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What is the central question of this study? Is 1 week of exercise training sufficient to reduce local and systemic inflammation? Do obesity and short-term concurrent aerobic and resistance exercise training alter skeletal muscle extracellular vesicle (EV) contents? What is the main finding and its importance? Obesity alters skeletal muscle small EV microRNAs targeting inflammatory and growth pathways. Exercise training alters skeletal muscle small EV microRNAs targeting inflammatory pathways, indicative of reduced inflammation. Our findings provide support for the hypotheses that EVs play a vital role in intercellular communication during health and disease and that EVs mediate many of the beneficial effects of exercise.Obesity is associated with chronic inflammation characterized by increased levels of inflammatory cytokines, whereas exercise training reduces inflammation. Small extracellular vesicles (EVs; 30-150 nm) participate in cell-to-cell communication in part through microRNA (miRNA) post-transcriptional regulation of mRNA. We examined whether obesity and concurrent aerobic and resistance exercise training alter skeletal muscle EV miRNA content and inflammatory signalling. Vastus lateralis biopsies were obtained from sedentary individuals with (OB) and without obesity (LN). Before and after 7 days of concurrent aerobic and resistance training, muscle-derived small EV miRNAs and whole-muscle mRNAs were measured. Pathway analysis revealed that obesity alters small EV miRNAs that target inflammatory (SERPINF1, death receptor and Gα

Obesity is associated with numerous changes in skeletal muscle including greater muscle mass and muscle fiber cross sectional area (FCSA), yet fasted muscle protein synthesis is lower. Activation of the IGF-1/Akt/mTOR pathway is critical for muscle mass maintenance, muscle hypertrophy, and muscle protein regulation. Resistance exercise (RE) increases muscle mass, FCSA, and IGF-1. Persons with obesity have greater skeletal muscle mass and larger skeletal muscle fiber cross sectional area. The IGF-1/Akt/mTOR pathway is critical for the regulation of skeletal muscle mass. Our study found men and women with obesity have lower skeletal muscle IGF-1 mRNA and protein and higher expression of miR-206 an epigenetic regulator of IGF-1, at rest and following an acute bout of resistance exercise. Despite this, Akt mediated signaling was maintained and maintenance of phosphorylation does not appear to be accounted for by compensatory pathways. Our findings suggest a possible negative feedback mechanism via increased miR-206 and in turn decreased IGF-1 to limit further skeletal muscle hypertrophy in persons with obesity. The current work investigated if: (1) obesity dysregulates basal skeletal muscle IGF-1 pathways; and (2) obesity augments the muscle IGF-1 pathway responses to acute RE. Eight sedentary (no self-reported physical activity), lean (LN) and eight sedentary subjects with obesity (OB) had vastus lateralis biopsies taken at rest, and 15 min and 3 h post-acute RE for the measurement of the IGF-1 pathway and muscle FCSA. Type II FCSA was larger in OB vs. LN. Skeletal muscle IGF-1 mRNA and IGF-1 protein were lower in OB vs. LN at rest and post-exercise. Acute RE increased IGF-1 protein similarly in both groups. The expression of miR-206, a post-transcriptional inhibitor of IGF-1 expression, was higher in OB vs. LN and linked with lower IGF-1 mRNA (r = – 0.54). In spite of greater muscle FCSA, muscle IGF-1 expression was lower in obesity suggesting negative feedback may be limiting muscle mass expansion in obesity.

New findings What is the central question of this study? What is the impact of acute aerobic and aerobic + resistance (concurrent) exercise on the regulation of multivesicular body formation in human skeletal muscle? What is the main finding and its importance? Gene expression for proteins associated with multivesicular body biogenesis was increased in response to concurrent exercise, and gene expression of microRNA processing (genetic information) was increased in response to aerobic and concurrent exercise. A greater understanding of the processing of multivesicular bodies in response to acute exercise may lead to novel treatments focused on intercellular communication pathways. Abstract Regular aerobic exercise (AEx) and resistance exercise (REx) promote many beneficial adaptations. Skeletal muscle participates in intercellular communication in part through the release of myokines and extracellular vesicles including exosomes (EXOs), the latter containing mRNA, microRNA (miRNA), lipids and proteins. Exercise-induced regulation of skeletal muscle multivesicular body (MVB) biogenesis leading to EXO formation and release is poorly understood. We hypothesized that acute exercise would increase skeletal muscle MVB biogenesis and EXO release pathways with a greater response to aerobic + resistance exercise (A+REx) than to AEx alone. Twelve sedentary, healthy male subjects exercised on a cycle ergometer for 45 min (AEx) followed by single leg, knee extensor, resistance exercise (A+REx). Vastus lateralis biopsies were obtained at rest and 1 h post-exercise. Key components of the MVB biogenesis, EXO biogenesis and release, and miRNA processing pathways were analysed. Clathrin and Alix mRNA (MVB biogenesis) were increased by A+REx, while DICER and exportin mRNA (miRNA processing) were increased by AEx and A+REx. There were positive relationships between MVBs and miRNA processing genes following both AEx and A+REx consistent with coordinated regulation of these interrelated processes (Alix mRNA increased with Drosha, exportin and Dicer mRNA). Acute exercise increases the regulation of components of MVB and EXO pathways as well as miRNA processing components. A greater understanding of the production and packaging of skeletal muscle MVBs, EXOs and mature miRNA could lead to novel treatments focused on intercellular communication.

What is the central question of this study? Do obesity and acute resistance exercise alter the regulation of muscle intercellular communication pathways consistent with inadequate compensatory angiogenesis in response to muscle loading present in individuals with obesity? What is the main finding and its importance? Obesity is associated with differences in both pro- and anti-angiogenic signalling consistent with lower muscle capillarization. Acute resistance exercise increases the release of skeletal muscle small extracellular vesicles independent of body mass. These results identify new cellular factors associated with impaired angiogenesis in obesity and the positive effects of acute resistance exercise in lean and obese skeletal muscle.Obesity (OB) impairs cell-to-cell communication signalling. Small extracellular vesicles (EVs), which include exosomes, are released by skeletal muscle and participate in cell-to-cell communication, including the regulation of angiogenesis. Resistance exercise (REx) increases muscle fibre size and capillarization. Although obesity increases muscle fibre size, there is an inadequate increase in capillarization such that capillary density is reduced. It was hypothesized that REx-induced angiogenic signalling and EV biogenesis would be lower with obesity. Sedentary lean (LN) and OB subjects (n = 8 per group) performed three sets of single-leg knee-extension REx at 80% of maximum. Muscle biopsies were obtained at rest, 15 min and 3 h postexercise and analysed for angiogenic and EV biogenesis mRNA and protein. In OB subjects, muscle fibre size was ∼20% greater and capillary density with type II fibres ∼25% lower compared with LN subjects (P 0.001). In response to REx, the increase in VEGF mRNA (pro-angiogenic) was similar (3-fold) between groups, while thrombospondin-1 (TSP-1) mRNA (anti-angiogenic) increased ∼2.5-fold in OB subjects only (P = 0.010). miR-130a (pro-angiogenic) was ∼1.4-fold (P = 0.011) and miR-503 (anti-angiogenic) ∼1.8-fold (P = 0.017) greater in OB compared with LN subjects at all time points. In both groups, acute REx decreased the EV surface protein Alix by ∼50%, consistent with the release of exosomes (P = 0.016). Acute REx appears to induce the release of skeletal muscle small EVs independent of body mass. However, with obesity there is predominantly impaired angiogenic signalling, consistent with inadequate angiogenesis in response to basal muscle hypertrophy.

New findings What is the central question of this study? Capillary rarefaction is found in diabetic and aged muscle, whereas exercise increases skeletal muscle angiogenesis. The association implies a crosstalk between muscle cells and endothelial cells. The underlying mechanisms mediating the crosstalk between these cells remains to be elucidated fully. What is the main finding and its importance? Endothelial cell functions are regulated by skeletal muscle cell-derived exosomes via a vascular endothelial growth factor-independent pathway. This study reveals a new mechanism mediating the crosstalk between skeletal muscle cells and endothelial cells. Abstract Loss of skeletal muscle capillarization, known as capillary rarefaction, is found in type 2 diabetes, chronic heart failure and healthy ageing and is associated with impaired delivery of substrates to the muscle. However, the interaction and communication of skeletal muscle with endothelial cells in the regulation of capillaries surrounding the muscle remains elusive. Exosomes are a type of secreted extracellular vesicle containing mRNAs, proteins and, especially, microRNAs that exert paracrine and endocrine effects. In this study, we investigated whether skeletal muscle-derived exosomes (SkM-Exo) regulate the endothelial cell functions of angiogenesis. We demonstrated that C2C12 myotube-derived exosomes improved endothelial cell functions, assessed by the proliferation, migration and tube formation of human umbilical vein endothelial cells (HUVECs), which were increased by 20, 23 and 40%, respectively, after SkM-Exo exposure. The SkM-Exo failed to activate HUVEC vascular endothelial growth factor (VEGF) signalling. The SkM-Exo increased HUVEC reactive oxygen species and activated the nuclear factor-κB pathway, suggesting that SkM-Exo-induced angiogenesis was mediated by a VEGF-independent pathway. In addition, several angiogenic microRNAs were packaged in SkM-Exo, with miR-130a being particularly enriched and successfully transferred from SkM-Exo to HUVECs. Delivery of miRNAs into endothelial cells might explain the enhancement of reactive oxygen species production and angiogenesis by SkM-Exo. The potential angiogenic effect of SkM-Exo could provide an effective therapy for promoting skeletal muscle angiogenesis in diseases characterized by capillary rarefaction or inadequate angiogenesis.

Skeletal muscles contain heterogeneous myofibers that are different in size and contractile speed, with type IIb myofiber being the largest and fastest. Here, we identify methyltransferase-like 21e (Mettl21e), a member of newly classified nonhistone methyltransferases, as a gene enriched in type IIb myofibers. The expression of Mettl21e was strikingly up-regulated in hypertrophic muscles and during myogenic differentiation in vitro and in vivo. Knockdown (KD) of Mettl21e led to atrophy of cultured myotubes, and targeted mutation of Mettl21e in mice reduced the size of IIb myofibers without affecting the composition of myofiber types. Mass spectrometry and methyltransferase assay revealed that Mettl21e methylated valosin-containing protein (Vcp/p97), a key component of the ubiquitin-proteasome system. KD or knockout of Mettl21e resulted in elevated 26S proteasome activity, and inhibition of proteasome activity prevented atrophy of Mettl21e KD myotubes. These results demonstrate that Mettl21e functions to maintain myofiber size through inhibiting proteasome-mediated protein degradation.—Wang, C., Zhang, B., Ratliff, A. C., Arrington, J., Chen, J., Xiong, Y., Yue, F., Nie, Y., Hu, K., Jin, W., Tao, W. A., Hrycyna, C. A., Sun, X., Kuang, S. Methyltransferase-like 21e inhibits 26S proteasome activity to facilitate hypertrophy of type IIb myofibers.

Acute exercise can increase skeletal muscle citrate synthase (CS) enzyme activity and resting skeletal muscle mitochondrial enzyme activity has been linked to maximal oxygen consumption ( $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ ). We investigated: (1) if acute aerobic exercise (AEx) increases muscle metabolic enzyme activities other than CS; (2) if the addition of acute resistance exercise (REx) enhances the response to AEx (A + REx); and (3) if post-exercise muscle metabolic enzyme activity was related to $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ . Twelve young, sedentary men completed 45 min of two-legged cycle ergometry at 55% of $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ and 3 sets of 8–12 repetitions of one-leg knee extensor at 55% of 1 repetition maximum (1-RM). Vastus lateralis biopsies were taken prior to and 1 h post AEx and A + REx for the measurement of phosphofructokinase (PFK), 3-l-hydroxyacyl CoA dehydrogenase (β-HAD), succinate dehydrogenase (SDH) and CS. As a group, there was no effect of acute AEx or A + REx on muscle PFK, β-HAD, CS, and SDH activities. Post exercise muscle PFK, β-HAD, CS, and SDH activities were related to higher $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ (r = 0.62–0.74). With participants grouped by $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ (LOW, 50th %), acute exercise-induced changes in muscle PFK, β-HAD, CS, and SDH were greater in NORM compared to LOW. These findings suggest acute exercise muscle metabolic enzyme activities are predictive of $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ and possibly supportive of higher $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ . Also, low $$\dot {V}{{\text{O}}_{2\hbox{max} }}$$ (below 30th percentile) appears to impair skeletal muscle metabolic enzyme responses to acute exercise.

Flame brush thickness is a significant parameter that demonstrates the evolution of turbulent premixed flame. In this paper, the flame height and flame brush thickness of lean turbulent premixed Bunsen CH4/air and C3H8/air flames were investigated under low turbulence intensities. The turbulence flow field is generated and controlled by changing the outlet velocity and perforated plates, and it is measured using a hot wire anemometry system. The turbulent flame fronts are detected by OH-PLIF technique. Results show that under fuel-lean conditions, the characteristic turbulent Bunsen flame height and centerline flame brush thickness decrease with increasing equivalence ratio, while they increase with the augment of outlet velocity. Turbulence intensity has marginal effect on these two parameters. The variations of flame brush thickness indicate that there are three regions during the whole development of turbulent Bunsen flames under low turbulence intensity. The first is the turbulence dominating region, where the variations of horizontal flame brush thickness follow the turbulence diffusion theory. The second is the non-local effect manifesting region, where the equivalence ratio, which controls the flame instability wavelength, influences the horizontal flame brush thickness. The third is the flame tip region where the flamelets intersect and make the radial wrinkles offset while the axial wrinkles superimpose. This unique tip region of turbulent Bunsen flame can be demonstrated by the linear relationship between its flame height and centerline flame brush thickness. Turbulent premixed Bunsen flames suffer different non-local effects during the flame development to downstream. To minimize the non-local effect on the statistics of turbulent flame front structure, a new method based on the development time period is proposed. Compared with statistical analysis based on the whole development time, this method can better demonstrate the flame-turbulence interaction.

Adult stem cells maintain regenerative tissue structure and function by producing tissue-specific progeny, but the factors that preserve their tissue identities are not well understood. The small and large intestines differ markedly in cell composition and function, reflecting their distinct stem cell populations. Here we show that SATB2, a colon-restricted chromatin factor, singularly preserves LGR5+ adult colonic stem cell and epithelial identity in mice and humans. Satb2 loss in adult mice leads to stable conversion of colonic stem cells into small intestine ileal-like stem cells and replacement of the colonic mucosa with one that resembles the ileum. Conversely, SATB2 confers colonic properties on the mouse ileum. Human colonic organoids also adopt ileal characteristics upon SATB2 loss. SATB2 regulates colonic identity in part by modulating enhancer binding of the intestinal transcription factors CDX2 and HNF4A. Our study uncovers a conserved core regulator of colonic stem cells able to mediate cross-tissue plasticity in mature intestines.

The characteristics of local extinction on the tip of syngas premixed Bunsen flames with different hydrogen fractions have been investigated experimentally with OH-PLIF technique and numerical simulation. Coupled effects of stretch and diffusion on the syngas Bunsen flame with highly negative curvature were studied. Local Karlovitz number was introduced to indicate the onset of local extinction at the Bunsen flame tip. Results showed that the onset equivelance ratio of local extinction at the syngas Bunsen flame tip does not change with the outlet velocity. The coupled effects of stretch and diffusion result in the decrease of flame reaction rate along the syngas Bunsen flame at fuel-lean condition. When the flame reaction rate decrease to a critical extent, local extinction of Bunsen flame tip occures. The local Karlovitz number can be considered as a parameter of the critical condition. The syngas Bunsen flame tip will extinguish when the local Karlovitz number reaches around 4, irrespective of hydrogen fractions and outlet velocity. This means that Bunsen flame tip starts to appear local extinction when the tangential flow time is comparable to normal chemical reaction time, which is the same as counterflow flames with positive stretch. Furthermore, results also showed that there is no H2 leakage even the syngas Bunsen flame tip extinguishes at the numerical study conditions.

Stress responses promote obesity and insulin resistance, in part, by activating the stress-responsive mitogen-activated protein kinases (MAPKs), p38 MAPK, and c-Jun NH2-terminal kinase (JNK). Stress also induces expression of MAPK phosphatase-1 (MKP-1), which inactivates both JNK and p38 MAPK. However, the equilibrium between JNK/p38 MAPK and MKP-1 signaling in the development of obesity and insulin resistance is unclear. Skeletal muscle is a major tissue involved in energy expenditure and glucose metabolism. In skeletal muscle, MKP-1 is upregulated in high-fat diet–fed mice and in skeletal muscle of obese humans. Mice lacking skeletal muscle expression of MKP-1 (MKP1-MKO) showed increased skeletal muscle p38 MAPK and JNK activities and were resistant to the development of diet-induced obesity. MKP1-MKO mice exhibited increased whole-body energy expenditure that was associated with elevated levels of myofiber-associated mitochondrial oxygen consumption. miR-21, a negative regulator of PTEN expression, was upregulated in skeletal muscle of MKP1-MKO mice, resulting in increased Akt activity consistent with enhanced insulin sensitivity. Our results demonstrate that skeletal muscle MKP-1 represents a critical signaling node through which inactivation of the p38 MAPK/JNK module promotes obesity and insulin resistance.

Sucrose nonfermenting-related kinase (SNRK) is a member of the AMPK-related kinase family, and its physiological role in adipose energy homeostasis and inflammation remains unknown. We previously reported that SNRK is ubiquitously and abundantly expressed in both white adipose tissue (WAT) and brown adipose tissue (BAT), but SNRK expression diminishes in adipose tissue in obesity. In this study we report novel experimental findings from both animal models and human genetics. SNRK is essential for survival; SNRK globally deficient pups die within 24 h after birth. Heterozygous mice are characterized by inflamed WAT and less BAT. Adipocyte-specific ablation of SNRK causes inflammation in WAT, ectopic lipid deposition in liver and muscle, and impaired adaptive thermogenesis in BAT. These metabolic disorders subsequently lead to decreased energy expenditure, higher body weight, and insulin resistance. We further confirm the significant association of common variants of the SNRK gene with obesity risk in humans. Through applying a phosphoproteomic approach, we identified eukaryotic elongation factor 1δ and histone deacetylase 1/2 as potential SNRK substrates. Taking these data together, we conclude that SNRK represses WAT inflammation and is essential to maintain BAT thermogenesis, making it a novel therapeutic target for treating obesity and associated metabolic disorders.

For better control of a lean premixed combustion system, it is necessary to understand the mechanism of unsteady cellular flame. The objective of the present study is to investigate how heat loss influences the cellular instability of lean premixed flame. The lean premixed CH4/H2/air cellular flames generated on a McKenna burner were studied, and heat loss was responsible for the manifestation of unsteady cellular structure. A method of applying proper orthogonal decomposition in scale analysis is presented, which has proven helpful in characterizing the cellular flame. The multiple scale nature of unstable cellular flame was identified, and the small scales were found to act in the random movement of cells. The contribution of each scale is obtained. The energy distribution on modes indicates that the energy on higher-order modes, equivalently the contribution of smaller scales, is decreased with decreasing heat loss. Through analyzing the principal modes and temporal coefficients, heat loss is f...

Brown adipose tissue (BAT) represents a promising agent to ameliorate obesity and other metabolic disorders. However, the abundance of BAT decreases with age and BAT paucity is a common feature of obese subjects. As brown adipocytes and myoblasts share a common Myf5 lineage origin, elucidating the molecular mechanisms underlying the fate choices of brown adipocytes versus myoblasts may lead to novel approaches to expand BAT mass. Here we identify MyoD as a key negative regulator of brown adipocyte development. CRISPR/CAS9-mediated deletion of MyoD in C2C12 myoblasts facilitates their adipogenic transdifferentiation. MyoD knockout downregulates miR-133 and upregulates the miR-133 target Igf1r, leading to amplification of PI3K–Akt signaling. Accordingly, inhibition of PI3K or Akt abolishes the adipogenic gene expression of MyoD null myoblasts. Strikingly, loss of MyoD converts satellite cell-derived primary myoblasts to brown adipocytes through upregulation of Prdm16, a target of miR-133 and key determinant of brown adipocyte fate. Conversely, forced expression of MyoD in brown preadipocytes blocks brown adipogenesis and upregulates the expression of myogenic genes. Importantly, miR-133a knockout significantly blunts the inhibitory effect of MyoD on brown adipogenesis. Our results establish MyoD as a negative regulator of brown adipocyte development by upregulating miR-133 to suppress Akt signaling and Prdm16., Highlights • Loss of MyoD facilitates adipogenic transdifferentiation of myoblasts. • Overexpression of MyoD transdifferentiate brown preadipocytes to myoblasts. • MyoD acts partially through miR-133 to suppress brown adipocyte cell fate. Brown fat burns fat to produce heat, and represents a promising agent to treat obesity and its related disorders. Brown fat cells and muscle cells share a common origin, but what controls the developmental separation of the two cell types is not well understood. This study reports that inhibition of “MyoD” gene in muscle progenitors promotes their differentiation into brown fat cells in mice. Conversely, forced expression of MyoD in brown fat progenitors converts them into muscle cells. This work suggests that inhibition of MyoD may represent a future direction to expand brown fat and alleviate obesity in humans.

Muscle development and regeneration require delicate cell cycle regulation of embryonic myoblasts and adult muscle satellite cells (MuSCs). Through analysis of the Polo-like kinase (Plk) family cell-cycle regulators in mice, we show that Plk1’s expression closely mirrors myoblast dynamics during embryonic and postnatal myogenesis. Cell-specific deletion of Plk1 in embryonic myoblasts leads to depletion of myoblasts, developmental failure and prenatal lethality. Postnatal deletion of Plk1 in MuSCs does not perturb their quiescence but depletes activated MuSCs as they enter the cell cycle, leading to regenerative failure. The Plk1-null MuSCs are arrested at the M-phase, accumulate DNA damage, and apoptose. Mechanistically, Plk1 deletion upregulates p53, and inhibition of p53 promotes survival of the Plk1-null myoblasts. Pharmacological inhibition of Plk1 similarly inhibits proliferation but promotes differentiation of myoblasts in vitro, and blocks muscle regeneration in vivo. These results reveal for the first time an indispensable role of Plk1 in developmental and regenerative myogenesis., eLife digest Muscles have their own population of stem cells, called muscle satellite cells. These cells are essential for muscle growth and repair. In healthy adult muscles, they spend most of their time inactive, but when there is an injury, they reawaken and start dividing. Some of the new cells return to an inactive stem cell state to await the next injury. The rest mature into new muscle cells or join with damaged muscle fibres to help them repair. The cell cycle is the series of events that a cell goes through from its birth until it divides. In muscle satellite cells, progression through the cell cycle is tightly controlled to ensure they divide and grow the correct amount. One of the proteins responsible for controlling the cell cycle is Polo-Like Kinase 1 (PLK1), but studying this protein is difficult. A common way to investigate a protein's effect is to delete the gene that makes it and observe the consequences. However, PLK1 is so essential to life that yeast, flies, zebrafish and mice all die when the gene is missing. Jia et al. deleted the gene that makes PLK1 only in mouse muscle satellite cells to find out the role this protein plays in controlling the cell cycle in stem cells. Deleting the gene that codes for PLK1 before the mice were born was lethal. The embryos failed to develop mature muscle fibres, and they died. But deleting the gene after the mice were born had a different effect. The muscles developed normally, but they were unable to heal when injured. The same healing problem also happened when healthy mice received a drug that blocked the function of PLK1 protein. A closer look at the muscle satellite cells revealed the source of the problem. Without PLK1, the cells got stuck part way through their cell cycle, just before they were due to divide. They tried to become muscle cells, but they did not make it. Instead, the muscle satellite cells started to act as though their DNA had been damaged, and then they self-destructed. Muscle satellite cells become less able to divide as we get older. They can also malfunction in some types of degenerative muscle diseases. Understanding how muscle satellite cells control their cell cycle could help us to find out what causes them to go wrong. Further work to understand PLK1 also has potential implications for cancer treatment. PLK1 blockers have been used to stop cancer cells from dividing, but Jia et al.’s findings show that this kind of drug may also hamper the ability of muscle to repair damage.

Key points Cellular communication occurs between endothelial cells and skeletal muscle satellite cells and is mitogenic for both cell types under normal conditions. Skeletal muscle atrophy and endothelial cell dysfunction occur in tandem in cardiovascular disease, type II diabetes and ageing. The present study investigated how induction of endothelial cell dysfunction via high glucose treatment impacts growth and differentiation of human skeletal muscle satellite cells in vitro. Secreted factors from high glucose treated endothelial cells impaired satellite cell expansion and differentiation via decreased proliferation and dysregulation of p38 mitogen-activated protein kinase in satellite cells committed to myogenesis. These findings highlight a novel potential role for endothelial cells in the development and pathology of skeletal muscle atrophy, which is common in patients with endothelial dysfunction related pathologies. Abstract Cross-talk between endothelial cells (ECs) and skeletal muscle satellite cells (MuSC) has been identified as an important regulator of cellular functions in both cell types. In healthy conditions, EC secreted factors promote MuSC growth and differentiation. Endothelial and satellite cell dysfunction occur in tandem in many disease states; however, no data exist examining the impact of dysfunctional EC signalling on satellite cells. Therefore, the present study aimed to evaluate the effect that factors secreted from high glucose (HG) treated ECs have on the growth and differentiation of human satellite cells (HMuSC) using a conditioned medium (CM) cell culture model. Satellite cells were isolated from human skeletal muscle and grown in CM from normal or HG treated human umbilical vein ECs (HUVECs). Satellite cells grown in CM from HG treated HUVECs reduced growth (25%), differentiation (25%) and myonuclear fusion (35%). These responses were associated with increased superoxide (50%) and inflammatory cytokines (25-50%) in HG treated HUVECs and HG-CM. Decreased expansion of HG-CM treated HMuSCs was driven by a decrease in proliferation. Impaired gene expression and protein content of myogenic differentiation factors were preceded by decreased phosphorylation of p38 mitogen-activated protein kinase in HMuSC treated with CM from HG treated HUVECs. The results obtained in the present study are the first to show that factors secreted from HG treated ECs cause impairments in human muscle satellite cell growth and differentiation in vitro, highlighting endothelial cell health and secretion as a potential target for treating vascular disease-associated skeletal muscle dysfunction.

Exercise promotes multiple beneficial effects on muscle function, including induction of mitochondrial biogenesis. miR-133a is a muscle-enriched microRNA that regulates muscle development and function. The role of miR-133a in exercise tolerance has not been fully elucidated. In the current study, mice that were deficient in miR-133a demonstrated low maximal exercise capacity and low resting metabolic rate. Transcription of the mitochondrial biogenesis regulators peroxisome proliferator-activated receptor-γ coactivator 1-α, peroxisome proliferator-activated receptor-γ coactivator 1-β, nuclear respiratory factor-1, and transcription factor A, mitochondrial were lower in miR-133a–deficient muscle, which was consistent with lower mitochondrial mass and impaired exercise capacity. Six weeks of endurance exercise training increased the transcriptional level of miR-133a and stimulated mitochondrial biogenesis in wild-type mice, but failed to improve mitochondrial function in miR-133a–deficient mice. Further mechanistic analysis showed an increase in the miR-133a potential target, IGF-1 receptor, along with hyperactivation of Akt signaling, in miR-133a–deficient mice, which was consistent with lower transcription of the mitochondrial biogenesis regulators. These findings indicate an essential role of miR-133a in skeletal muscle mitochondrial biogenesis, exercise tolerance, and response to exercise training.—Nie, Y., Sato, Y., Wang, C., Yue, F., Kuang, S., Gavin, T. P. Impaired exercise tolerance, mitochondrial biogenesis, and muscle fiber maintenance in miR-133a–deficient mice.

An experimental study on the cellular instability of spherical propagation syngas flames with H2O addition was performed in a constant volume combustion bomb over a wide range of CO/H2 ratios at elevated pressures. Schlieren images were recorded to observe and analyze the flame front evolution of spherical propagation flames. Results show that the cellular instability of syngas flames is promoted at atmospheric pressure and suppressed at elevated pressure with H2O addition for syngas mixtures with CO/H2 ratio of 80/20. At elevated pressure, critical radius, beyond which flame front starts to accelerate, increases with H2O addition for syngas mixtures with higher CO/H2 ratios of 65/35 and 50/50, while decreases for syngas mixtures with lower CO/H2 ratios of 20/80 and 0/100. Critical Peclet number decreases with H2O addition for all of the mixtures. The acceleration exponent increases with the flame radius and reaches a constant value at some points. This implies that both self-acceleration and self...

The intrinsic instability of laminar flame is one of the classic subjects in flame dynamics, which presents the dynamic response of laminar premixed flamelets to weak perturbations. The instabilities of laminar premixed CH4/H2/air flames stabilized on a non-adiabatic McKenna burner were investigated. The quantitative parameters of flame front were measured by OH-PLIF (Planar Laser Induced Fluorescence) technique. Combining theoretical analysis, a non-adiabatic linear dispersion relationship was derived by introducing the effect of heat loss and buoyance. The controlling factors on the cellular scale were further investigated. Heat loss was found to be the dominant factor in determining the cellular scale. High rate of heat loss can result in small cellular scale. On the other hand, the properties of mixtures, Lewis number Le and Markstein number Ma, also played an important role. If Le is more away from unit (Le < 1), the thermal-diffusive instability can be enhanced and the cells tend to exhibit a large scale. While with a larger (smaller) Ma, the stretch-induced counteract effect is stronger (weaker), which results in a less (more) irregular flame front and minor (larger) cellular scale. These three main factors were coupled to determine the final structure of cellular flame.

Cellular instability of non-adiabatic laminar premixed flat methane/hydrogen oxy-fuel flames highly diluted with CO2 was studied systematically. The CH4/H2/CO2/O2 flames were stabilized on a McKenna burner at atmospheric pressure. The CH4/H2/CO2/O2 flames with varied equivalence ratios and hydrogen mole fractions were investigated. Quantitative structure parameters of the cellular flame fronts were obtained with OH-PLIF technique. On the basis of experimental results, theoretical analysis was made to clarify the effect of mixture properties and heat loss on flame cellular instability. Results showed that cellular flame structure appeared near the lean limit combustion condition. Cell number of flame and its hydrodynamic factor, ωDL, present a positive correlation. The average stand-off distance, L, is well identified to characterize the cellular flames. Average cell amplitude, A, and wavelength, λ, increased with the increase of L. A good indicator of instability level, A·λ, was defined. The RS model was used in a linear stability analysis, and the onset of cellular flames was found to be governed by both hydrodynamic and thermal-diffusive instability mechanisms, while the heat loss made the flame more stable. The RS model is not an appropriate model to predict critical wavenumber for the non-adiabatic flame.

Context Almost 50% of type 2 diabetic (T2D) patients are poorly controlled [glycated hemoglobin (HbA1c) ≥ 7%]; however, the mechanisms responsible for progressively worsening glycemic control are poorly understood. Lower skeletal muscle mitochondrial respiratory capacity is associated with low insulin sensitivity and the development of T2D. Objective We investigated if skeletal muscle insulin sensitivity (SI) was different between well-controlled T2D (WCD) and poorly controlled T2D (PCD) and if the difference was associated with differences resulting from mitochondrial respiratory function. Design Vastus lateralis muscle mitochondrial respiration, mitochondrial content, mitochondrial enzyme activity, and fatty acid oxidation (FAO) were measured. SI and the acute response to glucose (AIRg) were calculated by MINMOD analysis from glucose and insulin obtained during a modified, frequently sampled, intravenous glucose tolerance test. Results SI and AIRg were lower in PCD than WCD. Muscle incomplete FAO was greater in PCD than WCD and greater incomplete FAO was associated with lower SI and higher HbA1c. Hydroxyacyl-coenzyme A dehydrogenase expression and activity were greater in PCD than WCD. There was no difference in maximal mitochondrial respiration or content between WCD and PCD. Conclusion The current results suggest that greater skeletal muscle incomplete FAO in poorly controlled T2D is due to elevated β oxidation and is associated with worsening muscle SI.

Myogenic regulatory factors (MRFs) including Myf5, MyoD and Myog are muscle-specific transcriptional factors orchestrating myogenesis. Although MRFs are essential for myogenic commitment and differentiation, timely repression of their activity is necessary for self-renewal and maintenance of muscle stem cells (satellite cells). Here we define a novel inhibitor of MRFs: the achaete-scute homologue 2 (Ascl2). During development, Ascl2 is transiently detected in a subpopulation of Pax7+MyoD+ progenitors (myoblasts) that become Pax7+MyoD− satellite cells prior to birth, but not detectable in postnatal satellite cells. Knockout of Ascl2 in embryonic myoblasts decreases both the number of Pax7+ cells and the proportion of Pax7+MyoD− cells. Conversely, overexpression of Ascl2 inhibits the proliferation and differentiation of cultured myoblasts, and impairs regeneration of injured muscles. At the molecular level, Ascl2 competes with MRFs for binding to E-boxes in the promoters of muscle genes, without activating gene transcription. Ascl2 also forms heterodimer with classical E-proteins to sequester their transcriptional activity on MRFs. Accordingly, MyoD or Myog expression rescues myogenic differentiation despite Ascl2 overexpression. Finally, Ascl2 expression is regulated by Notch signaling, a key governor of satellite cell self-renewal. These data together demonstrate that Ascl2 inhibits myogenic differentiation by targeting MRFs, and facilitates generation of postnatal satellite cells.

Low-intensity electric fields can induce changes in cell differentiation and cytoskeletal stresses that facilitate manipulation of osteoblasts and mesenchymal stem cells; however, the application times (tens of minutes) are of the order of physiological mechanisms, which can complicate treatment consistency. Intense nanosecond pulsed electric fields (nsPEFs) can overcome these challenges by inducing similar stresses on shorter timescales while additionally inducing plasma membrane nanoporation, ion transport and intracellular structure manipulation. This paper shows that treating myoblasts and osteoblasts with five 300 ns PEFs with intensities from 1.5 to 25 kV cm −1 increased proliferation and differentiation. While nsPEFs above 5 kV cm −1 decreased myoblast population growth, 10 and 20 kV cm −1 trains increased myoblast population by approximately fivefold 48 h after exposure when all cell densities were set to the same level after exposure. Three trials of the PEF-treated osteoblasts showed that PEF trains between 2.5 and 10 kV cm −1 induced the greatest population growth compared to the control 48 h after treatment. Trains of nsPEFs between 1.5 and 5 kV cm −1 induced the most nodule formation in osteoblasts, indicating bone formation. These results demonstrate the potential utility for nsPEFs to rapidly modulate stem cells for proliferation and differentiation and motivate future experiments to optimize PEF parameters for in vivo applications.

Heat therapy has been shown to promote capillary growth in skeletal muscle and in the heart in several animal models, but the effects of this therapy on angiogenic signaling in humans are unknown. We evaluated the acute effect of lower body heating (LBH) and unilateral thigh heating (TH) on the expression of angiogenic regulators and heat shock proteins (HSPs) in healthy young individuals. Exposure to LBH ( n = 18) increased core temperature (Tc) from 36.9 ± 0.1 to 37.4 ± 0.1°C ( P < 0.01) and average leg skin temperature (Tleg) from 33.1 ± 0.1 to 39.6 ± 0.1°C ( P < 0.01), but did not alter the levels of circulating angiogenic cytokines and bone marrow-derived proangiogenic cells (CD34+CD133+). In skeletal muscle, the change in mRNA expression from baseline of vascular endothelial growth factor (VEGF), angiopoietin 2 (ANGPT2), chemokines CCL2 and CX3CL1, platelet factor-4 (PF4), and several members of the HSP family was higher 30 min after the intervention in the individuals exposed to LBH ( n = 11) compared with the control group ( n = 12). LBH also reduced the expression of transcription factor FOXO1 ( P = 0.03). Exposure to TH ( n = 14) increased Tlegfrom 32.8 ± 0.2 to 40.3 ± 0.1°C ( P < 0.05) but Tcremained unaltered (36.8 ± 0.1°C at baseline and 36.9 ± 0.1°C at 90 min). This intervention upregulated the expression of VEGF, ANGPT1, ANGPT2, CCL2, and HSPs in skeletal muscle but did not affect the levels of CX3CL1, FOXO-1, and PF4. These findings suggest that both LBH and TH increase the expression of factors associated with capillary growth in human skeletal muscle.

Indirect calorimeter is a powerful tool to monitor resting energy metabolism through the measurement of oxygen (O2) consumption and carbon dioxide (CO2) production. From the measurement of VO2 and VCO2, the respiratory exchange ratio (RER) can be calculated to assess energy fuel utilization and energy expenditure (Evan et al., 2012). Previously, indirect calorimeter has been widely used in metabolic disease research in mice to reveal the potential roles of specific genes or treatments in regulating energy metabolism (for example: Bi et al., 2014; Feng et al., 2014). Here, we described a protocol to evaluate the resting energy metabolism of C57BL/6 mice during dark and light cycles using the Oxymax Open Circuit indirect calorimeter.

Glucose-dependent insulinotropic peptide (GIP) exerts multiple biological effects via the G-protein-coupled receptor GIPR, including glucose-stimulated insulin production and secretion, cell proliferation, and antiapoptosis in pancreatic β-cells. In an obese state, the circulating level of GIP is elevated. GIPR-knockout mice are resistant to high-fat-diet-induced obesity. The rising evidence suggests a potential role of GIP in adipocyte biology and lipid metabolism. In our study, we overexpressed GIPR in 3T3-L1 CAR adipocytes and demonstrated that GIP impaired the physiological functions of adipocytes as a consequence of increased production of inflammatory cytokines and chemokines and phosphorylation of IkB kinase (IKK)-β through activation of the cAMP-PKA pathway. Activation of Jun N-terminal kinase (JNK) pathway was also observed during GIP-induced inflammatory responses in adipocytes. The inhibition of JNK blocked GIP-stimulated secretion of inflammatory cytokines and chemokines, as well as phosphorylation of IKKβ. In addition, GIP-induced inflammatory response increased basal glucose uptake but inhibited insulin-stimulated glucose uptake. Moreover, GIP-induced adipocyte inflammation impaired insulin signaling in adipocytes as demonstrated by a reduction of AKT phosphorylation. Our results suggest that GIP might be one of the stimuli attributable to obesity-induced insulin resistance via the induction of adipocyte inflammation.

Estrogen-related receptor alpha (ERRalpha) is thought to primarily regulate lipid oxidation and control the transcription of genes in the oxidative phosphorylation pathway in skeletal and cardiac muscles. However, its role in white adipose tissue is not well studied. In this study, we aimed to establish a role for ERRalpha in adipocytes by down-regulating its activity through its inverse agonist XCT-790 in differentiated 3T3-L1 adipocytes. We found that XCT-790 differentially reduced the expression of ERRalpha target genes. Specifically, XCT-790 reduced the expressions of peroxisome proliferator-activated receptor gamma co-activator-1beta (PGC-1beta), resulting in reductions of mitochondrial biogenesis, adiogenesis and lipogeneis. Through suppressing the expression of another ERRalpha target gene pyruvate dehydrogenase kinase 2 (PDK2), we found that XCT-790 not only enhanced the conversion of pyruvate to acetyl-CoA and hyper-activated the tricarboxylic acid (TCA) cycle, but also led to higher levels of mitochondrial membrane potential and reactive oxidant species (ROS) production. Additionally, XCT-790 treatment also resulted in enhanced rates of glycolysis and basal glucose uptake. Therefore, ERRalpha stands at the crossroad of glucose and fatty acid utilization and acts as a homeostatic switch to regulate the flux of TCA cycle, mitochondrial membrane potential and glycolysis to maintain a steady level of ATP production, particularly, when mitochondrial biogenesis is reduced.

In this study, the role of sucrose non-fermenting–related kinase (SNRK) in white adipocyte biology was investigated. SNRK is abundantly expressed in adipose tissue, and the expression level is decreased in obese mice. SNRK expression is repressed by inflammatory signals but increased by insulin sensitizer in cultured adipocytes. In vivo, adipose tissue SNRK expression can be decreased by lipid injection but enhanced by macrophage ablation. Knocking down SNRK in cultured adipocytes activates both JNK and IKKβ pathways as well as promotes lipolysis. Insulin-stimulated Akt phosphorylation and glucose uptake are impaired in SNRK knockdown adipocytes. Phosphoproteomic analysis with SNRK knockdown adipocytes revealed significantly decreased phosphorylation of 49 proteins by 25% or more, which are involved in various aspects of adipocyte function with a clear indication of attenuated mTORC1 signaling. Phosphorylation of 43 proteins is significantly increased by onefold or higher, among which several proteins are known to be involved in inflammatory pathways. The inflammatory responses in SNRK knockdown adipocytes can be partially attributable to defective mTORC1 signaling, since rapamycin treatment activates IKKβ and induces lipolysis in adipocytes. In summary, SNRK may act as a suppressor of adipocyte inflammation and its presence is necessary for maintaining normal adipocyte function.

The IκB kinase β (IKKβ) is a master kinase involved in obesity-related inflammation and insulin resistance through nuclear factor κB dependent and independent pathways. However, the effect of IKKβ activation in adipose tissue, the organ critical for storage of excessive energy and initiation of inflammatory responses in the context of obesity, on systemic insulin sensitivity and metabolism, has not been investigated. In our study, we found that mice overexpressing the constitutively active IKKβ in adipose tissue under the control of murine adipocyte fatty acid binding protein (aP2) promoter were protected from age-related and diet-induced body weight gains, despite increased food intake. The aP2-IKKβ SE mice have significantly reduced weights in all white adipose tissue depots and reduced triglyceride contents in adipose tissue, liver, and muscle. Despite increased systemic and tissue inflammation, aP2-IKKβ SE mice displayed decreased blood glucose levels, improved glucose, and insulin tolerance. This may be at least partially attributable to increased energy expenditure. Histological analysis revealed presence of many small adipocytes in white adipose tissue of aP2-IKKβ SE mice fed on high-fat diet. Furthermore, transgenic expression of IKKβ in adipose tissue improved high-fat diet-induced hepatosteatosis. Collectively, increased energy expenditure and reduced plasma free fatty acid levels may contribute to enhanced systemic insulin sensitivity in aP2-IKKβ SE mice. Our study demonstrates that presence of inflammation in adipose tissue before the development of obesity has beneficial effect on metabolism.