proportions of cells in G1 and G2 phases differed in proliferating myoblasts when comparing donors (p < 0.05 to p < 0.005), but following KOSR treatment, the proportion of cells in G1 (p = 0.558), S (p = 0.606) and G2 phases (p = 0.884) were equivalent between donors. When cultured in the quiescence-induction media, expression of CD34 and Myf5 remained constant above > 98% over time from day 0 to day 10. In contrast activation (CD56), proliferation (Ki67) and myogenic marker MyoD decreased, indicated de-differentiation. Induction of quiescence was accompanied in all three clones by fold change in p21 mRNA greater than 3.5 and up to tenfold. After induction of quiescence, differentiation into myotubes was not affected. In conclusion, we describe a method to induce quiescence in PHMs from different donors.

Interleukin-6 Induces Myogenic Differentiation via JAK2-STAT3 Signaling in Mouse C2C12 Myoblast Cell Line and Primary Human Myoblasts.

Postnatal muscle growth and exercise- or injury-induced regeneration are facilitated by myoblasts. Myoblasts respond to a variety of proteins such as cytokines that activate various signaling cascades. Cytokines belonging to the interleukin 6 superfamily (IL-6) influence myoblasts' proliferation but their effect on differentiation is still being researched. The Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway is one of the key signaling pathways identified to be activated by IL-6. The aim of this study was to investigate myoblast fate as well as activation of JAK-STAT pathway at different physiologically relevant IL-6 concentrations (10 pg/mL; 100 pg/mL; 10 ng/mL) in the C2C12 mouse myoblast cell line and primary human myoblasts, isolated from eight young healthy male volunteers. Myoblasts' cell cycle progression, proliferation and differentiation in vitro were assessed. Low IL-6 concentrations facilitated cell cycle transition from the quiescence/Gap1 (G0/G1) to the synthesis (S-) phases. Low and medium IL-6 concentrations decreased the expression of myoblast determination protein 1 (MyoD) and myogenin and increased proliferating cell nuclear antigen (PCNA) expression. In contrast, high IL-6 concentration shifted a larger proportion of cells to the pro-differentiation G0/G1 phase of the cell cycle, substantiated by significant increases of both MyoD and myogenin expression and decreased PCNA expression. Low IL-6 concentration was responsible for prolonged JAK1 activation and increased suppressor of cytokine signaling 1 (SOCS1) protein expression. JAK-STAT inhibition abrogated IL-6-mediated C2C12 cell proliferation. In contrast, high IL-6 initially increased JAK1 activation but resulted in prolonged JAK2 activation and elevated SOCS3 protein expression. High IL-6 concentration decreased interleukin-6 receptor (IL-6R) expression 24 h after treatment whilst low IL-6 concentration increased IL-6 receptor (IL-6R) expression at the same time point. In conclusion, this study demonstrated that IL-6 has concentration- and time-dependent effects on both C2C12 mouse myoblasts and primary human myoblasts. Low IL-6 concentration induces proliferation whilst high IL-6 concentration induces differentiation. These effects are mediated by specific components of the JAK/STAT/SOCS pathway.

C-Reactive Protein Is Elevated Only in High Creatine Kinase Responders to Muscle Damaging Exercise.

The purpose of this study was to investigate if exertional rhabdomyolysis induced by an acute bout of plyometric exercise in untrained individuals was associated with histological characteristics of skeletal muscle, creatine kinase (CK) polymorphism or secondary damage. Twenty-six healthy male untrained individuals completed a bout of plyometric exercise (10 sets of 10 maximal squat jumps, with each standardized to achieve at least 95% of individual maximal jump height). Blood samples were taken, and perceived pain was scored immediately before the exercise intervention and 6 h, 1, 2, and 3 days post-intervention. Muscle biopsies were collected 9 or 4 days before (baseline) and 3 days after plyometric jumps. Subjects were divided into two groups, high (n = 10) and low responders (n = 16), based on a cut-off limit for exertional rhabdomyolysis of peak CK activity ≥ 1000 U/L in any post-exercise blood sample. Perceived pain was more severe assessed in squat than standing position. Low responders perceived more pain at 6 h and 1 day, while high responders perceived more pain than low responders on days three and four after exercise; structural (dystrophin staining) and ultra-structural (transmission electron microscopy) analysis of muscle fibers revealed no baseline pathology; damage was evident in all individuals in both groups, with no difference between high and low responders in either damage or fiber type proportion. High responders had significantly higher total white blood cell and neutrophil counts 6 h and significantly higher C-reactive protein (CRP) 6 h and days one and two after exercise compared to low responders. High responders had significantly greater muscle myeloperoxidase (MPO) levels in baseline and 3 day post-exercise biopsies compared to baseline of low responders. MLCK C49T single polymorphism was present in 26% of volunteers, whose CK responses were not higher than those with MLCK CC or CT genotype. In conclusion, perceived pain is more effectively assessed with potentially affected muscle under eccentric strain, even if static. High CK responders also have pronounced CRP responses to unaccustomed plyometric exercise intervention. Exertional rhabdomyolysis after unaccustomed eccentric exercise may be related to underlying inability to resolve intramuscular MPO.

Investigation of Circulating Extracellular Vesicle MicroRNA Following Two Consecutive Bouts of Muscle-Damaging Exercise.

Background: Extracellular vesicles (EVs) are nano-sized vesicles that are known to be powerful mediators of intercellular communication via their microRNA (miR) content. A paucity of information on EV-mediated communication arising from skeletal muscle (SkM) in response to exercise-induced muscle damage is present in the published literature. Lack of such information inhibits our understanding of muscle injury and repair processes. Aims: To assess circulating EV levels and selected miR content within them, in response to two consecutive bouts of muscle-damaging exercise. Methods: Serum creatine kinase activity (CK) and EVs were analyzed from the blood of 9 healthy, untrained males at baseline, and at 2 and 24 h post-exercise. The exercise regimen consisted of a combination of plyometric jumping and downhill running. Perceived muscle pain (PMP) was assessed on a scale from 1 to 10. Plasma EVs were isolated using size exclusion columns and visualized with transmission electron microscopy (TEM). EV size and number were quantified using nanoparticle tracking analysis (NTA). miR expression was quantified using qPCR, with normalization to an exogenous control (cel-miR-39). Results: PMP and CK were significantly elevated post-exercise compared to baseline levels, providing indirect evidence for muscle damage. EV visualization using TEM revealed an abundant and heterogeneously sized pool of intact particles within the exosome size range (30-150 nm). No significant change in mean EV size or number was seen over time. The SkM-specific miR-206 in EVs was found to be variable among participants and no significant change occurred in SkM-important miRs; 1, 133a, 133b, 486, and 499a. However, EV miR-31 decreased from baseline to 24 h post-exercise (p = 0.027). Conclusion: Mild to moderate exercise-induced muscle damage altered the miR-31 profile of circulating EVs within the first 24 h post-exercise, but not that of myomiRs in EVs. These data demonstrate that EVs carry selectively packaged cargo which can be affected by exercise. Future research into the total miR content of EVs in response to exercise-induced muscle damage may reveal other miRs responsive to this relatively mild perturbation. More time points post-muscle-damaging exercise would provide a better understanding of the temporal EV myomiR response.

Muscle satellite cells increase during hibernation in ground squirrels.

UNLABELLED: Skeletal muscle satellite cells (SCs) are involved in muscle growth and repair. However, clarification of their behavior in hibernating mammals is lacking. The aim of this study was to quantify SCs and total myonuclei in hibernator muscle during different phases of the torpor-arousal cycle. Skeletal muscle was collected from thirteen-lined ground squirrels, Ictidomys tridecemlineatus, at five timepoints during hibernation: control euthermic [CON, stable body temperature (Tb)], early torpor (ET, within 24h), late torpor (LT, 5+ consecutive days), early arousal (EA, increased respiratory rate >60 breaths/min, Tb 9-12°C) and interbout arousal (IA, euthermic Tb). Protein levels of p21, Myf5, Wnt4, and ß-catenin were determined by western blotting. SCs (Pax7(+)) and myonuclei were identified using immunohistochemistry. Over the torpor-arousal cycle, myonuclei/fiber remained unchanged. However, the percentage of SCs increased significantly during ET (7.35±1.04% vs. CONTROL: 4.18±0.58%; p<0.05) and returned to control levels during LT. This coincided with a 224% increase in p21 protein during ET. Protein levels of Wnt4 did not change throughout, whereas Myf5 was lower during EA (p<0.08) and IA (p<0.05). Compared to torpor, ß-catenin increased by 247% and 279% during EA and IA, respectively (p<0.05). In conclusion, SCs were not dormant during hibernation and increased numbers of SC during ET corresponded with elevated amounts of p21 suggesting that cell cycle control may explain the SC return to baseline levels during late torpor. Despite relatively low Tb during early arousal, active control of quiescence by Myf5 is reduced.

Polyphenol supplementation: benefits for exercise performance or oxidative stress?

Supplement use among athletes is widespread, including non-traditional and biological compounds. Despite increasing research, a comprehensive and critical review on polyphenol supplementation and exercise is still lacking. This review is relevant for researchers directly involved in the topic, as well as those with a broad interest in athletic performance enhancement and sports nutrition. The purpose of this review is to present background information on groups of polyphenols and their derivatives because their differing chemical structures influence mechanisms of action; to discuss the potential of plant, fruit and vegetable-based biological supplements, high in polyphenol content, to affect exercise performance and biomarkers of oxidative stress and exercise-induced muscle damage; and to critically discuss the exercise studies and biomarkers used. Subjects in the studies reviewed were either sedentary, healthy individuals, or active, recreationally trained or well-trained athletes. Polyphenol supplementation in exercise studies included mainly extracts (multicomponent or purified), juices, infusions or an increased intake of polyphenol-rich foods. This review includes details of supplement doses and exercise test protocols. Many studies considered only the performance or one or two selected biomarkers of antioxidant capacity instead of a comprehensive choice of biomarkers to assess damage to lipids or proteins. Evidence is insufficient to make recommendations for or against the use of polyphenol supplementation (neither specific polyphenols nor specific doses) for either recreational, competitive or elite athletes. Polyphenols have multiple biological effects, and future exercise studies must be designed appropriately and specifically to determine physiological interactions between exercise and the selected supplement, rather than considering performance alone.

Skeletal muscle wasting with disuse atrophy is multi-dimensional: the response and interaction of myonuclei, satellite cells and signaling pathways.

Maintenance of skeletal muscle is essential for health and survival. There are marked losses of skeletal muscle mass as well as strength and physiological function under conditions of low mechanical load, such as space flight, as well as ground based models such as bed rest, immobilization, disuse, and various animal models. Disuse atrophy is caused by mechanical unloading of muscle and this leads to reduced muscle mass without fiber attrition. Skeletal muscle stem cells (satellite cells) and myonuclei are integrally involved in skeletal muscle responses to environmental changes that induce atrophy. Myonuclear domain size is influenced differently in fast and slow twitch muscle, but also by different models of muscle wasting, a factor that is not yet understood. Although the myonuclear domain is 3-dimensional this is rarely considered. Apoptosis as a mechanism for myonuclear loss with atrophy is controversial, whereas cell death of satellite cells has not been considered. Molecular signals such as myostatin/SMAD pathway, MAFbx, and MuRF1 E3 ligases of the ubiquitin proteasome pathway and IGF1-AKT-mTOR pathway are 3 distinctly different contributors to skeletal muscle protein adaptation to disuse. Molecular signaling pathways activated in muscle fibers by disuse are rarely considered within satellite cells themselves despite similar exposure to unloading or low mechanical load. These molecular pathways interact with each other during atrophy and also when various interventions are applied that could alleviate atrophy. Re-applying mechanical load is an obvious method to restore muscle mass, however how nutrient supplementation (e.g., amino acids) may further enhance recovery (or reduce atrophy despite unloading or ageing) is currently of great interest. Satellite cells are particularly responsive to myostatin and to growth factors. Recently, the hibernating squirrel has been identified as an innovative model to study resistance to atrophy.

Contusion injury with chronic in vivo polyphenol supplementation: leukocyte responses.

INTRODUCTION: In vivo, daily proanthocyanidolic oligomer (PCO) supplementation before and after experimental skeletal muscle contusion injury has been shown to result in a blunted neutrophil response in tissue, quicker macrophage infiltration into muscle, and faster recovery due to a left shift in time course of inflammation. The current study investigated effects of PCO on circulatory neutrophils and macrophage subpopulations as well as in vitro neutrophil migration. METHODS: Primary cultured neutrophils obtained from control animals were incubated in media with 20% conditioned plasma. To obtain conditioned media, male Wistar rats were supplemented with PCO (20 mg·kg(-1)d(-1)) or placebo (PLA) for 2 wk before a mass-drop contusion injury. Conditioned plasma was prepared from blood collected at different time points after injury (12 h, 1 d, 3 d, and 5 d). Macrophage subpopulation distribution, inflammatory cytokine, and myeloperoxidase levels were assessed for all time points. RESULTS: On day 1 postinjury, circulating neutrophil numbers were significantly lower in PLA than PCO, suggesting that extravasation from the blood was reduced by PCO. Concurrently, neutrophil migration in vitro was blunted in the presence of conditioned plasma from PCO supplemented rats compared with PLA supplemented rats. Plasma M1 and M2c macrophage numbers differed over time and between groups. M1 macrophage numbers peaked on day 3 with PCO supplementation, followed by a rise in M2c macrophages on day 5, when M1 macrophages numbers were still high in PLA. CONCLUSIONS: We conclude that PCO supplementation limits neutrophil migration capacity in vitro despite a chemotactic gradient. Furthermore, the earlier appearance of type M2 macrophages suggests a switch to an anti-inflammatory phenotype after injury even in circulation.